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PI3K/AKT/mTOR pathway is one of the frequently disrupted signaling pathways in renal cell carcinoma (RCC) that plays a significant role in tumor formation, disease progression and therapeutic resistance. Therefore, novel natural molecules targeting the critical proteins of this pathway will provide the best alternative to existing drugs, which are toxic and develops resistance. Recent studies have recognized the anti-cancer therapeutic potential of mycocompounds. The current study is focused on screening various mycocompounds from

Neural tube defects (NTDs) are significant congenital deformities of the central nervous system among which spina bifida is the most common form that occurs due to defect in the neurulation process of embryogenesis. NTDs are among the most common type of birth defects occurring at a range of 0.5–10 in every 1000 live births worldwide and are thought to have multifactorial etiology, including multigenetic and epigenetic notions. Epigenetic regulations control differential gene expression in normal and disease phenotypes. DNA methylation is a significant epigenetic process, guided by DNMT1, one of the most important maintenance methylating agents. However, the relationship between DNMT1 and NTDs had always been inconclusive and poorly understood. In the present study, by utilizing in silico methodologies we tried to figure out potent single nucleotide variants (SNVs) that could play roles in generating functional differences in DNMT1 expression and we also tried to check (by in vitro method) if there is any connection between DNMT1 expression and spina bifida condition. A number of coding and non-coding (both intragenic and intergenic) SNVs of DNMT1 were found (using the in silico methods) that have potentials to alter its expression. From the in vitro experimentations, differential DNMT1 RNA expressions were found between spina bifida affected newborns and their respective mothers when compared with controls. It is the first report of NTD from Eastern India precisely showing inverse correlation between DNMT1 expression and occurrence of NTD. The findings of the present study could be further considered for early prognosis and future experimental designs.

Kidney stone disease (KSD) or nephrolithiasis (NL), is a serious clinical concern that gradually poses threat to global health and economy. The frequency of KSD is rapidly increasing and about 12% of the Indian population are suffering from this disease. In this scenario, we studied the incidence and age group stratification of KSD in the eastern part of India. Furthermore, patients with GRHPR mutation were highly predominant. Furthermore, real time quantitative PCR expression and protein expression through immunoblotting along with protein activity & computational analysis were carried out in patients with GRHPR mutation. Our findings revealed that the candidates of the early age group are prone to hyperoxaluria moreover a potent and frequent mutation in the GRHPR gene is widely present in this age group. Further, biochemical tests such as serum creatine, serum urea, serum and urinary calcium were carried out for the patients along with age matched controls. Moreover, the actual underlying cause of nephrolithiasis for lower age groups still remains unclear. The study revealed that mutations were more often found in GRHPR, AGXT, and HOGA1 genes in the study cohort.

As SARS-CoV-2 variants emerge, there is a critical need to understand the effectiveness of serum elicited by different SARS-CoV-2 vaccines. A reference reagent comprised of post-vaccination sera from recipients of different vaccines allows evaluation of in vitro variant neutralization, and provides a reference for comparing assay results across laboratories. We prepared and pooled >1 L serum from donors who received the SARS-CoV-2 mRNA vaccines (BNT162b2, Pfizer and mRNA-1273, Moderna), a replication-incompetent adenovirus type 26 vaccine (Ad26.COV2.S, Johnson and Johnson), or recombinant spike protein expressed by baculovirus incorporated into a nanoparticle vaccine plus Matrix-M adjuvant (NVX-CoV2373, Novavax). Twice frozen sera were aliquoted and are available for distribution to the research community (BEI Resources). The calculated WHO titer of pooled sera to spike protein was 1,312, 1,447, 1,936, and 587 and the reciprocal RBD binding to ACE-2 IC90-titers were 60, 64, 118, and 46 for BNT162b2, mRNA1273, Ad26.CoV2373, and NVX-CoV2373 sera, respectively.

Gastric cancer has emerged as a key challenge in oncology research as a malignant tumour with advanced stage detection. Apart from surgical management, a pharmacotherapeutic approach to stomach cancer treatment is an appealing option to consider. Andrographolide has been shown to have anticancer and chemosensitizer properties in a variety of solid tumors, including stomach cancer but the exact molecular mechanism is skeptical. In this study, we identified and validated pharmacological mechanism involved in the treatment of GC with integrated approach of network pharmacology and molecular docking. The targets of andrographolide and GC were obtained from databases. The intersected targets between andrographolide and GC-related genes were used to construct protein–protein interaction (PPI) network. Furthermore, mechanism of action of the targets was predicted by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Finally, these results were validated by molecular docking experiments, mRNA and protein expression level. A total of 197 targets were obtained for andrographolide treating GC. Functional enrichment analysis revealed that the target genes were exerted promising therapeutic effects on GC by HIF-1 and PI3K-Akt signaling pathway. The possible mechanism of action is by inactivation of HIF-1 signaling pathway which is dependent on the inhibition of upstream PI3K-AKT pathway. The PPI network identified SRC, AKT1, TP53, STAT3, PIK3CA, MAPK1, MAPK3, VEGFA, JUN and HSP90AA1 as potential hub targets. In addition, these results were further validated with molecular docking experiments. Survival analysis indicated that the expression levels of the hub genes were significantly associated with the clinical prognosis of GC. This study provided a novel approach to reveal the therapeutic mechanisms of andrographolide on GC, making future clinical application of andrographolide in the treatment of GC.

