Depicting how Eucalyptus globulus survives drought: involvement of redox and DNA methylation events.
Eucalyptus globulus Labill. is widely cultivated and used by industry but its productivity is currently restricted by drought events, so research focussing on supporting programs to breed adapted germplasm is needed. In the present work we monitored severity of acute drought stress (7 and 11 days after water withholding) and relief (2h and 3 days after rewatering) by quantifying several biochemical markers of oxidative stress and DNA methylation patterns in leaves. Water withholding imposed a mild oxidative stress as estimated by redox shifts in the major antioxidant pools and increased lipid peroxidation. At the DNA level, global 5-methylcytosine distribution increased over the dehydration period especially in vascular tissue as estimated by immunolocalisation. Using methylation-sensitive RAPD analysis, which discriminates methylation changes occurring in specific DNA sequences, we found a high number of specific demethylation events also taking place. Immunolocalisation indicated a rapid reduction in global DNA methylation 2h after rehydration; however, a large number of de novo methylation events were still detected by methylation-sensitive RAPD. These events were associated with decreased lipid peroxidation and high cellular GSH pools relative to unstressed plants. Our results indicate the parallel induction of redox and complex DNA methylation changes occurring during stress imposition and relief.


Observational prospective study on Lactobacillus plantarum P 17630 in the prevention of vaginal infections, during and after systemic antibiotic therapy or in women with recurrent vaginal or genitourinary infections.
We performed a prospective cohort parallel observational study on the use of Lactobacillus plantarum P 17630 in the prevention of vaginal infections. Eligible were women with a diagnosis of bacterial vaginosis (<15 days) and documented history of recurrent vaginal infections; and/or cystitis (<15 days); and/or treatment with antibiotics for bacterial respiratory tract infections during the week before the study entry. Study subjects were prescribed Lactobacillus plantarum P 17630 > 100.000.000 UFC one vaginal capsule per day for 6 days, then a capsule per week for 16 weeks. Eligible subjects were enrolled in two parallel cohorts: 85 women using (group A) and 39 not using (group B) Lactobacillus plantarum P 17630. The risk of recurrent infection within 4 months from the study entry, was higher among untreated women: multivariate OR 2.6 (95%CI 0.7-9.4). The modification of presence/intensity or symptoms was significant in both the study groups (p < .001). Impact statement What is already known on this subject? The Lactobacillus plantarum P 17630 has been shown to be active in the treatment of bacterial vaginosis and vaginal candidiasis. No data are available on its efficacy in the prevention of recurrent vaginal or urological infection or as a prevention strategy during systemic treatment with antibiotics. What do the results of this study add? This observational study suggests that Lactobacillus plantarum given for 4 months may lower the risk of recurrent infection in women with recurrent vaginal or genitourinary infection or after antibiotic systemic treatment for bacterial respiratory tract infection. The finding, however, is not statistically significant, possibly due to the lower than expected rate of infection observed in our population and consequently the limited power of the study. What are the implications of these findings for clinical practice and/or further research? New studies are needed in order to evaluate in different populations the role of Lactobacillus plantarum in lowering the risk of recurrent infection in a high-risk populations.


Evolving status of operative versus nonoperative management of coronary arterial obstruction.
The past 15 years has witnessed great change in the treatment of ischemic heart disease. The next decade should continue to show equally promising new approaches to diagnosis, management, and interventional therapy. The relative roles of surgery and angioplasty should become better defined. Clinical advances in the medical and surgical intervention for acute myocardial infarction will receive continued emphasis. The results of using the internal mammary artery as a preferred conduit for multiple bypasses should be available and data to prove or disprove percutaneous angioplasty as an enduring procedure will be known. It should be a stimulating period for all involved in the diagnosis and treatment of patients with coronary artery disease.


Oro-dental features as useful diagnostic tool in Rubinstein-Taybi syndrome.
Rubinstein-Taybi syndrome (RTS; OMIM # 180849) is a well-known disorder characterized by mental and growth retardation, broad thumbs and great toes, and unusual facial characteristics. We studied oro-dental findings in a group of RTS patients: 12 from the UK, 2 from Greece, and 26 from France. All were examined by two investigators, using the Diagnosing Dental Defects Database record form to document these. Various oro-dental features were found: small mouth, retrognathia, micrognathia, highly arched and narrow palate, talon cusps, expressed crowding, screwdriver incisors, cross bites, and enamel hypoplasia. Eruption was usually normal. Specific attention for these anomalies should facilitate diagnosis and help adequate management.


Effect of stand types on understory vegetation.
The objective of this study was to compare density vegetative cover, basal area, height growth, aboveground biomass production, frequency floristic composition, and species diversity of understory vegetation growing in Hungarian oak (Quercus frainetto Ten.), Oriental beech (Fagus orientalis L.) and a mixed stand of both species (90% Hungarian oak and 10% Oriental beech) with the same crown closure. Understory vegetation was sampled with a quadrat of 0.5 m x 1 m along the 25 m-long transect lines in each stand. A total of seventeen perennial plant species from thirteen different families were found in the three stand types and all of them were present in Hungarian oak stand, eleven in Oriental beech stand, and seven in the mixed stand of Hungarian oak and Oriental beech trees. Results showed that stand types had a statistically significant effect on density, vegetative cover, basal area, height growth, biomass production, frequency floristic composition, and species diversity of understory vegetation. Total understory vegetation density was the highest for Hungarian oak stand with about 136 plants m(-2) and the lowest for the mixed stand of Hungarian oak and Oriental beech with 44 plants m(-2). In addition, the Hungarian oak stand had the most diverse understory vegetation with about 4.3 different plant m(-2), followed by the Oriental beech stand with 3.7 plants m(-2), and the mixed stand of the Hungarian oak and Oriental beech with 2.7 plants m(-2). Species density and diversity were the highest in Hungarian oak stand whereas aboveground biomass production and height growth of understory vegetation were highest in the mixed stand.


Pathogenic germline variants in BRCA1 and TP53 increase lung cancer risk in Chinese.
Multiple studies have identified pathogenic germline variants in cancer susceptibility genes (CSGs) in Chinese lung cancer patients; however, accurate assessment of these variants' contributions to cancer predisposition is always hampered by the absence of data on the prevalence of these variants in the general population. It is necessary to conduct a large-scale case-control study to identify CSGs that significantly increase the risk of lung cancer. We performed targeted sequencing of a CSGs panel in 1117 lung cancer patients and 16,327 controls from the general Chinese population. In comparison to controls, lung cancer patients had a considerably higher prevalence of pathogenic and likely pathogenic (P/LP) variations. Among lung cancer patients, 72% of P/LP variants carriers did not have a family cancer history, who would be ignored if germline testing was only provided for patients meeting family history-based criteria. Furthermore, compared to individuals with late-onset lung cancer, patients with early-onset lung cancer had a considerably higher prevalence of P/LP variations. With odds ratios (ORs) ranging from 4-fold (BRCA1: OR, 4.193; 95%CI, 1.382-10.768) to 29-fold (TP53: OR, 29.281; 95%CI, 1.523-1705.506), P/LP variants in the BRCA1 and TP53 genes were discovered to be strongly related to increased lung cancer risk. Additionally, with ORs ranging from 7.322-fold to infinity, we discovered 23 variations previously categorized as non-P/LP variants were highly enriched in lung cancer patients. Our findings indicated that P/LP variants in BRCA1 and TP53 conferred increased risk of lung cancer in Chinese.


Association between financial strain, social network and five-year recovery from first episode psychosis.
Despite much effort to positively affect long-term outcome in psychosis and schizophrenia many patients are still facing a poor outcome with persistent psychotic symptoms and decline in social functioning. The aim of this study was to examine the relationship between financial strain and social network and five-year outcome of first episode psychosis (FEP). FEP patients were divided into recovered (n = 52) and non-recovered (n = 19). Each person was matched according to age and gender with four persons (n = 284) from a longitudinal population-based study. All persons had answered an extensive questionnaire including social network, quantitative and qualitative, financial strain and mental health. Linear regression analysis showed that both financial strain and social network were associated, and had a unique contribution, to outcome. The results indicate that FEP patients might benefit from interventions that reduce financial strain thus facilitating daily life and cultural and social activities.


Mechanisms of osteoblastic metastases: role of endothelin-1.
Certain solid tumors metastasize to bone, causing an osteoblastic response. The mechanisms by which tumor cells stimulate this new bone formation are not understood completely. We identified three breast cancer lines that cause osteoblastic metastases in female nude mice and provide evidence that tumor-produced endothelin-1 (ET-1) mediates the osteoblastic response. Tumor-conditioned media and exogenous ET-1 stimulated osteoblast proliferation and new bone formation in cultures of calvarias from mice. These effects were blocked by endothelin A (ETA) but not by ETB receptor antagonists. Mice inoculated with the ZR-75-1 breast cancer line and treated with a selective ETA receptor antagonist (ABT-627) had significantly fewer osteoblastic bone metastases and less tumor burden compared with untreated mice. In contrast, there was no effect of ABT-627 on osteolytic bone metastases caused by ET-1-negative breast cancer, MDA-MB-231. ABT-627 had no effect on cell growth in vitro or at the orthotopic site (mammary fat pad) of ZR-75-1, or MDA-MB-231 cells. Collectively, the data suggest that tumor-produced ET-1 mediates osteoblastic bone metastases by stimulating osteoblast proliferation and new bone formation. Endothelin A receptor blockade may be useful for the prevention and treatment of osteoblastic bone metastases attributable to breast or prostate cancer.


Thrombotic microangiopathies (TTP, HUS, HELLP).
Thrombocytopenia, strictly defined as a platelet count less than 150,000, is common in the emergency department. Recognition, diagnostic investigation, and proper disposition of a thrombocytopenic patient are imperative. One group of disorders leading to thrombocytopenia is the thrombotic microangiopathies, hallmarked by platelet destruction. These thrombotic microangiopathies include thrombotic thrombocytopenic purpura (TTP), hemolytic uremic syndrome (HUS) and hemolysis, elevated liver enzyme levels, low platelet count (HELLP), which should be distinguished from similar disease processes such as immune thrombocytopenia (ITP), disseminated intravascular coagulation (DIC) and heparin induced thrombocytopenia (HIT). In this article, clinical presentations, pathophysiology, diagnostic workup, management plans, complications, and dispositions are addressed for this complex group of platelet disorders.


