Kinetics of bicarbonate transport in human red blood cell membranes at body temperature.
We studied unidirectional [14C]HCO3- efflux from human resealed red cell ghosts with 1 mM acetazolamide under self-exchange conditions at pH = pH(i = o) 7.4-9.0 and 0-38 degrees C by means of the Millipore-Swinnex and continuous flow tube filtering techniques. 14CO2 loss from cells to efflux medium and further to the atmosphere was insignificant. [14C]HCO3- efflux was determined at pH 7.8, 38 degrees C under symmetric variation of the HCO3- concentrations (C(i = o)), and asymmetric conditions: C(i) varied, C(o) constant, or C(o) varied, C(i) constant. MM-fit, Jeff = Jmaxeff x C x (C + K1/2)-1, used to describe the concentration dependence of Jeff,o when only C(o) varied, yields at C(i) = 50 mM: K1/2o = 3.8 mMJ, Jmaxeff.o = 20 nmol cm-2 s-1; at C(i) = 165 mM: K1/2o = 10 mM, Jmaxeff.o = 32 nmol cm-2 s-1. When C(i) varied, noncompetitive self inhibition by HCO3- binding (inhibitor constant K1) to an intracellular site was included (MS-fit). Under conditions of (a) symmetry: C(i = o) = 9-600 mM, K1/2s = 173 mM, K1 = 172 mM, and Jmaxeff,s = 120 nmol cm-2 s-1, (b) asymmetry: C(o) = 50 mM, K1/2i = 116 mM, K1 = 136 mM, and Jmaxeff,i = 92 nmol cm-2 s-1. All flux parameters accord with the ping-pong model for anion exchange. The data for C(i) < 200 mM also fit well to the MM equation, but K1/2 and Jmaxeff are different from the MS-fit and are inconsistent with the ping-pong model. Thus, self-inhibition (MS-fit) must be included even at low concentrations. As at 0 degree C, the system is asymmetric: 8-10 times more unloaded transport sites face inward than outward when C(i = o). Jeff,s was not mono-exponentially dependent on temperature at 0-38 degrees C, indicating that the transmembrane anion transport is controlled by several rate constants with different temperature dependencies. Jeff,s was not significantly affected by increasing pH(i = o) from 7.4 to 7.8, but it decreased by 50% when pH was raised to 9.0.


A modified technique to extract fractured femoral stem in revision total hip arthroplasty: A report of two cases.
The removal of well-fixed broken femoral component and cement mantle can be extremely demanding, time consuming and potentially damaging to the host bone. Different methods have been described to extract broken femoral stem yet this remains one of the most challenging prospect to the revision hip surgeon. The authors present two cases underwent a modified sliding cortical window technique utilising a tungsten carbide drill, Charnley pin retractor and an orthopaedic mallet to aid extraction of a fractured cemented femoral stem in revision total hip arthroplasty. The modified technique offers a simple and controlled method in extracting a well fixed fractured cemented femoral stem. It has the advantage of retaining the cement mantle with subsequent good seal of the femoral cortical window secured with cable ready system. Furthermore, tungsten carbide drill bit and Charnley pin retractor are relatively readily available to aid the extraction of the broken stem. Finally, it yields the option of implanting a standard femoral stem and obviates the need for bypassing the cortical window with long revision femoral component. Fractured femoral stem is a rare yet a complex and very demanding prospect to both patients and hip surgeons. The sliding cortical window technique utilising tungsten carbide drill and Charnley pin retractor is technically easy and most importantly; preserves host bone stock with cement-in-cement revision hip arthroplasty. We believe this technique can be added to the armamentarium of revision hip surgeon when faced with the challenge of extracting a fractured cemented femoral stem.


A fluorescence-based high-throughput screening assay for the identification of T-type calcium channel blockers.
T-type voltage-gated Ca(2+) channels have been implicated in contributing to a broad variety of human disorders, including pain, epilepsy, sleep disturbances, cardiac arrhythmias, and certain types of cancer. However, potent and selective T-type Ca(2+) channel modulators are not yet available for clinical use. This may in part be due to their unique biophysical properties that have delayed the development of high-throughput screening (HTS) assays for identifying blockers. One notable challenge is that at the normal resting membrane potential (V(m)) of cell lines commonly utilized for drug screening purposes, T-type Ca(2+) channels are largely inactivated and thus cannot be supported by typical formats of functional HTS assays to both evoke and quantify the Ca(2+) channel signal. Here we describe a simple method that can successfully support a fluorescence-based functional assay for compounds that modulate T-type Ca(2+)channels. The assay functions by exploiting the pore-forming properties of gramicidin to control the cellular V(m) in advance of T-type Ca(2+) channel activation. Using selected ionic conditions in the presence of gramicidin, T-type Ca(2+) channels are converted from the unavailable, inactivated state to the available, resting state, where they can be subsequently activated by application of extracellular K(+). The fidelity of the assay has been pharmacologically characterized with sample T-type Ca(2+) channel blockers whose potency has been determined by conventional manual patch-clamp techniques. This method has the potential for applications in high-throughput fluorometric imaging plate reader (FLIPR(R), Molecular Devices, Sunnyvale, CA) formats with cell lines expressing either recombinant or endogenous T-type Ca(2+) channels.


Epidemiological and clinical aspects of leprosy in Indian armed forces.
Epidemiological and clinical aspects of leprosy in 1911 cases of the disease in armed forces personnel were studied. Typewise distribution of cases was Tuberculoid 53.53%, Lepromatous 20.57%, Indeterminate 8.74%, Borderline 11.67% and Polyneuritic 5.49%. Maximum leprosy cases occurred in those belonging to Uttar Pradesh 17.11%. Maximum number of cases were detected (88.55%) in age group 20-39 years. Incidence of leprosy increased with increased years of service. 11.82% patients were illiterate. 89.85% patients earned Rs. 200 to 499 per month. 56.08% patients had no landed property. Houses of 47.29% cases were located in congested areas. 68.23% patients had to support larger families. In 95.94% cases no family members were examined for leprosy. Diet, smoking and alcohol appear to have had no relation to disease in cases studied. Clinical presentation of cases was classical and typespecific. Skin eruption and loss of sensation were the commonest symptoms. Leprosy lesions were detected on almost all parts of body. Thermal sensation was the commonest modality lost. Ulnar, lateral popliteal and greater auricular were the frequently affected nerves. Amongst complications paralytic deformities were common (16.09%). 545 complications were detected in 1911 cases. While 84.29% patients had put in more than 4 years of service, the source of infection was known only in 0.57% positively (intrafamilial).


The Portuguese generic medicines market: a policy analysis.
This study aims to conduct a descriptive analysis of the policy environment surrounding the generic medicines retail market in Portugal. The policy analysis focuses on supply-side measures (i.e. market access, pricing, reference-pricing and reimbursement of generic medicines) and demand-side measures (i.e. incentives for physicians to prescribe, for pharmacists to dispense and for patients to use generic medicines). The policy analysis was based on an international literature review. Also, a simulation exercise was carried out to compute potential savings from substituting generic for originator medicines in Portugal using IMS Health data. Portugal has developed a successful generic medicines market by increasing reimbursement of generic medicines (until October 2005), by introducing a reference-pricing system, by encouraging physicians to prescribe by international non-proprietary name (INN), and by allowing generic substitution by pharmacists. However, the development of the generic medicines market has been hindered by the existence of copies, pricing regulation, certain features of the reference-pricing system, weak incentives for physicians to prescribe generic medicines and a financial disincentive for pharmacists to dispense generic medicines. Increased generic substitution would be expected to reduce public expenditure on originator medicines by 45%. The development of the Portuguese generic medicines market has mainly been fuelled by supply-side measures. To support the further expansion of the market, policy makers need to strengthen demand-side measures inciting physicians to prescribe, pharmacists to dispense and patients to use generic medicines.


Mortality and predictors of mortality in rheumatoid arthritis--a role for mannose-binding lectin?
Patients with rheumatoid arthritis (RA) have increased overall and cardiovascular mortality. Mannose-binding lectin (MBL) may play differentiated roles in the pathogenesis of RA. We had observed that high serum levels of MBL increased the risk of ischemic heart disease in patients with RA. In this followup study we describe the mortality in a cohort of 229 Danish patients with RA. We examine if previously reported factors and MBL influence the risk of overall death and death due to cardiovascular disease. Known predictors of RA mortality were assessed. MBL extended genotypes (YA/YA, YA/XA, XA/XA, YA/YO, XA/YO, YO/YO) were determined; MBL serum concentrations were measured. The vital status and causes of death were assessed in a prospective study. The median followup was 10.3 years. The overall risk of death was 4% per year. Comparing mortality in the RA cohort with mortality in an age- and sex-matched cohort based on the general Danish population, we found significantly increased overall mortality [standardized mortality ratio (SMR) 1.5, 95% CI 1.2-1.9, and cardiovascular mortality (SMR 1.7, 95% CI 1.3-2.6)]. In multivariate analysis, significant predictors of overall death were extraarticular manifestations, positive rheumatoid factor, increased C-reactive protein (CRP), poor nutritional state, and serum MBL. Predictors of cardiovascular death were Health Assessment Questionnaire score, increased CRP, poor nutritional state, and the high-producing MBL genotype YA/YA. Both overall and cardiovascular mortality were increased in Danish patients with RA. In our cohort, states of high MBL production and several previously reported factors contributed significantly to this increased risk of overall death and cardiovascular death.


Absorption enhancer approach for protein delivery by various routes of administration: a rapid review.
As bioactive molecules, peptides and proteins are essential in living organisms, including animals and humans. Defects in their function lead to various diseases in humans. Therefore, the use of proteins in treating multiple diseases, such as cancers and hepatitis, is increasing. There are different routes to administer proteins, which have limitations due to their large and hydrophilic structure. Another limitation is the presence of biological and lipophilic membranes that do not allow proteins to pass quickly. There are different strategies to increase the absorption of proteins from these biological membranes. One of these strategies is to use compounds as absorption enhancers. Absorption enhancers are compounds such as surfactants, phospholipids and cyclodextrins that increase protein passage through the biological membrane and their absorption by different mechanisms. This review focuses on using other absorption enhancers and their mechanism in protein administration routes.


