Linking CRISPR-Cas9 interference in cassava to the evolution of editing-resistant geminiviruses.
Geminiviruses cause damaging diseases in several important crop species. However, limited progress has been made in developing crop varieties resistant to these highly diverse DNA viruses. Recently, the bacterial CRISPR/Cas9 system has been transferred to plants to target and confer immunity to geminiviruses. In this study, we use CRISPR-Cas9 interference in the staple food crop cassava with the aim of engineering resistance to African cassava mosaic virus, a member of a widespread and important family (Geminiviridae) of plant-pathogenic DNA viruses. Our results show that the CRISPR system fails to confer effective resistance to the virus during glasshouse inoculations. Further, we find that between 33 and 48% of edited virus genomes evolve a conserved single-nucleotide mutation that confers resistance to CRISPR-Cas9 cleavage. We also find that in the model plant Nicotiana benthamiana the replication of the novel, mutant virus is dependent on the presence of the wild-type virus. Our study highlights the risks associated with CRISPR-Cas9 virus immunity in eukaryotes given that the mutagenic nature of the system generates viral escapes in a short time period. Our in-depth analysis of virus populations also represents a template for future studies analyzing virus escape from anti-viral CRISPR transgenics. This is especially important for informing regulation of such actively mutagenic applications of CRISPR-Cas9 technology in agriculture.


[Alcohol and adolescents: study to implement municipal policies].
To build students' profile regarding alcohol consumption and risk behavior. A total of 1,990 students were included in the study, aged 11 to 21 years old, from both genders, enrolled in public and private schools, in Southeastern Brazil, in 2004. A self-administered questionnaire was answered in the classroom without the presence of the teacher. The questionnaire also assessed the perception of how easy it was to get alcoholic beverages, the contexts where they drunk, and the consequences of drinking. Prevalence of lifetime alcohol use was 62.2%. Regarding consumption in the last 30 days, 17.3% of students reported at least one episode of binge drinking (five or more drinks). Adolescents reported that they had gotten alcoholic drinks very easily from shops, and also in social contexts with relatives and friends. Only 1% of underaged reported that they had tried and could not buy alcoholic beverages. As negative consequences of alcohol use in the last 12 months, students reported feeling sick due to drinking (17.9%), regret for doing something under the influence of alcohol (11%), blackouts (9.8%), and getting involved in a fight after drinking (5%). Over half of the students (55%) reported knowing someone who had been involved in a car accident because of a drunk driver. Data showed high prevalence of alcohol use among adolescents studied and how easy access to alcoholic beverages is, including to underaged people. Youngsters put themselves at risk and presented negative consequences of alcohol consumption. Prompt actions regarding public alcohol policies in Brazil are needed.


Molecular systematics of armadillos (Xenarthra, Dasypodidae): contribution of maximum likelihood and Bayesian analyses of mitochondrial and nuclear genes.
The 30 living species of armadillos, anteaters, and sloths (Mammalia: Xenarthra) represent one of the three major clades of placentals. Armadillos (Cingulata: Dasypodidae) are the earliest and most speciose xenarthran lineage with 21 described species. The question of their tricky phylogeny was here studied by adding two mitochondrial genes (NADH dehydrogenase subunit 1 [ND1] and 12S ribosomal RNA [12S rRNA]) to the three protein-coding nuclear genes (alpha2B adrenergic receptor [ADRA2B], breast cancer susceptibility exon 11 [BRCA1], and von Willebrand factor exon 28 [VWF]) yielding a total of 6869 aligned nucleotide sites for thirteen xenarthran species. The two mitochondrial genes were characterized by marked excesses of transitions over transversions-with a strong bias toward CT transitions for the 12S rRNA-and exhibited two- to fivefold faster evolutionary rates than the fastest nuclear gene (ADRA2B). Maximum likelihood and Bayesian phylogenetic analyses supported the monophyly of Dasypodinae, Tolypeutinae, and Euphractinae, with the latter two armadillo subfamilies strongly clustering together. Conflicting branching points between individual genes involved relationships within the subfamilies Tolypeutinae and Euphractinae. Owing to a greater number of informative sites, the overall concatenation favored the mitochondrial topology with the classical grouping of Cabassous and Priodontes within Tolypeutinae, and a close relationship between Euphractus and Chaetophractus within Euphractinae. However, low statistical support values associated with almost equal distributions of apomorphies among alternatives suggested that two parallel events of rapid speciation occurred within these two armadillo subfamilies.


Self-assembly of random co-polymers for selective binding and detection of peptides.
Amphiphilic random co-polymers, which form stable reverse micelle-type assemblies, have been designed and synthesized. We demonstrate that the reverse micelles, formed by these co-polymers are capable of selectively binding peptides through electrostatic interactions, indicating that these random polymers can self-organize into functionally selective materials. Moreover, these random co-polymers also enable the ordered co-crystallization of matrix and extracted guest molecules, giving rise to substantial signal enhancements during MALDI-MS detection. Together, these observations represent an excellent example of how random polymers can self-assemble into ordered, functional materials.


Abrogation of transforming growth factor-alpha/epidermal growth factor receptor autocrine signaling by an RXR-selective retinoid (LGD1069, Targretin) in head and neck cancer cell lines.
Clinical studies have demonstrated that retinoids, including retinol (Vitamin A) and its synthetic derivatives, can eradicate leukoplakia and suppress the formation of squamous cell carcinoma of the head and neck (SCCHN). Nonselective retinoids have been shown to abrogate transcriptional activation of transforming growth factor-alpha (TGF-alpha) and epidermal growth factor receptor (EGFR), which characterize SCCHN. LGD1069 (Targretin) is a potent RXR-selective retinoic acid agonist with a reduced toxicity profile compared with other nonselective retinoids. We examined the effect of LGD1069 (10 microm) on cellular proliferation and expression of putative intermediate biomarker genes including TGF-alpha, EGFR, and RAR-beta in seven SCCHN cell lines. A quantitative reverse transcription-PCR assay using a novel "primer dropping" method was used to determine expression levels of EGFR, TGF-alpha, and RAR-beta before and after treatment with LGD1069 (10 microM). SCCHN proliferation was reduced by a mean of 50% at 4 days in seven SCCHN cell lines after LGD1069 treatment (P < or = 0.05). EGFR expression levels were decreased by a mean of 58.4% (P = 0.007), TGF-alpha levels were decreased by a mean of 28.8% (P = 0.01), and RAR-beta levels were increased by a mean of 60% (P = 0.03). TGF-alpha stimulation of EGFR is associated with constitutive signal transducer and activator of transcription 3 (Stat3) activation in SCCHN. Abrogation of constitutive Stat3 activation was seen with LGD1069 treatment. These results suggest that an RXR-selective retinoic acid decreases SCCHN proliferation in part by interfering with TGF-alpha/EGFR autocrine signaling.


Population-Based Surveillance of Neisseria meningitidis Antimicrobial Resistance in the United States.
Background. Antimicrobial treatment and chemoprophylaxis of patients and their close contacts is critical to reduce the morbidity and mortality and prevent secondary cases of meningococcal disease. Through the 1990's, the prevalence of antimicrobial resistance to commonly used antimicrobials among Neisseria meningitidis was low in the United States. Susceptibility testing was performed to ascertain whether the proportions of isolates with reduced susceptibility to antimicrobials commonly used for N meningitidis have increased since 2004 in the United States. Methods. Antimicrobial susceptibility testing was performed by broth microdilution on 466 isolates of N meningitidis collected in 2004, 2008, 2010, and 2011 from an active, population-based surveillance system for susceptibility to ceftriaxone, ciprofloxacin, penicillin G, rifampin, and azithromycin. The molecular mechanism of reduced susceptibility was investigated for isolates with intermediate or resistant phenotypes. Results. All isolates were susceptible to ceftriaxone and azithromycin, 10.3% were penicillin G intermediate (range, 8% in 2008-16.7% in 2010), and <1% were ciprofloxacin, rifampin, or penicillin G resistant. Of the penicillin G intermediate or resistant isolates, 63% contained mutations in the penA gene associated with reduced susceptibility to penicillin G. All ciprofloxacin-resistant isolates contained mutations in the gyrA gene associated with reduced susceptibility. Conclusions. Resistance of N meningitidis to antimicrobials used for empirical treatment of meningitis in the United States has not been detected, and resistance to penicillin G and chemoprophylaxis agents remains uncommon. Therapeutic agent recommendations remain valid. Although periodic surveillance is warranted to monitor trends in susceptibility, routine clinical testing may be of little use.


Folate Receptor-Targeted Albumin Nanoparticles Based on Microfluidic Technology to Deliver Cabazitaxel.
Microfluidic technology (MF) has improved the formulation of nanoparticles (NPs) by achieving uniform particle size distribution, controllable particle size, and consistency. Moreover, because liquid mixing can be precisely controlled in the pores of the microfluidic chip, maintaining high mixing efficiency, MF exerts higher of NP encapsulation efficiency (EE) than conventional methods. MF-NPs-cabazitaxel (CTX) particles (MF-NPs-CTX) were first prepared by encapsulating CTX according to MF. Folate (FA)- Polyethylene glycol (PEG)-NPs-CTX particles (FA-PEG-NPs-CTX) were formulated by connecting FA to MF-NPs-CTX to endow NPs with targeted delivery capability. Accordingly, the mean particle size of FA-PEG-NPs-CTX increased by approximately 25 nm, as compared with MF-NPs-CTX. Upon morphological observation of FA-PEG-NPs-CTX and MF-NPs-CTX by transmission electron microscopy (TEM), all NPs were spherical and particle size distribution was uniform. Moreover, the increased delivery efficiency of CTX in vitro and its strong tumor inhibition in vivo indicated that FA-PEG-NPs-CTX had a powerful tumor-suppressive effect both in vitro and in vivo. In vivo imaging and pharmacokinetic data confirmed that FA-PEG-NPs-CTX had good drug delivery efficiency. Taken together, FA-PEG-NPs-CTX particles prepared using MF showed high efficient and targeted drug delivery and may have a considerable driving effect on the clinical application of targeting albumin NPs.