Participants' Role Expectations in Genetics Research and Re-consent: Revising the Theory and Methods of Mental Models Research Relating to Roles.
The rise of large cohort-based health research that includes genetic components has increased the communication challenges for researchers. Controversies have been amplified over requirements for re-consent, return of results, and privacy protections, among other issues. This study extended research on the impact that the perceived role of "research participant" might have on communication expectations to illuminate research participants' preferences for re-consent. The study employed an online survey of participants in a long-standing cancer genetics registry. Results confirmed previous exploratory findings that research participants endorse multiple mental models of participant roles in research (doctor-patient, collaborator, donor, legal contract, etc.). Regression analyses indicated that high and low salience of different models of the role of research participant are related to different communication expectations. However, the pattern of relationships among roles is relevant. The results of the regression analysis also indicated that preference for mandatory re-consent and its relationship to mental models of roles are related to attitudes of trust, benefits, and informational risks. The discussion identifies implications as including the use of explicit approaches to address role relationships in communication with research participants. It also points to implications for methodological approaches in mental model research.


Characterization of periostin isoform pattern in non-small cell lung cancer.
The extracellular matrix N-glycoprotein periostin (OSF-2, POSTN) is a major constituent of the desmoplastic stroma around solid tumors. It promotes tumor invasion and metastasis via epithelial-mesenchymal transition (EMT). In this study we investigated periostin expression at both RNA and protein level as well as the expression pattern of its splice isoforms in non-small cell lung cancer (NSCLC). Thirty fresh frozen and corresponding formalin-fixed NSCLC tissues (adeno- and squamous cell carcinoma subtype, each n=15) and their matched non-neoplastic tissues were investigated. Periostin mRNA levels were analyzed by quantitative RT-PCR. The EMT-markers periostin and vimentin were analyzed by immunohistochemistry. Laser capture microdissection allowed for analysis of periostin expression in tumor epithelia and stroma, separately. Isoform patterns were investigated by isoform-specific PCR following sequencing in NSCLC, fetal and adult normal lung tissue. The qRT-PCR analysis showed periostin mRNA up-regulation in NSCLC tissue in relation to normal lung, with significantly higher levels in the adeno-compared to the squamous cell subtype (p<0.05). However, protein levels in both tumor epithelia and stroma correlated with squamous cell carcinoma (p<0.001) and larger tumor size (p<0.05). Further, periostin tumor epithelia expression, correlated with higher tumor grade (p<0.05). Sequence analysis detected eight periostin isoforms in fetal lung, but only five in both NSCLC and matched normal lung tissue. Among the eight isoforms, four are new and were labelled 5, 7, 8 and 9. The exclusive presence of isoforms 1 and 9 in fetal tissue suggests splice-specific regulation during lung embryogenesis. Finally, laser capture microdissection demonstrated that both tumor epithelia and stromal cells can be a source of periostin production in NSCLC. This study represents the first analysis of periostin isoform expression patterns in NSCLC and a characterization of periostin expression in cancer versus stromal cells at both RNA and protein level.


The role of microRNAs in the pancreatic differentiation of pluripotent stem cells.
The generation of β-cells in vitro is an attractive option for cell therapy treatments for type 1 diabetes and also for the development of more accurate disease models. A number of studies have demonstrated that insulin-expressing cells can be generated by the in vitro differentiation of human pluripotent stem cells. However, to date, these differentiation protocols are often inefficient, time-consuming and highly variable. In many cases, this is a result of an incomplete understanding of the regulatory processes involved in the differentiation of human pluripotent stem cells. One such process is the control of gene expression by microRNAs (miRNAs). Given that miRNAs have the potential to influence cell fate, we present in this short review the evidence that a further understanding of the role of miRNAs in pancreatic development and function may be important in the on-going quest to generate insulin-secreting cells from pluripotent stem cells.


Molecular characterisation of group A streptococci from invasive and non-invasive disease episodes in Belgium during 1993-1994.
Five hundred clinical group A streptococcal (GAS) isolates were collected in Belgium during the period 1 Nov. 1993 to 31 Oct. 1994. Clinical and laboratory data were recorded and isolates were characterised. The presence of the genes encoding streptococcal pyrogenic exotoxin types A (speA), B (speB), C (speC), F (speF) and streptococcal superantigen (ssa) were determined by PCR to target specific sequences. These isolates were also emm-typed and analysed by pulsed-field gel electrophoresis (PFGE) of genomic macrorestriction fragments with the enzyme SmaI. In total, 136 unrelated GAS PFGE types were identified and genetic diversity was clearly demonstrated. Two GAS PFGE types predominated; a first PFGE type comprised 66 (13.2%) emm1 isolates characterised by speA-, speB+, speC-, speF+ and ssa-; the second PFGE type comprised 44 (8.8%) emm12 isolates characterised by speA-, speB+, speC+ (or speC-), speF+ and ssa-. Indistinguishable PFGE types were observed among both invasive and non-invasive isolates. Ten different PFGE types were found among 11 streptococcal toxic shock syndrome (STSS) isolates, and five of these lacked speA. Twenty-five (34.7%) of 72 invasive isolates gave negative results for speA, speC and ssa. This retrospective study confirmed the observation that the dissemination of one specific clone cannot be associated with invasive GAS disease and posed a question regarding the role of SPE A as a major virulence factor. Other streptococcal virulence factors in conjunction with host factors may determine the outcome of invasive GAS infection.


Adsorptive potential of Zea mays tassel activated carbon towards the removal of metformin hydrochloride from pharmaceutical effluent.
In the present study, Zea mays tassel which is a zero-value agricultural waste was used to produce a low-cost activated carbon using phosphoric acid as the activating agent. The prepared Z. mays tassel activated carbon (ZMTAC) was characterized by Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD). The adsorbent was applied for adsorption of an emerging contaminant, metformin hydrochloride (MH) from pharmaceutical effluent. The effects of solution pH, contact time, adsorbent dosage, and initial MH concentration and their interactions were investigated using a response surface methodology following a central composite experimental design (CCD). The optimum experimental conditions were as follows: pH 9.5, contact time 67.50 min, dosage 0.5750 g, and MH concentration 152.50 mg/L. The isotherm data followed Langmuir isotherm model (R2 = 0.979; sum of square deviation, SSD = 0.321). The saturation adsorption capacity of ZMTAC was 44.84 mg/g at 20 °C. MH adsorption process followed pseudo-second-order kinetics (higher R2 and smaller SSD values). The thermodynamic properties obtained showed that the adsorption process was feasible, endothermic and spontaneous. Consequently, the study demonstrated that Z. mays tassel is a potential precursor for preparation of adsorbents for the removal of the MH from pharmaceutical effluent.


Aortic arch reconstruction: deep and moderate hypothermic circulatory arrest with selective antegrade cerebral perfusion.
To compare the effects of moderate and deep hypothermic circulatory arrest (DHCA) with selective antegrade cerebral perfusion (SACP) during aortic arch surgery in adult patients and to offer the evidence for the detection of the temperature which provides best brain protection in the subjects who accept aortic arch reconstruction surgery. A total of 109 patients undergoing surgery of the aortic arch were divided into the moderate hypothermic circulatory arrest group (Group I) and the deep hypothermic circulatory arrest group (Group II). We recorded the data of the patients and their cardiopulmonary bypass (CPB) time, aortic clamping time, SACP time and postoperative anesthetized recovery time, tracheal intubation time, time in the intensive care unit (ICU) and postoperative neurologic dysfunction. Patient characteristics were similar in the two groups. There were four patients who died in Group II and 1 patient in Group I. There were no significant differences in aortic clamping time of each group (111.4±58.4 vs. 115.9±16.2) min; SACP time (27.4±5.9 vs. 23.5±6.1) min of the moderate hypothermic circulatory arrest group and the deep hypothermic circulatory arrest group; there were significant differences in cardiopulmonary bypass time (207.4±20.9 vs. 263.8±22.6) min, postoperative anesthetized recovery time (19.0±11.1 vs. 36.8±25.3) hours, extubation time (46.4±15.1 vs. 64.4±6.0) hours; length of stay in the intensive care unit (ICU) (4.7±1.7 vs. 8±2.3) days and postoperative neurologic dysfunction in the two groups. Compared to deep hypothermic circulatory arrest, moderate hypothermic circulatory arrest can provide better brain protection and achieve good clinical results.


Secretomes: the fungal strike force.
Microorganisms, although being very diverse because they comprise prokaryotic organisms such as bacteria or eukaryotic organisms such as fungi, all share an essential exodigester function. The consequence is their essential need to have a secretome adapted to their environment. The selection pressure exerted by environmental constraints led to the emergence of species with varying complexity in terms of composition of their secretomes. This review on fungal secretomes highlights the extraordinary variability among these organisms, even within the same species, and hence the absolute necessity to fully characterize all their components in the aims of understanding the fundamental mechanisms responsible for secretome plasticity and developing applications notably toward a better control of diseases caused by these pathogens.


HRD gene dependence of endoplasmic reticulum-associated degradation.
Work from several laboratories has indicated that many different proteins are subject to endoplasmic reticulum (ER) degradation by a common ER-associated machinery. This machinery includes ER membrane proteins Hrd1p/Der3p and Hrd3p and the ER-associated ubiquitin-conjugating enzymes Ubc7p and Ubc6p. The wide variety of substrates for this degradation pathway has led to the reasonable hypothesis that the HRD (Hmg CoA reductase degradation) gene-encoded proteins are generally involved in ER protein degradation in eukaryotes. We have tested this model by directly comparing the HRD dependency of the ER-associated degradation for various ER membrane proteins. Our data indicated that the role of HRD genes in protein degradation, even in this highly defined subset of proteins, can vary from absolute dependence to complete independence. Thus, ER-associated degradation can occur by mechanisms that do not involve Hrd1p or Hrd3p, despite their apparently broad envelope of substrates. These data favor models in which the HRD gene-encoded proteins function as specificity factors, such as ubiquitin ligases, rather than as factors involved in common aspects of ER degradation.


Responses of Heterodera glycines and Meloidogyne incognita Infective Juveniles to Root Tissues, Root Exudates, and Root Extracts from Three Plant Species.
The infective juvenile (J2) stage of endoparasitic plant nematodes uses plant chemical signals, released from roots, to localize and infect hosts. We examined the behaviors of soybean cyst nematode (Heterodera glycines) and root-knot nematode (Meloidogyne incognita) J2 in the presence of root signals from marigold (Tagetes patula), soybean (Glycine max), and pepper (Capsicum annuum). Signals were obtained from sources commonly used in phytoparasitic nematode chemotaxis studies: root tips, root exudates, and root extracts. Root tips from each plant species attracted M. incognita but H. glycines was attracted only to soybean. In contrast, root exudates prepared from marigold, pepper, or soybean seedlings were attractive to H. glycines but were repellent to M. incognita. Root extracts had the same effect as exudates. Fractionation of exudates by reversed-phase high-performance liquid chromatography (HPLC) (acetonitrile [CH3CN] and 0.1% trifluoroacetic acid) revealed highly polar and less polar components affecting behaviors. Fractions eluting at 12% CH3CN from all three plants attracted H. glycines and repelled M. incognita. None of the less polar HPLC fractions (>15% CH3CN) affected H. glycines but those from G. max and T. patula repelled M. incognita. Differences among exudates and effects of fractionation on behavior are discussed.