Complementary therapies in substance use recovery with pregnant women and girls.
Supporting women and girls who are pregnant and experiencing substance use challenges is a high priority for researchers, clinicians, and policymakers. Complementary therapies (CTs) can be effective forms of treatment in some contexts and populations; however, research on the use of CTs in substance use recovery with pregnant women and girls is scarce. To fill this gap, we conducted a mixed methods study using survey data collected at a women's recovery center in Canada. Our objectives were to describe CTs provided at the program; identify what CTs are perceived by participants as most/least positive; and explore factors that may impact participant experiences with CTs. We analyzed feedback responses from 255 women and girls (Mage = 27.5 years, range 15-64) using Pearson chi-square tests, logistic regression, and inductive content analysis. The most frequently provided CTs were yoga, energy-related activities (e.g. reiki, reflexology), and meditation. Among the most common CTs, participants provided the highest endorsements for massage and physical activity, and the lowest endorsements for yoga and drumming. Across CTs, whether participants looked forward to an activity contributed significantly to whether they found it helpful, would like to do it again, and planned to continue engaging in the activity after leaving the program. Four broad contextual factors were identified that may impact experiences and perspectives about CTs: (1) goodness of fit, (2) self-awareness, (3) growth, and (4) healing and holistic wellbeing. This study provides novel evidence on the potential impacts of CTs in substance use treatment for pregnant women and girls, and important contextual factors to consider when implementing these approaches.


Nanocrystalline TiO2/Ti3C2Tx MXene composites with a tunable work function prepared using atmospheric pressure oxygen plasma.
Composites of TiO2 and Ti3C2Tx MXene are of great interest because they combine superior TiO2 photocatalytic activity with excellent MXene conductivity. As these composites have conventionally been prepared using methods requiring high temperatures, a process for producing similar materials at reduced temperature could be beneficial for applications in flexible and printed electronics. Therefore, we used low-temperature dielectric barrier discharge to develop a method for forming crystalline TiO2 by treating Ti3C2Tx MXene surfaces with high-power-density oxygen plasma comprising various energetic and reactive oxygen species, which oxidize MXene surfaces and form TiO2 nanoparticles on disordered graphitic carbon sheets within a few seconds. Scanning electron microscopy was used to observe the plasma-induced morphological changes to elucidate the TiO2 formation mechanism. The MXene surface chemistry was studied in detail using X-ray photoelectron spectroscopy and ab initio modelling. The crystalline phase of TiO2 was further studied using transmission electron microscopy and Raman spectroscopy. The results presented here suggest the formation of small anatase nanoparticles on the surface of MXenes within just seconds of plasma exposure. Nanoparticles grew with prolonged plasma treatment and a transition from anatase to rutile was observed. Considering that the temperature of plasma was always below 70 °C, the oxygen plasma process for the preparation of TiO2/Ti3C2Tx composites is an excellent candidate for application on temperature-sensitive substrates in flexible and printed electronics.


Antagonism of sexual behavior in female rats by ventromedial hypothalamic implants of antiestrogen.
The present experiments sought to identify brain regions in which implants of an antiestrogen would antagonize the ability of a systemic estradiol treatment to activate sexual behavior in female rats. In experiment 1, ovariectomized female rats were implanted subcutaneously with 5-mm Silastic capsules containing a 5% concentration of estradiol and injected with 500 micrograms progesterone 2 days later, 4-5 h before testing for sexual behavior. Bilateral intracranial implants of 1% crystalline concentrations of the high-affinity antiestrogens monohydroxytamoxifen (TAM) or keoxifene placed into the ventromedial nucleus of the hypothalamus (VM) 24 h prior to estradiol treatment significantly reduced lordosis responsivity compared with control females receiving empty cannulae. Implants of 1% TAM into the medial preoptic area or medial amygdala 24 h prior to estradiol that no significant effect on lordosis. Similarly, implants of 1% TAM into the VM 12 h after estradiol had no effect on lordosis. In experiment 2, lordosis was activated by subcutaneous implants of Silastic capsules containing 1% estradiol plus 500 micrograms progesterone. In this experiment, implants of 1% TAM into the VM 24 h prior to estradiol significantly reduced lordosis only if both cannulae tips were in, or adjacent to, the VM. Females receiving intracranial 1% TAM, but whose cannulae (even unilaterally) were outside the VM, had levels of lordosis equivalent to those of control females. Increasing the concentration of intracranial TAM to 10% virtually eliminated lordosis in females with bilateral implants in the VM, whereas females receiving intracranial 10% TAM in the region of, but outside, the VM showed no evidence of a lordosis deficit.(ABSTRACT TRUNCATED AT 250 WORDS)


Analysis of phases of contraction during graded acute myocardial ischemia.
The quantitative effects of graded ischemia on the phases of contraction in ischemic and nonischemic myocardium were studied in 10 open-chest dogs. During progressive coronary stenosis, an index of the overall severity of ischemia, total myocardial blood flow deficit (TMFD), was measured using microspheres. Segment shortening (% delta L) during the isovolumic and ejection phases of systole in the ischemic and nonischemic zones was obtained with ultrasonic crystals. In the ischemic zone, total systolic and ejection phase % delta L decreased progressively as stenosis increased; isovolumic phase % delta L showed segment elongation (bulging) during mild ischemia, which was unchanged as ischemia became severe. Total % delta L in the nonischemic zone increased progressively with increasing stenosis, reaching 132 +/- 20% of the control value when TMFD was 24.8 +/- 6.5% of total expected left ventricular (LV) myocardial blood flow. The increase in total % delta L was entirely attributable to increased ejection % delta L and correlated with indices of preload (multivariate r = 0.65) and afterload (multivariate r = 0.62). Multiple linear regression analysis revealed that augmented contraction in the nonischemic zone was best described (multivariate r = 0.94) by a combination of four hemodynamic variables estimating LV preload and afterload, which correlated in turn with the overall severity of ischemia. Thus systolic bulging in the ischemic zone is primarily an isovolumic phase phenomenon that develops initially with mild ischemia. Little additional change occurs in the isovolumic phase as ischemic severity increases, although further reductions in shortening occur in the ejection phase.(ABSTRACT TRUNCATED AT 250 WORDS)


Instability of the cervical spine after decompression in patients who have Arnold-Chiari malformation.
Stability of the cervical spine was studied in two groups of children who had myelomeningocele. Group I consisted of twenty children who had an Arnold-Chiari Type-II malformation in whom a suboccipital craniectomy (partial occipital craniectomy through the suboccipital route) and cervical laminectomy was done to decompress the brain stem. The average duration of follow-up in this group (excluding one patient) was 4.7 years (range, 2.3 to 10.4 years) after the operation. Group II consisted of twenty children who had myelomeningocele but had not had an operation for decompression. Lateral radiographs of the cervical spine in flexion and extension showed no instability between the occiput and atlas or between the atlas and axis in either group. In contrast, translation between the second and third cervical vertebrae averaged four millimeters in Group I and one millimeter in Group II (p less than 0.01), and angulation between the third and fourth cervical vertebrae averaged 17 degrees in Group I and 6 degrees in Group II (p less than 0.01). Nineteen of the twenty patients in whom a suboccipital craniectomy and cervical laminectomy (Group I) had been done had instability of the cervical spine.


Endocytosis and degradation of chondroitin sulphate by liver endothelial cells.
Intravenously administered chondroitin sulphate, chemically labelled by [3H]acetylation of partially deacetylated polysaccharide, was taken up and degraded by the non-parenchymal cells of the liver. Studies using primary monolayer cultures of pure Kupffer cells, liver endothelial cells and parenchymal cells revealed that [3H]chondroitin sulphate was taken up and degraded by the liver endothelial cells only. Binding studies at 4 degrees C with [3H]chondroitin sulphate and 125I-chondroitin sulphate proteoglycan indicated that the glycosaminoglycan and the proteoglycan are recognized by the same binding sites on the liver endothelial cells. The ability of hyaluronic acid to compete with the labelled ligands for binding suggested that the binding site is identical with the recently described hyaluronate receptor on the liver endothelial cells [Smedsrød, Pertoft, Eriksson, Fraser & Laurent (1984) Biochem. J. 223, 617-626]. Fluorescein-labelled chondroitin sulphate proteoglycan accumulated in perinuclear vesicles of the liver endothelial cells, indicating that the proteoglycan is internalized and transported to the lysosomes. The finding that [3H]chondroitin sulphate and 125I-chondroitin sulphate proteoglycan were degraded by the liver endothelial cells to low-molecular-mass radioactive products suggested that both the polysaccharide chain and the core protein were catabolized by the cells.


Probing dimerization and intraprotein fluorescence resonance energy transfer in a far-red fluorescent protein from the sea anemone Heteractis crispa.
Proteins from Anthozoa species are homologous to the green fluorescent protein (GFP) from Aequorea victoria but with absorption/emission properties extended to longer wavelengths. HcRed is a far-red fluorescent protein originating from the sea anemone Heteractis crispa with absorption and emission maxima at 590 and 650 nm, respectively. We use ultrasensitive fluorescence spectroscopic methods to demonstrate that HcRed occurs as a dimer in solution and to explore the interaction between chromophores within such a dimer. We show that red chromophores within a dimer interact through a Forster-type fluorescence resonance energy transfer (FRET) mechanism. We present spectroscopic evidence for the presence of a yellow chromophore, an immature form of HcRed. This yellow chromophore is involved in directional FRET with the red chromophore when both types of chromophores are part of one dimer. We show that by combining ensemble and single molecule methods in the investigation of HcRed, we are able to sort out subpopulations of chromophores with different photophysical properties and to understand the mechanism of interaction between such chromophores. This study will help in future quantitative microscopy investigations that use HcRed as a fluorescent marker.


The role of integrins in melanoma: a review.
Integrins are the major family of cell adhesion receptors in humans and essential for a wide range of normal physiology, including formation and maintenance of tissue structure integrity, cell migration, proliferation, and differentiation. Integrins also play a prominent role in tumor growth and metastasis. Translational research has tried to define the contribution of integrins to the phenotypic aggressiveness of melanoma because such knowledge is clinically useful. For example, differential expression of integrins in primary cutaneous melanoma can be used to distinguish indolent from aggressive, prometastatic melanoma. Recent studies have shown that gene expression-based testing of patient-derived melanoma tissue is feasible, and molecular tests may fully replace interventional surgical methods such as sentinel lymph node biopsies in the future. Because of their central role in mediating invasion and metastasis, integrins are likely to be useful biomarkers. Integrins are also attractive candidate targets for interventional therapy. This article focuses on the role of integrins in melanoma and highlights recent advances in the field of translational research.