Oropharyngeal flora and chest infection after upper abdominal surgery.
The oropharyngeal flora was determined before and after operation in 127 patients undergoing upper abdominal surgery. Swabs of the oropharynx were obtained on the day before operation and on the first, third, and fifth postoperative days. Isolation of Haemophilus influenzae, Streptococcus pneumoniae, and coliforms was noted. In the 108 patients with the full series of throat swabs the incidence of oropharyngeal colonisation by H influenzae was 16% and was unchanged after operation. S pneumoniae was present in only 5.6 (six patients) before operation and the incidence fell to 1.9% (two patients). There was a transient rise in coliform colonisation postoperatively. Twenty four patients developed a chest infection. In eight a bacterial cause was established, in six H influenzae and in two S pneumoniae. There was a significant relation between the carriage of H influenzae before operation and development of a chest infection. H influenzae was also found more often in cigarette smokers. The presence of S pneumoniae or coliform organisms before surgery was not related to the development of infection. The high incidence of postoperative chest infection in cigarette smokers appears to be due in part to preoperative colonisation of the oropharynx by H influenzae.


Enantioselective synthesis of β-amino acid derivatives via nickel-promoted regioselective carboxylation of ynamides and rhodium-catalyzed asymmetric hydrogenation.
We succeeded in the development of a new method for enantioselective synthesis of α-substituted-β-amino acid derivatives. Thus, nickel(0)-promoted carboxylation of ynamide gave the α-substituted-β-aminoacrylate derivative in a highly regioselective manner. Then, rhodium-catalyzed asymmetric hydrogenation of the α-substituted β-aminoacrylate produced the corresponding α-substituted β-amino acid derivative as an optically active form.


Strong antibacterial effect of miswak against oral microorganisms associated with periodontitis and caries.
The chewing stick (miswak) is used for oral hygiene in many parts of the world. In addition to the mechanical removal of plaque, an antibacterial effect has been postulated; however, tests of miswak extract from Salvadora persica (Arak) disclosed only low to moderate antibacterial effects. This may be attributable to the extraction process. Our aim was to test in vitro the antibacterial effect of miswak pieces, without extraction, on bacteria implicated in the etiology of periodontitis and caries. Miswak pieces were standardized by size and weight (0.07 and 0.14 g) and tested against Streptococcus mutans, Lactobacillus acidophilus, Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans), Porphyromonas gingivalis, and, as a reference, Haemophilus influenzae. The miswak pieces were tested in two ways: embedded in the agar plate or suspended above the agar plate. The inhibitory effect was most pronounced on P. gingivalis, A. actinomycetemcomitans, and H. influenzae, less on S. mutans, and least on L. acidophilus. Suspended miswak had comparable or stronger effects than miswak embedded in agar. The 0.14-g suspended miswak exhibited significantly greater inhibition on A. actinomycetemcomitans and H. influenzae than the 0.14-g miswak embedded in agar (P<0.01 and P<0.001, respectively). Miswak embedded in agar or suspended above the agar plate had strong antibacterial effects against all bacteria tested. The antibacterial effect of suspended miswak pieces suggests the presence of volatile active antibacterial compounds.


Interactions among resource partitioning, sampling effect, and facilitation on the biodiversity effect: a modeling approach.
Resource partitioning, facilitation, and sampling effect are the three mechanisms behind the biodiversity effect, which is depicted usually as the effect of plant-species richness on aboveground net primary production. These mechanisms operate simultaneously but their relative importance and interactions are difficult to unravel experimentally. Thus, niche differentiation and facilitation have been lumped together and separated from the sampling effect. Here, we propose three hypotheses about interactions among the three mechanisms and test them using a simulation model. The model simulated water movement through soil and vegetation, and net primary production mimicking the Patagonian steppe. Using the model, we created grass and shrub monocultures and mixtures, controlled root overlap and grass water-use efficiency (WUE) to simulate gradients of biodiversity, resource partitioning and facilitation. The presence of shrubs facilitated grass growth by increasing its WUE and in turn increased the sampling effect, whereas root overlap (resource partitioning) had, on average, no effect on sampling effect. Interestingly, resource partitioning and facilitation interacted so the effect of facilitation on sampling effect decreased as resource partitioning increased. Sampling effect was enhanced by the difference between the two functional groups in their efficiency in using resources. Morphological and physiological differences make one group outperform the other; once these differences were established further differences did not enhance the sampling effect. In addition, grass WUE and root overlap positively influence the biodiversity effect but showed no interactions.


In Inflamed Intestinal Tissues and Epithelial Cells, Interleukin 22 Signaling Increases Expression of H19 Long Noncoding RNA, Which Promotes Mucosal Regeneration.
Inflammation affects regeneration of the intestinal epithelia; long noncoding RNAs (lncRNAs) regulate cell functions, such as proliferation, differentiation, and migration. We investigated the mechanisms by which the lncRNA H19, imprinted maternally expressed transcript (H19) regulates regeneration of intestinal epithelium using cell cultures and mouse models of inflammation. We performed RNA-sequencing transcriptome analyses of intestinal tissues from mice with lipopolysaccharide (LPS)-induced sepsis to identify lncRNAs associated with inflammation; findings were confirmed by quantitative real-time polymerase chain reaction and in situ hybridization analyses of intestinal tissues from mice with sepsis or dextran sulfate sodium (DSS)-induced mucosal wound healing and patients with ulcerative colitis compared to healthy individuals (controls). We screened cytokines for their ability to induce expression of H19 in HT-29 cells and intestinal epithelial cells (IECs), and confirmed findings in crypt epithelial organoids derived from mouse small intestine. IECs were incubated with different signal transduction inhibitors and effects on H19 lncRNA levels were measured. We assessed intestinal epithelial proliferation or regeneration in H19ΔEx1/+ mice given LPS or DSS vs wild-type littermates (control mice). H19 was overexpressed in IECs using lentiviral vectors and cell proliferation was measured. We performed RNA antisense purification, RNA immunoprecipitation, and luciferase reporter assays to study functions of H19 in IECs. In RNA-sequencing transcriptome analysis of lncRNA expression in intestinal tissues from mice, we found that levels of H19 lncRNA changed significantly with LPS exposure. Levels of H19 lncRNA increased in intestinal tissues of patients with ulcerative colitis, mice with LPS-induced and polymicrobial sepsis, or mice with DSS-induced colitis, compared with controls. Increased H19 lncRNA localized to epithelial cells in the intestine, regardless of Lgr5 messenger RNA expression. Exposure of IECs to interleukin 22 (IL22) increased levels of H19 lncRNA with time and dose, which required STAT3 and protein kinase A activity. IL22 induced expression of H19 in mouse intestinal epithelial organoids within 6 hours. Exposure to IL22 increased growth of intestinal epithelial organoids derived from control mice, but not H19ΔEx1/+ mice. Overexpression of H19 in HT-29 cells increased their proliferation. Intestinal mucosa healed more slowly after withdrawal of DSS from H19ΔEx1/+ mice vs control mice. Crypt epithelial cells from H19ΔEx1/+ mice proliferated more slowly than those from control mice after exposure to LPS. H19 lncRNA bound to p53 and microRNAs that inhibit cell proliferation, including microRNA 34a and let-7; H19 lncRNA binding blocked their function, leading to increased expression of genes that promote regeneration of the epithelium. The level of lncRNA H19 is increased in inflamed intestinal tissues from mice and patients. The inflammatory cytokine IL22 induces expression of H19 in IECs, which is required for intestinal epithelial proliferation and mucosal healing. H19 lncRNA appears to inhibit p53 protein and microRNA 34a and let-7 to promote proliferation of IECs and epithelial regeneration.


HIV infection, malaria, and pregnancy: a prospective cohort study in Kigali, Rwanda.
In order to study the relation between human immunodeficiency virus (HIV) infection and malaria in women, during and after pregnancy, a prospective cohort study was initiated at the Centre Hospitalier de Kigali in Rwanda through routine voluntary and confidential HIV screening in antenatal clinics. At inclusion in the cohort of all HIV-positive and an equivalent number of HIV-negative pregnant women, between 21 and 28 weeks of gestation, sociodemographic characteristics and medical history during the current pregnancy were collected; screening for malaria (tick blood smear) and anemia and a CD4 lymphocyte count were systematically performed. Each woman enrolled had a monthly follow-up until 6 months after delivery. A clinic was implemented that was accessible and free of charge to every woman during the study period between scheduled visits. Malaria infection was systematically screened in case of fever or other compatible symptoms. The cohort included 228 HIV-positive and 229 HIV-negative women. At inclusion, malaria prevalence was 8.0% in HIV-positive women and 3.5% in HIV-negative women (P < 0.04). Over the study period, the incidence of malaria was 6.2 per 100 women-months in the HIV-positive group and 3.5 in the HIV-negative group (relative risk [RR] = 1.7, 95% confidence interval [CI] = 1.4-2.3). The bulk of the difference occurred postpartum. The Kaplan-Meier 9-month probability of remaining free of malaria infection was 51.8% in HIV-positive women and 65.2% in HIV-negative women (P = 0.013). When taking account in the same multivariate model (including HIV infection, primiparity, CD4 lymphocytes, anemia, and education level), positive HIV serostatus remained the only factor significantly associated with malaria infection (RR = 1.4, CI = 1.1-1.6; P = 0.016). Our study prospectively documents the association between malaria and maternal HIV infection and highlights the increased risk of malaria occurrence in all HIV-infected women. Strategies to reduce the malaria morbidity during pregnancy should be reinforced in areas of high HIV seroprevalence.


Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing.
CRISPR/Cas9 uses a guide RNA (gRNA) molecule to execute sequence-specific DNA cleavage and it has been widely used for genome editing in many organisms. Modifications at either end of the gRNAs often render Cas9/gRNA inactive. So far, production of gRNA in vivo has only been achieved by using the U6 and U3 snRNA promoters. However, the U6 and U3 promoters have major limitations such as a lack of cell specificity and unsuitability for in vitro transcription. Here, we present a versatile method for efficiently producing gRNAs both in vitro and in vivo. We design an artificial gene named RGR that, once transcribed, generates an RNA molecule with ribozyme sequences at both ends of the designed gRNA. We show that the primary transcripts of RGR undergo self-catalyzed cleavage to generate the desired gRNA, which can efficiently guide sequence-specific cleavage of DNA targets both in vitro and in yeast. RGR can be transcribed from any promoters and thus allows for cell- and tissue-specific genome editing if appropriate promoters are chosen. Detecting mutations generated by CRISPR is often achieved by enzyme digestions, which are not very compatible with high-throughput analysis. Our system allows for the use of universal primers to produce any gRNAs in vitro, which can then be used with Cas9 protein to detect mutations caused by the gRNAs/CRISPR. In conclusion, we provide a versatile method for generating targeted mutations in specific cells and tissues, and for efficiently detecting the mutations generated.