Effect of molecular structure on the properties of naphthalene-dipeptide hydrogelators.
Dipeptide-conjugates can be efficient low molecular weight hydrogelators. However, the effective design of a gelator for a specific application is compromised by the lack of a clear understanding of the design rules that govern assembly and hence gelation. Here, we report a library of naphthalene-dipeptides, their physical chemical properties, and gelation ability. We have varied both the amino acids in the dipeptides and the substitution on the naphthalene ring to allow variation of the structure throughout the molecule. We have examined the effects of these permutations on the critical micelle concentration and air-water partition coefficient at high pH and the apparent pK(a). We show that there is a clear link between these properties and the predicted hydrophobicity of the overall conjugates, rather than the properties varying with, for example, the dipeptide sequence. The majority of these dipeptide-conjugates are effective hydrogelators, although there is no apparent link between the solution properties and whether or not a conjugate is a hydrogelator. Nevertheless, where gelation occurs, the link between hydrophobicity and apparent pK(a) allows the prediction of the pH at which a gel will be formed and hence informed choice of gelator for specific applications.


Cell cycle-regulated trafficking of Chs2 controls actomyosin ring stability during cytokinesis.
Cytokinesis requires the coordination of many cellular complexes, particularly those involved in the constriction and reconstruction of the plasma membrane in the cleavage furrow. We have investigated the regulation and function of vesicle transport and fusion during cytokinesis in budding yeast. By using time-lapse confocal microscopy, we show that post-Golgi vesicles, as well as the exocyst, a complex required for the tethering and fusion of these vesicles, localize to the bud neck at a precise time just before spindle disassembly and actomyosin ring contraction. Using mutants affecting cyclin degradation and the mitotic exit network, we found that targeted secretion, in contrast to contractile ring activation, requires cyclin degradation but not the mitotic exit network. Analysis of cells in late anaphase bearing exocyst and myosin V mutations show that both vesicle transport and fusion machineries are required for the completion of cytokinesis, but this is not due to a delay in mitotic exit or assembly of the contractile ring. Further investigation of the dynamics of contractile rings in exocyst mutants shows these cells may be able to initiate contraction but often fail to complete the contraction due to premature disassembly during the contraction phase. This phenotype led us to identify Chs2, a transmembrane protein targeted to the bud neck through the exocytic pathway, as necessary for actomyosin ring stability during contraction. Chs2, as the chitin synthase that produces the primary septum, thus couples the assembly of the extracellular matrix with the dynamics of the contractile ring during cytokinesis.


The use of high-frequency skin ultrasound in the diagnosis of lipodermatosclerosis.
Lipodermatosclerosis is a symptom of severe venous insufficiency, the diagnosis of which is based on the clinical picture. Although the histopathology of the skin and the subcutaneous tissue allows for the most reliable diagnosis, it is not recommended due to healing disorders. The aim of this study was to assess the usefulness of high-frequency ultrasound in the diagnosis of lipodermatosclerosis. The study included 10 patients with lipodermatosclerosis who underwent Duplex Doppler ultrasound of lower limb veins, high-frequency ultrasound of the tibial skin, and radiography of the lower leg, all of which were analyzed in correlation with clinical symptoms. The study group included 9 women and 1 man aged 39-81 years. Manifestations of lipodermatosclerosis were detected in 14 limbs. High-frequency ultrasound showed that the mean dermis thickness at the affected sites was 2.63 mm, and was significantly thicker compared to healthy skin (1.45 mm) (p = 0.00002). Higher echogenicity was detected in the affected body regions in 85.7% of cases for the skin and 92.9% of cases for the subcutaneous tissue. Subcutaneous and vascular wall calcifications were detected in 92.9% and 78.6% of cases, respectively. Fibrosis was observed in all limbs, and compression sonoelastography showed that the compliance of the subcutaneous tissue was lower than that of muscles. The border between the skin and the subcutaneous tissue was blurred in 57.1% of cases. Radiography revealed thickening of the affected skin regions in all limbs, with calcifications detected in 85.7% of cases. A blurred border between the skin and the subcutaneous tissue was observed in 35.7% of limbs. High-frequency ultrasonography of the skin and the subcutaneous tissue in the lower legs supported with radiological findings is highly useful in the diagnosis of lipodermatosclerosis.


Modulation of the Association between the PEPD Variant and the Risk of Type 2 Diabetes by n-3 Fatty Acids in Chinese Hans.
Type 2 diabetes (T2D) is modulated by the interactions between genetic and dietary factors. This study sought to examine whether the associations of genome-wide association study (GWAS)-identified genetic variants with T2D risk were modulated by n-3 fatty acids in Chinese Hans. Six hundred and twenty-two T2D patients and 293 healthy controls were recruited. Erythrocyte phospholipid fatty acids were measured by standard methods. Nine GWAS-identified T2D-related single-nucleotide polymorphisms (SNPs) were genotyped. These SNPs were all identified in GWAS of Asian populations with a high minor allele frequency (>0.2). Among the 9 SNPs, only rs3786897 at PEPD (peptidase D) showed a significant interaction with n-3 fatty acids (p(interaction) after Bonferroni correction = 0.027). The rs3786897 A allele was associated with a higher risk of T2D [GA+AA vs. GG: odds ratio (OR) = 2.16, 95% confidence interval (CI) 1.32-3.55] when n-3 fatty acids were lower than the population median, but no significant association (GA+AA vs. GG: OR = 0.63, 95% CI 0.35-1.12) was observed when n-3 fatty acids were higher than the median. The association between the PEPD genetic variant and the risk of T2D was modulated by n-3 fatty acids. Higher n-3 fatty acids may abolish the adverse effect of the risk allele at PEPD for T2D.


Postoperative patellar tendon healing: an ultrasound study. VIS Tendon Study Group.
The patellar tendon donor site of 20 patients who underwent anterior cruciate ligament (ACL) reconstruction using the patellar tendon tissue as autograft was examined with high resolution 7.5 MHz ultrasound. The patients were randomly divided into four groups and studied at 3, 6, 9 or 12 months postoperatively. The size of the postoperative tendon defect was measured just distal to the lower pole of the patella. The size of the tendon defect diminished progressively from a mean of 109 mm2 at 3 months to a mean of 23 mm2 at 12 months. Increasing echogenicity was first noticed 12 months after tendon repair. Seven patients developed clinical features postoperatively of jumper's knee (patellar tendinosis). There were no ultrasound signs that differentiated these patients from asymptomatic patients. It is concluded that ultrasound provides objective evidence of patellar tendon healing after ACL reconstruction: the surgical defect diminished in size and became echogenic after a period of 12 months.


Mathematical model of the roles of T cells in inflammatory bowel disease.
Gut mucosal homeostasis depends on complex interactions among the microbiota, the intestinal epithelium, and the gut associated immune system. A breakdown in some of these interactions may precipitate inflammation. Inflammatory bowel diseases, Crohn's disease, and ulcerative colitis are chronic inflammatory disorders of the gastrointestinal tract. The initial stages of disease are marked by an abnormally high level of pro-inflammatory helper T cells, Th1. In later stages, Th2 helper cells may dominate while the Th1 response may dampen. The interaction among the T cells includes the regulatory T cells (Treg). The present paper develops a mathematical model by a system of differential equations with terms nonlocal in the space spanned by the concentrations of cytokines that represents the interaction among T cells through a cytokine signaling network. The model demonstrates how the abnormal levels of T cells observed in inflammatory bowel diseases can arise from abnormal regulation of Th1 and Th2 cells by Treg cells.


Vitamin D₃ metabolites enhance the NLRP3-dependent secretion of IL-1β from human THP-1 monocytic cells.
Vitamin D3 has emerged as an important regulator of the immune system. With metabolic enzymes for vitamin D3 activation and vitamin D receptors (VDR) now identified in a variety of immune cells, the active vitamin D3 metabolite 1,25(OH)2D3, is thought to possess immunomodulatory properties. We examined whether 1,25(OH)2D3 might also enhance the NLRP3-dependent release of mature IL-1β from macrophages. PMA-differentiated THP-1 cells were stimulated with vitamin D3 metabolites and assessed for CYP27, CYP24, NLRP3, ASC, pro-caspase-1 expression by western blot and real-time qPCR as well as inflammasome activation with pro-inflammatory cytokine IL-1β release measured by ELISA. Exposure to 1,25(OH)2D3 had no effect on the basal expression levels of VDR; however, CYP27A1 transcript was suppressed and CYP24A1 transcript was substantively elevated. Both 1,25(OH)2D3 - and 25(OH)D3 induced IL-1β release from THP-1 cells, and these effects were blocked with application of the caspase-1 inhibitor YVAD and the NLRP3 inhibitors glyburide and Bay 11-7082. Interestingly, 1,25 (OH)2D3 exposure reduced NLRP3 protein expression but had no effect on ASC or pro-caspase-1 protein levels. The increase in mature IL-1β elicited by 1,25(OH)2D3 was modest compared to that found for ATP or C. difficile toxins. However, co-treatment of THP-1 cells with ATP and 1,25(OH)2D3 resulted in more IL-1β secretion than ATP or 1,25(OH)2D3 alone.


The Role of Heart-Rate Variability Parameters in Activity Recognition and Energy-Expenditure Estimation Using Wearable Sensors.
Human-activity recognition (HAR) and energy-expenditure (EE) estimation are major functions in the mobile healthcare system. Both functions have been investigated for a long time; however, several challenges remain unsolved, such as the confusion between activities and the recognition of energy-consuming activities involving little or no movement. To solve these problems, we propose a novel approach using an accelerometer and electrocardiogram (ECG). First, we collected a database of six activities (sitting, standing, walking, ascending, resting and running) of 13 voluntary participants. We compared the HAR performances of three models with respect to the input data type (with none, all, or some of the heart-rate variability (HRV) parameters). The best recognition performance was 96.35%, which was obtained with some selected HRV parameters. EE was also estimated for different choices of the input data type (with or without HRV parameters) and the model type (single and activity-specific). The best estimation performance was found in the case of the activity-specific model with HRV parameters. Our findings indicate that the use of human physiological data, obtained by wearable sensors, has a significant impact on both HAR and EE estimation, which are crucial functions in the mobile healthcare system.


