Discrete Control of TRPV4 Channel Function in the Distal Nephron by Protein Kinases A and C

We have recently documented that the Ca²⁺-permeable TRPV4 channel, which is abundantly expressed in distal nephron cells, mediates cellular Ca²⁺ responses to elevated luminal flow. In this study, we combined Fura-2-based [Ca²⁺]_i imaging with immunofluorescence microscopy in isolated split-opened distal nephrons of C57BL/6 mice to probe the molecular determinants of TRPV4 activity and subcellular distribution. We found that activation of the PKC pathway with phorbol 12-myristate 13-acetate significantly increased [Ca²⁺]_i responses to flow without affecting the subcellular distribution of TRPV4. Inhibition of PKC with bisindolylmaleimide I diminished cellular responses to elevated flow. In contrast, activation of the PKA pathway with forskolin did not affect TRPV4-mediated [Ca²⁺]_i responses to flow but markedly shifted the subcellular distribution of the channel toward the apical membrane. These actions were blocked with the specific PKA inhibitor H-89. Concomitant activation of the PKA and PKC cascades additively enhanced the amplitude of flow-induced [Ca²⁺]_i responses and greatly increased basal [Ca²⁺]_i levels, indicating constitutive TRPV4 activation. This effect was precluded by the selective TRPV4 antagonist HC-067047. Therefore, the functional status of the TRPV4 channel in the distal nephron is regulated by two distinct signaling pathways. Although the PKA-dependent cascade promotes TRPV4 trafficking and translocation to the apical membrane, the PKC-dependent pathway increases the activity of the channel on the plasma membrane.     

Association of PON1 and PON2 Polymorphisms with PON1 Activity and Significant Coronary Stenosis in a Tunisian Population

PON1 and PON2 have attracted considerable attention as candidate genes for coronary heart disease because their enzymes function as key factors in lipoprotein catabolism pathways. We studied the distribution of PON1 and PON2 polymorphisms, including genotyping, lipid profile, and PON1 activity, and their association with PON1 activity and significant coronary stenosis (SCS) in a Tunisian population. PON1 activity was lower in patients with SCS than in controls. It increased with the R allele (QQ < QR < RR) in PON1-192 genotypes and with the L allele (MM < ML < LL) in PON1-55 genotypes. In the presence of metabolic syndrome and diabetes, PON1-192RR and PON2-311CC were associated with an increased risk of SCS and PON1-55MM seems to have lower risk. This association was evident among nonsmokers for PON1-55MM and among smokers for PON1-192RR and PON2-311CC. The GTGC haplotype seemed to increase the risk of SCS compared with the wild haplotype in a Tunisian population.     

Rye Pm8 and wheat Pm3 are orthologous genes and show evolutionary conservation of resistance function against powdery mildew

The improvement of wheat through breeding has relied strongly on the use of genetic material from related wild and domesticated grass species. The 1RS chromosome arm from rye was introgressed into wheat and crossed into many wheat lines, as it improves yield and fungal disease resistance. Pm8 is a powdery mildew resistance gene on 1RS which, after widespread agricultural cultivation, is now widely overcome by adapted mildew races. Here we show by homology‐based cloning and subsequent physical and genetic mapping that Pm8 is the rye orthologue of the Pm3 allelic series of mildew resistance genes in wheat. The cloned gene was functionally validated as Pm8 by transient, single‐cell expression analysis and stable transformation. Sequence analysis revealed a complex mosaic of ancient haplotypes among Pm3‐ and Pm8‐like genes from different members of the Triticeae. These results show that the two genes have evolved independently after the divergence of the species 7.5 million years ago and kept their function in mildew resistance. During this long time span the co‐evolving pathogens have not overcome these genes, which is in strong contrast to the breakdown of Pm8 resistance since its introduction into commercial wheat 70 years ago. Sequence comparison revealed that evolutionary pressure acted on the same subdomains and sequence features of the two orthologous genes. This suggests that they recognize directly or indirectly the same pathogen effectors that have been conserved in the powdery mildews of wheat and rye.     

Lipid and fatty acid variations in muscle tissues of the ‘yellow’ stage of the European eel (Anguilla anguilla) during short-term adaptation to freshwater and seawater under food deprivation

Changes in body mass and water content (WC), lipids, and fatty acid (FA) composition of muscle were determined in European eels (Anguilla anguilla) in the ‘yellow’ stage (sexually immature). The animals were held in either freshwater (FW) or seawater (SW) and deprived of food for four weeks. During this period, body mass decreased steadily. This decrease was accompanied by reduced muscle WC and total lipid in both SW and FW. ‘Yellow’ eels appeared to use lipids in muscle to provide metabolic and osmoregulatory energy. However, they maintained relative constancy in the FA profiles, so that total FA, saturated, monounsaturated, and polyunsaturated FA, were similar between salinities during short-term starvation.     