Fifty moves a year: is there an association between joint physical custody and psychosomatic problems in children?
In many Western countries, an increasing number of children with separated parents have joint physical custody, that is, live equally much in their parent's respective homes. In Sweden, joint physical custody is particularly common and concerns between 30% and 40% of the children with separated parents. It has been hypothesised that the frequent moves and lack of stability in parenting may be stressful for these children. We used data from a national classroom survey of all sixth and ninth grade students in Sweden (N=147839) to investigate the association between children's psychosomatic problems and living arrangements. Children in joint physical custody were compared with those living only or mostly with one parent and in nuclear families. We conducted sex-specific linear regression analyses for z-transformed sum scores of psychosomatic problems and adjusted for age, country of origin as well as children's satisfaction with material resources and relationships to parents. Clustering by school was accounted for by using a two-level random intercept model. Children in joint physical custody suffered from less psychosomatic problems than those living mostly or only with one parent but reported more symptoms than those in nuclear families. Satisfaction with their material resources and parent-child relationships was associated with children's psychosomatic health but could not explain the differences between children in the different living arrangements. Children with non-cohabitant parents experience more psychosomatic problems than those in nuclear families. Those in joint physical custody do however report better psychosomatic health than children living mostly or only with one parent. Longitudinal studies with information on family factors before and after the separation are needed to inform policy of children's postseparation living arrangements.


[A case of osteoclast-like giant cell tumor of the pancreas].
Osteoclast-like giant cell tumor of the pancreas is a very rare tumor. Despite their striking morphologic resemblance to certain mesenchymal tumors of bone and tendon sheath, it has been suggested that these tumors may arise from epithelial precursors. This unusual tumor presents in the 6th or 7th decade with a nearly equal gender ratio. Pure forms of osteoclast-like giant cell tumor have a better prognosis because they have a predilection to local spread, are slower to metastasize and rarely metastasize to lymph nodes, but these forms are very rare. We present an osteoclast-like giant cell tumor arising in the body of the pancreas in a 71 year-old male patient. The tumor was composed of two major cell types: atypical mononuclear cells and abundant osteoclast-like multinucleated giant cells. Immunohistochemical studies showed that atypical cells were strongly reactive for vimentin and focally reactive for cytokeratin. In contrast, the giant cells were immunoreactive for CD68, but negative for cytokeratin. Three months later, the tumor size increased and liver metastasis was newly developed. He died at 11 months after the diagnosis.


What is a clinical pathway? Refinement of an operational definition to identify clinical pathway studies for a Cochrane systematic review.
Clinical pathways (CPWs) are a common component in the quest to improve the quality of health. CPWs are used to reduce variation, improve quality of care, and maximize the outcomes for specific groups of patients. An ongoing challenge is the operationalization of a definition of CPW in healthcare. This may be attributable to both the differences in definition and a lack of conceptualization in the field of clinical pathways. This correspondence article describes a process of refinement of an operational definition for CPW research and proposes an operational definition for the future syntheses of CPWs literature. Following the approach proposed by Kinsman et al. (BMC Medicine 8(1):31, 2010) and Wieland et al. (Alternative Therapies in Health and Medicine 17(2):50, 2011), we used a four-stage process to generate a five criteria checklist for the definition of CPWs. We refined the operational definition, through consensus, merging two of the checklist's criteria, leading to a more inclusive criterion for accommodating CPW studies conducted in various healthcare settings. The following four criteria for CPW operational definition, derived from the refinement process described above, are (1) the intervention was a structured multidisciplinary plan of care; (2) the intervention was used to translate guidelines or evidence into local structures; (3) the intervention detailed the steps in a course of treatment or care in a plan, pathway, algorithm, guideline, protocol or other 'inventory of actions' (i.e. the intervention had time-frames or criteria-based progression); and (4) the intervention aimed to standardize care for a specific population. An intervention meeting all four criteria was considered to be a CPW. The development of operational definitions for complex interventions is a useful approach to appraise and synthesize evidence for policy development and quality improvement.


Induction of micronuclei in human lymphocytes exposed in vitro to microwave radiation.
Increasing applications of electromagnetic fields are of great concern with regard to public health. Several in vitro studies have been conducted to detect effects of microwave exposure on the genetic material leading to negative or questionable results. The micronucleus (MN) assay which is proved to be a useful tool for the detection of radiation exposure-induced cytogenetic damage was used in the present study to investigate the genotoxic effect of microwaves in human peripheral blood lymphocytes in vitro exposed in G(0) to electromagnetic fields with different frequencies (2.45 and 7.7GHz) and power density (10, 20 and 30mW/cm(2)) for three times (15, 30 and 60min). The results showed for both radiation frequencies an induction of micronuclei as compared to the control cultures at a power density of 30mW/cm(2) and after an exposure of 30 and 60min. Our study would indicate that microwaves are able to cause cytogenetic damage in human lymphocytes mainly for both high power density and long exposure time.


Tunable one-step double functionalization of graphene based on fluorographene chemistry.
Double functionalized graphene derivatives were synthetized by a one-pot reaction of fluorographene with organometallic nucleophiles. Their nucleophilicity governed the preference for grafting and was utilized for tuning the functionalization. This approach paves the way toward the facile, up-scalable and controllable multifunctionalization of graphene.


A study of some infectious causes of reproductive disorders in cattle owned by resource-poor farmers in Gauteng Province, South Africa.
Two hundred and thirty-nine cattle from Gauteng Province in South Africa were tested for various pathogens causing reproductive diseases includingbovine viral diarrhoea/mucosal disease (BVD/MD) virus, infectious bovine rhinotracheitis/infectious pustular vulvovaginitis (IBR/IPV) virus, Neospora caninum and Brucella abortus usingvarious tests. For BVD/MD virus, 49.37% tested positive, 74.47% for IBR/IPV virus, 8.96% for Neospora caninum and 3.8% for Brucella abortus. The result for Brucella abortus is higher than the national average, possibly due to the small sample size. A high seroprevalence of antibodies to both BVD/MD virus and IBR/IPV virus was evident. These 2 viruses should be considered, in addition to Brucella abortus, when trying to establish causes of abortion in cattle. The clinical significance of Neospora caninum as a cause of abortion in Gauteng needs further investigation. One hundred and forty-three bulls were tested for Campylobacter fetus and Trichomonas fetus, and a low prevalence of 1.4% and 2.1% respectively was found in this study. The clinical implications of these findings are discussed.


Regional dissociations within the hippocampus--memory and anxiety.
The amnestic effects of hippocampal lesions are well documented, leading to numerous memory-based theories of hippocampal function. It is debatable, however, whether any one of these theories can satisfactorily account for all the consequences of hippocampal damage: Hippocampal lesions also result in behavioural disinhibition and reduced anxiety. A growing number of studies now suggest that these diverse behavioural effects may be associated with different hippocampal subregions. There is evidence for at least two distinct functional domains, although recent neuroanatomical studies suggest this may be an underestimate. Selective lesion studies show that the hippocampus is functionally subdivided along the septotemporal axis into dorsal and ventral regions, each associated with a distinct set of behaviours. Dorsal hippocampus has a preferential role in certain forms of learning and memory, notably spatial learning, but ventral hippocampus may have a preferential role in brain processes associated with anxiety-related behaviours. The latter's role in emotional processing is also distinct from that of the amygdala, which is associated specifically with fear. Gray and McNaughton's theory can in principle incorporate these apparently distinct hippocampal functions, and provides a plausible unitary account for the multiple facets of hippocampal function.


Genetic stability of Brucella abortus isolates from an outbreak by multiple-locus variable-number tandem repeat analysis (MLVA16).
Brucellosis caused by Brucella abortus is one of the most important zoonoses in the world. Multiple-locus variable-number tandem repeat analysis (MLVA16) has been shown be a useful tool to epidemiological traceback studies in B. abortus infection. Thus, the present study aimed (i) to evaluate the genetic diversity of B. abortus isolates from a brucellosis outbreak, and (ii) to investigate the in vivo stability of the MLVA16 markers. Three-hundred and seventy-five clinical samples, including 275 vaginal swabs and 100 milk samples, were cultured from a brucellosis outbreak in a cattle herd, which adopted RB51 vaccination and test-and-slaughter policies. Thirty-seven B. abortus isolates were obtained, eight from milk and twenty-nine from post-partum/abortion vaginal swabs, which were submitted to biotyping and genotyping by MLVA16. Twelve B. abortus isolates obtained from vaginal swabs were identified as RB51. Twenty four isolates, seven obtained from milk samples and seventeen from vaginal swabs, were identified as B. abortus biovar 3, while one isolate from vaginal swabs was identified as B. abortus biovar 1. Three distinct genotypes were observed during the brucellosis outbreak: RB observed in all isolates identified as RB51; W observed in all B. abortus biovar 3 isolates; and Z observed in the single B. abortus biovar 1 isolate. Epidemiological and molecular data show that the B. abortus biovar 1 genotype Z strain is not related to the B. abortus biovar 3 genotype W isolates, and represents a new introduction B. abortus during the outbreak. The results of the present study on typing of multiple clinical B. abortus isolates from the same outbreak over a sixteen month period indicate the in vivo stability of MLVA16 markers, a low genetic diversity among B. abortus isolates and the usefulness of MLVA16 for epidemiological studies of bovine brucellosis.


The State of the Job Market for Pathologists: Evidence From the College of American Pathologists Practice Leader Survey.
Disagreement exists within the pathology community about the status of the job market for pathologists. Although many agree that jobs in pathology were harder to come by earlier this decade, recent evidence suggests improvement is occurring. To assess the state of the job market for pathologists. We analyzed data from the 2018 College of American Pathologists Practice Leader Survey. This survey contains data from 253 practice leaders on practices' hiring (and retrenchments) in 2017, the skills and level of experience being sought, success in filling those positions, and expectations for hiring in the next 3 years. Among the surveyed practice leaders, 115 (45.5%) sought to hire at least 1 pathologist in 2017, and together tried to fill 246 full-time equivalent positions that year, of which 93.5 full-time equivalents (38%) were newly created. This hiring was not limited to larger, academic-based practices, but also occurred among smaller practices and practices based in nonacademic hospitals, independent laboratories, and other settings. Although some practices retrenched (60 full-time equivalents in 2017), the net increase was a healthy 187 full-time equivalents. Practices most frequently sought pathologists who had at least 2 years of experience, but the level of experience identified with the "optimal" candidate varied by desired areas of subspecialty expertise. Practice leaders also reported expected growth in hiring, with the number of positions they hope to fill in the next 3 years exceeding those vacated by retirement. Our findings support the proposition that the demand for pathologists is strong, at least at the current time.