Elective genetic testing: Genetics professionals' perspectives and practices.
Elective genetic testing (EGT) to identify disease risk in individuals who may or may not meet clinical criteria for testing is increasingly being offered in clinical practice. However, little is known about how EGT is currently implemented and how genetics professionals perceive this type of testing. We conducted a mixed-methods survey study to evaluate genetics professionals' perspectives and attitudes about EGT and describe the current landscape of EGT practices in the United States (U.S.) and Canada. Six clinical geneticists and 131 genetic counselors responded to the online survey, among whom 44% reported offering EGT in their practice. Over 84% of survey respondents agreed that EGT may improve health outcomes and understanding of genotype-phenotype correlations, and 85% agreed that potential risks include result misinterpretation and contribution to economic health disparities. Though most respondents felt comfortable providing pretest (77%) and post-test (86%) counseling for EGT, lack of provider resources (such as time and personnel) and prioritization of diagnostic testing were cited most frequently in free-text responses as reasons for not offering EGT. Of those offering EGT, 88% reported positive overall experiences. Qualitative analysis of open-ended questions identified benefits of EGT as expanding access to genetic testing, providing potential health benefits, and providing psychological benefits for patients. Disadvantages included prohibitive costs, limited clinical utility, and strain on resources. Overall, we found that genetics providers perceive both potential benefits and harms of EGT and that those offering this testing had generally positive experiences, although ethical reservations and practical limitations exist.


Factors Associated with Apical Periodontitis: A Multilevel Analysis.
Ignoring the cluster effect is a common statistical oversight that is also observed in endodontic research. The aim of this study was to explore the use of multilevel modeling in investigating the effect of tooth-level and patient-level factors on apical periodontitis (AP). A random sample of digital panoramic radiographs from the database of a dental hospital was evaluated. Two calibrated examiners (κ ≥ 0.89) assessed the technical quality of the root fillings and the radiographic periapical health status by using the periapical index. Descriptive statistical analysis was carried out, followed by multilevel modeling by using tooth-level and patient-level predictors. Model fit information was obtained, and the findings of the best-fit model were reported. A total of 6409 teeth were included in the analysis. The predicted probability of a tooth having AP was 0.42%. There was a statistically significant variability between patients (P < .05). Approximately 53.16% of the variability was accounted for by the patients, leaving 46.84% of the variability to teeth or other factors. Posterior tooth, inadequate root filling, and age were found to be significantly associated with AP (P < .05). Multilevel modeling is a valid and efficient statistical method in analyzing AP data. The predicted probability of a tooth having AP was generally small, but there was great variation between individuals. Posterior teeth and those with poor quality root filling were found to be more frequently associated with AP. On the patient level, advancing age was a factor significantly associated with AP.


Highway to health: on-line health education.
The TeleHealth Project is a model telecommunications program that provides health education and consultation on the Internet. The TeleHealth Project delivers health information to rural ninth-grade students and their communities on the North Carolina Information Highway through interactive distance learning. The "trip" is a two-way adventure for students, educators, and health professionals. The Eastern Area Health Education Center (EAHEC), associated with East Carolina University (ECU) in Greenville, North Carolina, produces the TeleHealth Program. This collaborative model was built on prior successful relationships between the school system and EAHEC, and university faculty who had worked on interdisciplinary projects related to adolescents and the professional preparation of health providers in school settings.


Different effects of in vivo ouabain and digoxin on renal artery function and blood pressure in the rat.
To investigate vascular mechanisms in hypertension, we isolated renal arterial rings from rats with ouabain-dependent hypertension and studied their function. In rats infused with ouabain for 5 weeks, systolic and mean blood pressures (BP) were increased relative to controls. Contractions evoked by high KCl solutions were greater in rings from ouabain-infused rats whereas the threshold concentrations and EC50s for KCl and the peak caffeine contractures were not different. KCl contractures were not affected by 5 microM prazosin. Phenylephrine contractures were increased marginally in ouabain-infused rats, while acetylcholine-induced relaxation was normal. In vitro superfusion of rings with 10 nM ouabain or digoxin did not affect the measured parameters. Plasma ouabain, BP, and all evoked responses were normal one week following interruption of the ouabain infusion. In a second study, BP increased in ouabain (15 microg/kg/day, n= 23), but not digoxin (30 microg/kg/day, n=12), or vehicle-infused (n=16) rats. KCl contractures were greater in rings from ouabain-but decreased in rings from digoxin-infused rats, respectively and correlated with systolic and mean BP (r=0.69, n=30, p<0.005). Peak caffeine (25 mM) responses were similar but the area under the contraction was reduced in the vessels from ouabain-infused rats and correlated inversely with MBP (r=-0.47, n=33, p<0.02). We conclude that a voltage-dependent component of tone in the rat renal artery is reversibly and specifically augmented by in vivo administration of ouabain whereas it is diminished by in vivo digoxin. Vascular production of and response to nitric oxide does not appear to be impaired in the ouabain model. Alterations of intracellular Ca2+ storage and Ca2+ influx in response to in vivo ouabain may underlie the increase in renal vascular resistance and hypertension in this model. The opposite effects of ouabain and digoxin on the hemodynamic and vascular parameters in this study indicate that these agents have novel mechanisms of action in vivo that may not be mediated exclusively by sodium-potassium pumps.


Oxygen is required for the L-cysteine-mediated decomposition of protein-bound dinitrosyl-iron complexes.
Increasing evidence suggests that iron-sulfur proteins are the primary targets of nitric oxide (NO). Exposure of Escherichia coli cells to NO readily converts iron-sulfur proteins to protein-bound dinitrosyl-iron complexes (DNICs). Although the protein-bound DNICs are stable in vitro under aerobic or anaerobic conditions, they are efficiently repaired in aerobically growing E. coli cells even without new protein synthesis. The cellular repair mechanism for the NO-modified iron-sulfur proteins remains largely elusive. Here we report that, unlike aerobically growing E. coli cells, starved E. coli cells fail to reactivate the NO-modified iron-sulfur proteins. Significantly, the addition of L-cysteine, but not other related biological thiols, results in decomposition of the protein-bound DNICs in starved E. coli cells and in cell extracts under aerobic conditions. However, under anaerobic conditions, L-cysteine has little or no effect on the protein-bound DNICs in starved E. coli cells or in vitro, suggesting that oxygen is required for the L-cysteine-mediated decomposition of the protein-bound DNICs. Additional studies reveal that L-cysteine is able to release the DNIC from the protein and bind to it, and the L-cysteine-bound DNICs are rapidly disrupted by oxygen, resulting in the eventual decomposition of the protein-bound DNICs under aerobic conditions.


Treatment of early epithelial ovarian cancer with chemotherapy and abdominopelvic radiotherapy: results of a prospective treatment protocol.
To test the hypothesis that the combination of adjuvant chemotherapy and abdominopelvic radiation (APRT) improves the outcome of patients with early ovarian cancer compared to treatment with APRT alone. Between 1991 and 1994, 93 patients with Stage I to III, optimally cytoreduced, invasive, epithelial ovarian cancer were treated with sequential chemotherapy and APRT. Treatment was assigned using a prognostic classification that was derived from previous cohorts of patients. Low-risk patients (n = 9) received APRT alone, intermediate-risk patients (n = 66) received two courses of cisplatin followed by APRT, and high-risk patients (n = 18) received 6 courses of cisplatin and cyclophosphamide followed by APRT. Disease recurred in 22 patients, and was confined to the pelvis or abdomen in 15. Nine patients died and the remainder were alive with disease after receiving salvage chemotherapy. The 3-year disease-free and overall survivals were 78% and 91%, respectively. The prognostic classification used to assign treatment was the only factor that predicted disease-free survival (83% and 59% at 3 years for low/intermediate- and high-risk patients, respectively; p = 0.03). There was no detectable difference in outcome between the present series and an historical control group treated with APRT alone. Treatment was well tolerated and only 2 patients (2.5%) developed serious complications. APRT is an effective adjuvant treatment for carefully selected patients with early ovarian cancer. The addition of chemotherapy as used in this study to APRT does not significantly improve outcome compared to APRT alone.


[Endolymphatic chemotherapy of cervix neoplasms].
In the gynecological department of Rostov Oncological Institute endolymphatic polychemotherapy with a preliminary determination of tumor sensitivity to chemotherapy was employed in 54 patients with cervical cancer, 3 of them in stage III and IV having contraindications to radiotherapy. A combination of chemical substances and the dosage of single administration were as follows: thio-TEPA--100--150 mg + methotrexate 75--100 mg; thio-TEPA--100--150 mg + 5-fluoruracil--1000 mg; cyclophosphane--2000 mg + methotrexate--75--100 mg; cyclophosphane--1600 mg + methotrexate--75--100 mg + 5-fluoruracil--750 mg. Lymphoinfusion was made in the lower extremity, and after 7--10 days it was repeated in the second one. Visual disappearance of the tumor was noted in 17 patients, lessening of the tumor size by two-thirds--in 16, a 1/2 decrease--in 20. Among 23 patients in stage III after the treatment parametrial infiltrations disappeared entirely in 11 cases and only in pelvic walls--in 12. In 40 cases an extensive extirpation of the uterus with adnexa was performed. In all cases degenerative changes in tumor tissues were observed, whereas in 7 cases no tumor cells were found.


Cardiovascular and metabolic risk profiles in young and old patients with type 2 diabetes.
Young patients (aged < 40 years) with type 2 diabetes (T2D) have a high lifetime risk of developing cardiovascular disease (CVD). However, little is known about the CVD risk profile of this cohort in the UK primary care setting. To determine CVD risk profile of young patients with T2D without CVD compared to older (aged >40 years) subjects. A cross-sectional study using The Health Improvement Network (THIN) database, which contains anonymized patient information from more than 300 general practices throughout England and Wales. T2D subjects above the age of 18 years without previous CVD and not on lipid or blood pressure lowering therapy were randomly selected. Data on glycaemic control and CVD risk factors [weight, body mass index (BMI), lipid profile] were collected. A total of 49,919 patients with T2D were identified, of whom 2756 (0.5%) and 47,163 (99.5%) were aged below and above 40 years, respectively. Despite being at least 30 years younger (mean age: early vs. later onset; 33.8 vs. 66.9 years, P < 0.001), the proportions of adverse CVD risk profiles for young patients were similar to the older cohort with T2D. For young vs. old patients: the prevalence of BMI >25: 84.4% vs. 85.3%, P = 0.77; total cholesterol >4 mmol/l: 53.4% vs. 53.8%, P = 0.76; systolic hypertension: 58.2 vs. 58.4%, P = 0.36 and diastolic hypertension: 28.1 vs. 28.5%, P = 0.73). Glycaemic controls were similarly suboptimal between the two groups (mean HbA1c: young vs. old; 7.6% vs. 7.5%, P = 0.49). The prevalence of risk factor clustering were also similar between young vs. old patients with T2D. Young T2D subjects possess risk factors that confer high lifetime risk for macrovascular complications, and therefore merits aggressive cardioprotective treatment.