Assessment of Ki-67 as a potential biomarker in patients with breast cancer

Breast cancer is the most common cancer in females, it accounts for one third of all malignancies affecting women. Appropriate biomarkers play significant role in predicting the prognosis and decide the specific therapy to each patient. In this study we aimed at evaluating the value of Ki-67 as a prognostic marker in breast cancer patients and to analyze the associations between Ki-67 and their clinicopathological parameters. This study included 92 patients with developed non metastatic breast cancer and 10 women had benign breast tumor served as controls. We measured the serum level by ELISA technique and tissue expression of Ki-67 by immunohistochemical technique. Our results showed that there were no statistically significant differences in serum Ki-67 levels between the two studied groups. As for Ki-67expression in breast cancer cells, the score increases with increase of tumor size, grade, premenopausal, Ki-67 expression in estrogen and progesterone receptor positive tumors showed lower values than estrogen and progesterone negative tumors, while higher Ki-67 expression was more frequently associated with HER2-positive. In conclusion; our study supports the finding that tissue Ki-67 expression may add prognostic information to that obtained from classical prognostic factors and can provide data of significant value to other important prognostic indicators such as pathological grading, and axillary lymph node involvement.     

Early Detrimental Metabolic Outcomes of rs17300539‐A Allele of ADIPOQ Gene Despite Higher Adiponectinemia

Minor allele A of single‐nucleotide polymorphism (SNP) 11391 G/A of ADIPOQ gene (rs17300539) has been consistently associated with higher adiponectin levels in adults and children. The aim of this study was to investigate the metabolic role of this variant in a large cohort of children of European origin. A total of 1,852 children from two general populations in Verona and in Fleurbaix–Laventie and from the Lille childhood obesity cohort, were genotyped and pooled together after checking for the absence of genetic heterogeneity for rs17300539 between Italian and French children. The genotype of rs17300539 was studied in relation to circulating adiponectin levels, BMI, fasting plasma glucose, fasting serum insulin (FSI), insulin resistance index (homeostasis model assessment of insulin resistance (HOMA_IR)), high‐density lipoprotein cholesterol, and triglycerides. After adjustment for known confounders, rs17300539 GA+AA carriers had 1.6 µg/ml higher adiponectin levels (P = 6 × 10^?8) than GG carriers. They also showed higher BMI (B = 0.97, P = 0.015) and higher prevalence of obesity (OR = 1.35 (1.06–1.85), P = 0.015) than GG carriers. Before adjusting for obesity status, GA+AA carriers had higher FSI (B = 1.10, P = 0.040) and higher HOMA_IR (B = 0.31, P = 0.020) than GG carriers. After adjustment for obesity status, they did not differ from GG carriers for any metabolic parameter, either among obese or nonobese children. The rs17300539‐A variant, though consistently associated with higher adiponectin levels, does not exert any appreciable protective metabolic effect in children, either in the presence or absence of obesity. In contrast, this SNP may increase the risk for childhood obesity and related insulin resistance.     

Diagnosis of Coronary Heart Diseases Using Gene Expression Profiling; Stable Coronary Artery Disease,Cardiac Ischemia with and without Myocardial Necrosis

Cardiovascular disease (including coronary artery disease and myocardial infarction) is one of the leading causes of death in Europe, and is influenced by both environmental and genetic factors. With the recent advances in genomic tools and technologies there is potential to predict and diagnose heart disease using molecular data from analysis of blood cells. We analyzed gene expression data from blood samples taken from normal people (n = 21), non-significant coronary artery disease (n = 93), patients with unstable angina (n = 16), stable coronary artery disease (n = 14) and myocardial infarction (MI; n = 207). We used a feature selection approach to identify a set of gene expression variables which successfully differentiate different cardiovascular diseases. The initial features were discovered by fitting a linear model for each probe set across all arrays of normal individuals and patients with myocardial infarction. Three different feature optimisation algorithms were devised which identified two discriminating sets of genes, one using MI and normal controls (total genes = 6) and another one using MI and unstable angina patients (total genes = 7). In all our classification approaches we used a non-parametric k-nearest neighbour (KNN) classification method (k = 3). The results proved the diagnostic robustness of the final feature sets in discriminating patients with myocardial infarction from healthy controls. Interestingly it also showed efficacy in discriminating myocardial infarction patients from patients with clinical symptoms of cardiac ischemia but no myocardial necrosis or stable coronary artery disease, despite the influence of batch effects and different microarray gene chips and platforms.     