Truncating plakophilin-2 mutations in arrhythmogenic cardiomyopathy are associated with protein haploinsufficiency in both myocardium and epidermis.
Arrhythmogenic cardiomyopathy (AC) is a hereditary cardiac condition associated with ventricular arrhythmias, heart failure, and sudden death. The disease is most often caused by mutations in the desmosomal gene for plakophilin-2 (PKP2), which is expressed in both myocardial and epidermal tissue. This study aimed to investigate protein expression in myocardial tissue of patients with AC carrying PKP2 mutations and elucidate whether keratinocytes of the same individuals exhibited a similar pattern of protein expression. Direct sequencing of 5 AC genes in 71 unrelated patients with AC identified 10 different PKP2 mutations in 12 index patients. One patient, heterozygous for a PKP2 nonsense mutation, developed severe heart failure and underwent cardiac transplantation. Western blotting and immunohistochemistry of the explanted heart showed a significant decrease in PKP2 protein expression without detectable amounts of truncated PKP2 protein. Cultured keratinocytes of the patient showed a similar reduction in PKP2 protein expression. Nine additional PKP2 mutations were investigated in both cultured keratinocytes and endomyocardial biopsies from affected individuals. It was evident that PKP2 mutations introducing a premature termination codon in the reading frame were associated with PKP2 transcript and protein levels reduced to ≈50%, whereas a missense variant did not seem to affect the amount of PKP2 protein. The results of this study showed that truncating PKP2 mutations in AC are associated with low expression of the mutant allele and that the myocardial protein expression of PKP2 is mirrored in keratinocytes. These findings indicate that PKP2 haploinsufficiency contributes to pathogenesis in AC.


Total Hip Arthroplasty by the Direct Anterior Approach Using a Neck-preserving Stem: Safety, efficacy and learning curve.
The concept of femoral neck preservation in total hip replacement (THR) was introduced in 1993. It is postulated that retaining cortical bone of the femoral neck offers triplanar stability, uniform stress distribution, and accommodates physiological anteversion. However, data on safety, efficacy and learning curve are lacking. We prospectively assessed all patients who were operated for a THR with a short neck preserving stem (MiniHip) between 2012 and 2014. The safety and learning curve were assessed by recording operative time; stem size; and adverse events including periprosthetic fracture; paresthesia; and limb length discrepancy (LLD). The cohort was divided into equal groups to assess the learning curve effect, and the cumulative sums (CUSUM) test was performed to monitor intraoperative neck fractures. For assessment of efficacy, Oxford Hip Score (OHS) and Short Form-36 (SF-36) scores were compared preoperatively and postoperatively. 138 patients with median age 62 years (range 35-82 years) were included with a median followup of 42 months (range 30-56 months). The minimum followup was 2.5 years. The OHS, SF-36 (physical and mental component) scores improved by a mean score of 26, 28, and 27 points, respectively. All patients had LLD of <10 mm (1.9 mm ± 1.3). Adverse events included intraoperative neck fracture (n = 6), subsidence (n = 1), periprosthetic fracture (n = 1), paresthesia (n = 12), and trochanteric bursitis (n = 2). After early modification of the technique to use a smaller finishing broach, the CUSUM test demonstrated acceptable intraoperative neck fracture risk. The second surgery group had a reduced risk of intraoperative neck fracture (5/69 vs. 1/69 P = 0.2), reduced operative time (66 vs. 61 min, P = 0.06), and increased stem size (5 vs. 6, P = 0.09) although these differences were not statistically significant. The MiniHip stem is safe alternative to standard THR with good functional outcomes but with a learning curve for the surgical technique, implants sizing, and the risk of intraoperative neck fractures.


A performance comparison of anthropomorphic model observers for breast cone beam CT images: A single-slice and multislice study.
In this work, we investigate single-slice and multislice model observers which can predict human observer performance for simulated single-slice and multislice breast cone beam computed tomography (CBCT) images with a constant internal noise level. Breast background is generated based on a power spectrum of mammograms, and breast mass is modeled by a spherical signal. Human observer performance is evaluated for detecting 1 and 2 mm signals in different noise structures stemming from different reconstruction filters and image planes in a Feldkamp-Davis-Kress reconstruction. To predict human observer performance, we use single-slice channelized Hotelling observer (i.e., ssCHO) and multislice CHO (i.e., msCHOa and msCHOb) with dense difference-of-Gaussian and Gabor channels. In addition, we use single-slice nonprewhitening observer with an eye-filter (i.e., ssNPWE) and multislice NPWE (i.e., msNPWEa and msNPWEb), where ms-a model estimates the template for each image slice and ms-b model estimates the template for the central slice. For NPWE, we use the most common eye-filter with a peak value at a frequency of 4 cyc/deg. In addition, we propose an eye-filter with a peak value at a frequency of 7 cyc/deg which shows good correlation with human observer performance in single-slice breast CBCT images. Channel and decision variable internal noise are used for CHO, and decision variable internal noise is used for NPWE. The internal noise level is determined by comparing human and model observer performance for single-slice images, after which the same level is used for the multislice model observers. For single-slice images, all model observers predict human observer performance well. When the same internal noise level for the single-slice model observer is used for the multislice model observer, CHO with channel internal noise produces a higher performance than the human observer. In contrast, msCHO and msNPWEb with decision variable internal noise produce a similar performance to the human observer. Especially, ssNPWE and msNPWEb with the proposed eye-filter predict the human observer performance better than the other model observers for different noise structures. ssCHO/ssNPWE and msCHO/msNPWEb with decision variable internal noise can predict human observer performance for single-slice and multislice images with the same internal noise level. In the presence of breast anatomical background, ssNPWE and msNPWEb with the proposed eye-filter predict human observer performance better than the other model observers for different noise structures.


Optical antenna of comb-shaped split ring architecture for increased field localization in NIR and MIR.
We propose and demonstrate novel designs of optical antennas based on comb-shaped split ring architecture that display multi resonance field intensity enhancement spectrum. These nanoantennas achieve substantially increased field localization at longer wavelengths than that of a single or an array of dipoles with the same side length. With these optical antennas, localizing near infrared (NIR) and mid infrared (MIR) lights within a region of tens of nanometers at an intensity enhancement level of the order of thousands of magnitude can be accomplished.


Structure of the cytochrome b6f complex of oxygenic photosynthesis: tuning the cavity.
The cytochrome b6f complex provides the electronic connection between the photosystem I and photosystem II reaction centers of oxygenic photosynthesis and generates a transmembrane electrochemical proton gradient for adenosine triphosphate synthesis. A 3.0 angstrom crystal structure of the dimeric b6f complex from the thermophilic cyanobacterium Mastigocladus laminosus reveals a large quinone exchange cavity, stabilized by lipid, in which plastoquinone, a quinone-analog inhibitor, and a novel heme are bound. The core of the b6f complex is similar to the analogous respiratory cytochrome bc1 complex, but the domain arrangement outside the core and the complement of prosthetic groups are strikingly different. The motion of the Rieske iron-sulfur protein extrinsic domain, essential for electron transfer, must also be different in the b6f complex.


Rehabilitation of limb apraxia improves daily life activities in patients with stroke.
We randomly assigned 33 patients with left hemisphere stroke, limb apraxia, and aphasia to an apraxia or a control (aphasia) treatment group. Before and after each treatment, patients underwent a comprehensive neuropsychological testing battery and a caregiver evaluation of patient's activities of daily life (ADL) independence. Apraxia severity was related with ADL independence. Control (aphasia) treatment improved patients' language and intelligence performance. Apraxia treatment specifically improved praxic function and ADL.


Cross- and Co-Packaging of Retroviral RNAs and Their Consequences.
Retroviruses belong to the family Retroviridae and are ribonucleoprotein (RNP) particles that contain a dimeric RNA genome. Retroviral particle assembly is a complex process, and how the virus is able to recognize and specifically capture the genomic RNA (gRNA) among millions of other cellular and spliced retroviral RNAs has been the subject of extensive investigation over the last two decades. The specificity towards RNA packaging requires higher order interactions of the retroviral gRNA with the structural Gag proteins. Moreover, several retroviruses have been shown to have the ability to cross-/co-package gRNA from other retroviruses, despite little sequence homology. This review will compare the determinants of gRNA encapsidation among different retroviruses, followed by an examination of our current understanding of the interaction between diverse viral genomes and heterologous proteins, leading to their cross-/co-packaging. Retroviruses are well-known serious animal and human pathogens, and such a cross-/co-packaging phenomenon could result in the generation of novel viral variants with unknown pathogenic potential. At the same time, however, an enhanced understanding of the molecular mechanisms involved in these specific interactions makes retroviruses an attractive target for anti-viral drugs, vaccines, and vectors for human gene therapy.


The role of cooperatives in sustaining the livelihoods of rural communities: The case of rural cooperatives in Shurugwi District, Zimbabwe.
The main focus of the research was to analyse the role of cooperatives in sustaining the livelihoods of local rural communities in Shurugwi District in Zimbabwe. Descriptive survey design was used in this mixed method approach to the study. A questionnaire, interviews and observation methods were employed as the main research instruments. Purposive sampling technique was adopted and data were collected from government officials and from members of the six cooperatives in Shurugwi District. A total of 50 research participants were involved in the study. It was found that cooperatives were established as a strategy to sustain livelihoods of rural communities. With the adoption of cooperatives, people in the rural communities managed to generate employment, boost food production, empower the marginalised, especially women, and promote social cohesion and integration, thereby improving their livelihoods and reducing poverty. Most cooperatives face a number of challenges that include lack of financial support, poor management and lack of management skills, and lack of competitive markets to sell their produce. The study recommends that the government and the banking sector render financial support to cooperatives in rural communities to allow them to expand and diversify their business operations; constant training on leadership and management skills is provided to cooperatives' members. There is also a need for cooperatives, especially those in the agricultural sector, to form some producer associations so as to easily market their produce. Lastly, the study recommends that future research should focus on investigating issues that hinder the growth of the cooperative movement in rural communities of Zimbabwe. It is hoped that policy-makers, the academia and communities would benefit from the study.