Assessing uncertainty in dynamic functional connectivity.
Functional connectivity (FC) - the study of the statistical association between time series from anatomically distinct regions (Friston, 1994, 2011) - has become one of the primary areas of research in the field surrounding resting state functional magnetic resonance imaging (rs-fMRI). Although for many years researchers have implicitly assumed that FC was stationary across time in rs-fMRI, it has recently become increasingly clear that this is not the case and the ability to assess dynamic changes in FC is critical for better understanding of the inner workings of the human brain (Hutchison et al., 2013; Chang and Glover, 2010). Currently, the most common strategy for estimating these dynamic changes is to use the sliding-window technique. However, its greatest shortcoming is the inherent variation present in the estimate, even for null data, which is easily confused with true time-varying changes in connectivity (Lindquist et al., 2014). This can have serious consequences as even spurious fluctuations caused by noise can easily be confused with an important signal. For these reasons, assessment of uncertainty in the sliding-window correlation estimates is of critical importance. Here we propose a new approach that combines the multivariate linear process bootstrap (MLPB) method and a sliding-window technique to assess the uncertainty in a dynamic FC estimate by providing its confidence bands. Both numerical results and an application to rs-fMRI study are presented, showing the efficacy of the proposed method.


Perceptions of cannabis health information labels among people who use cannabis in the U.S. and Canada.
The emergence of legal cannabis industries poses a new public health challenge. Health information labels are part of the public health strategy for tobacco and alcohol, but there is limited research on cannabis-related messaging. This study explored perceptions of cannabis health information labels among people who used cannabis in the last 12 months residing in the U.S. and Canada. The Global Drug Survey (GDS) is a large anonymous cross-sectional web-survey. In GDS2019, respondents were presented with six labels with cannabis-related health information (dependence; driving stoned; harms of smoking; harms to developing brain; lack of motivation; effects on memory), and asked if information was new, believed, would it change behavior, and about acceptability of having health labels on legal products. This paper includes 1,275 respondents from Canada and 2,224 from U.S. states where cannabis was legal at the time of the survey, and 5,230 from other U.S. states. Few respondents said that the information was new (6.6-24.6%). Most said the information was believable (63.5-72.0%) other than for the dependence message (28.1% new, 56.8% believed), which was perceived to be the least likely to change behavior (10.2%). Driving stoned was the message perceived to be the most likely to change behavior (58.5%). Respondents living in Canada were less likely to say information was new and rated most messages more believable than those in the U.S. Respondents from legal U.S. states were less likely to say information was new compared to other states. Respondents who used cannabis daily rated acceptability of labels lower (27.8%) than those using 1-48 days (40.6%). Novel, believable information may be more effective at changing behavior. Regular consumers may be less susceptible to messages. Information focusing on safer use strategies and benefits of reducing use may be more acceptable and should be assessed in future research.


Meiosis drives extraordinary genome plasticity in the haploid fungal plant pathogen Mycosphaerella graminicola.
Meiosis in the haploid plant-pathogenic fungus Mycosphaerella graminicola results in eight ascospores due to a mitotic division following the two meiotic divisions. The transient diploid phase allows for recombination among homologous chromosomes. However, some chromosomes of M. graminicola lack homologs and do not pair during meiosis. Because these chromosomes are not present universally in the genome of the organism they can be considered to be dispensable. To analyze the meiotic transmission of unequal chromosome numbers, two segregating populations were generated by crossing genetically unrelated parent isolates originating from Algeria and The Netherlands that had pathogenicity towards durum or bread wheat, respectively. Detailed genetic analyses of these progenies using high-density mapping (1793 DArT, 258 AFLP and 25 SSR markers) and graphical genotyping revealed that M. graminicola has up to eight dispensable chromosomes, the highest number reported in filamentous fungi. These chromosomes vary from 0.39 to 0.77 Mb in size, and represent up to 38% of the chromosomal complement. Chromosome numbers among progeny isolates varied widely, with some progeny missing up to three chromosomes, while other strains were disomic for one or more chromosomes. Between 15-20% of the progeny isolates lacked one or more chromosomes that were present in both parents. The two high-density maps showed no recombination of dispensable chromosomes and hence, their meiotic processing may require distributive disjunction, a phenomenon that is rarely observed in fungi. The maps also enabled the identification of individual twin isolates from a single ascus that shared the same missing or doubled chromosomes indicating that the chromosomal polymorphisms were mitotically stable and originated from nondisjunction during the second division and, less frequently, during the first division of fungal meiosis. High genome plasticity could be among the strategies enabling this versatile pathogen to quickly overcome adverse biotic and abiotic conditions in wheat fields.


Aspiration thrombectomy of M2 segment in acute ischemic stroke: The clinical reality in a neurovascular reference center.
Our study aimed to evaluate safety, efficacy and clinical outcomes in patients with acute ischemic stroke with occlusion of M2 segment treated with thromboaspiration. A retrospective study was conducted in patients with ischemic stroke of M2 segment undergoing endovascular thromboaspiration. The time period analyzed was from October 2015 until February 2021. Thromboaspiration was performed with AXS Catalyst 5 (Stryker) or AXS Catalyst 6 (Stryker) catheters. The following parameters were assessed: risk factors for ischemic stroke, National Institutes of Health Stroke Scale (entry and discharge), pre-procedural fibrinolysis, pre-procedural and 24-h Alberta Stroke Program Early CT Score, recanalization time, number of passages for recanalization, Thrombolysis in cerebral infarction scale score, periprocedural complications, Modified Rankin Scale score at 90 days from procedure and mortality. 90 patients were included in the study. The mean age was 75 ± 11.1 with National Institutes of Health Stroke Scale at entry 13 ± 5 and 8 ± 4 at discharge. Pre-procedural fibrinolysis were performed in 40 patients. Pre-procedural Alberta Stroke Program Early CT Score were 8.8 ± 1.3 and 6.9 ± 2.4 after 24 h from the procedure. Time of recanalization from onset of symptoms was 300 ± 82 min. Number of passages for recanalization were 1.8 ± 1.1. Thrombolysis in cerebral infarction scale score ≥ 2b were obtained in 90 % of procedures. After 90 days 33 % of patient obtained an Modified Rankin Scale between 0 and 1 (between 0 and 2 was 40 %). We didn't detect any complication in 64 % of cases (subarachnoid haemorrhage in 2 %, HI1 and HI 2 in 15 %, PH1 in 9 % of patients, PH2 in 6 % of patients). This paper confirms the usefulness and safety of thrombospiration in patients with ischemic stroke in the M2 segment with low intra-operative risks, high technical success and positive impact on the outcome of the patients.


[Ultrastructure of the osphradium of Siphonaria grisea (Mollusca, Pulmonata)].
By means of scanning and transmissive electron microscopic methods osphradium of Siphonaria grisea has been studied. The osphradium of the animal is presented as a small torulus formed by supporting ciliated cells. Among them bodies of receptory cells are situated; they are of smaller size and decorated with a bundle of microvilli. Central processes of the receptory cells, penetrating through a thick layer of the connective tissue, reach the subepithelial neural trunk. Concentration of the neural cells in the periphery of the trunk is small, and in the central area no chemical synapses are revealed. The data presented demonstrate a primitive structure of the osphradial chemoreceptory organ in Siphonaria.


Airway inflammation and remodelling changes in patients with chronic cough: do they tell us about the cause of cough?
Airway mucosal changes have been reported in chronic cough. In cough variant asthma and in eosinophilic bronchitis, there is evidence of eosinophil infiltration and sub-basement membrane thickening. In non-asthmatic cough, an increase of bronchoalveolar mast cells, mucosal mononuclear cells, and epithelial shedding have been reported. In a more recent study, evidence of airway wall remodelling has been observed in both asthmatic and non-asthmatic cough, such as an increase in sub-basement membrane thickness, goblet cell area, vascularity and vessel size. Smooth muscle area was increased in non-asthmatic coughers. Heightened cough sensitivity in non-asthmatic coughers was related to the degree of goblet cell hyperplasia and epithelial shedding. Cough reflex may be heightened by increased production of growth factors that might be further enhanced by the physical effects of cough on the airways. Mast cells may participate in the cough pathophysiology through release of growth factors as well as tussive mediators. Changes in the airway wall mucosa and epithelium may be important in the pathogenesis of cough receptor sensitization.


Internet Resources for Gene Expression Analysis in Arabidopsis thaliana.
The number of online databases and web-tools for gene expression analysis in Arabidopsis thaliana has increased tremendously during the last years. These resources permit the database-assisted identification of putative cis-regulatory DNA sequences, their binding proteins, and the determination of common cis-regulatory motifs in coregulated genes. DNA binding proteins may be predicted by the type of cis-regulatory motif. Further questions of combinatorial control based on the interaction of DNA binding proteins and the colocalization of cis-regulatory motifs can be addressed. The database-assisted spatial and temporal expression analysis of DNA binding proteins and their target genes may help to further refine experimental approaches. Signal transduction pathways upstream of regulated genes are not yet fully accessible in databases mainly because they need to be manually annotated. This review focuses on the use of the AthaMap and PathoPlant((R)) databases for gene expression regulation analysis and discusses similar and complementary online databases and web-tools. Online databases are helpful for the development of working hypothesis and for designing subsequent experiments.


Notch regulation of tumor angiogenesis.
The growth of new blood vessels (angiogenesis) is critical for tumor growth and progression. The highly conserved Notch signaling pathway is involved in a variety of cell fate decisions and regulates many cellular biological processes, including angiogenesis. Aberrant Notch signaling has also been implicated in tumorigenesis. Notch ligands and receptors are expressed on many different cell types present within the tumor, including tumor cells and the stromal compartment. This article highlights in particular the various mechanisms by which Notch signaling can mediate tumor angiogenesis. The most studied Notch ligands, Delta-like 4 and Jagged1, competitively regulate tumor angiogenesis. Studies have demonstrated that Delta-like 4 functions as a negative regulator of tumor angiogenesis, whereas Jagged1 promotes angiogenesis. Understanding the implications of Notch signaling in various tumor backgrounds will enable the effects of specific Notch signaling inhibition on tumor angiogenesis and growth to be evaluated as a potential for a novel antiangiogenic therapy in the clinic.


