Expression of non-neuronal cholinergic system in maxilla of rat in vivo

BACKGROUND: Acetylcholine (ACh) is known to be a key neurotransmitter in the central and peripheral nervous systems, which is also produced in a variety of non-neuronal tissues and cell. The existence of ACh in maxilla in vivo and potential regulation role for osteogenesis need further study. RESULTS: Components of the cholinergic system (ACh, esterase, choline acetyltransferase, high-affinity choline uptake, n- and mAChRs) were determined in maxilla of rat in vivo, by means of Real-Time PCR and immunohistochemistry. Results showed RNA for CarAT, carnitine/acylcarnitine translocase member 20 (Slc25a20), VAChT, OCTN2, OCT1, OCT3, organic cation transporter member 4 (Slc22a4), AChE, BChE, nAChR subunits α1, α2, α3, α5, α7, α10, ß1, ß2, ß4, γ and mAChR subunits M1, M2, M3, M4, M5 were detected in rat's maxilla. RNA of VAChT, AChE, nAChR subunits α2, ß1, ß4 and mAChR subunits M4 had abundant expression (2(-ΔCt) > 0.03). Immunohistochemical staining was conducted for ACh, VAChT, nAChRα7 and AChE. ACh was expressed in mesenchymal cells, chondroblast, bone and cartilage matrix and bone marrow cells, The VAChT expression was very extensively while ACh receptor α7 was strongly expressed in newly formed bone matrix of endochondral and bone marrow ossification, AchE was found only in mesenchymal stem cells, cartilage and bone marrow cells. CONCLUSIONS: ACh might exert its effect on the endochondral and bone marrow ossification, and bone matrix mineralization in maxilla.

A Case of Exercise-induced Acute Renal Failure with G774A Mutation in SCL22A12 Causing Renal Hypouricemia

Acute renal failure with severe loin pain which develops after anaerobic exercise is rare. One of predisposing factors of exercise-induced acute renal failure is renal hypouricemia. Idiopathic renal hypouricemia is a genetic disorder characterized by hypouricemia with abnormally high renal tubular uric acid excretion. The mutation in SCL22A12 gene which encodes renal uric acid transporter, URAT1, is the known major cause of this disorder. We here described a 25-yr-old man showing idiopathic renal hypouricemia with G774A mutation in SCL22A12 who presented exercise-induced acute renal failure. There have been a few reports of mutational analysis in Korean idiopathic renal hypouricemia without acute renal failure. This is the first report of genetically diagnosed idiopathic renal hypouricemia with exercise-induced acute renal failure in Korea.

Identification of a kidney-specific mouse organic cation transporter like-1 (mOCTL1)

Organic ion transporters are expressed in various tissues that transport endogenous and exogenous compounds including their metabolites. There are organic anion transporter (OAT), organic cation transporter (OCT), organic anion transporter like protein (OATLP) and organic cation transporter like (OCTL). Considering the variety of charged organic ionic compounds, the existence of numerous isoforms of organic ion transporters can be assumed. In the present study, we have searched for a new isoform in the expressed sequence tag (EST) database using human organic anion transporter 4 (hOAT4) amino acid sequence as a "query". We found a candidate clone (BC021449) from the mouse kidney cDNA library. This clone was identified as an ortholog of ORCTL3 or OCTL-1. The mOCTL1 cDNA consists of 2016 base pairs encoding 551 amino acid residues with 12 putative transmembrane domains. The deduced amino acid sequence of mOCTL1 showed 35 to 40% identity to those of the other members of the OATs and OCTs. According to the tissue distribution, examined by Northern blot analysis, about a 2.4-kb transcript of mOCTL1 was observed in the kidney. About a 90-kDa band was detected when Western blot analysis in the mouse kidney was done by using antibody against synthesized oligopeptide of mOCTL1. The immunohistochemical result showed that mOCTL1 was stained at the glomerulus (the parietal epithelial cells and podocytes), pars recta of proximal tubule, distal convoluted tubules, connecting tubules and collecting tubules. From these results, we conclude that mOCTL1 may be a candidate for an organic ion transporter isoform in the mouse kidney.

Effect of splice-site polymorphisms of the TMPRSS4, NPHP4 and ORCTL4 genes on their mRNA expression.

Genetic polymorphisms associated with structural changes of their gene product are important in terms of their potential relation with diseases. Therefore, in this study, splice-site variants of the transmembrane serine protease gene TMPRSS4, nephronophthisis gene NPHP4, and organic-cation transporter gene ORCTL4, were selected from the dbSNP single nucleotide polymorphism database as candidates to identify genetic polymorphisms associated with a structural change in their mRNA transcripts. The allele frequencies of the TMPRSS4 c.4-7A>G, NPHP4 c.2818-2A>T, and ORCTL4 c.517-2A>C polymorphisms in a Japanese population were determined to be 0.42, 0.10, and 0.27, respectively, by PCR-SSCP analysis. Next, the effect of these polymorphisms on the mode of pre-mRNA splicing was investigated by RT-PCR analysis followed by sequencing analysis. The TMPRSS4, NPHP4, and ORCTL4 polymorphisms were associated with the production of the r.4-6_4-1ins transcript, the r.2818_2823del and r.2818_2859del transcripts, and the r.517-94_517-1ins; r.517-2a>c and r.517_620del transcripts, respectively. Since the proteins encoded by all these transcripts are associated with relatively significant structural changes in the form amino acid insertion/deletion and premature termination, their functional ability may be greatly reduced. Our demonstration of structural changes in mRNA transcripts as a result of splice-site polymorphisms implies that they may be of biological significance in certain pathological conditions.