The polymorphic variation of MTR and MTRR genes can influence the folate metabolism and homeocysteineremethylation processes and consequently has strong impact on Neural Tube Defects (NTD). The SNP of MTR and MTRR genes has been reported in some populations, but in the Indian population has never been explored. This study is focused on the frequency distributions of rs1805087 (D919G/A2756G) and rs1801394 (I22M/A66G) of these genes in the West Bengal population, India for the first time. The G allele frequencies of rs1085087 and rs1801394 were 8% and 47% respectively. The G allele frequency of MTR rs1085087 in Indian population is less than the other major population; such as, in European (17%), American (18%), African (28%) and South Asian (32%) population. The G allele frequency of MTRR rs1801394 is quite like the European and south Asian population (G=52%) but greatly differs from African (25%), East Asian (26%) and mixed American (28%) population. This observation might help us to understand pattern of genetic alteration and development of single screening protocol for early detection and better prognosis in future.

Renal tubular acidosis (RTA) is a heterogenous group of relatively uncommon tubular disorders which are characterised by hyperchloremic metabolic acidosis with a normal anion gap. It usually worsen...

Background The aim of this study was to evaluate the risk of gastric cancer associated with individual or combined glutathione S-transferases (GSTs) polymorphism and their interaction with environmental factors. Materials and Methods Genotyping by PCR was carried out for 80 cases and controls each for GSTM1, GSTT1, and GSTP1 polymorphism and mapped for gene—environment association studies. The samples were subjected to pathogen detection and GSTP1 expression for analyzing their association with different genotypes. Logistic regression analyses were conducted to compute the influence of both genetic and environmental factors for gastric cancer. MDR analysis was performed to assess the risk of gastric cancer by studying the gene–gene and gene–environment effect on the basis of GST genotyping and GSTP1 gene expression. Results Infection with Helicobacter pylori and CagA+ strains was more frequent in patients with GSTM1/T1 null genotype. Intake of high fermented fat and smoked meat was found to be significantly associated with gastric cancer. The G/G, A/G (rs1695), and T/T (rs1138272) were found to be significantly associated with low expression of GSTP1 gene in cancer tissue. Conclusion Presence of H. pylori with CagA genotype showed significant individual effect with GSTT1 polymorphism as well as strong synergistic effect in gastric cancer risk. Majority of the gastric cancer samples showed significant negative expression in G/G, A/G (rs1695), and T/T (rs1138272) genotypes. This study shows that GST gene polymorphism was significantly relevant for determining the individual susceptibility to gastric cancer.

Genetically caused nonsyndromic hearing loss is highly heterogeneous. Inspite of this large heterogeneity, mutations in the genes GJB2, GJB6 and SLC26A4 are major contributors. The mutation spectrum of these genes varies among different ethnic groups. Only a handful of studies focused on the altered genetic signature of these genes in different demographic regions of India but never focused on the eastern part of the country. Our study for the first time aimed to characterize the mutation profile of these genes in hearing loss patients of West Bengal state, India. Mutations in GJB2, GJB6 and SLC26A4 genes were screened by bidirectional sequencing from 215 congenital nonsyndromic hearing loss patients. Radiological diagnosis was performed in patients with SLC26A4 mutations by temporal bone CT scan. The study revealed that 4.65% and 6.97% patients had monoallelic and biallelic GJB2 mutations respectively. Six mutations were identified, p.W24X being the most frequent one accounting for 71.05% of the mutated alleles. Mutations in GJB6 including the previously identified deletion mutation (GJB6-D13S1830) were not identified in our study. Further, no patients harbored biallelic mutations in the SLC26A4 gene or the common inner ear malformation Enlarged Vestibular Aqueduct (EVA). The mutation profile of GJB2 in our study is distinct from other parts of India, suggesting that the mutation spectrum of this gene varies with ethnicity and geographical origin. The absence of GJB6 mutations and low frequency of SLC26A4 mutations suggest that additional genetic factors may also contribute to this disease.

Alcohol is eliminated from the body by various metabolic mechanisms. The primary enzymes in such mechanism involved are alcohol dehydrogenase, aldehyde dehydrogenase, cytochrome P450 2E1, and catalase. Variations in the genes for these enzymes have been found to influence alcohol consumption. The consequences of alcohol metabolism include oxygen deficits (i.e., hypoxia) in the liver, resulting in the formation of harmful compounds (i.e., adducts) and highly reactive oxygen-containing molecules (i.e., reactive oxygen species) that can damage cell components. Approximately, worldwide 3.6 % of cancers derive from chronic alcohol drinking, including those of the upper aerodigestive tract, the liver, the colorectum and the breast. Although the mechanisms for alcohol-associated carcinogenesis are not completely understood, recent findings have focused on acetaldehyde, the first and most toxic ethanol metabolite, as a cancer-causing agent. Alcohol-related carcinogenesis may aggravate due to other factors such as smoking and being triggered by genetic susceptibility. Besides, the role of genetic polymorphisms of the alcohol-metabolizing enzymes could not be ruled out.