Emulsifier for intravenous cyclosporin inhibits neurite outgrowth, causes deficits in rapid axonal transport and leads to structural abnormalities in differentiating N1E.115 neuroblastoma.
The emulsifier for cyclosporin in clinical i.v. formulations, Cremophor EL, has recently come into question as a possible source of neurotoxic side effects in immunosuppressant therapy. To address this issue we tested Cremophor EL and cyclosporin on an in vitro neuronal model, the differentiating N1E.115 neuroblastoma cell. In terms of effects on elaboration of neurites by these cells, Cremophor accounted for nearly all the neurotoxicity of clinically formulated cyclosporin. At a concentration of 0.005% (v/v), Cremophor EL halved the number of cells that extended neurites after 48 hr in serum-free medium. Average neurite length was also reduced substantially. Inhibition of neurite outgrowth first became apparent 24 hr after exposure to Cremophor EL. Neurites that did grow in the presence of Cremophor were disfigured by a series of regularly spaced, gross dilatations (beads) filled with large (0.2-0.5 microns) lipid vesicles. Abnormalities of rapid axonal transport were documented in the beaded neurites by means of video-enhanced contrast, differential interference-contrast microscopy. Velocity of retrograde transport remained normal, but the velocity of anterograde transport and the total bidirectional flux of organelles were both reduced. It seems likely that the inhibition of neurite outgrowth, the swellings of the neurites and the abnormalities of transport are interrelated phenomena.


Finite-size critical scaling in Ising spin glasses in the mean-field regime.
We study in Ising spin glasses the finite-size effects near the spin-glass transition in zero field and at the de Almeida-Thouless transition in a field by Monte Carlo methods and by analytical approximations. In zero field, the finite-size scaling function associated with the spin-glass susceptibility of the Sherrington-Kirkpatrick mean-field spin-glass model is of the same form as that of one-dimensional spin-glass models with power-law long-range interactions in the regime where they can be a proxy for the Edwards-Anderson short-range spin-glass model above the upper critical dimension. We also calculate a simple analytical approximation for the spin-glass susceptibility crossover function. The behavior of the spin-glass susceptibility near the de Almeida-Thouless transition line has also been studied, but here we have only been able to obtain analytically its behavior in the asymptotic limit above and below the transition. We have also simulated the one-dimensional system in a field in the non-mean-field regime to illustrate that when the Imry-Ma droplet length scale exceeds the system size one can then be erroneously lead to conclude that there is a de Almeida-Thouless transition even though it is absent.


[Treatment of pancreaticopleural fistulas].
Pancreaticopleural fistulas are rare but serious complications of chronic or recurrent inflammatory pancreatic disease. Massive unilateral or bilateral pleural effusion may occur in association with rupture of pancreatic duct or pseudocyst into the pleural cavity. In the past decade we have treated 10 patients with pancreaticopleural fistulas. Eight of them had a previous history of inflammatory pancreatic disease. Diagnosis was made by finding a markedly elevated amylase level (10 pts) in the pleural fluid. US and CT examination enabled the establishment of the diagnosis of pancreatic origin, showing chronic pancreatic (9 pts) and pancreatic pseudocyst (6 pts). Pancreaticopleural fistula was successfully demonstrated by ERCP in four patients. Initial treatment was non-operative using total parenteral or jejunal nutrition and multiple thoracocentesis or thoracic drainage. Anti-secretory octreotide therapy was used in all patients. This conservative treatment was successful in three patients (3/10). Septic complication (1 pt) and unsuccessful medical therapy (6 pts) recommended surgical intervention. Decompression procedure (4 pts) and resection (3 pts) were performed. Surgery was successful in all seven patients. We lost no patients and none of them required subsequent surgical treatment.


Are the University Hospitals in Puerto Rico following Current Guidelines for Cervical Cancer Screening?
Cervical cancer is the fifth most common cancer among women in Puerto Rico (PR). Information about which obstetricians and gynecologists in PR are following current cervical cytology guidelines is not available. Our two objectives were to determine whether current guidelines for abnormal cervical cytology management were being followed adequately by OB-GYN residents and faculty at 3 university hospitals and, in addition, to evaluate the knowledge of the current guidelines that is possessed by these individuals. A retrospective medical record review (records from January 2009 through December 2010) of patients with abnormal cervical cytology results (n=166) to evaluate the management they were given and a cross-sectional survey of OB-GYN residents (n=34) and faculty (n=46) to evaluate their knowledge of cervical cytology guidelines were both performed. One hundred and sixty-six medical records were reviewed. The level of management for 45% of the patients was found to have been optimal."Thirty-three percent were lost to follow-up. Eleven percent of the patients were managed sub optimally and the management of 12% of the patients was considered to have been poor. The survey showed that the faculty were less updated with regard to current guidelines for the management of cervical cytology than were the residents. Study limitations were the lack of electronic medical records in the hospital--which makes access to patient information more difficult--and the low response rate of the faculty. OB-GYN physicians and residents were not consistent with their management, according to current guidelines, of cervical cytological abnormalities. The high-risk nature of the patients and their poor adherence to treatment (loss to follow-up) may have been the cause of this inconsistency. Meetings aimed at addressing the lack of compliance by local OB-GYN physicians with the guidelines for cervical cancer screening have been Better screening strategies are also needed if we are to improve patient compliance in our population.


The risk of a second hip fracture in patients after their first hip fracture.
We investigated the incidence of additional fractures and the rate of prescription of osteoporotic pharmacotherapy after an initial hip fracture. We surveyed female patients aged 65 and over who sustained their first hip fracture between January 1, 2006, and December 31, 2007, treated at 25 hospitals in five geographic areas in Japan. Data for 1 year after the first hip fracture were collected from medical records, and questionnaires were mailed to all patients. In total, 2,663 patients were enrolled, and 335 patients were excluded based on exclusion criteria. The analysis was performed on 2,328 patients. During the 1-year follow-up period 160 fractures occurred in 153 patients and 77 subsequent hip fractures occurred in 77 patients. The incidence of all additional fractures among patients who sustained their first hip fracture was 70 (per 1,000 person-year) and that for second hip fracture was 34. In comparison to the general population, women ≥65 years of age who sustained an initial hip fracture were four times as likely to sustain an additional hip fracture. Antiosteoporosis pharmacotherapy was prescribed for 436 patients (18.7%), while 1,240 patients (53.3%) did not receive any treatment during the 1-year period. Patients who have sustained one hip fracture have a higher risk of a second hip fracture compared to the general population, and most of these women receive no pharmaceutical treatment for osteoporosis.


Effects on 4080 rats of chronic ingestion of N-nitrosodiethylamine or N-nitrosodimethylamine: a detailed dose-response study.
Four thousand eighty inbred rats were maintained from weaning on various different concentrations of N-nitrosodiethylamine (NDEA) or N-nitrosodimethylamine (NDMA). The principal aim was to characterize the dose-response relationship for the effects of these agents on esophageal cancer (NDEA) or on various types of liver cancer (NDEA and NDMA), although NDEA also caused a few tumors of the nasopharynx and NDMA also caused a few tumors of the lung. The numbers of tumors of mesenchymal and Kupffer cells in the liver were too few to allow easy characterization of the dose-response relationships, and although NDMA induced large numbers of bile duct neoplasms, NDEA did not. Thus, the four principal dose-response relationships studied were of NDEA on esophageal or liver cells and of NDMA on bile duct or liver cells. At doses sufficiently high for the median time to death from the disease of interest to be estimated, relationships were observed of the general form (Dose rate) x (median)n = constant where n was about 2.3 for the first three relationships and about 1 for the last one (NDMA on liver cell tumors). By contrast, at doses sufficiently low for longevity to be nearly normal (median survival about 2.5 years), there remained no material dependence on the dose rate of the age distribution of the induced neoplasms. At these low dose rates, the number of liver (but not of esophageal) neoplasms induced by treatment was simply proportional to the dose rate. This finding is not surprising, since the background incidence of liver (but not of esophageal) neoplasms was appreciable. The linear relationship observed at low dose rates (below 1 ppm) suggests that under these experimental conditions, among rats allowed to liver their natural life span, a dose of 1 ppm of NDEA or NDMA in the drinking water will cause about 25% to develop a liver neoplasm, a dose of 0.1 ppm will cause about 2.5% to do so, and a dose of 0.01 ppm will cause about 0.25% to do so, etc., with no indication of any "threshold." (At these low dose rates, the incidence of liver neoplasms appears likely to exceed greatly that of esophageal neoplasms.) In addition, even quite low dose rates of the test agents caused a variety of nonneoplastic liver abnormalities (e.g., hyperplastic nodules, or shrinkage of hepatocytes) at a frequency roughly proportional to the dose rate.


HDAC inhibitors tune miRNAs in extracellular vesicles of dystrophic muscle-resident mesenchymal cells.
We show that extracellular vesicles (EVs) released by mesenchymal cells (i.e., fibro-adipogenic progenitors-FAPs) mediate microRNA (miR) transfer to muscle stem cells (MuSCs) and that exposure of dystrophic FAPs to HDAC inhibitors (HDACis) increases the intra-EV levels of a subset of miRs, which cooperatively target biological processes of therapeutic interest, including regeneration, fibrosis, and inflammation. Increased levels of miR-206 in EVs released by FAPs of muscles from Duchenne muscular dystrophy (DMD) patients or mdx mice exposed to HDACi are associated with enhanced regeneration and decreased fibrosis. Consistently, EVs from HDACi-treated dystrophic FAPs can stimulate MuSC activation and expansion ex vivo, and promote regeneration, while inhibiting fibrosis and inflammation of dystrophic muscles, upon intramuscular transplantation in mdx mice, in vivo. AntagomiR-mediated blockade of individual miRs reveals a specific requirement of miR-206 for EV-induced expansion of MuSCs and regeneration of dystrophic muscles, and indicates that cooperative activity of HDACi-induced miRs accounts for the net biological effect of these EVs. These data point to pharmacological modulation of EV content as novel strategy for therapeutic interventions in muscular dystrophies.


von Willebrand disease in the pediatric and adolescent population.
Recent studies indicate that bleeding disorders, particularly von Willebrand disease (VWD) is more prevalent than previously thought in adolescents with menorrhagia. Menorrhagia management in undiagnosed disorders of hemostasis may be associated with unwanted risks and complications. The prevalence of symptomatic VWD in the pediatric primary care setting appears to be 0.11% (95% CI, 0.04-0.25%). Studies evaluating the prevalence of VWD in adolescents with menorrhagia have included over 500 patients with a prevalence range from 3 to 36% depending on the clinical setting studied, with the highest prevalence seen in adolescents referred to an outpatient Hemophilia Center, while the lowest prevalence is seen in the acute hospital setting. Recently, the diagnosis of VWD has been facilitated by the use of pediatric bleeding questionnaires that have proved useful in quantifying the severity of bleeding symptoms. Treatment of VWD is often complex because a combination of therapies is often required. Potential treatment options include estrogen-progesterone preparations, desmopressin, antifibrinolytic agents and von Willebrand factor concentrates. More research is needed to evaluate the effectiveness of the various treatment modalities in the adolescent population.