Rumination and Impaired Prospective Memory.
Prospective memory (PM), remembering to remember, is crucial to everyday functioning. Understanding factors associated with PM impairments is thus important. One likely factor is rumination: a common cognitive process comprising repetitive self-focused thoughts. We investigated whether rumination is associated with impaired PM, and whether any associated impairment is exacerbated with negative stimuli. A sentence-rating task with sentences varying in valence was used with embedded PM cues in a non-clinical sample (N = 60). State rumination, two trait rumination subtypes (reflective pondering and brooding), and mood were measured in relation to PM cue detection and response times. Results showed that state rumination was associated with impaired PM cue detection and slower response times to PM cues embedded in negative sentences (not positive or neutral). Trait brooding (not reflective pondering) was associated with slower PM response times. These findings indicate that state rumination and trait brooding are associated with dissociable PM impairments.


Computational and experimental characterization of physically clustered simple sequence repeats in plants.
The type and frequency of simple sequence repeats (SSRs) in plant genomes was investigated using the expanding quantity of DNA sequence data deposited in public databases. In Arabidopsis, 306 genomic DNA sequences longer than 10 kb and 36,199 EST sequences were searched for all possible mono- to pentanucleotide repeats. The average frequency of SSRs was one every 6.04 kb in genomic DNA, decreasing to one every 14 kb in ESTs. SSR frequency and type differed between coding, intronic, and intergenic DNA. Similar frequencies were found in other plant species. On the basis of these findings, an approach is proposed and demonstrated for the targeted isolation of single or multiple, physically clustered SSRs linked to any gene that has been mapped using low-copy DNA-based markers. The approach involves sample sequencing a small number of subclones of selected randomly sheared large insert DNA clones (e.g., BACs). It is shown to be both feasible and practicable, given the probability of fortuitously sequencing through an SSR. The approach is demonstrated in barley where sample sequencing 34 subclones of a single BAC selected by hybridization to the Big1 gene revealed three SSRs. These allowed Big1 to be located at the top of barley linkage group 6HS.


Enhancement of drug-specific lymphocyte proliferation using CD25(hi)-depleted CD3(+) effector cells.
The lymphocyte transformation test (LTT) is used for in vitro diagnosis of drug hypersensitivity reactions. While its specificity is over 90%, sensitivity is limited and depends on the type of reaction, drug and possibly time interval between the event and analysis. Removal of regulatory T cells (Treg/CD25(hi)) from in vitro stimulated cell cultures was previously reported to be a promising method to increase the sensitivity of proliferation tests. The aim of this investigation is to evaluate the effect of removal of regulatory T cells on the sensitivity of the LTT. Patients with well-documented drug hypersensitivity were recruited. Peripheral blood mononuclear cells, isolated CD3(+) and CD3(+) T cells depleted of the CD25(hi) fraction were used as effector cells in the LTT. Irrelevant drugs were also included to determine specificity. (3)H-thymidine incorporation was utilized as the detection system and results were expressed as a stimulation index (SI). SIs of 7/11 LTTs were reduced after a mean time interval of 10.5 months (LTT 1 vs. LTT 2). Removal of the CD25(hi) fraction, which was FOXP3(+) and had a suppressive effect on drug-induced proliferation, resulted in an increased response to the relevant drugs. Sensitivity was increased from 25 to 82.35% with dramatically enhanced SI (2.05 to 6.02). Specificity was not affected. Removal of Treg/CD25(hi) cells can increase the frequency and strengths of drug-specific proliferation without affecting specificity. This approach might be useful in certain drug hypersensitivity reactions with borderline responses or long time interval since the hypersensitivity reaction.


Role of gap junctional intercellular communication in radiation-induced bystander effects in human fibroblasts.
Involvement of gap junctional intercellular communication (GJIC) in bystander responses of confluent human fibroblasts irradiated with a carbon-ion beam was investigated. It was found that the lower the radiation dose, the higher the yield of radiation-induced micronuclei per nuclear traversal, suggesting the existence of bystander effects. This low-dose sensitivity was increased when GJIC was enhanced by treating cells with 8-Br-cAMP, but it was partly reduced by treating cells with DMSO, an effective scavenger of reactive oxygen species (ROS). Moreover, no low-dose sensitivity was observed when cells were treated with 100 micro M lindane, an inhibitor of GJIC. The survival of irradiated cells was increased by DMSO but was not influenced significantly by cAMP or lindane. On the other hand, G(1)-phase arrest was detected in the irradiated cells, and it was enhanced by cAMP. In contrast, this arrest was reduced or almost eliminated by DMSO or lindane, respectively, even when cells were irradiated with such a high dose that each cell received five nuclear traversals on average. Thus the bystander responses occurred after both low-dose and relatively high-dose irradiation. Our results indicated that both GJIC and ROS contributed to the radiation-induced bystander effect, but gap junctional channels might play an essential role by modulating the release of radiation-induced signaling factors.


Testing the relationship between intraspecific competition and individual specialization across both behavior and diet.
Individual specialization within populations is increasingly recognized as important in both ecology and evolution, but researchers working on intraspecific variation in behavior and diet infrequently interact. This may be because individual specialization on diet and behavior was historically difficult to investigate simultaneously on the same individuals. However, approaches like stable isotope analysis that allow hindcasting past field diets for laboratory organisms may provide opportunities to unite these areas of inquiry. Here, we tested the role of intraspecific competition on individual specialization through analysis of both behavior and diet simultaneously. We focused on intraspecific competition as a mechanism that might drive individual specialization of both diet and behavior. We conducted this study in Vilas County, Wisconsin, United States (US), using rusty crayfish Faxonius rusticus from six lakes across a relative abundance gradient. We conducted six assays to measure individual specialization of behavior and used stable isotope analysis to measure individual specialization of diet. We then related both measures of individual specialization to relative abundance of F. rusticus using linear and quadratic models. We found a unimodal relationship between intraspecific competition and individual specialization of diet for F. rusticus, likely because some preferred resources are unavailable to specialize on at the highest densities of this well-studied crayfish invader. Conversely, we found greater support for a linear relationship between individual specialization of behavior and intraspecific competition, perhaps because specialization by behavior is not inherently resource-limited. Our results show that dietary and behavioral specialization may exhibit different responses to increased intraspecific competition, and demonstrate a potential technique that can be used to investigate individual specialization of diet and behavior simultaneously for the same individuals and populations.


Children's estimates of food portion size: the effect of timing of dietary interview on the accuracy of children's portion size estimates.
For food intakes to be converted into nutrient intakes a measure or estimate of the amount of food consumed is required. A number of methods have been developed to assist subjects in providing an estimate of portion size. Children's ability to use perception, conceptualisation and memory skills to estimate food portion size has not been investigated systematically. The aim of the present study was to test the effect of the timing of a dietary interview on the accuracy of estimates of food portion sizes made by children, using food photographs, food models and an interactive portion size assessment system, developed for use with children and based on portion sizes of foods consumed by children. Children (n 108) aged 4-14 years were supplied with known quantities of foods and asked to estimate the portion size of each food using each of the three portion size assessment tools. Interviews took place (a) with the food in view, (b) just after the child had eaten the food or (c) 24 h after the child had eaten the food. There were no significant differences in children's ability to estimate food portion size (either as served or as eaten) with timing of interview. That is, children were as accurate in their estimates of portion size 24 h after consuming the food as when the food was in view. Under these conditions many children were able to estimate food portion size utilising perception, conceptualisation and memory skills.


Fish trypanosomes: their position in kinetoplastid phylogeny and variability as determined from 12S rRNA kinetoplast sequences.
Fish trypanosomes have traditionally been classified according to the host species from which they were isolated, each isolate being regarded as a distinct species. To test the soundness of this practice, the genetic variabilities of the kinetoplast 12S rRNA-encoding genes of different fish trypanosomes isolates were compared. The DNAs were extracted from trypanosomes cloned from blood samples of 15 donors representing ten different fish species in four orders from waters of three major river systems of Central and Northern Europe. Comparison with other trypanosomatid sequences revealed that the fish trypanosomes form a monophyletic group with Trypanosoma brucei as a sister group. Pairwise comparisons of genetic distances yielded a wide range of continuous variation with no indication of any discontinuities attributable to barriers to gene flow. The genetic distances did not correlate with either the identity of the host species or geography. The host specificity of fish trypanosomes appears to be limited.


An eleven-year case report of an avulsed maxillary central incisor after delayed replantation.
A 12 year-old girl presented with an avulsed maxillary central incisor for a trauma occurred two days earlier. The avulsed tooth was endodontically treated and replanted. Regardless of the ankylosis and replacement root resorption the tooth was retained for a very favourable nine-year period after trauma. In addition no severe infraposition occurred. When the extraction was necessary, despite the slight atrophy, as a result of compromised alveolar bone development due to ankylosis, an implant-supported crown was placed as definitive rehabilitation without recourse to bone augmentation procedures. In conclusion, delayed replantation in children will require long-term therapeutic commitment. In favourable cases, tooth with delayed replantation might be retained for a very long time thus avoiding the bone atrophy that would have accompanied early tooth loss and, consequently, reducing grafting requirements when a definitive implant-supported restoration will be required.


Results of an epidemiologic study of postdate pregnancy.
Obstetric and neonatal data on 18,610 deliveries between 1975 and 1982 were analyzed to determine the risks associated with postdate pregnancy in an era of routine fetal heart rate monitoring. Thirteen percent of the deliveries occurred between 42 and 43 weeks' gestation, and 1% occurred at 44 weeks or later. There was a nonsignificant rise in the perinatal mortality rates for 38-41-week deliveries as compared to those delivered at 42 weeks or later. However, the perinatal mortality rate was six times higher for small-for-gestational-age infants. There was a significant rise in the perinatal morbidity in infants delivered at or beyond 42 weeks (5% incidence). This finding implies that the sensitivity of currently used screening tests for perinatal mortality is good but that it is unsatisfactory for the detection and prevention of morbidity.