Kidney Stone Disease (KSD) is a complex urologic disorder with strong genetic constituent. Earlier association studies have indicated that the genetic polymorphisms are the potential cause of stone materialization; however unfortunately, the actual genetic signature is still unknown. Therefore, present study was aimed to investigate the potential contribution of two important polymorphisms of calcitonin receptor gene (CALCR): (i) rs1801197 (Leu447Pro) and (ii) rs1042138 (3′UTR + 18C > T) in renal stone formation. Accordingly, we enrolled 152 patients registered with calcium-rich stone in kidney (case) and 144 corresponding age, sex and ethnicity matched healthy individuals (controls). Epidemiological and clinical data were recorded as well as peripheral blood sample was collected from each individual. Serum creatinine and urinary calcium level was found high in patients, compared to controls. Out of two studied polymorphisms, we have not found any significant association against the rs1042138 with KSD, nonetheless, significant high frequency (p = 0.001; Odds ratio = 1.81; 95% CI: 1.28–2.55) of risk allele T against the rs1801197 (T > C) in patient was noted. Moreover, significant association with KSD was noted by genotypic analysis of rs1801197 (Leu447Pro) in our population. Interestingly, male patients carrying TT genotype was found to be at high risk of stone formation, while no such association was observed in female patients. Altogether, present study indicated that the rs1042138 might not be used as a useful marker for susceptibility of kidney stone formation, whereas, the rs1801197 could definitely be considered as one of the risk factors for KSD in Indian population at least in West Bengal in particular.

Primers using for amplification of SNPs of ADH1A, ADH1B, ADH1C and ALDH2 gene, Description of data- list all primers used in the study. (DOCX 14Â kb)

Gastric cancer patient report, Description of data- participant questionnaire used in the study. (DOCX 16Â kb)

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disorder with no effective medical treatments available. The generation of myofibroblasts, which are critical for fibrogenesis, requires both a mechanical signal and activated TGF-β; however, it is not clear how fibroblasts sense and transmit the mechanical signal(s) that promote differentiation into myofibroblasts. As transient receptor potential vanilloid 4 (TRPV4) channels are activated in response to changes in plasma membrane stretch/matrix stiffness, we investigated whether TRPV4 contributes to generation of myofibroblasts and/or experimental lung fibrosis. We determined that TRPV4 activity is upregulated in lung fibroblasts derived from patients with IPF. Moreover, TRPV4-deficient mice were protected from fibrosis. Furthermore, genetic ablation or pharmacological inhibition of TRPV4 function abrogated myofibroblast differentiation, which was restored by TRPV4 reintroduction. TRPV4 channel activity was elevated when cells were plated on matrices of increasing stiffness or on fibrotic lung tissue, and matrix stiffness-dependent myofibroblast differentiation was reduced in response to TRVP4 inhibition. TRPV4 activity modulated TGF-β1-dependent actions in a SMAD-independent manner, enhanced actomyosin remodeling, and increased nuclear translocation of the α-SMA transcription coactivator (MRTF-A). Together, these data indicate that TRPV4 activity mediates pulmonary fibrogenesis and suggest that manipulation of TRPV4 channel activity has potential as a therapeutic approach for fibrotic diseases.

Chronic kidney disease (CKD) is a worldwide public health threat and is still unrecognized by many low-income countries as a potentially devastating cause of morbidity and mortality in their population. Many other causes of morbidity and mortality compete for limited country resources both financially and for trained health personnel. Many low-income countries lack data about the rising burden of CKD. In a low-resource setting, where measures of serum creatinine may be unavailable, a simple CKD-screening program could include measures of proteinuria, but, can also include measures of CKD risk factors including blood glucose, blood pressure and body mass index. Screening programs can be coupled with both treatment and patient education programs where resources permit. Many low-income countries have no readily accessible and affordable renal replacement program so patients with ESKD die or must travel outside the country for treatment. Programs that focus both on educating nephrology health-care professionals and on prevention of CKD are necessary. Significant effort will be necessary to develop these resources and increased access to low-cost renal replacement therapy options will be important in the next decade.

Loss of superoxide dismutase (SOD) activity is a defining biochemical feature of asthma. However, mechanisms for the reduced activity are unknown. We hypothesized that loss of asthmatic SOD activity is due to greater susceptibility to oxidative inactivation.Activity assays of blood samples from asthmatics and healthy controls revealed impaired dismutase activity of copper-zinc SOD (CuZnSOD) in asthma. CuZnSOD purified from erythrocytes or airway epithelial cells from asthmatic was highly susceptible to oxidative inactivation. Proteomic analyses identified that inactivation was related to oxidation of cysteine 146 (C146), which is usually disulfide bonded to C57. The susceptibility of cysteines pointed to an alteration in protein structure, which is likely related to the loss of disulfide bond. We speculated that a shift to greater intracellular reducing potential might account for the change. Strikingly, measures of reduced and oxidized glutathione confirmed greater reducing intracellular state in asthma, compared with controls. Similarly, greater free thiol in CuZnSOD was confirmed by ~2-fold greater N-ethylmaleimide binding to C146 in asthma as compared with controls.Greater reducing potential under a chronic inflammatory state of asthma, thus, leads to susceptibility of CuZnSOD to oxidative inactivation due to cleavage of C57-C146 disulfide bond and exposure of usually unavailable cysteines.Vulnerability of CuZnSOD influenced by redox likely amplifies injury and inflammation during acute asthma attacks when reactive oxygen species are explosively generated. Overall, this study identifies a new paradigm for understanding the chemical basis of inflammation, in which redox regulation of thiol availability dictates protein susceptibility to environmental and endogenously generated reactive species.