Achalasia combined with esophageal cancer treated by concurrent chemoradiation therapy.
Achalasia is a rare neurological deficit of the esophagus that produces an impaired relaxation of the lower esophageal sphincter and decreased motility of the esophageal body. Achalasia is generally accepted to be a pre-malignant disorder, since, particularly in the mega-esophagus, chronic irritation by foods and bacterial overgrowth may contribute to the development of dysplasia and carcinoma. We present a case of a 51-year-old man with achalasia combined with esophageal cancer who has had dysphagia symptoms for more than 20 years. Since there was a clinically high possibility of supraclavicular lymph node metastasis, concurrent chemoradiation therapy was scheduled. After the third cycle of chemoradiation therapy, transthoracic esophageolymphadenectomy was performed. Histopathological examination of the main esophagus specimen revealed no residual carcinoma. And the entire regional lymph node areas were free of carcinoma except for one azygos metastatic lymph node. In summary, achalasia is a predisposing factor for esophageal squamous cell carcinoma. Although surveillance endoscopy in achalasia patients is still controversial, periodic screening for cancer development in long-standing achalasia patients might be advisable.


Comparison of 2 different regimens for reactogenicity, safety, and immunogenicity of the live attenuated oral rotavirus vaccine RIX4414 coadministered with oral polio vaccine in South African infants.
A phase II, randomized, double-blind, placebo-controlled study was conducted in South Africa during 2003-2004 to evaluate the safety, reactogenicity, and immunogenicity of 2 regimens of the live attenuated oral human rotavirus vaccine RIX4414 when coadministered with the Expanded Program on Immunization childhood vaccines, including oral polio vaccine. Healthy infants were randomized (2:2:1) to receive either 2 doses of RIX4414 (n = 190; at 10 and 14 weeks, with placebo at 6 weeks), 3 doses of RIX4414 (n = 189; at 6, 10, and 14 weeks), or 3 doses of placebo (n = 96), all with concomitant routine vaccinations. The antirotavirus IgA seroconversion rate was assessed using enzyme-linked immunosorbent assay at 2 months after the last dose of RIX4414 or placebo. Antipolio types 1, 2, and 3 antibodies were measured using a virus neutralization assay. Solicited symptoms were recorded for 15 days after each dose. The antirotavirus IgA seroconversion rates were similar in the RIX4414 2- and 3-dose groups (44.3% and 44.4%, respectively; P = .544, by 1-sided Fisher exact test) and antirotavirus IgA geometric mean concentrations were also comparable. Seroprotection rates for antipolio types 1, 2, and 3 antibodies were high (93%-100%) and were not significantly different among groups. Solicited symptoms reported within 15 days after vaccination were similar in all groups. The immune seroconversion response to the RIX4414 vaccine with 3 doses was not superior to the 2-dose regimen. There was no interference by either regimen with antibody response to oral polio vaccine, and RIX4414 was well tolerated when given with routine vaccinations.


Affinity of aporphines for the human 5-HT2A receptor: insights from homology modeling and molecular docking studies.
Analogs of nantenine were docked into a modeled structure of the human 5-HT(2A) receptor using ICM Pro, GLIDE, and GOLD docking methods. The resultant docking scores were used to correlate with observed in vitro apparent affinity (K(e)) data. The GOLD docking algorithm when used with a homology model of 5-HT(2A), based on a bovine rhodopsin template and built by the program MODELLER, gives results which are most in agreement with the in vitro results. Further analysis of the docking poses among members of a C1 alkyl series of nantenine analogs, indicate that they bind to the receptor in a similar orientation, but differently than nantenine. Besides an important interaction between the protonated nitrogen of the C1 alkyl analogs and residue Asp155, we identified Ser242, Phe234, and Gly238 as key residues responsible for the affinity of these compounds for the 5-HT(2A) receptor. Specifically, the ability of some of these analogs to establish a H-bond with Ser242 and hydrophobic interactions with Phe234 and Gly238 appears to explain their enhanced affinity as compared to nantenine.


An instrument to evaluate the process of psychiatric care in ambulatory settings.
As part of an evaluation of the Utah Prepaid Mental Health Plan, the Process of Care Review Form was developed to assess the quality of the process of psychiatric care provided by Utah's community mental health centers (CMHCs) to clients with schizophrenia. This article briefly reviews issues in the measurement of quality of care and describes the development and implementation of the form. The 67-item form was designed for use by a trained abstracter to gather data from CMHC medical records in six areas: general management of the patient, medication management, medical management, social support, psychiatric hospitalization, and psychiatric assessment. A 59-item version of the form that omits the section on psychiatric assessment has been used in three waves of data collection to document data spanning five years (1990-1994) in the evaluation of the process of psychiatric care in the Utah plan. It is currently being used longitudinally to examine differences between Utah CMHCs receiving capitated payments and those paid on a fee-for-service basis by Medicaid.


A 'joint venture' model of recontacting in clinical genomics: challenges for responsible implementation.
Advances in genomics often lead healthcare professionals (HCPs) to learn new information, e.g., about reinterpreted variants that could have clinical significance for patients seen previously. A question arises of whether HCPs should recontact these former patients. We present some findings interrogating the views of patients (or parents of patients) with a rare or undiagnosed condition about how such recontacting might be organised ethically and practically. Forty-one interviews were analysed thematically. Participants suggested a 'joint venture' model in which efforts to recontact are shared with HCPs. Some proposed an ICT-approach involving an electronic health record that automatically alerts them to potentially relevant updates. The need for rigorous privacy controls and transparency about who could access their data was emphasised. Importantly, these findings highlight that the lack of clarity about recontacting is a symptom of a wider problem: the lack of necessary infrastructure to pool genomic data responsibly, to aggregate it with other health data, and to enable patients/parents to receive updates. We hope that our findings will instigate a debate about the way responsibilities for recontacting under any joint venture model could be allocated, as well as the limitations and normative implications of using ICT as a solution to this intractable problem. As a first step to delineating responsibilities in the clinical setting, we suggest HCPs should routinely discuss recontacting with patients/parents, including the new information that should trigger a HCP to initiate recontact, as part of the consent process for genetic testing.


[Is the tuberculin skin test still suitable to diagnose tuberculosis infection ?].
Tuberculosis (TB) infection is currently being diagnosed by the tuberculin skin test (TST) with PPD. Some Mycobacterium tuberculosis PPD components are present in BCG, which can be the cause of false positive TST results in BCG vaccinated persons. New IFN-g release assays (IGRAs) are based on the ex vivo release of IFN-g by peripheral blood cells in presence of M. tuberculosis antigens ESAT-6 and CFP-10, which should be absent in BCG. These assays consist in either quantifying released IFN-g or enumerating IFN-g producing cells. In principle, IGRAs should differentiate true TB infection from vaccination and results of several studies suggest that these assays display lower positivity than TST. Whether the lower positivity could be attributed to higher specificity or to lower sensitivity as compared with PPD is still unclear. BCG vaccination, if not recently applied, cannot be blamed for false positive TST reactions. Strong TST reactions (> or = 10 mm or > or =15 mm) are highly correlated with TB infection. In settings where TB continues being a serious health problem, cost-effectiveness evaluations would privilege TST under certain conditions: supply of quality-assured PPD reagent, standardized criteria for TST application, reading and interpretation, and regular availability in health centers countrywide. In view of current limitations in the supply of imported PPD in Argentina, its production/quality assurance should be considered a public health priority. Still, key questions remain to be addressed concerning the role of IGRAs and TST in predicting risk of TB disease, in other words, in identifying persons who will benefit most from chemoprophylaxis.


Fast protein classification with multiple networks.
Support vector machines (SVMs) have been successfully used to classify proteins into functional categories. Recently, to integrate multiple data sources, a semidefinite programming (SDP) based SVM method was introduced. In SDP/SVM, multiple kernel matrices corresponding to each of data sources are combined with weights obtained by solving an SDP. However, when trying to apply SDP/SVM to large problems, the computational cost can become prohibitive, since both converting the data to a kernel matrix for the SVM and solving the SDP are time and memory demanding. Another application-specific drawback arises when some of the data sources are protein networks. A common method of converting the network to a kernel matrix is the diffusion kernel method, which has time complexity of O(n(3)), and produces a dense matrix of size n x n. We propose an efficient method of protein classification using multiple protein networks. Available protein networks, such as a physical interaction network or a metabolic network, can be directly incorporated. Vectorial data can also be incorporated after conversion into a network by means of neighbor point connection. Similar to the SDP/SVM method, the combination weights are obtained by convex optimization. Due to the sparsity of network edges, the computation time is nearly linear in the number of edges of the combined network. Additionally, the combination weights provide information useful for discarding noisy or irrelevant networks. Experiments on function prediction of 3588 yeast proteins show promising results: the computation time is enormously reduced, while the accuracy is still comparable to the SDP/SVM method. Software and data will be available on request.