High prevalence of cattle fascioliasis in coastal areas of Thua Thien Hue province, Vietnam.
In Vietnam, especially central Vietnam, patients with fascioliasis are increasingly being reported. Since the fascioliasis is zoonotic, survey on the cattle fascioliasis should be informative for the control of human fascioliasis. In this study, the prevalence of cattle fascioliasis as well as the density of the intermediate host snails, Lymnaea swinhoei and L. viridis, were studied in Thua Thien Hue (TTH) province during 2014-2015. A total of 572 cattle feces were examined from 27 communes in 9 districts. Fasciola eggs were detected in cattle from 24 communes with an average prevalence of 23.4% (134/ 572). The highest prevalence was detected in cattle in the coastal plain terrain (31.0%) followed by plain (25.5%), mountain (21.7%), and low hilly (16.2%) terrains. The highest proportion of heavy infection (>200 EPG) was observed in the coastal plain terrain (36.1%), followed by mountains (20.0%), low hills (13.0%), and plains (8.9%). Low number of heavy infection, as well as relatively low prevalence in low hills and plains were associated with the extensive use of anti-fluke treatments. High number of intermediate host snails in low hilly and plain terrains also indicate high risk of fascioliasis. In this study, the density of Lymnaea snails in the coastal plain terrain was found to be very high (17.3 snails/m2) compared to that in previous studies. This is the first report indicating the recent expansion of cattle fascioliasis in the coastal region in Vietnam.


Calcium antagonists and islet function. VIII. The effect of magnesium.
Mg2+ in high concentration (10 to 20 mEq/l) caused a dose-related inhibition of glucose-induced 45calcium net uptake and subsequent insulin release in isolated islets. Experiments performed with the isolated perfused pancreas indicated that the inhibitory effect of Mg2+ upon insulin secretion was rapid and reversible. Although excess Mg2+ reduced lactate production by the islets, the process of glucose recognition by the B-cell was apparently unaltered as judged by the inhibitory effect of the hexose upon 45calcium efflux from perifused islets and the resultant accumulation of 45calcium in the islets. Mg2+ in high concentration failed to facilitate 45calcium efflux from the islets, whether in the presence or absence of glucose. These data suggest that the major effect of Mg2+ upon islet function is to inhibit Ca2+ entry in the B-cell.


Infrared hyperspectral imaging polarimeter using birefringent prisms.
A compact short-wavelength and middle-wavelength infrared hyperspectral imaging polarimeter (IHIP) is introduced. The sensor includes a pair of sapphire Wollaston prisms and several high-order retarders to form an imaging Fourier transform spectropolarimeter. The Wollaston prisms serve as a birefringent interferometer with reduced sensitivity to vibration versus an unequal path interferometer, such as a Michelson. Polarimetric data are acquired through the use of channeled spectropolarimetry to modulate the spectrum with the Stokes parameter information. The collected interferogram is Fourier filtered and reconstructed to recover the spatially and spectrally varying Stokes vector data across the image. The IHIP operates over a ±5° field of view and implements a dual-scan false signature reduction technique to suppress polarimetric aliasing artifacts. In this paper, the optical layout and operation of the IHIP sensor are presented in addition to the radiometric, spectral, and polarimetric calibration techniques used with the system. Spectral and spectropolarimetric results from the laboratory and outdoor tests with the instrument are also presented.


In vitro and in vivo efficacy of sulfo-carrabiose, a sugar-based cosmetic ingredient with anti-cellulite properties.
Most of adult women exhibit cellulite on the hips, buttock and thighs. Although extracellular matrix and lymphatic system disorders can increase its appearance, cellulite basically results from an excessive fat storage in the adipose tissue which exerts considerable pressure on the surrounding skin tissue and creates a dimpled irregular appearance. Caffeine, the most widely used anti-cellulite ingredient, favours fat break-down by inhibiting the phosphodiesterase enzyme and encouraging a high intracellular level of cAMP. A series of studies has shown that spermine and spermidine, two ubiquitous polyamines, encouraged fat storage and slowed fat break-down in the adipose tissue. Besides, it was shown that heparan sulfate glycosaminoglycans had a strong affinity for polyamines. To design a new cosmetic ingredient with anti-cellulite properties, we used molecular modelling to screen several ingredients with a structure similar to that of heparan sulfate glycosaminoglycans. This way, we identified sulfo-carrabiose as a potent molecule for trapping spermine and spermidine. These virtual results were first confirmed in tubo where sulfo-carrabiose was shown to dose-dependently inactivate spermine and spermidine. In vitro, adipocytes cultured with sulfo-carrabiose exhibited a significant reduction of lipogenesis and a significant increase of lipolysis. When sulfo-carrabiose was incorporated in a cosmetic formula, significant improvements were observed in thigh circumference, with better results than those obtained with caffeine after 28 days of use. Furthermore, a combination of caffeine and sulfo-carrabiose led to results significantly better than those obtained with caffeine alone. As measured by fringe projection, thigh volume was also significantly reduced after sulfo-carrabiose treatment. Finally, the appearance of cellulite assessed by clinical evaluation was also significantly reduced within 28 days.


High prevalence of nasopharyngeal colonization by Staphylococcus aureus among children with HIV-1 infection in extreme southern Brazil.
To compare nasopharyngeal colonization between children with HIV-1 infection and those without HIV-1 infection, with special emphasis on nasopharyngeal carriage of Staphylococcus aureus. This hospital-based cross-sectional study was carried out in the Paediatric Day Hospital of a teaching hospital. Nasopharyngeal swabs were collected in 93 children aged up to 18 years old born to HIV-positive mothers (31 children with HIV-1 infection and 62 age-matched non-infected children). The prevalence of nasopharyngeal colonization by S. aureus was higher among children with HIV-1 infection compared with those without HIV-1 infection (45.16% vs. 12.9%, p = 0.001). After adjusting all potential confounders, HIV-1 infection was an independent risk factor for nasopharyngeal colonization by S. aureus, with a prevalence ratio of 4.29 (95% confidence interval: 1.72-10.70). Children with HIV-1 infection had a higher prevalence of nasopharyngeal colonization by S. aureus than children without HIV-1 infection. Most of the isolated strains of S. aureus were methicillin-susceptible.


Gut-derived uremic toxin handling in vivo requires OAT-mediated tubular secretion in chronic kidney disease.
The role of the renal organic anion transporters OAT1 (also known as SLC22A6, originally identified as NKT) and OAT3 (also known as SLC22A8) in chronic kidney disease (CKD) remains poorly understood. This is particularly so from the viewpoint of residual proximal tubular secretion, a key adaptive mechanism to deal with protein-bound uremic toxins in CKD. Using the subtotal nephrectomy (STN) model, plasma metabolites accumulating in STN rats treated with and without the OAT inhibitor, probenecid, were identified. Comparisons with metabolomics data from Oat1-KO and Oat3-KO mice support the centrality of the OATs in residual tubular secretion of uremic solutes, such as indoxyl sulfate, kynurenate, and anthranilate. Overlapping our data with those of published metabolomics data regarding gut microbiome-derived uremic solutes - which can have dual roles in signaling and toxicity - indicates that OATs play a critical role in determining their plasma levels in CKD. Thus, the OATs, along with other SLC and ABC drug transporters, are critical to the movement of uremic solutes across tissues and into various body fluids, consistent with the remote sensing and signaling theory. The data support a role for OATs in modulating remote interorganismal and interorgan communication (gut microbiota-blood-liver-kidney-urine). The results also have implications for understanding drug-metabolite interactions involving uremic toxins.


Dok-1 is a positive regulator of IL-4 signalling and IgE response.
Interleukin-4 (IL-4) plays an essential role in the control of humoral immunity by regulating lymphocyte proliferation and differentiation, including the T helper type 2 lineage commitment of CD4(+) T cells as well as the isotype switching to IgE in B cells. The adaptor protein Dok-1 is known to have an essential role in the negative regulation of a variety of cytokine signalling events. However, here we have found that the loss of Dok-1 impaired the proliferative response of CD4(+) T cells and B cells to IL-4. Conversely, the forced expression of Dok-1 in the myeloid cell line 32D augmented the IL-4-induced proliferation, indicating a positive role for Dok-1. Tyrosine phosphorylation, and thereby the activation of Stat6 and IRS-2, is critical for IL-4 signalling; however, only the activation of Stat6, not the IRS-2-dependent phosphorylation of Akt, was perturbed in Dok-1-deficient cells stimulated with IL-4. Furthermore, mice lacking Dok-1 showed an impaired IgE response to thymus-dependent antigen. Thus, Dok-1 is a positive regulator of IL-4 signalling and IgE response.


[Genetics of nephroblastoma. II. The incidence of developmental defects and dysplastic and asymmetrical traits in children with nephroblastoma].
The frequency of developmental defects and displasticity characters in children with nephroblastoma was determined in two groups--with early, up to 2 years manifestation of the tumor (ENB), in 40 patients, and with advanced nephroblastoma (ANB), in 59 patients (manifestation of the disease after 2 years). The data were correlated with control groups adequate for age (130 children). The frequency of serious developmental defects was higher in ANB group (20%) than in ENB group (7%). Over a half of developmental defects were hemihypertrophy and doubling of organs. One case of a child with the combination of nephroblastoma and Smith-Lemli-Opitz syndrome was defined. In ANB group an increased frequency of asymmetry in the conjugate organs (foot, hand, middle finger, ears) was found. Its direction is correlated with tumor localization (tumor site). In ENB group no analogous effect was shown. The data obtained present phenotype characteristics of groups with early and late manifestation of the disease which are probably different in their genesis and thus, their identification is necessary for the adequate medicogenetic consultation (examination).