Background Gastric cancer (GC) is one of the most frequently diagnosed digestive tract cancers and carries a high risk of mortality. Acetaldehyde (AA), a carcinogenic intermediate of ethanol metabolism contributes to the risk of GC. The accumulation of AA largely depends on the activity of the major metabolic enzymes, alcohol dehydrogenase and aldehyde dehydrogenase encoded by the ADH (ADH1 gene cluster: ADH1A, ADH1B and ADH1C) and ALDH2 genes, respectively. This study aimed to evaluate the association between genetic variants in these genes and GC risk in West Bengal, India. Methods We enrolled 105 GC patients (cases), and their corresponding sex, age and ethnicity was matched to 108 normal individuals (controls). Genotyping for ADH1A (rs1230025), ADH1B (rs3811802, rs1229982, rs1229984, rs6413413, rs4147536, rs2066702 and rs17033), ADH1C (rs698) and ALDH2 (rs886205, rs968529, rs16941667 and rs671) was performed using DNA sequencing and RFLP. Results Genotype and allele frequency analysis of these SNPs revealed that G allele of rs17033 is a risk allele (A vs G: OR = 3.67, 95% CI = 1.54–8.75, p = 0.002) for GC. Significant association was also observed between rs671 and incidence of GC (p = 0.003). Moreover, smokers having the Lys allele of rs671 had a 7-fold increased risk of acquiring the disease (OR = 7.58, 95% CI = 1.34–42.78, p = 0.009). Conclusion In conclusion, rs17033 of ADH1B and rs671 of ALDH2 SNPs were associated with GC risk and smoking habit may further modify the effect of rs671. Conversely, rs4147536 of ADH1B might have a protective role in our study population. Additional studies with a larger patient population are needed to confirm our results. Electronic supplementary material The online version of this article (10.1186/s12885-017-3713-7) contains supplementary material, which is available to authorized users.

High levels of arginine metabolizing enzymes, including inducible nitric oxide synthase (iNOS) and arginase (ARG), are typical in asthmatic airway epithelium; however, little is known about the metabolic effects of enhanced arginine flux in asthma. Here, we demonstrated that increased metabolism sustains arginine availability in asthmatic airway epithelium with consequences for bioenergetics and inflammation. Expression of iNOS, ARG2, arginine synthetic enzymes, and mitochondrial respiratory complexes III and IV was elevated in asthmatic lung samples compared with healthy controls. ARG2 overexpression in a human bronchial epithelial cell line accelerated oxidative bioenergetic pathways and suppressed hypoxia-inducible factors (HIFs) and phosphorylation of the signal transducer for atopic Th2 inflammation STAT6 (pSTAT6), both of which are implicated in asthma etiology. Arg2-deficient mice had lower mitochondrial membrane potential and greater HIF-2α than WT animals. In an allergen-induced asthma model, mice lacking Arg2 had greater Th2 inflammation than WT mice, as indicated by higher levels of pSTAT6, IL-13, IL-17, eotaxin, and eosinophils and more mucus metaplasia. Bone marrow transplants from Arg2-deficient mice did not affect airway inflammation in recipient mice, supporting resident lung cells as the drivers of elevated Th2 inflammation. These data demonstrate that arginine flux preserves cellular respiration and suppresses pathological signaling events that promote inflammation in asthma.

The impairment of vasodilator nitric oxide (NO) production is well accepted as a typical marker of endothelial dysfunction in vascular diseases, including in the pathophysiology of pulmonary arterial hypertension (PAH), but the molecular mechanisms accounting for loss of NO production are unknown. We hypothesized that low NO production by pulmonary arterial endothelial cells in PAH is due to inactivation of NO synthase (eNOS) by aberrant phosphorylation of the protein. To test the hypothesis, we evaluated eNOS levels, dimerization, and phosphorylation in the vascular endothelial cells and lungs of patients with PAH compared with controls. In mechanistic studies, eNOS activity in endothelial cells in PAH lungs was found to be inhibited due to phosphorylation at T495. Evidence pointed to greater phosphorylation/activation of protein kinase C (PKC) α and its greater association with eNOS as the source of greater phosphorylation at T495. The presence of greater amounts of pT495-eNOS in plexiform lesions in lungs of patients with PAH confirmed the pathobiological mechanism in vivo. Transfection of the activating mutation of eNOS (T495A/S1177D) restored NO production in PAH cells. Pharmacological blockade of PKC activity by β-blocker also restored NO formation by PAH cells, identifying one mechanism by which β-blockers may benefit PAH and cardiovascular diseases through recovery of endothelial functions.

Accurate Species diagnosis is the key element for biodiversity studies and conservation planning. Conventionally, morphological characters are used to identify a species. But, this approach needs a thorough expertise in identifying the external features which often leads to narrowing down of specialization with regard to ascertaining a species within a limited group of taxa. The approach may be particularly valuable for species identification of organisms that are rare, fragile, and/or small, especially when morphological identification is problematic and errors are likely due to simple or evolutionarily conserved body plans. However, each time a new technique has been introduced in science it was accompanied by some debate and distress, and DNA barcoding was no exception. Therefore, more collaborative efforts are needed to explore the potentialities of DNA barcoding in proper species identification across all taxa. At the same time, we need to set a threshold of the genetic variation in species delimitation to find out the cryptic species. It is also an important point to know that the benefits of DNA barcoding are not restricted to taxonomic or systematic research only. The discovery of high-throughput sequencing technologies are going to change the dimension of these techniques in the years to come.