Impact of early life adversity on reward processing in young adults: EEG-fMRI results from a prospective study over 25 years.
Several lines of evidence have implicated the mesolimbic dopamine reward pathway in altered brain function resulting from exposure to early adversity. The present study examined the impact of early life adversity on different stages of neuronal reward processing later in life and their association with a related behavioral phenotype, i.e. attention deficit/hyperactivity disorder (ADHD). 162 healthy young adults (mean age = 24.4 years; 58% female) from an epidemiological cohort study followed since birth participated in a simultaneous EEG-fMRI study using a monetary incentive delay task. Early life adversity according to an early family adversity index (EFA) and lifetime ADHD symptoms were assessed using standardized parent interviews conducted at the offspring's age of 3 months and between 2 and 15 years, respectively. fMRI region-of-interest analysis revealed a significant effect of EFA during reward anticipation in reward-related areas (i.e. ventral striatum, putamen, thalamus), indicating decreased activation when EFA increased. EEG analysis demonstrated a similar effect for the contingent negative variation (CNV), with the CNV decreasing with the level of EFA. In contrast, during reward delivery, activation of the bilateral insula, right pallidum and bilateral putamen increased with EFA. There was a significant association of lifetime ADHD symptoms with lower activation in the left ventral striatum during reward anticipation and higher activation in the right insula during reward delivery. The present findings indicate a differential long-term impact of early life adversity on reward processing, implicating hyporesponsiveness during reward anticipation and hyperresponsiveness when receiving a reward. Moreover, a similar activation pattern related to lifetime ADHD suggests that the impact of early life stress on ADHD may possibly be mediated by a dysfunctional reward pathway.


Therapeutic antibodies for the treatment of pancreatic cancer.
Pancreatic cancer is a devastating disease with the worst mortality rate and an overall 5-year survival rate lower than 5%. In the U.S., this disease is the fourth leading cause of death and represents 6% of all cancer-related deaths. Gemcitabine, the current standard first-line treatment, offers marginal benefits to patients in terms of symptom control and prolongation of life. Since 1996, about 20 randomized phase III trials have been performed to improve the efficacy of gemcitabine, with little success regarding a significant improvement in survival outcomes. The need for novel therapeutic strategies, such as target therapy, is obvious. Monoclonal antibodies have finally come of age as therapeutics and several molecules are now approved for cancer therapies. This review aims to give a general view on the clinical results obtained so far by antibodies for the treatment of pancreatic cancer and describes the most promising avenues toward a significant improvement in the treatment of this frustrating disease.


Evaluation of environmental, nutritional, and host factors in cats with hyperthyroidism.
The pathologic changes associated with hyperthyroidism (adenomatous hyperplasia, adenoma of the thyroid gland) have been well characterized in cats, but the pathogenesis of these changes remains unclear. In this research, we undertook a case-control study to search for potential risk factors for this disease. Owners of 379 hyperthyroid and 351 control cats were questioned about their cats' exposure to potential risk factors including breed, demographic factors, medical history, indoor environment, chemicals applied to the cat and environment, and diet. The association between these hypothesized risk factors and outcome of disease was evaluated by conditional logistic regression. Two genetically related cat breeds (ie, Siamese and Himalayan) were found to have diminished risk of developing hyperthyroidism. Cats that used litter had higher risk of developing hyperthyroidism than those that did not. Use of topical ectoparasite preparations was associated with increased risk of developing hyperthyroidism. Compared with cats that did not eat canned food, those that ate commercially prepared canned food had an approximate 2-fold increase in risk of disease. When these 4 variables (breed, use of cat litter, consumption of canned cat food, and use of topical ectoparasite preparations) from the univariate analysis were selected for further study as candidate risk factors and analyzed by multivariate conditional logistic regression, a persistent protective effect of breed (ie, Siamese or Himalayan) was found. In addition, results suggested a 2- to 3-fold increase in risk of developing hyperthyroidism among cats eating a diet composed mostly of canned cat food and a 3-fold increase in risk among those using cat litter. In contrast, the use of commercial flea products did not retain a strong association. The results of this study indicate that further research into dietary and other potentially important environmental factors (eg, cat litter) is warranted.


Effect of patient socioeconomic status on access to early-phase cancer trials.
Little is known about the influence of socioeconomic factors on patient access to cancer trials. Differences should be considered to ensure generalizability of trial results and equality of access. Phase I trials unit referrals at our center over 5 years, from 2007 to 2012, were reviewed. Socioeconomic status was defined by the Index of Multiple Deprivation (IMD; 1, least deprived; 5, most deprived). Multivariate analysis was performed comparing incident cancer cases with referred patients and those ultimately enrolled onto a trial. Four hundred thirty patients were referred (median age, 62 years). Compared with 10,784 incident cases, referral was less likely for patients in the more-deprived quintiles compared with the least deprived (IMD 5: odds ratio [OR], 0.53; 95% CI, 0.38 to 0.74). Once reviewed in the unit, enrollment onto a trial was not affected (IMD 5: OR, 0.81; 95% CI, 0.40 to 1.63). Ethnicity analysis showed the nonwhite population was less likely to be recruited (OR, 0.48; 95% CI, 0.26 to 0.88). This relationship was lost with adjustment for age, sex, cancer type, and deprivation index. We show for the first time to our knowledge that socioeconomic status affects early-phase cancer trial referrals. The least-deprived patients are almost twice as likely to be referred compared with the most deprived. This may be because more-deprived patients are less suitable for a trial-as a result of comorbidities, for example-or because of inequalities that could be addressed by patient or referrer education. Once reviewed at the unit, enrollment onto a trial is not affected by deprivation.


Efficient cross-effect dynamic nuclear polarization without depolarization in high-resolution MAS NMR.
Dynamic nuclear polarization (DNP) has the potential to enhance the sensitivity of magic-angle spinning (MAS) NMR by many orders of magnitude and therefore to revolutionize atomic resolution structural analysis. Currently, the most widely used approach to DNP for studies of chemical, material, and biological systems involves the cross-effect (CE) mechanism, which relies on biradicals as polarizing agents. However, at high magnetic fields (≥5 T), the best biradicals used for CE MAS-DNP are still far from optimal, primarily because of the nuclear depolarization effects they induce. In the presence of bisnitroxide biradicals, magic-angle rotation results in a reverse CE that can deplete the initial proton Boltzmann polarization by more than a factor of 2. In this paper we show that these depolarization losses can be avoided by using a polarizing agent composed of a narrow-line trityl radical tethered to a broad-line TEMPO. Consequently, we show that a biocompatible trityl-nitroxide biradical, TEMTriPol-1, provides the highest MAS NMR sensitivity at ≥10 T, and its relative efficiency increases with the magnetic field strength. We use numerical simulations to explain the absence of depolarization for TEMTriPol-1 and its high efficiency, paving the way for the next generation of polarizing agents for DNP. We demonstrate the superior sensitivity enhancement using TEMTriPol-1 by recording the first solid-state 2D 13C-13C correlation spectrum at natural isotopic abundance at a magnetic field of 18.8 T.


Histomorphological analysis of gastric polyps.
Incidence of gastric carcinoma and gastric polyps is on rise all over the world. Chronic atrophic gastritis to intestinal metaplasia progressing to adenocarcinoma has been documented pathway for gastric carcinogenesis. Another pathway for gastric carcinoma is adenoma carcinoma sequence similar to colon cancer. To study prevalence, endoscopic, and histomorphological features of gastric polyps. This was retrospective analysis of gastric polyps from 2012 to 2019 in consecutive 10,800 upper gastrointestinal endoscopies. Demographic, endoscopic, and histopathological data were obtained from hospital records. All gastric polyps were classified as per standard histologic criteria. Additional histological features noted were presence of dysplasia, focus of adenoma, or malignancy. The prevalence of gastric polyps was 434 (4%) of 10,800 upper gastrointestinal endoscopies. Majority of polyps were found in the last 4 years (277: 63.8%). Mean age was 55.4 years with male to female ratio 1:1.2. Most of the polyps (94.9%) were less than 1 cm, located in gastric antrum. Multiple polyps were seen in 20.9% cases. On histopathology, fundic gland polyps were most common (147: 33.8%), followed by hyperplastic (128: 29.4%) polyps. Adenomatous polyps were nine (2%); of these, two cases of hyperplastic polyps and one each of fundic gland polyp and benign epithelial polyp showed adenomatous foci. Fundic gland polyps were the most common polyps. With rising incidence of gastric carcinoma, identification of gastric polyps on endoscopy with biopsy can prevent progression to carcinogenesis.


Effects of Polar Bear and Killer Whale Derived Contaminant Cocktails on Marine Mammal Immunity.
Most controlled toxicity studies use single chemical exposures that do not represent the real world situation of complex mixtures of known and unknown natural and anthropogenic substances. In the present study, complex contaminant cocktails derived from the blubber of polar bears (PB; Ursus maritimus) and killer whales (KW; Orcinus orca) were used for in vitro concentration-response experiments with PB, cetacean and seal spp. immune cells to evaluate the effect of realistic contaminant mixtures on various immune functions. Cytotoxic effects of the PB cocktail occurred at lower concentrations than the KW cocktail (1 vs 16 μg/mL), likely due to differences in contaminant profiles in the mixtures derived from the adipose of each species. Similarly, significant reduction of lymphocyte proliferation occurred at much lower exposures in the PB cocktail (EC50: 0.94 vs 6.06 μg/mL; P < 0.01), whereas the KW cocktail caused a much faster decline in proliferation (slope: 2.9 vs 1.7; P = 0.04). Only the KW cocktail modulated natural killer (NK) cell activity and neutrophil and monocyte phagocytosis in a concentration- and species-dependent manner. No clear sensitivity differences emerged when comparing cetaceans, seals and PB. Our results showing lower effect levels for complex mixtures relative to single compounds suggest that previous risk assessments underestimate the effects of real world contaminant exposure on immunity. Our results using blubber-derived contaminant cocktails add realism to in vitro exposure experiments and confirm the immunotoxic risk marine mammals face from exposure to complex mixtures of environmental contaminants.


Caffeine for the management of apnea in preterm infants.
Considerable uncertainty persists regarding the efficacy and safety of methylxanthines (caffeine, theophylline - in intravenous form named aminophylline) for the prevention and treatment of infant apnea. To help inform national guideline development in Kenya we undertook structured literature searches to identify current evidence on caffeine therapy for infant apnea. Available evidence shows that caffeine is as effective as intravenous theophylline (aminophylline), but is safer and easier to give and has better therapeutic properties. It is therefore recommended for the treatment of apnea of prematurity. Caffeine is also the preferred drug if clinicians plan to provide apnea prophylaxis. As prematurity is likely to result in more than 1 million deaths a year, mostly in resource-poor settings, greater efforts need to be made to ensure interventions such as caffeine, currently unavailable in countries such as Kenya, are made more widely available.