Simultaneous splenectomy improves outcomes after adult living donor liver transplantation.
Small-for-size graft (SFSG) syndrome is a major cause of graft loss after living donor liver transplantation (LDLT). Splenectomy (Spx) is an option to prevent this catastrophic complication, but its effect remains controversial. Herein, we aimed to elucidate the effect of simultaneous Spx on graft function and long-term outcomes after LDLT. Three hundred and twenty patients were divided into 2 groups: those undergoing (n = 258) and those not undergoing (n = 62) simultaneous Spx. To overcome selection bias, propensity score matching (PSM) was performed (n = 50 in each group). Before PSM, recipients undergoing simultaneous Spx showed better graft function on post-operative day (POD) 7 and 14, as well as lower sepsis frequency within 6 months after LDLT and better graft survival rates compared to those not undergoing Spx. After PSM, compared to patients not undergoing Spx, those undergoing Spx had a lower frequency of early graft dysfunction on POD 7 (p = 0.04); a lower frequency of SFSG syndrome (p = 0.01), lower serum total bilirubin levels (p = 0.001), and lower international normalized ratio (p = 0.004) on POD 14; lower sepsis frequency within 6 months after LDLT (p = 0.02), and better graft survival rates (p = 0.04). Univariate analysis revealed that not undergoing Spx (hazard ratio 3.06; 95% CI 1.07-11.0; p = 0.037) was the only risk factor for graft loss after LDLT. Simultaneous Spx may prevent SFSG syndrome and is a predictive factor for graft survival after LDLT. Simultaneous Spx is recommended when a small graft (≤35% of standard liver weight) is predicted preoperatively, or for patients with portal hypertension or high portal pressure (above 20 mmHg) after reperfusion in LDLT. Living donor liver transplantation (LDLT) for patients with acute or chronic liver failure is an alternative to overcome the deceased donor shortage. The potential mismatch between graft and body size is a problem that needs to be solved for LDLT recipients. Herein, we evaluated the impact of simultaneous splenectomy and showed that it was associated with favorable outcomes in patients undergoing LDLT.


Transcription factors in ferroptotic cell death.
Ferroptosis, a form of regulated cell death, is characterized by an excessive degree of iron accumulation and lipid peroxidation. Although it was originally identified only in cells expressing a mutant RAS oncogene, ferroptosis has also been found in normal cells following treatment by small molecules (e.g., erastin and RSL3) or drugs (e.g., sulfasalazine, sorafenib, and artesunate), which target antioxidant enzyme systems, especially the amino acid antiporter system xc- and the glutathione peroxidase GPX4. Dysfunctional ferroptosis is implicated in various physiological and pathological processes (e.g., metabolism, differentiation, and immunity). Targeting the ferroptotic network appears to a new treatment option for diseases or pathological conditions (e.g., cancer, neurodegeneration, and ischemia reperfusion injury). While the molecular machinery of ferroptosis remains largely unknown, several transcription factors (e.g., TP53, NFE2L2/NRF2, ATF3, ATF4, YAP1, TAZ, TFAP2C, SP1, HIF1A, EPAS1/HIF2A, BACH1, TFEB, JUN, HIC1, and HNF4A) play multiple roles in shaping ferroptosis sensitivity through either transcription-dependent or transcription-independent mechanisms. In this review, we summarize recent progress in understanding the transcriptional regulation underlying ferroptotic cell death, and discuss how it has provided new insights into cancer therapy.


Epidemiological analysis of a methicillin-resistant Staphylococcus aureus outbreak using restriction fragment length polymorphisms of genomic DNA.
The genomic DNA of 58 isolates of methicillin-resistant Staphylococcus aureus (MRSA) obtained during an infection outbreak at two major Canberra hospitals was analysed for restriction fragment length polymorphism (RFLP) by digestion with the endonuclease SmaI and resolution of the fragments by pulsed-field gel electrophoresis. Based on the fraction of common fragments generated by the endonuclease, DNA similarities among the isolates were estimated. Distance matrix analysis showed that the MRSA isolates could be divided into two major clusters (RFLP types I and II) and one minor one (type 46). A fourth group of miscellaneous isolates was found to be heterogeneous in terms of DNA sequence similarity. The epidemiological data indicated that RFLP type I was most common in the intensive care units in the two hospitals, with particular subtypes of RFLP type I concentrated in individual units. RFLP type II and the miscellaneous group were more generally distributed. Type 46 isolates appear to be related to a group which was present in epidemics in Melbourne hospitals in the early 1980s. Using the standard phage set, the RFLP type I group was largely untypable. However, type II isolates were all phage typable, with a shared susceptibility to phages 29/85/95/90; type 46 isolates had a shared susceptibility to phages 85/90. The miscellaneous isolates were of variable phage types.


The P. falciparum alternative histones Pf H2A.Z and Pf H2B.Z are dynamically acetylated and antagonized by PfSir2 histone deacetylases at heterochromatin boundaries.
The Plasmodium falciparum alternative histones Pf H2A.Z and Pf H2B.Z are enriched in the same nucleosomes in intergenic euchromatin but depleted from heterochromatin. They occupy most promoters but are only dynamically associated with expression at var genes. In other organisms, acetylation of H2A.Z is important for its functions in gene expression and chromatin structure. Here, we show that acetylated Pf H2A.Z and Pf H2B.Z are dynamically associated with gene expression at promoters. In addition, acetylated Pf H2A.Z and Pf H2B.Z are antagonized by the sirtuin class III histone deacetylases (HDAC) PfSir2A and B at heterochromatin boundaries and encroach upon heterochromatin in parasites lacking PfSir2A or B. However, the majority of acetylated Pf H2A.Z and Pf H2B.Z are deacetylated by class I or II HDACs. Acetylated Pf H2A.Z and Pf H2B.Z are also dynamically associated with promoter activity of both canonical upstream var gene promoters and var gene introns. These findings suggest that both acetylated Pf H2A.Z and Pf H2B.Z play critical roles in gene expression and contribute to maintenance of chromatin structure at the boundaries of subtelomeric, facultative heterochromatin, critical for the variegated expression of genes that enable rapid adaptation to altered host environments.IMPORTANCEThe malaria parasite Plasmodium falciparum relies on variant expression of members of multi-gene families as a strategy for environmental adaptation to promote parasite survival and pathogenesis. These genes are located in transcriptionally silenced DNA regions. A limited number of these genes escape gene silencing, and switching between them confers variant fitness on parasite progeny. Here, we show that PfSir2 histone deacetylases antagonize DNA-interacting acetylated alternative histones at the boundaries between active and silent DNA. This finding implicates acetylated alternative histones in the mechanism regulating P. falciparum variant gene silencing and thus malaria pathogenesis. This work also revealed that acetylation of alternative histones at promoters is dynamically associated with promoter activity across the genome, implicating acetylation of alternative histones in gene regulation genome wide. Understanding mechanisms of gene regulation in P. falciparum may aid in the development of new therapeutic strategies for malaria, which killed 619,000 people in 2021.


The application of Sn(IV)Cl2 and In(III)Cl porphyrin-dyed TiO2 nanofibers in photodynamic antimicrobial chemotherapy for bacterial inactivation in water.
The World Health Organization has reported that antimicrobial resistance is one of the top 10 health threats that humanity faces today. Due to this, alternative therapies to the common antimicrobials are being explored and among these is photodynamic antimicrobial chemotherapy, where a combination of light, a photosensitizer and reactive oxygen species can be used to target microbial cells. In this research, free base, tin (IV) and indium (III) tetramethoxyporphyrins photosensitizers are adsorbed onto inorganic titanium dioxide nanofibers in an effort to create reusable fibers that are effective against Staphylococcus aureus. The photodynamic antimicrobial chemotherapy studies indicate that the metalloporphyrin adsorbed nanofibers exhibit good photodynamic antimicrobial activity against Staphylococcus aureus where the Cl2Sn(IV) tetramethoxyporphyrin dyed TiO2 exhibited 100% bacterial inhibition after a 30 min irradiation period.


G protein beta 5 is targeted to D2-dopamine receptor-containing biochemical compartments and blocks dopamine-dependent receptor internalization.
G beta 5 (Gbeta5, Gβ5) is a unique G protein β subunit that is thought to be expressed as an obligate heterodimer with R7 regulator of G protein signaling (RGS) proteins instead of with G gamma (Gγ) subunits. We found that D2-dopamine receptor (D2R) coexpression enhances the expression of Gβ5, but not that of the G beta 1 (Gβ1) subunit, in HEK293 cells, and that the enhancement of expression occurs through a stabilization of Gβ5 protein. We had previously demonstrated that the vast majority of D2R either expressed endogenously in the brain or exogenously in cell lines segregates into detergent-resistant biochemical fractions. We report that when expressed alone in HEK293 cells, Gβ5 is highly soluble, but is retargeted to the detergent-resistant fraction after D2R coexpression. Furthermore, an in-cell biotin transfer proximity assay indicated that D2R and Gβ5 segregating into the detergent-resistant fraction specifically interacted in intact living cell membranes. Dopamine-induced D2R internalization was blocked by coexpression of Gβ5, but not Gβ1. However, the same Gβ5 coexpression levels had no effect on agonist-induced internalization of the mu opioid receptor (MOR), cell surface D2R levels, dopamine-mediated recruitment of β-arrestin to D2R, the amplitude of D2R-G protein coupling, or the deactivation kinetics of D2R-activated G protein signals. The latter data suggest that the interactions between D2R and Gβ5 are not mediated by endogenously expressed R7 RGS proteins.


Lessons learnt from metal-on-metal hip arthroplasties will lead to safer innovation for all medical devices.
Metal-on-metal bearings were re-popularised in the late 1990s with the introduction of modern hip resurfacing. Large diameter (LD) metal-on-metal (MoM) hips became more prevalent and have been the least successful group of hip implants ever used. They were rapidly adopted from 2004 until the British Hip Society stopped their use in 2012. Well functioning MoM hip results (including the BHR and Metasul) are hidden in the mire of poor results from the group of all MoM bearings.We have reviewed what happened and we make 3 observations. Firstly, collaboration between surgeons and then between surgeons and other disciplines, first identified and then solved the clinical management problems. Secondly, the problems with MoM hips occurred because hip simulation was inadequate at predicting performance in patients. They gave no indications of the biological effects of wear in the human environment. Lastly, retrieval of failed implants was essential to understanding why failure occurred.These lessons must never be forgotten and must form the basis by which new or altered implants are introduced and how they should be monitored. This will enable safer innovation for patients, surgeons and manufacturers. The problems with MoM hips will not have been in vain.


Facile synthesis of metal-organic framework-derived SiW12@Co3O4 and its peroxidase-like activity in colorimetric assay.
Over the past few years, artificial enzymes have attracted enormous attention due to their high stabilities and cost-effective productions. In this work, metal-organic framework-derived SiW12@Co3O4 was synthesized in large quantities by stirring the mixture at ambient temperature and calcination. The obtained SiW12@Co3O4 exhibited a highly inherent peroxidase-like activity and excellent stability. Kinetic studies demonstrated that the synthesized SiW12@Co3O4 had a strong binding affinity to 3,3',5,5'-tetramethylbenzidine (TMB), stronger than HRP had. Specifically, the peroxidase-like activity of SiW12@Co3O4 in an aqueous solution was well maintained after incubation at an elevated temperature, at an extreme pH and for a long time. A SiW12@Co3O4-based method was further developed for H2O2 and one-pot glucose detection with good sensitivity and reliability. The facile synthesis approach is expected to facilitate the practical use of metal-organic frameworks and their derivatives as enzyme mimics in the future.