Asthma, a chronic airway inflammatory disease is typically associated with high levels of exhaled nitric oxide (NO). Over the past decades, extensive research has revealed that NO participates in a number of metabolic pathways that contribute to animal models of asthma and human asthma. In asthmatic airway, high levels of NO lead to greater formation of reactive nitrogen species (RNS), which modify proteins adversely affecting functional activities. In contrast, high levels of NO are associated with lower than normal levels of S-nitrosothiols, which serve a bronchodilator function in the airway. Detailed mechanistic studies have enabled the development of compounds that target NO metabolic pathways, and provide opportunities for novel asthma therapy. This review discusses the role of NO in asthma with the primary focus on therapeutic opportunities developed in recent years.

Data on the burden of acute kidney injury (AKI) in resource-poor countries such as Tanzania are minimal because of a lack of nephrology services and an inability to recognize and diagnose AKI with any certainty. In the few published studies, high morbidity and mortality are reported. Improved nephrology care and dialysis may lower the mortality from AKI in these settings. Hemodialysis is expensive and technically challenging in resource-limited settings. The technical simplicity of peritoneal dialysis and the potential to reduce costs if consumables can be made locally, present an opportunity to establish cost-effective programs for managing AKI. Here, we document patient outcomes in a pilot peritoneal dialysis program established in 2009 at a referral hospital in Northern Tanzania.

Iron homeostasis influences the development of pulmonary arterial hypertension (PAH) associated with hypoxia or hematologic disorders. To investigate whether severity of idiopathic PAH (IPAH) is impacted by alterations in iron metabolism, we assessed iron metabolic markers, including levels of zinc‐protoporphyrin (Zn‐pp), transferrin receptor, and red blood cell numbers and morphology in IPAH, associated PAH and sleep apnea‐induced pulmonary hypertension patients in comparison to healthy controls and asthmatics. Despite similarly normal measures of iron metabolism, Zn‐pp levels in IPAH and sleep apnea patients were elevated approximately twofold, indicating deficient iron incorporation to form heme and levels were closely related to measures of disease severity. Consistent with high Zn‐pp, PAH patients had increased red cell distribution width (RDW). In an expanded cohort including patients with IPAH and familial disease, the RDW was validated and related to clinical parameters of severity; including pulmonary artery pressures and 6‐minute walk distances. These results reveal an increased prevalence of subclinical functional iron deficiency in primary forms of PAH that is quantitatively related to disease severity. This suggests that altered iron homeostasis influences disease progression and demonstrates the importance of closely monitoring iron status in PAH patients. Clin Trans Sci 2011; Volume 4: 253–258

The low barometric pressure at high altitude causes lower arterial oxygen content among Tibetan highlanders, who maintain normal levels of oxygen use as indicated by basal and maximal oxygen consumption levels that are consistent with sea level predictions. This study tested the hypothesis that Tibetans resident at 4,200 m offset physiological hypoxia and achieve normal oxygen delivery by means of higher blood flow enabled by higher levels of bioactive forms of NO, the main endothelial factor regulating blood flow and vascular resistance. The natural experimental study design compared Tibetans at 4,200 m and U.S. residents at 206 m. Eighty-eight Tibetan and 50 U.S. resident volunteers (18–56 years of age, healthy, nonsmoking, nonhypertensive, not pregnant, with normal pulmonary function) participated. Forearm blood flow, an indicator of systemic blood flow, was measured noninvasively by using plethysmography at rest, after breathing supplemental oxygen, and after exercise. The Tibetans had more than double the forearm blood flow of low-altitude residents, resulting in greater than sea level oxygen delivery to tissues. In comparison to sea level controls, Tibetans had >10-fold-higher circulating concentrations of bioactive NO products, including plasma and red blood cell nitrate and nitroso proteins and plasma nitrite, but lower concentrations of iron nitrosyl complexes (HbFe II NO) in red blood cells. This suggests that NO production is increased and that metabolic pathways controlling formation of NO products are regulated differently among Tibetans. These findings shift attention from the traditional focus on pulmonary and hematological systems to vascular factors contributing to adaptation to high-altitude hypoxia.

Nitration is a posttranslational modification that can compromise protein function. We hypothesized that nitration of growth factors secreted in the lung may alter their interaction with their respective receptors and modulate the normal growth and differentiation program induced by ligand-receptor interaction. We tested this hypothesis in vitro by nitration of neuregulin-1's (NRG-1) EGF-like domain and studying the effect on NRG-1's activity. Nitration of NRG-1's (nNRG-1) EGF-like domain resulted in an inability to activate its receptor, the human epidermal growth factor receptors 2 and 3 (HER2/HER3) heterodimer, as defined by loss of HER2 tyrosine phosphorylation induced by nNRG-1 in MCF-7 cells. Receptor activation was not restored with increasing nNRG-1 concentration or exposure times. nNRG-1 did not compete with NRG-1 for HER2/HER3 binding in competition assays. In addition, nNRG-1 no longer induced proliferation of the MCF-7 cell line, as MCF-7 cells exposed to nNRG-1 and NRG-1 concurrently had the same proliferation rate as that induced by NRG-1 alone. Thus nitration of NRG-1's EGF-like domain caused it to lose its ability to bind and activate its receptor with loss of ligand-induced proliferation. Posttranslational nitration of growth factors in states where reactive nitrogen species are increased may be an important means of regulating growth factor receptor effects in the lung.