Differential expression of the adaptor protein HSH2 controls the quantitative and qualitative nature of the humoral response.
Endogenous expression of the adaptor protein hematopoietic Src homology 2-containing adaptor protein (HSH2) is regulated in a dynamic manner during B cell maturation and differentiation. Developing B cells lack detectable HSH2, whereas transitional 1 and 2 B cells in the periphery exhibit increasing levels of expression. Mature follicular B cells exhibit decreased expression of HSH2 compared with transitional 2 B cells, and expression is further downregulated in germinal center B cells. In contrast, marginal zone B cells and B1a/b B cells exhibit high-level HSH2 expression. Regulation of HSH2 expression plays a critical role in determining the outcome of the humoral immune response as demonstrated using HSH2 transgenic (Tg) mice. Constitutive expression of HSH2 in the B lineage at levels comparable to B1a/b B cells results in decreased serum Ig titers for all subclasses with the exception of IgA. HSH2 Tg mice immunized with T-dependent or T-independent Ags exhibit a moderate decrease in the production of Ag-specific IgM, whereas class-switched isotypes are decreased by ∼80-90% compared with control mice. Analysis of HSH2 Tg B cell activation in vitro demonstrated that HSH2 selectively regulates the B cell response to TNF family receptors (i.e., CD40 and BAFF-R), but not BCR- or TLR-dependent signals. These data demonstrate that changes in HSH2 expression have profound effects on the humoral immune response.


DNA surface exploration and operator bypassing during target search.
Many proteins that bind specific DNA sequences search the genome by combining three-dimensional diffusion with one-dimensional sliding on nonspecific DNA1-5. Here we combine resonance energy transfer and fluorescence correlation measurements to characterize how individual lac repressor (LacI) molecules explore the DNA surface during the one-dimensional phase of target search. To track the rotation of sliding LacI molecules on the microsecond timescale, we use real-time single-molecule confocal laser tracking combined with fluorescence correlation spectroscopy (SMCT-FCS). The fluctuations in fluorescence signal are accurately described by rotation-coupled sliding, in which LacI traverses about 40 base pairs (bp) per revolution. This distance substantially exceeds the 10.5-bp helical pitch of DNA; this suggests that the sliding protein frequently hops out of the DNA groove, which would result in the frequent bypassing of target sequences. We directly observe such bypassing using single-molecule fluorescence resonance energy transfer (smFRET). A combined analysis of the smFRET and SMCT-FCS data shows that LacI hops one or two grooves (10-20 bp) every 200-700 μs. Our data suggest a trade-off between speed and accuracy during sliding: the weak nature of nonspecific protein-DNA interactions underlies operator bypassing, but also speeds up sliding. We anticipate that SMCT-FCS, which monitors rotational diffusion on the microsecond timescale while tracking individual molecules with millisecond resolution, will be applicable to the real-time investigation of many other biological interactions and will effectively extend the accessible time regime for observing these interactions by two orders of magnitude.


Scalable Synthesis of Anomerically Pure Orthogonal-Protected GlcN3 and GalN3 from d-Glucosamine.
An improved and scalable synthesis of orthogonally protected d-glucosamine and d-galactosamine building blocks from inexpensive d-glucosamine has been developed. The key reaction is an inversion/migration step providing access to a fully orthogonal protecting group pattern, which is required for microbial oligosaccharide synthesis. The method can be carried out on a multigram scale as several of the reactions can be purified by crystallization to give anomerically pure products.


Impact of High-Molecular-Risk Mutations on Transplantation Outcomes in Patients with Myelofibrosis.
Mutational profiling has demonstrated utility in predicting the likelihood of disease progression in patients with myelofibrosis (MF). However, there is limited data regarding the prognostic utility of genetic profiling in MF patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HCT). We performed high-throughput sequencing of 585 genes on pre-transplant samples from 101 patients with MF who underwent allo-HCT and evaluated the association of mutations and clinical variables with transplantation outcomes. Overall survival (OS) at 5 years post-transplantation was 52%, and relapse-free survival (RFS) was 51.1 % for this cohort. Nonrelapse mortality (NRM) accounted for most deaths. Patient's age, donor's age, donor type, and Dynamic International Prognostic Scoring System score at diagnosis did not predict for outcomes. Mutations known to be associated with increased risk of disease progression, such as ASXL1, SRSF2, IDH1/2, EZH2, and TP53, did not impact OS or RFS. The presence of U2AF1 (P = .007) or DNMT3A (P = .034) mutations was associated with worse OS. A Mutation-Enhanced International Prognostic Scoring System 70 score was available for 80 patients (79%), and there were no differences in outcomes between patients with high risk scores and those with intermediate and low risk scores. Collectively, these data identify mutational predictors of outcome in MF patients undergoing allo-HCT. These genetic biomarkers in conjunction with clinical variables may have important utility in guiding transplantation decision making.


Lipoteichoic acid and interleukin 1 stimulate synergistically production of hepatocyte growth factor (scatter factor) in human gingival fibroblasts in culture.
Lipoteichoic acids (LTA) from various gram-positive bacteria, including oral streptococci such as Streptococcus sanguis, enhanced the production of hepatocyte growth factor (HGF) (scatter factor) by human gingival fibroblasts in culture, whereas lipopolysaccharides (LPS) from various gram-negative bacteria did not. In contrast, LPS induced interleukin 1 activity in human gingival epithelial cells in culture, while LTA had little effect. LTA and recombinant human interleukin 1 alpha enhanced synergistically the production of HGF/SF in human gingival fibroblast cultures. Recombinant human HGF, in turn, enhanced the proliferation of human gingival epithelial cells in culture.


Effects of competition, predation, and dispersal on species richness at local and regional scales.
This study explores the consequences of predator-mediated coexistence among competitors for patterns of incidence and diversity at local and regional scales. We develop a model that draws on elements of metapopulation models of competitors and food chains by allowing competitors to coexist locally in the presence of predators but not in their absence. The model predicts that predators promote regional coexistence by greatly expanding the range of conditions under which two competitors persist at equilibrium. Predators could have positive or negative effects on mean local diversity within the region depending on their dispersal rates, those of the prey, and their effects on prey extinction rates. The presence of predators increased the abundance of inferior competitors, thereby expanding the conditions for positive relationships between local and regional diversity. The model also predicted positive correlations between local diversity of predators and prey. These predictions were supported by patterns of phytoplankton, zooplankton, and fish species richness among lakes. The model may help to resolve the apparent contrast between linear patterns of local and regional richness and experimental evidence for strong invasion resistance and rapid dispersal in zooplankton.


Financing transformative health systems towards achievement of the health Sustainable Development Goals: a model for projected resource needs in 67 low-income and middle-income countries.
The ambitious development agenda of the Sustainable Development Goals (SDGs) requires substantial investments across several sectors, including for SDG 3 (healthy lives and wellbeing). No estimates of the additional resources needed to strengthen comprehensive health service delivery towards the attainment of SDG 3 and universal health coverage in low-income and middle-income countries have been published. We developed a framework for health systems strengthening, within which population-level and individual-level health service coverage is gradually scaled up over time. We developed projections for 67 low-income and middle-income countries from 2016 to 2030, representing 95% of the total population in low-income and middle-income countries. We considered four service delivery platforms, and modelled two scenarios with differing levels of ambition: a progress scenario, in which countries' advancement towards global targets is constrained by their health system's assumed absorptive capacity, and an ambitious scenario, in which most countries attain the global targets. We estimated the associated costs and health effects, including reduced prevalence of illness, lives saved, and increases in life expectancy. We projected available funding by country and year, taking into account economic growth and anticipated allocation towards the health sector, to allow for an analysis of affordability and financial sustainability. We estimate that an additional $274 billion spending on health is needed per year by 2030 to make progress towards the SDG 3 targets (progress scenario), whereas US$371 billion would be needed to reach health system targets in the ambitious scenario-the equivalent of an additional $41 (range 15-102) or $58 (22-167) per person, respectively, by the final years of scale-up. In the ambitious scenario, total health-care spending would increase to a population-weighted mean of $271 per person (range 74-984) across country contexts, and the share of gross domestic product spent on health would increase to a mean of 7·5% (2·1-20·5). Around 75% of costs are for health systems, with health workforce and infrastructure (including medical equipment) as the main cost drivers. Despite projected increases in health spending, a financing gap of $20-54 billion per year is projected. Should funds be made available and used as planned, the ambitious scenario would save 97 million lives and significantly increase life expectancy by 3·1-8·4 years, depending on the country profile. All countries will need to strengthen investments in health systems to expand service provision in order to reach SDG 3 health targets, but even the poorest can reach some level of universality. In view of anticipated resource constraints, each country will need to prioritise equitably, plan strategically, and cost realistically its own path towards SDG 3 and universal health coverage. WHO.


Personality disorders and perceived stress in major depressive disorder.
The investigation of comorbidity between major depressive disorder (MDD) and personality disorders (PDs) has attracted considerable interest. Whereas some studies found that the presence of PDs has adverse effects on the course and treatment of MDD, others have failed to demonstrate this link. These inconsistent findings suggest that specific PD comorbidity might affect the course of MDD by modulating factors that increase the overall risk of depression, including an elevated tendency to perceive stress. To investigate whether the presence of a specific PD cluster was associated with elevated levels of stress appraisal, we administered the Perceived Stress Scale (PSS) before and after treatment to 227 MDD outpatients enrolled in an 8-week open-label treatment with fluoxetine. Following treatment, multiple linear regression analyses revealed that the presence of Cluster A, but not Cluster B or C, was associated with higher levels of perceived stress, even after adjusting for baseline depression severity and PSS scores, as well as various sociodemographic variables. The presence of Cluster A PD comorbidity was uniquely associated with elevated stress appraisal after antidepressant treatment, raising the possibility that stress exacerbation might be an important factor linked to poor treatment outcome in MDD subjects with Cluster A pathology.


Framework for identifying quantitative trait loci in association studies using structural equation modeling.
In this article, we suggest a framework for identifying quantitative trait loci (QTL) in association studies using structural equation modeling. Two tests to detect QTLs and estimate the proportion of variance they explain are discussed. The first test assumes that there is no population admixture and only requires that the subjects are genotyped. The second one is a TDT-like test that cannot give false-positive results due to population admixture but requires that the parents of the subjects are genotyped as well and that subjects have at least one heterozygous parent. Power calculations showed that with the first test, 100 subjects were generally sufficient to detect a locus that explained 10% and less than 1,000 subject to detect a locus that explained 1% of the total variance. To obtain the same power, the TDT-like test required an initial sample that was on average 1.7 times larger. Calculations showed that the first test was quite robust against population admixture and that the power of tests to detect admixture was good. This suggested that in the extreme and very specific conditions in which population admixture may cause false-positive findings, admixture can often be detected.