Endoplasmic reticulum (ER) Ca2+-channel activity contributes to ER stress and cone death in cyclic nucleotide-gated channel deficiency.
Endoplasmic reticulum (ER) stress and mislocalization of improperly folded proteins have been shown to contribute to photoreceptor death in models of inherited retinal degenerative diseases. In particular, mice with cone cyclic nucleotide-gated (CNG) channel deficiency, a model for achromatopsia, display both early-onset ER stress and opsin mistrafficking. By 2 weeks of age, these mice show elevated signaling from all three arms of the ER-stress pathway, and by 1 month, cone opsin is improperly distributed away from its normal outer segment location to other retinal layers. This work investigated the role of Ca2+-release channels in ER stress, protein mislocalization, and cone death in a mouse model of CNG-channel deficiency. We examined whether preservation of luminal Ca2+ stores through pharmacological and genetic suppression of ER Ca2+ efflux protects cones by attenuating ER stress. We demonstrated that the inhibition of ER Ca2+-efflux channels reduced all three arms of ER-stress signaling while improving opsin trafficking to cone outer segments and decreasing cone death by 20-35%. Cone-specific gene deletion of the inositol-1,4,5-trisphosphate receptor type I (IP3R1) also significantly increased cone density in the CNG-channel-deficient mice, suggesting that IP3R1 signaling contributes to Ca2+ homeostasis and cone survival. Consistent with the important contribution of organellar Ca2+ signaling in this achromatopsia mouse model, significant differences in dynamic intraorganellar Ca2+ levels were detected in CNG-channel-deficient cones. These results thus identify a novel molecular link between Ca2+ homeostasis and cone degeneration, thereby revealing novel therapeutic targets to preserve cones in inherited retinal degenerative diseases.


Effective biomedical document classification for identifying publications relevant to the mouse Gene Expression Database (GXD).
The Gene Expression Database (GXD) is a comprehensive online database within the Mouse Genome Informatics resource, aiming to provide available information about endogenous gene expression during mouse development. The information stems primarily from many thousands of biomedical publications that database curators must go through and read. Given the very large number of biomedical papers published each year, automatic document classification plays an important role in biomedical research. Specifically, an effective and efficient document classifier is needed for supporting the GXD annotation workflow. We present here an effective yet relatively simple classification scheme, which uses readily available tools while employing feature selection, aiming to assist curators in identifying publications relevant to GXD. We examine the performance of our method over a large manually curated dataset, consisting of more than 25 000 PubMed abstracts, of which about half are curated as relevant to GXD while the other half as irrelevant to GXD. In addition to text from title-and-abstract, we also consider image captions, an important information source that we integrate into our method. We apply a captions-based classifier to a subset of about 3300 documents, for which the full text of the curated articles is available. The results demonstrate that our proposed approach is robust and effectively addresses the GXD document classification. Moreover, using information obtained from image captions clearly improves performance, compared to title and abstract alone, affirming the utility of image captions as a substantial evidence source for automatically determining the relevance of biomedical publications to a specific subject area. www.informatics.jax.org.


The use of host cell machinery in the pathogenesis of Listeria monocytogenes.
The bacterial pathogen, Listeria monocytogenes, exploits the host cell's machinery, enabling the pathogen to enter into cells and spread from cell to cell. Three bacterial surface proteins are crucial for these processes: internalin and InlB, which mediate entry into cells, and ActA, which induces actin polymerisation at one pole of the bacterium and promotes intracellular and intercellular motility. Recent studies have identified several of the cellular factors involved in the entry process and major discoveries have unravelled the mechanisms underlying the actin-based motility. Increasing evidence shows that many cellular genes are up- or down-regulated during infection and probably play a role in the establishment of infection, inflammation and induction of the host immune response.


Association of two resistance mechanisms in a clinical isolate of Enterobacter cloacae with high-level resistance to imipenem.
Carbapenem resistance was studied in a clinical isolate of Enterobacter cloacae, strain 201 (MIC of imipenem and meropenem, 16 micrograms/ml). This strain was analyzed comparatively with the carbapenem-susceptible parent strain 200, an equally susceptible revertant, 201-Rev, and in vitro-selected mutants with different levels of carbapenem resistance. All strains produced similarly high amounts of the same cephalosporinase (pIapp = 8.8). Strain 201 apparently lacked two major outer membrane proteins of ca. 37 and 38 kDa, while 201-Rev produced only the 37-kDa protein. The permeability coefficient, determined with cephaloridine, was reduced up to ninefold in the resistant strains which also showed a substantial reduction in the uptake of [14C]meropenem. The introduction of the plasmid-borne ampD gene (whose product decreases the expression of ampC) resulted in almost complete cessation of cephalosporinase production in all strains and a substantial decrease in the MICs of the carbapenems which remained, however, 8- to 16-fold higher than those determined for the susceptible strains containing the ampD gene. This "residual" resistance was attributed to reduced outer membrane permeability. The contribution of cephalosporinase production was verified in a reverse experiment, in which the introduction of ampC into a low-level cephalosporinase producer resulted in a fourfold increase in the carbapenem MICs. From these results, we infer that reduced outer membrane permeability and high-level cephalosporinase production can operate in conjunction in clinical isolates of E. cloacae to confer imipenem resistance.


Mutations in RABL3 alter KRAS prenylation and are associated with hereditary pancreatic cancer.
Pancreatic ductal adenocarcinoma is an aggressive cancer with limited treatment options1. Approximately 10% of cases exhibit familial predisposition, but causative genes are not known in most families2. We perform whole-genome sequence analysis in a family with multiple cases of pancreatic ductal adenocarcinoma and identify a germline truncating mutation in the member of the RAS oncogene family-like 3 (RABL3) gene. Heterozygous rabl3 mutant zebrafish show increased susceptibility to cancer formation. Transcriptomic and mass spectrometry approaches implicate RABL3 in RAS pathway regulation and identify an interaction with RAP1GDS1 (SmgGDS), a chaperone regulating prenylation of RAS GTPases3. Indeed, the truncated mutant RABL3 protein accelerates KRAS prenylation and requires RAS proteins to promote cell proliferation. Finally, evidence in patient cohorts with developmental disorders implicates germline RABL3 mutations in RASopathy syndromes. Our studies identify RABL3 mutations as a target for genetic testing in cancer families and uncover a mechanism for dysregulated RAS activity in development and cancer.


Ethical dilemmas in community-based participatory research: recommendations for institutional review boards.
National and international codes of research conduct have been established in most industrialized nations to ensure greater adherence to ethical research practices. Despite these safeguards, however, traditional research approaches often continue to stigmatize marginalized and vulnerable communities. Community-based participatory research (CBPR) has evolved as an effective new research paradigm that attempts to make research a more inclusive and democratic process by fostering the development of partnerships between communities and academics to address community-relevant research priorities. As such, it attempts to redress ethical concerns that have emerged out of more traditional paradigms. Nevertheless, new and emerging ethical dilemmas are commonly associated with CBPR and are rarely addressed in traditional ethical reviews. We conducted a content analysis of forms and guidelines commonly used by institutional review boards (IRBs) in the USA and research ethics boards (REBs) in Canada. Our intent was to see if the forms used by boards reflected common CBPR experience. We drew our sample from affiliated members of the US-based Association of Schools of Public Health and from Canadian universities that offered graduate public health training. This convenience sample (n = 30) was garnered from programs where application forms were available online for download between July and August, 2004. Results show that ethical review forms and guidelines overwhelmingly operate within a biomedical framework that rarely takes into account common CBPR experience. They are primarily focused on the principle of assessing risk to individuals and not to communities and continue to perpetuate the notion that the domain of "knowledge production" is the sole right of academic researchers. Consequently, IRBs and REBs may be unintentionally placing communities at risk by continuing to use procedures inappropriate or unsuitable for CBPR. IRB/REB procedures require a new framework more suitable for CBPR, and we propose alternative questions and procedures that may be utilized when assessing the ethical appropriateness of CBPR.


Artesunate and sorafenib: Combinatorial inhibition of liver cancer cell growth.
An antimalarial medication, artesunate (Art), has exhibited promising anticancer effects with excellent tolerability in various types of cancer, suggesting that it has the potential to be used in combination with sorafenib (Sora) in hepatocellular carcinoma (HCC) treatment. To determine the potency of this combination, the present study attempted to quantitatively measure the dose-effect relationship of each drug alone and in combination in liver cancer cells in vitro using Calcusyn software. Cell growth inhibition was determined using the CyQUANT proliferation assay in two liver cancer cell lines, HepG2 and Huh7. Drug combination and reduction indices and isobologram plots were used to assess drug interactions. Cell apoptosis was evaluated by measurements of the proportion of cells in the sub G0/G1 phase of the cell cycle, and determination of protein expression levels of cleaved poly ADP ribose polymerase and caspase-9. Additionally, a cell migration assay was conducted using Essen ImageLock plates with an IncuCyte Zoom imaging system. The results of the present study revealed that the inhibitory effect of Sora on cell growth was synergistically enhanced by the combination with Art in HepG2 and Huh7 cells. The combination index and dose reduction index were specific to each cell line. Furthermore, combination at a fixed ratio presented mutual enhancement with respect to apoptosis induction and suppression of in vitro liver cancer cell migration. Therefore, considering the low toxicity and well-defined clinical characteristics of Art, combination of Sora and Art may present an attractive therapeutic option in the development of clinical trials for HCC treatment.


THYROID DYSFUNCTION AND INFERTILITY TREATMENT.
To find out the relationship of thyroid hormone profile of females with outcomes after intra cytoplasmic sperm injection (ICSI). It was a cross sectional study conducted in Islamabad Clinic Serving Infertile Couple from June 2013 till August 2015. T3 (triiodo thyronine), T4 (thyroxine) and TSH (thyroid stimulating hormone) of 168 consented females was estimated after they underwent the first step of treatment protocol (ovarian down regulation) for ICSI. Pregnant group had ß hCG result more than 25 IU/mL while the rest were included in the non-pregnant group. Both groups were compared by using independent sample t-test. Pearson correlation coefficient was used to associate T3 and T4 with other pregnancy variables with their significance. Non pregnant women had significantly higher mean values for T3 and T4 as compared to pregnant women (p <0.05, p<0.01). Difference in mean TSH value between non-pregnant and pregnant women was not significant p=0.08. It was found that T4 gave significant negative association with grading of embryo-I, blastocysts formed, thickness of endometrium and number of gestational sacs. Disturbance in thyroid profile with raised T4 levels leads to alteration in endometrial thickness and quality of embryos required for implantation and hence conception.