Gastric cancer is one of the most common malignancies in India. DNA repair gene or xenobiotic pathway gene polymorphisms have recently been shown to affect individual susceptibility to gastric cancer. Here, the possible interaction between common polymorphisms in X-ray repair cross complementing group I (XRCC1) gene and glutathione S-transferase (GST) genes (GSTM1, GSTT1 and GSTP1), smoking and alcohol consumption and overall survival in gastric cancer patients were evaluated. In this population-based case control study, 70 gastric cancer patients and 82 healthy controls were enrolled. The epidemiological data were collected by a standard questionnaire, and blood samples were collected from each individual. XRCC1 Arg194Trp, Arg280His and Arg399Gln polymorphisms were determined by polymerase chain reaction and direct DNA sequencing. GSTM1 and GSTT1 null polymorphisms and GSTP1 Ile105Val polymorphism were identified by multiplex polymerase chain reaction and restriction fragment length polymorphism (RFLP), respectively. The risk of gastric cancer was significantly elevated in individuals with XRCC1 Arg/Gln +Gln/Gln (p = 0.031; odds ratio = 2.32; 95 % confidence interval (CI) 1.07-5.06) and GSTP1 Val/Val genotype (p = 0.009; odds ratio = 8.64; 95 % CI 1.84-40.55). An elevated risk for GC was observed in smokers and alcohol consumers carrying GSTP1 Ile/Val +Val/Val genotype (p = 0.041; odds ratio = 3.71; 95 % CI 0.98-14.12; p = 0.002; odds ratio = 12.31; 95 % CI 1.71-88.59). These findings suggest that XRCC1 rs25487 and GSTP1 rs1695 can be considered as a risk factor associated with gastric cancer and might be used as a molecular marker for evaluating the susceptibility of the disease.

Airway mucus in cystic fibrosis (CF) is highly elastic, but the mechanism behind this pathology is unclear. We hypothesized that the biophysical properties of CF mucus are altered because of neutrophilic oxidative stress. Using confocal imaging, rheology, and biochemical measures of inflammation and oxidation, we found that CF airway mucus gels have a molecular architecture characterized by a core of mucin covered by a web of DNA and a rheological profile characterized by high elasticity that can be normalized by chemical reduction. We also found that high levels of reactive oxygen species in CF mucus correlated positively and significantly with high concentrations of the oxidized products of cysteine (disulfide cross-links). To directly determine whether oxidation can cross-link mucins to increase mucus elasticity, we exposed induced sputum from healthy subjects to oxidizing stimuli and found a marked and thiol-dependent increase in sputum elasticity. Targeting mucin disulfide cross-links using current thiol-amino structures such as N-acetylcysteine (NAC) requires high drug concentrations to have mucolytic effects. We therefore synthesized a thiol-carbohydrate structure (methyl 6-thio-6-deoxy-α-D-galactopyranoside) and found that it had stronger reducing activity than NAC and more potent and fast-acting mucolytic activity in CF sputum. Thus, oxidation arising from airway inflammation or environmental exposure contributes to pathologic mucus gel formation in the lung, which suggests that it can be targeted by thiol-modified carbohydrates.

Kidney stone disease (KSD) is a major clinical problem imposing a large burden for both healthcare and economy globally. In India, the prevalence of kidney stone disease is rapidly increasing. This study aimed to evaluate the association between genetic defects in vitamin D receptor (VDR), calcium sensing receptor (CaSR) and claudin 14 (CLDN14) genes and kidney stone disease in patients from eastern India. We enrolled 200 consecutive kidney stone patients (age 18–60 years) (cases) and their corresponding sex and age matched 200 normal individuals (controls). To identify genetic variants responsible for KSD, we performed sequence analysis of VDR, CaSR and CLDN14 genes. Four non-synonymous (rs1801725, rs1042636, rs1801726 and rs2228570), one synonymous (rs219780) and three intronic single nucleotide polymorphisms (SNPs) (rs731236, rs219777 and rs219778) were identified. Genotype and allele frequency analysis of these SNPs revealed that, rs1801725 (Ala986Ser), rs1042636 (Arg990Gly) of CaSR gene and rs219778, rs219780 (Thr229Thr) of CLDN14 gene were significantly associated with KSD. Serum calcium levels were significantly higher in subjects carrying 986Ser allele and calcium excretion was higher in subjects bearing 990Gly allele. In conclusion, rs1801725, rs1042636, rs219778 and rs219780 SNPs were associated with kidney stone risk in patients from the eastern part of India.