Assisted reproductive technology, congenital malformations, and epigenetic disease.
In the 3 decades since the birth of the first in vitro fertilization-conceived child, assisted reproduction technologies were rapidly assimilated into clinical care and are now responsible for 1% to 3% of all livebirths in North America and Europe. The rate of congenital anomalies is low (4% to 6%), though it represents a modest increase over the background rate of major malformations (3%). Additionally, emerging studies of imprinting and epigenetic conditions, those genetic disorders due to changes in DNA transcription without change in DNA sequence, suggest that the preimplantation period is vulnerable to perturbations. Review of these studies provides clinically useful information and a basis for investigations of the long-term effects of assisted reproduction technology on the children conceived.


[Relationship between metabolic diseases and all-cause and cardiovascular death in an elderly male population during a 15-year follow-up].
To explore the relationship between metabolic diseases and death from all causes, cardiovascular diseases (CVD) in an elderly male population. A cohort of 1 447 elderly males was followed up for 15 years from 1996 to 2011. All of them received annual check-ups at our hospital. The Cox proportional hazard model was applied to multivariate survival analysis for all-cause and CVD mortality. And the cumulative survival rates were calculated by Kaplan-Meier method and log-rank test was used to compare the survival rates. During a 15-year follow-up, 639 subjects died, including 186 deaths from cardiovascular causes. Multivariate Cox regression analysis showed that age [relative risk (RR) = 1.131, 95% confidence interval (CI) 1.114-1.148], impaired glucose metabolism (RR = 1.344, 95% CI 1.139-1.585), hypertension (RR = 1.241, 95% CI 1.055-1.460) , elevated fasting glucose level (RR = 1.101, 95% CI 1.031-1.177) and lower body mass index (BMI) (RR = 0.968, 95% CI 0.943-0.993) increased the risks of all-cause mortality while age (RR = 1.119, 95% CI 1.086-1.153) , impaired glucose metabolism (RR = 1.856, 95% CI 1.386-2.458) and hypertension (RR = 1.699, 95% CI 1.242-2.324) elevated the risks of CVD mortality. The cumulative survival rates from all-cause and CVD mortality in impaired glucose regulation and diabetes group were significantly lower than those in normal glucose tolerance group (P < 0.01) . However, no difference existed between impaired glucose regulation (IGR) and diabetes groups. The cumulative survival rates from all-cause and CVD mortality significantly decreased in cases of impaired glucose metabolism and hypertension (P < 0.01) . The cumulative survival rates from all-cause mortality in low BMI group were significantly lower than those in normal and high BMI groups (P < 0.05) . A substantially higher risk of all-cause and CVD mortality was present in those with 2 or more metabolic disorders versus those with 0-1 metabolic disorder (P < 0.01). Malignant tumor and CVD are the main cause of death for Chinese elderly male population. Advanced age, impaired glucose metabolism, hypertension and 2 or more concurrent metabolic disorders are risk factors of all-cause and CVD mortality. And underweight is associated with an increased risk of death in elders.


Preliminary Results of Extracorporeal Membrane Oxygenation Assisted Tracheal Sleeve Pneumonectomy for Cancer.
Tracheal sleeve pneumonectomy is a challenge in lung cancer management and in achieving long-term oncological results. In November 2018, we started a prospective study on the role of extracorporeal membrane oxygenation (ECMO) in tracheal sleeve pneumonectomy. We aim to present our preliminary results. From November 2018 to November 2019, six patients (three men and three women; median age: 61 years) were eligible for tracheal sleeve pneumonectomy for lung cancer employing the veno-venous ECMO during tracheobronchial anastomosis. Only in one patient, an intrapericardial pneumonectomy without ECMO support was performed, but cannulas were maintained during surgery. The median length of surgery was 201 minutes (range: 162-292 minutes), and the average duration of the apneic phase was 38 minutes (range: 31-45 minutes). No complications correlated to the positioning of the cannulas were recorded. There was only one major postoperative complication (hemothorax). At the time of follow-up, all patients were alive; one patient alive with bone metastasis was being treated with radiotherapy. ECMO-assisted oncological surgery was rarely described, and its advantages include hemodynamic stability with low bleeding complications and a clean operating field. As suggested by our preliminary data, ECMO-assisted could be a useful alternative strategy in select lung cancer patients.


Differential projections of the infralimbic and prelimbic cortex in the rat.
The medial prefrontal cortex has been associated with diverse functions including attentional processes, visceromotor activity, decision-making, goal-directed behavior, and working memory. The present report compares and contrasts projections from the infralimbic (IL) and prelimbic (PL) cortices in the rat by using the anterograde anatomical tracer, Phaseolus vulgaris-leucoagglutinin. With the exception of common projections to parts of the orbitomedial prefrontal cortex, olfactory forebrain, and midline thalamus, PL and IL distribute very differently throughout the brain. Main projection sites of IL are: 1) the lateral septum, bed nucleus of stria terminalis, medial and lateral preoptic nuclei, substantia innominata, and endopiriform nuclei of the basal forebrain; 2) the medial, basomedial, central, and cortical nuclei of amygdala; 3) the dorsomedial, lateral, perifornical, posterior, and supramammillary nuclei of hypothalamus; and 4) the parabrachial and solitary nuclei of the brainstem. By contrast, PL projects at best sparingly to each of these structures. Main projection sites of PL are: the agranular insular cortex, claustrum, nucleus accumbens, olfactory tubercle, the paraventricular, mediodorsal, and reuniens nuclei of thalamus, the capsular part of the central nucleus and the basolateral nucleus of amygdala, and the dorsal and median raphe nuclei of the brainstem. As discussed herein, the pattern of IL projections is consistent with a role for IL in the control of visceral/autonomic activity homologous to the orbitomedial prefrontal cortex of primates, whereas those of PL are consistent with a role for PL in limbic-cognitive functions homologous to the dorsolateral prefrontal cortex of primates.


New and future approaches for spinal cord protection.
The major cause of spinal cord injury, during and after aortic surgery, is based on the occurrence of one or more of the three following events: (1) the duration and degree of ischemia;(2) failure to re-establish blood flow to the spinal cord after the repair; and (3) a biochemically mediated reperfusion injury. Clinically, this manifests either as permanent or reversible paraplegia or paraparesis, or a neurogenic bladder. For more than 40 years, numerous methods have been attempted to prevent paralysis, and some of the newer technical innovations include reducing the duration of ischemia, the use of newer centrifugal pump distal perfusion techniques, localized hypothermia, intrathecal maneuvers, pharmacological agents, angiography, somatosensory-evoked potential monitoring, spinal motor-evoked potential monitoring, hydrogen mapping, not resecting the posterior aortic wall, the use of stents, and a spectrum of various pharmacological agents to prevent reperfusion injury to the spinal cord. Some of these techniques and agent seem to be effective at reducing the risk of spinal cord injury.


Novel (Multilevel) Focus Group Training for a Transdisciplinary Research Consortium.
Health researchers are increasingly turning to qualitative research for a nuanced understanding of complex health phenomena. The quality and rigor of qualitative research relies on individual data collector skills, yet few guidelines exist for training multidisciplinary, multi-institution qualitative research teams. Specific guidance is needed on qualitative research practices that ensure scientific rigor by optimizing diverse experience and expertise across research centers. We describe our systematic approach to training a cohort of 15 focus group moderators from seven universities in the Prevention of Lower Urinary Tract Symptoms (PLUS) Research Consortium's Study of Habits, Attitudes, Realities, and Experiences (SHARE). SHARE's aim was to explore women and girls' experiences, perceptions, beliefs, knowledge, and behaviors related to bladder health and function across the life course. Drawing on adult education and action-learning best practices, a three-phase curriculum was designed to maximize moderator proficiency and qualitative research expertise. The phases involved online, interactive web-based education, in-person didactic training with experiential components, and tailored supplemental online training. Evaluative feedback was collected before, during, and after the training. Feedback was used to identify emergent training needs. This training approach may be used by transdisciplinary research teams conducting multisite research to assure qualitative research credibility and trustworthiness.


Astaxanthin-Shifted Gut Microbiota Is Associated with Inflammation and Metabolic Homeostasis in Mice.
Astaxanthin is a red lipophilic carotenoid that is often undetectable in human plasma due to the limited supply in typical Western diets. Despite its presence at lower than detectable concentrations, previous clinical feeding studies have reported that astaxanthin exhibits potent antioxidant properties. We examined astaxanthin accumulation and its effects on gut microbiota, inflammation, and whole-body metabolic homeostasis in wild-type C57BL/6 J (WT) and β-carotene oxygenase 2 (BCO2) knockout (KO) mice. Six-wk-old male and female BCO2 KO and WT mice were provided with either nonpurified AIN93M (e.g., control diet) or the control diet supplemented with 0.04% astaxanthin (wt/wt) ad libitum for 8 wk. Whole-body energy expenditure was measured by indirect calorimetry. Feces were collected from individual mice for short-chain fatty acid assessment. Hepatic astaxanthin concentrations and liver metabolic markers, cecal gut microbiota profiling, inflammation markers in colonic lamina propria, and plasma samples were assessed. Data were analyzed by 3-way ANOVA followed by Tukey's post hoc analysis. BCO2 KO but not WT mice fed astaxanthin had ∼10-fold more of this compound in liver than controls (P < 0.05). In terms of the microbiota composition, deletion of BCO2 was associated with a significantly increased abundance of Mucispirillum schaedleri in mice regardless of gender. In addition to more liver astaxanthin in male KO compared with WT mice fed astaxanthin, the abundance of gut Akkermansia muciniphila was 385% greater, plasma glucagon-like peptide 1 was 27% greater, plasma glucagon and IL-1β were 53% and 30% lower, respectively, and colon NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome activation was 23% lower (all P < 0.05) in male KO mice than the WT mice. Astaxanthin affects the gut microbiota composition in both genders, but the association with reductions in local and systemic inflammation, oxidative stress, and improvement of metabolic homeostasis only occurs in male mice.