Prevalence and Correlates of HIV and Hepatitis C Virus Infections and Risk Behaviors among Malaysian Fishermen

Fishermen in Southeast Asia have been found to be highly vulnerable to HIV, with research evidence highlighting the role of sexual risk behaviors. This study aims to estimate the rate of HIV as well as hepatitis C virus (HCV) infections among Malaysian fishermen, and the risky sexual and injection drug use behaviors that may contribute to these infections. The study also includes an assessment of socio-demographic, occupational and behavioral correlates of testing positive for HIV or HCV, and socio-demographic and occupational correlates of risk behaviors. The study had a cross-sectional design and recruited 406 fishermen through respondent-driven sampling (RDS). Participants self-completed a questionnaire and provided biological specimens for HIV and HCV testing. We conducted and compared results of analyses of both unweighted data and data weighted with the Respondent-Driven Sampling Analysis Tool (RDSAT). Of the participating fishermen, 12.4% were HIV positive and 48.6% had HCV infection. Contrary to expectations and findings from previous research, most fishermen (77.1%) were not sexually active. More than a third had a history of injection drug use, which often occurred during fishing trips on commercial vessels and during longer stays at sea. Of the fishermen who injected drugs, 42.5% reported unsafe injection practices in the past month. Reporting a history of injection drug use increased the odds of testing HIV positive by more than 6 times (AOR = 6.22, 95% CIs [2.74, 14.13]). Most fishermen who injected drugs tested positive for HCV. HCV infection was significantly associated with injection drug use, being older than 25 years, working on a commercial vessel and spending four or more days at sea per fishing trip. There is an urgent need to strengthen current harm reduction and drug treatment programs for Malaysian fishermen who inject drugs, especially among fishermen who work on commercial vessels and engage in deep-sea fishing.     

Fourier-transform infrared and Raman spectroscopic evidence for the incorporation of cinnamaldehydes into the lignin of transgenic tobacco (Nicotiana tabacum L.) plants with reduced expression of cinnamyl alcohol dehydrogenase

Xylem from stems of genetically manipulated tobacco plants which had had cinnamyl alcohol dehydrogenase (CAD; EC 1.1.1.195) activity down-regulated to a greater or lesser degree (clones 37 and 49, respectively) by the insertion of antisense CAD cDNA had similar, or slightly higher, lignin contents than xylem from wild-type plants. Fourier-transform infrared (FT-IR) microspectroscopy indicated that down-regulation of CAD had resulted in the incorporation of moieties with conjugated carbonyl groups into lignin and that the overall extent of cross-linking, particularly of guaiacyl (4-hydroxy-3-methoxyphenyl) rings, in the lignin had altered. The FT-Raman spectra of manipulated xylem exhibited maxima consistent with the presence of elevated levels of aldehydic groups conjugated to a carbon-carbon double bond and a guaiacyl ring. These maxima were particularly intense in the spectra of xylem from clone 37, the xylem of which exhibits a uniform red coloration, and their absolute frequencies matched those of coniferaldehyde. Furthermore, xylem from clone 37 was found to have a higher content of carbonyl groups than that of clone 49 or the wild-type (clone 37: clone 49: wild-type; 2.4:1.6:1.0) as measured by a degradative chemical method. This is the first report of the combined use of FT-IR and FT-Raman spectroscopies to study lignin structure in situ. These analyses provide strong evidence for the incorporation of cinnamaldehyde groups into the lignin of transgenic plants with down-regulated CAD expression. In addition, these non-destructive analyses also suggest that the plants transformed with antisense CAD, in particular clone 37, may contain lignin that is less condensed (cross-linked) than that of the wild-type. Received: 27 May 1996 / Accepted: 30 July 1996     

Temperature and pH interaction in NO3 uptake

Summary The interaction effects of temperature, pH and the presence of the poly-valent cation aluminium on nitrate uptake by excised barley roots have been studied. Results indicate that nitrate uptake increased with temperature at the two pH values studied. NO₃ ⁻ uptake was higher from pH6 than from pH4 at all temperatures in the absence of Al⁺³. The addition of Al⁺³ to the external solution modifies this effect. Different theories are given to explain these interactions.