To the Editor: Glutathione (l-γ-glutamyl-l-cysteinylglycine) is found at exceptionally high levels in the human lung (1–3), where it serves as the primary redox couple in the reduced GSH and oxidized GSSG forms. We have identified that the airway epithelium of patients with asthma is shifted to greater reducing potential, recognized by a higher ratio of reduced to oxidized glutathione (4), which occurs in response to the episodic oxidative inflammation of asthma (1, 5–9). On this basis, we hypothesized that regional airway inflammation in asthma may be quantitatively evaluated by a nuclear imaging strategy based on uptake of a radiopharmaceutical (99mTc-exametazime [HMPAO]) selectively retained in tissues depending on GSH levels (10–13) (see Figure E1 in the online supplement). To test this concept, airway redox, inflammation, and 99mTc-HMPAO uptake determined by single-photon emission computed tomography (SPECT) were evaluated in individuals with asthma and in control subjects (Table E1). Some of the results of these studies have been previously reported in the form of abstracts (14–16). 99mTc-HMPAO (Ceretec; GE Healthcare, Arlington Heights, IL) readily diffuses into cells, where it is reduced to a hydrophilic form in the presence of GSH and retained in tissues (Figure E1) (10–13). 99mTc-HMPAO accumulation in the lung was confirmed to be glutathione dependent as indicated by markedly less uptake in mice depleted of glutathione with diethyl maleate (17) as compared with naive mice (Figure E1). Many studies report an increase in total glutathione in asthmatic lungs that occurs in an adaptive response to the episodic asthmatic exacerbations that are characterized by profound oxidative bursts of inflammatory cells in the airways (1, 5–9). Here, in anticipation of imaging, intracellular glutathione was determined in airway epithelial cells obtained at bronchoscopy from patients with asthma and control subjects to assess the magnitude of difference in redox potential between asthma and control lungs. Airway epithelium was obtained by gently brushing the airway, and the freshly obtained cells were immediately processed in the bronchoscopy suite for determination of reduced and oxidized glutathione and reducing potential in human airway epithelium of control subjects and patients with asthma in vivo. Similar to a prior report (4), GSH tended to be higher but GSSG lower in asthma (GSH, μM: patients with asthma, 17 ± 3 [n = 3]; control subjects, 8.9 ± 0.5 [n = 3]; P = 0.14; GSSG, μM: patients with asthma, 0.38 ± 0.04; control subjects, 0.48 ± 0.08; P = 0.2), so that the ratio of GSH to GSSG in the airway epithelium of patients with asthma was double that of control subjects (GSH/GSSG: patients with asthma, 43 ± 6; control subjects, 20 ± 4; P = 0.05) (Figure 1A). The reduction potential (Eh) of the GSH:GSSG couple calculated by Nernst equation (18, 19) revealed a significantly greater intracellular reducing state in the airway epithelium of patients with asthma as compared with control subjects (Eh, mV: patients with asthma, –323 ± 4; control subjects, –306 ± 3; P = 0.04) (Figure 1A). On this basis, we tested whether redox-based imaging might differentiate asthmatic from control lungs, and used SPECT and coregistered computed tomography (CT) imaging to quantitate 99mTc-HMPAO concentration and to determine anatomic sites of 99mTc-HMPAO retention (detailed imaging methods are presented in the online supplement). 99mTc-HMPAO SPECT/CT imaging revealed visibly greater HMPAO uptake in asthmatic lungs as compared with control lungs (Figures 1C and 1D; Movie E1). Normalized uptake values (NUVs) were significantly higher in patients with asthma than control subjects within central and middle transverse areas of lower lung regions (Figure 1E). Interestingly, there was significant variance in the 99mTc-HMPAO NUV of upper and lower lung regions of patients with asthma. Control lung uptake was similar among upper and lower lungs, and across the central, middle, and peripheral transverse regions of the lung. In contrast, patients with asthma had significantly greater 99mTc-HMPAO uptake in lower lung as compared with upper lung regions, and the greatest uptake within central lung regions as compared with middle and peripheral transverse regions (Figure 1E and Table 1). The finding of greater uptake in central lung regions is consistent with a greater distribution of conducting airways in central regions, and the notion of greater reducing potential in asthmatic airways. Overall, the 99mTc-HMPAO uptake findings indicate a greater than normal reducing potential in asthmatic airways as compared with normal airways, as well as significant heterogeneity of lung redox, with the most striking changes in the lower and central regions of the asthmatic lung. In this context, the lower and central regions tend to be the most prominent regions of air trapping (20, 21), which also supports the idea of more severe inflammation/airway disease in those regions. Figure 1. Redox imaging of asthmatic lungs. (A) Glutathione, GSH/GSSG, and glutathione (GSH:GSSG) redox potential (Eh) are greater in the airway epithelium of patients with asthma (n = 3) as compared with control subjects (n = 3). (B–D) Quantitation of ... Table 1: Regional Distribution of Inflammation and 99mTc-HMPAO Uptake in Asthmatic Lung The greater 99mTc-HMPAO uptake in lower regions of asthmatic lungs also suggested that inflammation might be greater in lower lung regions. To investigate this, bronchoalveolar lavage (BAL) was performed in the upper and lower lobes of patients with asthma to compare inflammatory cells and cytokines. BAL volume return, protein concentration, and total cell counts were similar among upper and lower lobes (Table 1). Eosinophils and cytokines that typify helper T-cell type 2 (Th2) inflammation, IL-4 and IL-13, were present in greater concentrations in lower lobes as compared with upper lobes (Table 1). In contrast, IFN-γ, a prototypic Th1-type cytokine, tended to be greater in upper lobes (Table 1). The IL-13 levels in BAL were inversely related to IFN-γ levels (R = –0.483, P = 0.02). This letter reveals that there is a greater reducing potential in the asthmatic airway as compared with control lungs but that there is also substantial heterogeneity of reducing potential across the lung in asthma. This supports the general concept that functional imaging based on redox potential may be used to identify intensity and regionality of airway inflammatory disease. It is important to note that the greater uptake of HMPAO may not necessarily be due to the airway epithelium in asthma. In fact, the endothelium is also inflamed in asthma and might also account for the uptake of the HMPAO (22–26). However, vascular endothelial growth factor was similar among upper and lower lung regions in asthma (Table 1). Further, to assess whether the variation of HMPAO uptake in asthma was related to variation in vascular perfusion, the blood flow in the pulmonary circulation was measured in the supine position, using 99mTc-macroaggregated albumin SPECT/CT (patients with asthma, n = 3; control subjects, n = 3). Pulmonary blood flows were not different in upper and lower regions of the lungs (all P > 0.5). Because all subjects underwent 99mTc-HMPAO SPECT/CT in the supine position (during injection, uptake, and scan), this validated that blood flow to upper and lower parts of the lungs was equally distributed and unlikely to be the cause of variance in upper and lower lobe HMPAO uptake in asthma. Likewise, there may be potential effects and/or dependence of 99mTc-HMPAO uptake on participant age or sex. Although numbers are small, there was no apparent association among 99mTc-HMPAO uptake by age or sex (all P > 0.3). Other studies have identified heterogeneity of remodeling and airflow obstruction in the lungs of patients with asthma as identified by anatomical and functional imaging modalities, for example, high-resolution CT imaging and hyperpolarized helium magnetic resonance imaging (21, 27). The participants in this study were patients with stable asthma, and further investigation is necessary to determine whether 99mTc-HMPAO uptake changes with asthma exacerbations and/or with greater asthma severity. Resolution limitation of SPECT imaging precludes the assignment of uptake to specific airways, limiting the generalized application of 99mTc-HMPAO SPECT/CT imaging in asthma. However, future development of redox-imaging compounds detectable by superior resolution modalities, for example, positron emission tomography, would be a valuable approach to visualize sites of remodeling by coregistered CT images and amounts of inflammation by redox imaging. Current studies rely on invasive bronchoscopic approaches to investigate regional asthma inflammation. The invasive nature and risk of these approaches can preclude enrollment of those patients with severe airflow limitation. A noninvasive functional nuclear imaging approach might offer an alternative quantitative solution to advance research in asthma and assess new biologic-based therapies.