Short sleep duration as a risk factor for hypercholesterolemia: analyses of the National Longitudinal Study of Adolescent Health.
To explore the relationship between sleep duration in adolescence and hypercholesterolemia in young adulthood. Experimental sleep restriction has been shown to significantly increase total cholesterol and LDL cholesterol levels in women. Short sleep duration has been found in cross sectional studies to be associated with higher total cholesterol and lower HDL cholesterol levels. Sleep deprivation could increase the risk for hypercholesterolemia by increasing appetite and dietary consumption of saturated fats, decreasing motivation to engage in regular physical activity, and increasing stress and resultant catecholamine induced lipolysis. No previous published population studies have examined the longitudinal relationship between sleep duration and high cholesterol. Multivariate longitudinal analyses stratified by sex of the ADD Health using logistic regression. United States nationally representative, school-based, probability-based sample. Adolescents (n = 14,257) in grades 7 to 12 at baseline (1994-95) and ages 18 to 26 at follow-up (2001-02). Among females, each additional hour of sleep was associated with a significantly decreased odds of being diagnosed with high cholesterol in young adulthood (OR = 0.85, 95% CI 0.75-0.96) after controlling for covariates. Additional sleep was associated with decreased, yet not statistically significant, odds ratios for hypercholesterolemia in males (OR = 0.91, 95% CI 0.79-1.05). Short sleep durations in adolescent women could be a significant risk factor for high cholesterol. Interventions that lengthen sleep could potentially serve as treatments and as primary preventative measures for hypercholesterolemia.


Variations in respiratory rate do not reflect changes in tidal volume or minute ventilation after major abdominal surgery.
Monitoring of postoperative pulmonary function usually includes respiratory rate and oxygen saturation measurements. We hypothesized that changes in postoperative respiratory rate do not correlate with changes in tidal volume or minute ventilation. In addition, we hypothesized that variability of minute ventilation and tidal volume is larger than variability of respiratory rate. Respiratory rate and changes in tidal volume and in minute ventilation were continuously measured in 27 patients during 24 h following elective abdominal surgery, using an impedance-based non-invasive respiratory volume monitor (ExSpiron, Respiratory Motion, Waltham, MA, US). Coefficients of variation were used as a measure for variability of respiratory rate, tidal volume and minute ventilation. Data of 38,149 measurements were analyzed. We found no correlation between respiratory rate and tidal volume or minute ventilation (r2 = 0.02 and 0.01). Mean respiratory rate increased within the first 24 h after abdominal surgery from 13.9 ± 2.5 to 16.2 ± 2.4 breaths/min (p = 0.008), while tidal volume and minute ventilation remained unchanged (p = 0.90 and p = 0.18). Of interest, variability of respiratory rate (0.21 ± 0.06) was significantly smaller than variability of tidal volume (0.37 ± 0.12, p < 0.001) and minute ventilation (0.41 ± 0.12, p < 0.001). Changes in postoperative respiratory rate do not allow conclusions about changes in tidal volume or minute ventilation. We suggest that postoperative alveolar hypoventilation may not be recognized by monitoring respiratory rate alone. Variability of respiratory rate is smaller than variability in tidal volume and minute ventilation, suggesting that adaptations of alveolar ventilation to metabolic needs may be predominately achieved by variations in tidal volume.


Multiple types of data are required to identify the mechanisms influencing the spatial expansion of melanoma cell colonies.
The expansion of cell colonies is driven by a delicate balance of several mechanisms including cell motility, cell-to-cell adhesion and cell proliferation. New approaches that can be used to independently identify and quantify the role of each mechanism will help us understand how each mechanism contributes to the expansion process. Standard mathematical modelling approaches to describe such cell colony expansion typically neglect cell-to-cell adhesion, despite the fact that cell-to-cell adhesion is thought to play an important role. We use a combined experimental and mathematical modelling approach to determine the cell diffusivity, D, cell-to-cell adhesion strength, q, and cell proliferation rate, λ, in an expanding colony of MM127 melanoma cells. Using a circular barrier assay, we extract several types of experimental data and use a mathematical model to independently estimate D, q and λ. In our first set of experiments, we suppress cell proliferation and analyse three different types of data to estimate D and q. We find that standard types of data, such as the area enclosed by the leading edge of the expanding colony and more detailed cell density profiles throughout the expanding colony, does not provide sufficient information to uniquely identify D and q. We find that additional data relating to the degree of cell-to-cell clustering is required to provide independent estimates of q, and in turn D. In our second set of experiments, where proliferation is not suppressed, we use data describing temporal changes in cell density to determine the cell proliferation rate. In summary, we find that our experiments are best described using the range D=161-243μm2 hour-1, q=0.3-0.5 (low to moderate strength) and λ=0.0305-0.0398 hour-1, and with these parameters we can accurately predict the temporal variations in the spatial extent and cell density profile throughout the expanding melanoma cell colony. Our systematic approach to identify the cell diffusivity, cell-to-cell adhesion strength and cell proliferation rate highlights the importance of integrating multiple types of data to accurately quantify the factors influencing the spatial expansion of melanoma cell colonies.


Population distribution of the methionine allele at the PRNP codon 129 polymorphism in Europe and the Middle East.
Methionine homozygosity at codon 129 of the prion protein gene is a risk factor for Creutzfeldt-Jakob disease. Knowledge of M129V polymorphism in normal populations may contribute to a better understanding of prion diseases. M129V polymorphism was studied in 2201 normal subjects, originating from 15 populations from Europe and the Middle East. Mean heterozygosity in these populations is 38.9%, and there is some significant geographic heterogeneity between them. A comparison of M129 allele frequencies in these 15 populations to those already published for 8 European countries plus Turkey shows significant correlations with both latitude (r = -0.77) and longitude (r = 0.69). The geographic map of methionine allele frequencies indicates an east-west gradient of decreasing methionine allele values from the Middle East to Western Europe.


Species distribution and population connectivity of deep-sea mussels at hydrocarbon seeps in the Gulf of Mexico.
Hydrocarbon seepage is widespread and patchy in the Gulf of Mexico, and six species of symbiont containing bathymodiolin mussels are found on active seeps over wide and overlapping depth and geographic ranges. We use mitochondrial genes to discriminate among the previously known and a newly discovered species and to assess the connectivity among populations of the same species in the northern Gulf of Mexico (GoM). Our results generally validate the morphologically based distribution of the three previously known GoM species of Bathymodiolus, although we found that approximately 10% of the morphologically based identifications were incorrect and this resulted in some inaccuracies with respect to their previously assigned depth and geographical distribution patterns. These data allowed us to confirm that sympatry of two species of Bathymodiolus within a single patch of mussels is common. A new species of bathymodiolin, Bathymodiolus sp. nov., closely related to B. heckerae was also discovered. The two species live at the same depths but have not been found in sympatry and both have small effective population sizes. We found evidence for genetic structure within populations of the three species of Bathymodiolinae for which we had samples from multiple sites and suggest limited connectivity for populations at some sites. Despite relatively small sample sizes, genetic diversity indices suggest the largest population sizes for B. childressi and Tamu fisheri and the smallest for B. heckerae and B. sp. nov. among the GoM bathymodiolins. Moreover, we detected an excess of rare variants indicating recent demographic changes and population expansions for the four species of bathymodiolins from the Gulf of Mexico.


Cucumber malate decarboxylase, CsNADP-ME2, functions in the balance of carbon and amino acid metabolism in fruit.
Central metabolism produces carbohydrates and amino acids that are tightly correlated to plant growth and thereby crop productivity. Malate is reported to link mitochondrial respiratory metabolism with cytosolic biosynthetic pathways. Although the function of malate metabolism-related enzymes in providing carbon has been characterized in some plants, evidence for this role in the fleshy fruit of cucumber is lacking. Here, radiolabeled bicarbonate fed into the xylem stream from the cucumber roots was incorporated into amino acids, soluble sugars, and organic acids in the exocarp and vasculature of fruits. The activities of decarboxylases, especially decarboxylation from NADP-dependent malic enzyme (NADP-ME), were higher in cucumber fruit than in the leaf lamina. Histochemical localization revealed that CsNADP-ME2 was mainly located in the exocarp and vascular bundle system of fruit. Radiotracer and gas-exchange analysis indicated that overexpression of CsNADP-ME2 could promote carbon flux into soluble sugars and starch in fruits. Further studies combined with metabolic profiling revealed that the downregulation of CsNADP-ME2 in RNA interference (RNAi) lines caused the accumulation of its substrate, malate, in the exocarp. In addition to inhibition of glycolysis-related gene expression and reduction of the activities of the corresponding enzymes, increased amino acid synthesis and decreased sugar abundance were also observed in these lines. The opposite effect was found in CsNADP-ME2-overexpressing lines, suggesting that there may be a continuous bottom-up feedback regulation of glycolysis in cucumber fruits. Overall, our studies indicate that CsNADP-ME2 may play potential roles in both central carbon reactions and amino acid metabolism in cucumber fruits.


Diagnosing and managing mitral regurgitation.
Mitral regurgitation is the most prevalent valvular disorder in the United States. Patient presentation varies from asymptomatic to symptoms of severe heart failure. Primary care providers often are the first to detect mitral regurgitation as a result of careful auscultation. Medical management focuses on reducing cardiac workload, combating cardiac remodeling, and treating left ventricular failure. Surgical management is reserved for severe cases.


Effect of aldosterone on abundance and phosphorylation kinetics of Na-K-ATPase of toad urinary bladder.
The possibility was explored that aldosterone-dependent modulation of either the abundance or the kinetic properties of the Na+ pump (Na-K-ATPase) is involved in the mechanism of natriferic action on toad urinary bladder. Cell membranes from the epithelium of urinary bladders of toads were prepared by a sucrose-Ficoll step gradient method, which yielded a 10-fold increase in specific activity of plasma membrane marker enzymes and only minor contamination with other subcellular fractions. Phosphorylation of Na-K-ATPase by gamma-PO4 of [gamma-32P]ATP in the presence of Mg2+ was Na+ dependent, whereas dephosphorylation was K+ dependent. Km for phosphorylation by ATP was 3 X 10(-8) M and K1/2 (half-maximal stimulation) for Na+ was 13 +/- 2 mM. Pretreatment of cell membranes with sodium deoxycholate (DOC) increased the maximal extent of phosphorylation (Nmax) about threefold without changing the Km for ATP. Aldosterone (5 X 10(-8) M) stimulated transepithelial Na+ transport two- to threefold in 5 h but had no significant effect on Km for ATP in the phosphorylation reaction (with or without activation by DOC) or on the abundance of the enzyme inferred from Nmax of phosphorylation in the absence of activation by DOC. After pretreatment with DOC, average Nmax was 13% greater in the aldosterone-treated population. Regression analysis of these responses revealed no significant correlation between increments in short-circuit currents and Nmax values of phosphorylation with or without pretreatment with DOC.(ABSTRACT TRUNCATED AT 250 WORDS)