Prefrontal Cortex Oxygenation During Endurance Performance: A Systematic Review of Functional Near-Infrared Spectroscopy Studies.
Introduction: A myriad of factors underlie pacing-/exhaustion-decisions that are made during whole-body endurance performance. The prefrontal cortex (PFC) is a brain region that is crucial for decision-making, planning, and attention. PFC oxygenation seems to be a mediating factor of performance decisions during endurance performance. Nowadays, there is no general overview summarizing the current knowledge on how PFC oxygenation evolves during whole-body endurance performance and whether this is a determining factor. Methods: Three electronic databases were searched for studies related to the assessment of PFC oxygenation, through near-IR spectroscopy (NIRS), during endurance exercise. To express PFC oxygenation, oxygenated (HbO2) and deoxygenated hemoglobin (HHb) concentrations were the primary outcome measures. Results: Twenty-eight articles were included. Ten articles focused on assessing prefrontal oxygenation through a maximal incremental test (MIT) and 18 focused on using endurance tasks at workloads ranging from low intensity to supramaximal intensity. In four MIT studies measuring HbO2, an increase of HbO2 was noticed at the respiratory compensation point (RCP), after which it decreased. HbO2 reached a steady state in the four studies and increased in one study until exhaustion. All studies found a decrease or steady state in HHb from the start until RCP and an increase to exhaustion. In regard to (non-incremental) endurance tasks, a general increase in PFC oxygenation was found while achieving a steady state at vigorous intensities. PCF deoxygenation was evident for near-to-maximal intensities at which an increase in oxygenation and the maintenance of a steady state could not be retained. Discussion/Conclusion: MIT studies show the presence of a cerebral oxygenation threshold (ThCox) at RCP. PFC oxygenation increases until the RCP threshold, thereafter, a steady state is reached and HbO2 declines. This study shows that the results obtained from MIT are transferable to non-incremental endurance exercise. HbO2 increases during low-intensity and moderate-intensity until vigorous-intensity exercise, and it reaches a steady state in vigorous-intensity exercise. Furthermore, ThCox can be found between vigorous and near-maximal intensities. During endurance exercise at near-maximal intensities, PFC oxygenation increases until the value exceeding this threshold, resulting in a decrease in PFC oxygenation. Future research should aim at maintaining and improving PFC oxygenation to help in improving endurance performance and to examine whether PFC oxygenation has a role in other performance-limiting factors.


Predictors of anterior and posterior wall carotid intima media thickness progression in men and women at moderate risk of coronary heart disease.
The rate of carotid intima media thickness (CIMT) progression has been widely used in clinical trials as a surrogate marker for subclinical atherosclerosis. The aim of this study was to investigate relationships between coronary heart disease (CHD) risk markers and progression of CIMT in patients at moderate CHD risk. Participants included men (45-75 years) and women (55-74 years) in the control arm of a clinical trial. All had at least one major CHD risk factor and baseline posterior wall CIMT 0.7-2.0 mm, without significant stenosis. Posterior (n = 134) and anterior wall (in a subset, n = 72) CIMT were assessed with B-mode ultrasound at baseline and 12 and ∼18 months. Fasting lipoprotein lipid, apolipoprotein (Apo), inflammatory, and oxidative stress markers were evaluated. Baseline CIMT was inversely associated (P < .001) with CIMT progression. After adjustment for baseline CIMT, significant predictors of anterior wall CIMT progression in linear regression analyses included glucose (P = .044), high-density lipoprotein cholesterol (HDL-C, inverse, P = .006), triglycerides (TG, P = .006), and ratios of total cholesterol (TC)/HDL-C (P = .013), TG/HDL-C (P = .005), and Apo B/HDL-C (P = .021). Posterior wall CIMT progressed on average, whereas anterior wall CIMT regressed (0.0078 vs -0.0164 mm/year, P = .014). Significant baseline CIMT-adjusted predictors of posterior wall CIMT progression included TC (P = .028), low-density lipoprotein-C (P = .035), non-HDL-C (P = .004), TG (P = .016), Apo B (P = .005), and ratios of TC/HDL-C (P < .001), TG/HDL-C (P = .015), Apo B/Apo AI (P = .012) and Apo B/HDL-C (P = .004). The strongest predictors for CIMT progression in anterior and posterior walls were lower baseline CIMT, increased TG, and elevated ratios, including TC/HDL-C, TG/HDL-C and Apo B/HDL-C.


Bilateral renal descensus and intravesicular ureteroneocystostomy for treatment of bilateral ureteral ligation and transection that occurred during ovariohysterectomy in two cats.
6-month-old and 7-month-old spayed female domestic shorthair cats were referred because of complications associated with inadvertent bilateral ureteral ligation and transection during ovariohysterectomy. Both cats had a 1- to 2-day history of lethargy, inappetence, and vomiting. Initial exam findings included lethargy, signs of abdominal pain, anuria, and dehydration. Clinicopathologic testing revealed azotemia and hyperkalemia. Abdominal ultrasonography revealed peritoneal effusion and bilateral pyelectasia in both cats and retroperitoneal effusion in one. Fluid analysis in both cats supported a diagnosis of uroabdomen. Exploratory celiotomy was performed in both cats, and bilateral ureteral ligation and transection was confirmed. Bilateral renal descensus and ureteroneocystostomy with an intravesicular mucosal apposition technique was successfully performed in both cats. Clinicopathologic evaluation performed 1 day after surgery in one cat and 5 days after surgery in the other revealed complete resolution of azotemia. Ultrasonographic examination of the urogenital tract performed approximately 4 months after surgery in the first cat and 1 month after surgery in the second cat revealed complete resolution of renal pelvic dilation bilaterally. Bilateral intravesicular ureteroneocystostomy in conjunction with bilateral renal descensus was used successfully to treat bilateral ureteral transection that occurred in 2 cats during routine ovariohysterectomy. Limited treatment options currently exist for this serious complication, and euthanasia is often considered. This technique, which relies on the use of the natural surrounding tissues for successful treatment, can offer a potential treatment option to correct this uncommon but devastating complication.


The Effect of Mesoporous Structure of the Support on the Oxidation of Dibenzothiophene.
A source of Brønsted acid centers, generated on the surface of two mesoporous silica supports of different structures (SBA-15 and MCF), was 3-(trihydroxysilyl)-1-propanesufonic acid (TPS). The materials obtained were characterized and applied as catalysts for the oxidative desulfurization of dibenzothiophene (DBT) with hydrogen peroxide as a model ODS (oxidative desulfurization) process. The properties of the materials were examined via nitrogen physisorption, XRD (X-ray Diffraction) and elemental analysis showing the preservation of the support structure after modification with organosilane species. Due to the aggregation of catalyst particles in the reaction mixture, the SBA-15 based catalyst was not very effective in DBT oxidation. Contrary, TPS/MCF catalyst exhibited a very good activity (almost total conversion of DBT after 1 h in optimized reaction conditions) and stability in dibenzothiophene oxidation in mild reaction conditions.


The ketogenic diet and epilepsy.
The ketogenic diet has long been used to treat medically refractory epilepsy. The mechanisms underlying its clinical effects, however, have remained a mystery. The evidence to date suggests that a fundamental shift from glycolysis to intermediary metabolism induced by the ketogenic diet is necessary and sufficient for clinical efficacy. This notion is supported by a growing number of studies indicating that glucose restriction, ketone bodies and polyunsaturated fatty acids may all play mechanistic roles, possibly by enhancing mitochondrial respiration and ATP production, and decreasing reactive oxygen species production. Recent reports indicate that ketone bodies can reduce oxidative stress and that fatty acid-induced mitochondrial uncoupling may also yield similar protective effects. Ketone bodies may attenuate spontaneous firing of ATP-sensitive potassium channels in central neurons, and pharmacological inhibition of glycolysis has been shown to retard epileptogenesis in a rat kindling model. While the mechanisms underlying the broad clinical efficacy of the ketogenic diet remain unclear, there is growing evidence that the ketogenic diet alters the fundamental biochemistry of neurons in a manner that not only inhibits neuronal hyperexcitability but also induces a protective effect. Thus, the ketogenic diet may ultimately be useful in the treatment of a variety of neurological disorders.


A conditional immortalized mouse muller glial cell line expressing glial and retinal stem cell genes.
Müller glia have multiple functions in the retina, including synthesis of neurotrophic factors, uptake and metabolism of neurotransmitters, spatial buffering of ions, maintenance of the blood-retinal barrier, and response to injury. A population of Müller glia has some stem cell-like characteristics both in vivo and in vitro. The purpose of this study was to generate and characterize novel Müller glial cell lines from the postnatal mouse retina. Cells were cultured from postnatal day (P) 10 double heterozygous transgenic (H-2K(b)-tsA58/+; HRhoGFP/+) or C57BL/6 mice after papain dissociation. Interferon gamma (IFNγ) induction of the SV40 T-antigen (TAg) was assayed by immunohistochemistry and Western blot analysis. Proliferation was assayed by BrdU uptake and cell counts of calcein AM/ethidium bromide-stained cells. Gene expression was analyzed by RT-PCR and immunohistochemistry. Conditionally immortalized (ImM10 [Immortmouse Müller P10]) and spontaneously immortalized (C57M10 [C57BL/6 Müller P10]) Müller glial cell lines were selected by differential adherence to laminin; both consisted of adherent flat cells with large, diffusely staining nuclei and an epithelial morphology. TAg induction stimulated BrdU uptake by Müller glia in mixed retinal cultures from H-2K(b)-tsA58/+; HRhoGFP/+ mice and increased the proliferation of ImM10 cells. ImM10 and C57M10 cells expressed genes characteristic of Müller glia but not genes characteristic of differentiated retinal neurons. ImM10 cells also expressed retinal stem cell genes. The ImM10 cell line is a novel, conditionally immortalized Müller glial cell line isolated from the P10 mouse retina that expresses genes characteristic of Müller glial and retinal stem cells.


