Polymorphisms in CaSR and CLDN14 Genes Associated with Increased Risk of Kidney Stone Disease in Patients from the Eastern Part of India

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.     

Inhibitory action of spermidine on formyl-methionyl-leucyl-phenylalanine stimulated inositol phosphate production in human neutrophils

The effect of the polyamine, spermidine, on formyl-methionyl-leucyl-phenylalanine stimulated hydrolysis of polyphosphoinositides was examined in purified human neutrophils by measurement of inositol phosphate production from radioactively labelled inositol. Spermidine inhibited formyl-methionyl-leucyl-phenylalanine stimulated inositol phosphate production by neutrophil in a dose dependent manner. Inhibition of formyl-methionyl-leucyl-phenylalanine stimulated inositol phosphate accumulation by spermidine was maximal at 10 microM and the IC50 value for this effect was 4.2 microM spermidine. This action of spermidine, thought to be mediated by a membrane component other than phospholipase C, may reflect a control mechanism modulating the response of the polyphosphoinositide system.     

Binding of an antagonistic monoclonal antibody to an intact and fragmented EGF-receptor polypeptide

A murine monoclonal antibody (No. 425) raised against human A431 carcinoma cells specifically immunoprecipitates the 170,000 molecular weight epidermal growth factor (EGF)-receptor from extracts of A431 cells as well as from extracts of human placenta and cultured fibroblasts, but does not recognize the murine receptor. Binding to the external domain of the human EGF-receptor was indicated by indirect immunofluorescent staining of fixed nonpermeable cells. The antibody binds to both glyco- and aglycoreceptor forms, indicating that the epitope is a part of the polypeptide chain. Binding of the antibody to the receptor is conformation dependent; i.e., denatured receptors lacking EGF-binding activity are not recognized by the antibody. The results of antibody binding studies indicate that the epitope is closely linked to the EGF binding active site, and is common to both high- and low-affinity EGF-receptors. Interaction of this epitope with the antibody inhibits EGF binding and bioactivity, and triggers receptor down-regulation, but does not generate EGFlike kinase-stimulatory or mitogenic responses either in vitro or in vivo. The antibody was tested for its ability to bind to domain-sized fragments of the 170-kDa EGF-receptor. It can recognize both the proteolytically generated 110-kDa EGF binding peptide, and a soluble 100-kDa EGF-receptor secreted by A431 cells. This indicates that the epitope recognized this antibody retains its conformation after proteolytic separation of the EGF binding domain from the rest of the receptor molecule.     

Ubiquitin Hydrolase UCH-L1 Destabilizes mTOR Complex 1 by Antagonizing DDB1-CUL4-Mediated Ubiquitination of Raptor

Mammalian target of rapamycin (mTOR) is a serine/threonine kinase that regulates processes including mRNA translation, proliferation, and survival. By assembling with different cofactors, mTOR forms two complexes with distinct biological functions. Raptor-bound mTOR (mTORC1) governs cap-dependent mRNA translation, whereas mTOR, rictor, and mSin1 (mTORC2) activate the survival and proliferative kinase Akt. How the balance between the competing needs for mTORC1 and -2 is controlled in normal cells and deregulated in disease is poorly understood. Here, we show that the ubiquitin hydrolase UCH-L1 regulates the balance of mTOR signaling by disrupting mTORC1. We find that UCH-L1 impairs mTORC1 activity toward S6 kinase and 4EBP1 while increasing mTORC2 activity toward Akt. These effects are directly attributable to a dramatic rearrangement in mTOR complex assembly. UCH-L1 disrupts a complex between the DDB1-CUL4 ubiquitin ligase complex and raptor and counteracts DDB1-CUL4-mediated raptor ubiquitination. These events lead to mTORC1 dissolution and a secondary increase in mTORC2. Experiments in Uchl1-deficient and transgenic mice suggest that the balance between these pathways is important for preventing neurodegeneration and the development of malignancy. These data establish UCH-L1 as a key regulator of the dichotomy between mTORC1 and mTORC2 signaling.     

Adenosine transport by the lung

Adenosine, a nucleoside and potent vasodilator, has been found to be taken up by the lung and converted by deamination into inosine and hypoxanthine. In a single circulation through an isolated rat lung, 69.3 +/- 3.3% of infused [14C]adenosine (10 microM) was removed from the circulation. Uptake of [14C]adenosine remained unchanged when deamination of adenosine was inhibited by 8-azaguanine or coformycin. In a single passage of adenosine through the pulmonary artery, very little of the deaminated products appeared in the pulmonary circulation, but when adenosine was recirculated through the pulmonary circulation inosine and hypoxanthine appeared in the venous effluent. These adenosine metabolites were also taken up by the lung. A major portion of the circulating adenosine was transported into the lung, where it was used to synthesize adenine nucleotides. Inhibition of adenosine kinase by iodotubercidin resulted in reduced formation of ATP and ADP. Uptake of adenosine by the lung was saturable on a concentration gradient and was a passive process because it was not affected by the absence of glucose or the presence of ouabain. Km and Vmax for adenosine transport were 0.227 mM and 4.6 mumol.min-1.g lung-1, respectively. Adenosine transport was inhibited by adenosine analogues, and the inhibitions were found to be competitive in nature. These results suggest that a specific and rate-limiting transport system exists in the lung for adenosine.