The effects of hypothyroidism on parameters of oxidative stress and on intraneuronal distribution of neurofilaments have been investigated in the developing rat brain. Progressive hypothyroidism during the first 4 weeks of postnatal development led to an increase in superoxide dismutase and catalase activity, decline in the level of glutathione and mitochondrial cytochrome c oxidase activity and increase in the level of .OH radical along with enhanced protein carbonylation and lipid peroxidation. Immunocytochemical staining of cryostat sections of normal and hypothyroid cerebella from 25 day postnatal rats with anti neurofilament (NF) light chain (L) antibody showed aberrant accumulation of neurofilaments in the perikaryon of the hypothyroid Purkinje neurons in contrast to relatively uniform distribution in the controls. The morphological and biochemical alterations in the neurons of the developing hypothyroid brain are comparable to those seen in several neurodegenerative diseases.

The role of thyroid hormone (TH) in the expression of neurofilament (NF) genes during the first 3-4 postnatal weeks of rat brain development has been examined. I.p. administration of TH to 2-day-old hypothyroid rats resulted in a 2-fold increase in cerebral NF-M and NF-L mRNAs, while administration to 15-day-old hypothyroid rats led to a 1.5- to 3-fold increase in NF-H mRNA within 2-4 h of hormone injection. Comparison of the level of these mRNAs in cerebra from 5, 10, 15 and 20-day-old normal and hypothyroid rats by Northern blot analysis revealed that hypothyroidism declined the expression of all three mRNAs by 50-70% at all ages examined. Western blot analysis of total protein and cytoskeletal proteins isolated from cerebras of 5, 10, 15, 20 and 25-day-old normal and hypothyroid rats demonstrated an even greater reduction (60-90%) in the expression of NF proteins in the hypothyroid cerebra during the period examined. The overall results show that TH plays an important role in regulating the expression of all three NF genes during rat brain development.

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disorder with marginally effective medical treatment. Myofibroblasts are critical to the fibrogenic lung repair process; however, the mechanism whereby the mechanical signal that leads to myofibroblast generation is sensed and transmitted remains to be determined. As transient receptor potential vanilloid 4 (TRPV4) ion channels are activated by plasma membrane stretch/matrix stiffness, we investigated whether TRPV4 plays a role in myofibroblast differentiation and/or in vivo lung fibrosis. TRPV4 inhibition by small-molecule inhibitors almost completely abrogated both the calcium influx response to TPRV4 agonist in a dose-dependent manner and the myofibroblast differentiation response to transforming growth factor-β. Similar findings were noted on TRPV4 deletion with small interfering RNA or in TRPV4 KO murine lung fibroblasts. Further, TRPV4 exhibited its mechanosensing and myofibroblast-differentiating effect under conditions of matrix stiffness in the pathophysiological range (1–25 kPa). Mechanistically, TRPV4 activity modulates transforming growth factor β actions in a Smad-independent manner, in part through enhanced actomyosin remodeling with resultant nuclear translocation of the α–smooth muscle actin transcription coactivator (myocardin-related transcription factor-A). The myofibroblast differentiation response to actual fibrotic lung tissue was also completely inhibited on TRPV4 blockade. Furthermore, TRPV4 deficiency protects mice from fibrosis-inducing effects of bleomycin (1 or 4 U/kg dose) at the biochemical (75% less hydroxyproline; P < 0.05), histologic, and physiologic levels (57% less impairment of compliance; P < 0.05). These data identify TRPV4 as a long-elusive mechanosensor/transducer that mediates myofibroblast differentiation and in vivo pulmonary fibrogenesis. Successful manipulation of TRPV4 channel activity may be a novel therapeutic approach for fibrotic diseases of the lung and other organs.