| At the early stage of chronic renal failure when the GFR has reduced to 50mllmim, the level of serum parathyroid hormone(PTH) increases significantly because of hypocalcemia, hyperphosphatemia, deficiency of active vitamin D, downregulation of receptors for calcium and vitaminD. The high level of serum PTH not only affects the metablism of bone to severe renal osteopathy, but has negative effects on cardiovascularsystem, immune system and erythropoiesis so it has been a novel uremia toxin recently. However , there has little systematic study on the renal toxic effects of PTH till nowadays.As we all know, studies on ligand interactions with PTHR on the classical target tissues ( bone and renal tubule) of PTh reveals at least two second messenger signaling systems: the adenylyl cyclase/protein kinase A(AC/PKA) pathway and the phospholipase Cl protein kinase C( PLC/PKC) pathway, then leads to activation of cAMP or 1P3 and DAG respectively, finally regulates the metablism of calcium and phosphorum. Besides, PTH can stimulate the proliferation of osteoblast, induce theexpression of early response genes like c-fos, cjun and IL-6 on osteoblast andchondrocyte, as wel1 as increase the synthesis of IGF-1 and TGF- P in culturedoSteoblast, Whch suggests that PTH can activate some other pathWays to regulate cellproliferation and synthsis and secretion of some cytokine such as TGF(6.So the purpose of our St'Udy is to evaluate the toxic effects of PTH on residualrenal function by acting on mesangial cells. The contents of our study were: O Detectthe eXPression of receptors for PTH in cultUred rat mesangial ce1ls by reversetranscription polpoerase chain reaction (RT-PCR) method. @ Evaluate theproliferative effect of bPTHl-34 on cultUred rat mesangial cells by cel1 coUning,methylene blue assay and flow cytometer method. @ Examine the synthesis of TGF-5 1 and exPression of TGF~ 6 1InRNA from inesangial cells stinnilated by hPTHl-34using ELISA and semi-quantitative RT-PCR method. @ Examine the synthesis of F'Nin cultUred rat mesangial ceIls stimulated by bPml-34 using ELISA and evaluate itsprobable mechedsm invo1ved via TGF- P. @ Evaluate the effect of rPTH1-34intraPeritoneaJ1y injected into rats for 2 and 4 weeks on some blood and urineparametes as well as the eXPression of TGF-1 protein and gene on kidney tissue bydriunohistochemical method and semi-quamitive RT-PCR.The results of thes study were:l. The agarose gel electrophOresis of amP1ification produCts amPlified byPTH-R specific primers showed a single straP Which matched to our expectation, Whichsuggested that the recePtors fOr PTH exists on the rat mesangial cells.2. By using cell coUning, methylene b1ue assay and flow cytometermethod, it was suggested that hPrnl-34 stAnu1ated the proliferation of rat mesangialcells in a dose- and time-dependent maxmer with the peak at the concentration of10-omoUl (P<0.01). Besides, Whn at the concendation of 10-'moin, hPTH1-34 inducesaPoptosis or death of cel1s maybe because of its toxic effects.3. bPTHlo' stimulated the stwsis of TGF- P l frOm cultured rat mesangialcel1s in a dose- and time-dependellt manner with the peak at the concentr -ation of103mofl (P<0.01) by ELISA. Also the expression of TGF-扨 1mRNA was increased by hPTH1-34 dose- and time-dependently using semi-quantitive RT-PCR (P<0.05).4.The level of FN protein increased signigicantly after the mesangial cells were stimulated by hPTH1-34 with the peak at concentration of l0~8mol/l for 48 hours (113.03~50.64 ng/ml VS 74.65d3 1.07 nglml, P<0.0l). Anti-TGF- ~ inhibited the stimulative effect of hPTH1-34 on synthesis of FN significantly (without antibody54.63~28.3 lngIml vs with antibody 46.85~30.56ngIm1, P = 0.01).5.2 weeks or 4 weeks after the rats were injected intraperitoneally at the dose of 108g1(lOOgWtd), the level of hemoglobin decreased while serum creatinine, blood calcium increased slightly. Level of blood phosphorate decreased while urinary excretion of N-acetyl- ~ -D... |
PDF Full Text Request