Prevalence and risk factors for hair loss in outdoor-wintered beef cattle under cold weather conditions.
Keeping cattle outdoors year-around is considered an attractive alternative to indoor winter-housing, due to lower investment costs and better welfare. However, hair loss, attributed to lice, may impair cattle's thermal balance during harsh winters. During the winters of 2019-2021, outdoor cattle in Sweden were studied for the prevalence and development of hair loss, while surveys were conducted among the farmers on their perceptions and attitudes around hair loss. Of the 463 groups of cattle from 75 farms enrolled in a welfare control program issued by the Swedish Board of Agriculture, 25.7% (n = 119) had at least one animal with hair loss. When we followed up a subset of animals (n = 3673) which did not receive prophylactic delousing, 15.7% developed hair loss. Hair loss occurrence increased between visits within each winter in these animals, suggesting a contagious etiology. Logistic regression analyses, using the information collected in the control program and the hair loss outcome, showed that preventive delousing before November was effective, alongside keeping animals clean and the group size small. Meanwhile, being older (>2 years) and having access to bedding materials was shown to increase the risk at an animal-level. Some groups (n = 34) had no hair loss despite receiving no prophylactic delousing. Based on the survey conducted among the farmers (n = 15), groups with lower hair loss prevalence belonged to farmers who were more observant of hair loss and gave prompt treatment. This study provides knowledge useful to limit delousing interventions without compromising animal welfare.


Tissue-specific expression of the human type II collagen gene in mice.
Type II collagen is crucial to the development of form in vertebrates as it is the major protein of cartilage. To study the factors regulating its expression we introduced a cosmid containing the human type II collagen gene, including 4.5 kilobases of 5' and 2.2 kilobases of 3' flanking DNA, into embryonic stem cells in vitro. The transformed cells contribute to all tissues in chimeric mice allowing the expression of the exogenous gene to be studied in vivo. Human type II collagen mRNA is restricted to tissues showing transcription from the endogenous gene and human type II collagen is found in extracellular matrix surrounding chondrocytes in cartilage. The results indicate that the cis-acting requirements for correct temporal and spatial regulation of the gene are contained within the introduced DNA.


Human anti-NKp46 antibody for studies of NKp46-dependent NK cell function and its applications for type 1 diabetes and cancer research.
Natural killer (NK) cells are innate lymphocytes that efficiently eliminate cancerous and infected cells. NKp46 is an important NK activating receptor shown to participate in recognition and activation of NK cells against pathogens, tumor cells, virally infected cells, and self-cells in autoimmune conditions, including type I and II diabetes. However, some of the NKp46 ligands are unknown and therefore investigating human NKp46 activity and its critical role in NK cell biology is problematic. We developed a unique anti-human NKp46 monocloncal antibody, denoted hNKp46.02 (02). The 02 mAb can induce receptor internalization and degradation. By binding to a unique epitope on a particular domain of NKp46, 02 lead NKp46 to lysosomal degradation. This downregulation therefore enables the investigation of all NKp46 activities. Indeed, using the 02 mAb we determined NK cell targets which are critically dependent on NKp46 activity, including certain tumor cells lines and human pancreatic beta cells. Most importantly, we showed that a toxin-conjugated 02 inhibits the growth of NKp46-positive cells; thus, exemplifying the potential of 02 in becoming an immunotherapeutic drug to treat NKp46-dependent diseases, such as, type I diabetes and NK and T cell related malignancies.


Consensus-Based Recommendations on Priority Activities to Address Acute Kidney Injury in Children: A Modified Delphi Consensus Statement.
Increasing evidence indicates that acute kidney injury (AKI) occurs frequently in children and young adults and is associated with poor short-term and long-term outcomes. Guidance is required to focus efforts related to expansion of pediatric AKI knowledge. To develop expert-driven pediatric specific recommendations on needed AKI research, education, practice, and advocacy. At the 26th Acute Disease Quality Initiative meeting conducted in November 2021 by 47 multiprofessional international experts in general pediatrics, nephrology, and critical care, the panel focused on 6 areas: (1) epidemiology; (2) diagnostics; (3) fluid overload; (4) kidney support therapies; (5) biology, pharmacology, and nutrition; and (6) education and advocacy. An objective scientific review and distillation of literature through September 2021 was performed of (1) epidemiology, (2) risk assessment and diagnosis, (3) fluid assessment, (4) kidney support and extracorporeal therapies, (5) pathobiology, nutrition, and pharmacology, and (6) education and advocacy. Using an established modified Delphi process based on existing data, workgroups derived consensus statements with recommendations. The meeting developed 12 consensus statements and 29 research recommendations. Principal suggestions were to address gaps of knowledge by including data from varying socioeconomic groups, broadening definition of AKI phenotypes, adjudicating fluid balance by disease severity, integrating biopathology of child growth and development, and partnering with families and communities in AKI advocacy. Existing evidence across observational study supports further efforts to increase knowledge related to AKI in childhood. Significant gaps of knowledge may be addressed by focused efforts.


Cardiac reperfusion injury: aging, lipid peroxidation, and mitochondrial dysfunction.
Cardiac reperfusion and aging are associated with increased rates of mitochondrial free radical production. Mitochondria are therefore a likely site of reperfusion-induced oxidative damage, the severity of which may increase with age. 4-Hydroxy-2-nonenal (HNE), a major product of lipid peroxidation, increases in concentration upon reperfusion of ischemic cardiac tissue, can react with and inactivate enzymes, and inhibits mitochondrial respiration in vitro. HNE modification of mitochondrial protein(s) might, therefore, be expected to occur during reperfusion and result in loss in mitochondrial function. In addition, this process may be more prevalent in aged animals. To begin to test this hypothesis, hearts from 8- and 24-month-old rats were perfused in Langendorff fashion and subjected to periods of ischemia and/or reperfusion. The rate of state 3 respiration of mitochondria isolated from hearts exposed to ischemia (25 min) was approximately 25% less than that of controls, independent of age. Reperfusion (40 min) caused a further decline in the rate of state 3 respiration in hearts isolated from 24- but not 8-month-old rats. Furthermore, HNE modification of mitochondrial protein (approximately 30 and 44 kDa) occurred only during reperfusion of hearts from 24-month-old rats. Thus, HNE-modified protein was present in only those mitochondria exhibiting reperfusion-induced declines in function. These studies therefore identify mitochondria as a subcellular target of reperfusion damage and a site of age-related increases in susceptibility to injury.


Streptococcus mitis Expressing Pneumococcal Serotype 1 Capsule.
Streptococcus pneumoniae's polysaccharide capsule is an important virulence factor; vaccine-induced immunity to specific capsular polysaccharide effectively prevents disease. Serotype 1 S. pneumoniae is rarely found in healthy persons, but is highly invasive and a common cause of meningitis outbreaks and invasive disease outside of the United States. Here we show that genes for polysaccharide capsule similar to those expressed by pneumococci were commonly detected by polymerase chain reaction among upper respiratory tract samples from older US adults not carrying pneumococci. Serotype 1-specific genes were predominantly detected. In five oropharyngeal samples tested, serotype 1 gene belonging to S. mitis expressed capsules immunologically indistinct from pneumococcal capsules. Whole genome sequencing revealed three distinct S. mitis clones, each representing a cps1 operon highly similar to the pneumococcal cps1 reference operon. These findings raise important questions about the contribution of commensal streptococci to natural immunity against pneumococci, a leading cause of mortality worldwide.


N-terminal pro-B-type natriuretic peptide variability in stable dialysis patients.
Monitoring N-terminal pro-B-type natriuretic peptide (NT-proBNP) may be useful for assessing cardiovascular risk in dialysis patients. However, its biologic variation is unknown, hindering the accurate interpretation of serial concentrations. The aims of this prospective cohort study were to estimate the within- and between-person coefficients of variation of NT-proBNP in stable dialysis patients, and derive the critical difference between measurements needed to exclude biologic and analytic variation. Fifty-five prevalent hemodialysis and peritoneal dialysis patients attending two hospitals were assessed weekly for 5 weeks and then monthly for 4 months between October 2010 and April 2012. Assessments were conducted at the same time in the dialysis cycle and entailed NT-proBNP testing, clinical review, electrocardiography, and bioimpedance spectroscopy. Patients were excluded if they became unstable. This study analyzed 136 weekly and 113 monthly NT-proBNP measurements from 40 and 41 stable patients, respectively. Results showed that 22% had ischemic heart disease; 9% and 87% had left ventricular systolic and diastolic dysfunction, respectively. Respective between- and within-person coefficients of variation were 153% and 27% for weekly measurements, and 148% and 35% for monthly measurements. Within-person variation was unaffected by dialysis modality, hydration status, inflammation, or cardiac comorbidity. NT-proBNP concentrations measured at weekly intervals needed to increase by at least 46% or decrease by 84% to exclude change due to biologic and analytic variation alone with 90% certainty, whereas monthly measurements needed to increase by at least 119% or decrease by 54%. The between-person variation of NT-proBNP was large and markedly greater than within-person variation, indicating that NT-proBNP testing might better be applied in the dialysis population using a relative-change strategy. Serial NT-proBNP concentrations need to double or halve to confidently exclude change due to analytic and biologic variation alone.


Balanced nutrient requirements for maize in the Northern Nigerian Savanna: Parameterization and validation of QUEFTS model.
Establishing balanced nutrient requirements for maize (Zea mays L.) in the Northern Nigerian Savanna is paramount to develop site-specific fertilizer recommendations to increase maize yield, profits of farmers and avoid negative environmental impacts of fertilizer use. The model QUEFTS (QUantitative Evaluation of Fertility of Tropical Soils) was used to estimate balanced nitrogen (N), phosphorus (P) and potassium (K) requirements for maize production in the Northern Nigerian Savanna. Data from on-farm nutrient omission trials conducted in 2015 and 2016 rainy seasons in two agro-ecological zones in the Northern Nigerian Savanna (i.e. Northern Guinea Savanna "NGS" and Sudan Savanna "SS") were used to parameterize and validate the QUEFTS model. The relations between indigenous soil N, P, and K supply and soil properties were not well described with the QUEFTS default equations and consequently new and better fitting equations were derived. The parameters of maximum accumulation (a) and dilution (d) in kg grain per kg nutrient for the QUEFTS model obtained were respectively 35 and 79 for N, 200 and 527 for P and 25 and 117 for K in the NGS zone; 32 and 79 for N, 164 and 528 for P and 24 and 136 for K in the SS zone; and 35 and 79 for N, 199 and 528 for P and 24 and 124 for K when the data of the two zones were combined. There was a close agreement between observed and parameterized QUEFTS predicted yields in each of the agro-ecological zone (R2 = 0.69 for the NGS and 0.75 for the SS). Although with a slight reduction in the prediction power, a good fit between the observed and model predicted grain yield was also detected when the data for the two agro-ecological zones were combined (R2 = 0.67). Therefore, across the two agro-ecological zones, the model predicted a linear relationship between grain yield and above-ground nutrient uptake until yield reached about 50 to 60% of the yield potential. When the yield target reached 60% of the potential yield (i.e. 6.0 t ha-1), the model showed above-ground balanced nutrient uptake of 20.7, 3.4 and 27.1 kg N, P, and K, respectively, per one tonne of maize grain. These results suggest an average NPK ratio in the plant dry matter of about 6.1:1:7.9. We concluded that the QUEFTS model can be widely used for balanced nutrient requirement estimations and development of site-specific fertilizer recommendations for maize intensification in the Northern Nigerian Savanna.


Kinetics of endosome acidification in mutant and wild-type Chinese hamster ovary cells.
Acidification of endocytic compartments is necessary for the proper sorting and processing of many ligands and their receptors. Robbins and co-workers have obtained Chinese hamster ovary (CHO) cell mutants that are pleiotropically defective in endocytosis and deficient in ATP-dependent acidification of endosomes isolated by density centrifugation (Robbins, A. R., S. S. Peng, and J. L. Marshall. 1983. J. Cell Biol. 96:1064-1071; Robbins, A. R., C. Oliver, J. L. Bateman, S. S. Krag, C. J. Galloway, and I. Mellman. 1984. J. Cell Biol. 99:1296-1308). In this and the following paper (Yamashiro, D. J., and F. R. Maxfield. 1987. J. Cell Biol. 105:2723-2733) we describe detailed studies of endosome acidification in the mutant and wild-type CHO cells. Here we describe a new microspectrofluorometry method based on changes in fluorescein fluorescence when all cellular compartments are equilibrated to the same pH value. Using this method we measured the pH of endocytic compartments during the first minutes of endocytosis. We found in wild-type CHO cells that after 3 min, fluorescein-labeled dextran (F-Dex) was in endosomes having an average pH of 6.3. By 10 min, both F-Dex and fluorescein-labeled alpha 2-macroglobulin (F-alpha 2M) had reached acidic endosomes having an average pH of 6.0 or below. In contrast, endosome acidification in the CHO mutants DTG 1-5-4 and DTF 1-5-1 was markedly slowed. The average endosomal pH after 5 min was 6.7 in both mutant cell lines. At least 15 min was required for F-Dex and F-alpha 2M to reach an average pH of 6.0 in DTG 1-5-4. Acidification of early endocytic compartments is defective in the CHO mutants DTG 1-5-4 and DTF 1-5-1, but pH regulation of later compartments on both the recycling pathway and lysosomal pathway is nearly normal. The properties of the mutant cells suggest that proper functioning of pH regulatory mechanisms in early endocytic compartments is critical for many pH-mediated processes of endocytosis.


A simple method of mapping atrial premature depolarizations triggering atrial fibrillation.
Atrial premature depolarizations (APDs) originating from focal sites, particularly the pulmonary veins (PV), may become triggers of atrial fibrillation (AF). Accurate mapping of APDs with conventional methods may be time consuming and expose the patient to unnecessary instrumentation of the left atrium. We hypothesized that the atrial activation sequence recorded using a simple system that includes an esophageal catheter and a custom-made 16-electrode catheter with two sets of floating electrodes eight in the coronary sinus and eight in the high right atrium) could be sufficient to localize the APDs. The study included 29 patients with frequent APDs and AF refractory to antiarrhythmic medications. The APD site of origin was confirmed with single-point sequential mapping techniques using the CARTO system ten patients) or by placement of multielectrode catheters in the right and left PV (19 patients). Of the 29 patients, 20 patients had a single APD focus; 8 patients had two different APD morphologies; and 1 patient had three APD foci. Mapping for ablation of the APD foci showed earliest activation in the left superior PV in 12 patients, right superior PV in 15 patients, right middle PV in 4 patients, right inferior PV in 1 patient, the lingular branch of the left superior PV in 2 patients, left inferior PV in 2 patients, and right atrium along the crista terminalis in 3 patients. The activation sequence and relative timing of the recordings obtained with our catheter configuration was highly predictive of right and left atrial origin and, more importantly, of right and left PV foci.


A genetic record of population isolation in pocket gophers during Holocene climatic change.
A long-standing question in Quaternary paleontology is whether climate-induced, population-level phenotypic change is a result of large-scale migration or evolution in isolation. To directly measure genetic variation through time, ancient DNA and morphologic variation was measured over 2,400 years in a Holocene sequence of pocket gophers (Thomomys talpoides) from Lamar Cave, Yellowstone National Park, Wyoming. Ancient specimens and modern samples collected near Lamar Cave share mitochondrial cytochrome b sequences that are absent from adjacent localities, suggesting that the population was isolated for the entire period. In contrast, diastemal length, a morphologic character correlated with body size and nutritional level, changed predictably in response to climatic change. Our results demonstrate that small mammal populations can experience the long-term isolation assumed by many theoretical models of microevolutionary change.


Curcumin administration suppress acetylcholinesterase gene expression in cadmium treated rats.
Curcumin, the main polyphenolic component of turmeric (Curcuma longa) rhizomes have been reported to exert anticholinesterase potential with limited information on how they regulate acetylcholinesterase (AChE) gene expression. Hence, this study sought to evaluate the effect of curcumin on cerebral cortex acetylcholinesterase (AChE) activity and their mRNA gene expression level in cadmium (Cd)-treated rats. Furthermore, in vitro effect of different concentrations of curcumin (1-5μg/mL) on rat cerebral cortex AChE activity was assessed. Animals were divided into six groups (n=6): group 1 serve as control (without Cd) and receive saline/vehicle, group 2 receive saline plus curcumin at 25mg/kg, group 3 receive saline plus curcumin 50mg/kg, group 4 receive Cd plus vehicle, group 5 receive Cd plus curcumin at 25mg/kg and group 6 receive Cd plus curcumin at 50mg/kg. Rats received Cd (2.5mg/kg) and curcumin (25 and 50mg/kg, respectively) by oral gavage for 7days. Acetylcholinesterase activity was measured by Ellman's method and AChE expression was carried out by a quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) assay. We observed that acute administration of Cd increased acetylcholinesterase activity and in addition caused a significant (P<0.05) increase in AChE mRNA levels in whole cerebral cortex when compared to control group. However, co-treatment with curcumin inhibited AChE activity and alters AChE mRNA levels when compared to Cd-treated group. In addition, curcumin inhibits rat cerebral cortex AChE activity in vitro. In conclusion, curcumin exhibit anti-acetylcholinesterase activity and suppressed AChE mRNA gene expression level in Cd exposed rats, thus providing some biochemical and molecular evidence on the therapeutic effect of this turmeric-derived compound in treating neurological disorders including Alzheimer's disease.


[Vaginal ultrasound follow-up of suspicious smears of the cervix uteri].
Using transvaginal sonography (TVS) it is possible to place the scanner probe very close to the organs of the female small pelvis. The near position allows the employment of a higher ultrasound frequency with a better resolving power and thus with more information about the examined structures. A clinical study was performed to investigate whether it is possible to gain some more information about alterations of the uterine cervix surface, after a suspicious cytology result, by applying TVS. A total 59 women with a pathological swab were examined, followed by conisation (using an operating knife) and a D & C. Histological examination showed that in 21 cases with sonographically not suspicious results, 10 cancers in situ and 1 early invasive squamous cell cancer could not be detected. The determined sensitivity of the method was 30.8%; the specificity was 59%. Concluding from the result TVS can be applied only with restriction concerning detection of beginning cancer of the uterine cervix. A statistical significance of vaginosonographical results compared with the suspicious smear results could not be established.


Genetic, physiological and comparative genomic studies of hypertension and insulin resistance in the spontaneously hypertensive rat.
We previously mapped hypertension-related insulin resistance quantitative trait loci (QTLs) to rat chromosomes 4, 12 and 16 using adipocytes from F2 crosses between spontaneously hypertensive (SHR) and Wistar Kyoto (WKY) rats, and subsequently identified Cd36 as the gene underlying the chromosome 4 locus. The identity of the chromosome 12 and 16 genes remains unknown. To identify whole-body phenotypes associated with the chromosome 12 and 16 linkage regions, we generated and characterised new congenic strains, with WKY donor segments introgressed onto an SHR genetic background, for the chromosome 12 and 16 linkage regions. We found a >50% increase in insulin sensitivity in both the chromosome 12 and 16 strains. Blood pressure and left ventricular mass were reduced in the two congenic strains consistent with the congenic segments harbouring SHR genes for insulin resistance, hypertension and cardiac hypertrophy. Integrated genomic analysis, using physiological and whole-genome sequence data across 42 rat strains, identified variants within the congenic regions in Upk3bl, RGD1565131 and AABR06087018.1 that were associated with blood pressure, cardiac mass and insulin sensitivity. Quantitative trait transcript analysis across 29 recombinant inbred strains showed correlation between expression of Hspb1, Zkscan5 and Pdgfrl with adipocyte volume, systolic blood pressure and cardiac mass, respectively. Comparative genome analysis showed a marked enrichment of orthologues for human GWAS-associated genes for insulin resistance within the syntenic regions of both the chromosome 12 and 16 congenic intervals. Our study defines whole-body phenotypes associated with the SHR chromosome 12 and 16 insulin-resistance QTLs, identifies candidate genes for these SHR QTLs and finds human orthologues of rat genes in these regions that associate with related human traits. Further study of these genes in the congenic strains will lead to robust identification of the underlying genes and cellular mechanisms.


Time-Driven Activity-Based Costing in Fracture Care: Is This a More Accurate Way to Prepare for Alternative Payment Models?
To use surgical treatment of isolated ankle fractures as a model to compare time-driven activity-based costing (TDABC) and our institution's traditional cost accounting (TCA) method to measure true cost expenditure around a specific episode of care. Level I trauma center ankle fractures treated between 2012 and 2016 were identified through a registry. Inclusion criteria were age greater than 18 years and same-day ankle fracture operation. Exclusion criteria were pilon fractures, vascular injuries, soft-tissue coverage, and external fixation. Time for each phase of care was determined through repeated observations. The TCA method at our institution uses all hospital costs and allocates them to surgeries using a relative value method. A total of 35 patients met the inclusion/exclusion criteria, 18 were men and 17 were women. Age at time of surgery was 47 ± 15 years. Time from injury to surgery was 10 ± 4 days. Operative time was 86 ± 30 minutes, Post-anesthesia care unit (PACU) time was 87 ± 27 minutes, and secondary recovery time was 100 ± 56 minutes. Average cost was significantly lower for the TDABC method ($2792 ± 734) than the TCA method ($5782 ± 1348) (P < 0.001). There was no difference between methods for implant cost ($882 ± 507 for Traditional Accounting (TA) and $957 ± 651 for TDABC, P = 0.593). TCA produced a significantly greater cost (P < 0.01) in every other category. As orthopaedics transitions to alternative payment models, accurate costing will become critical to maintaining a successful practice. TDABC may provide a better estimate of the cost of the resources necessary to treat a patient.