Modulation effects of L-mevalonate on the inhibition effects of simvastatin on rat cardiac myocytes hypertrophy / 中国医师杂志

ardiac myocytes hypertrophy.

Role of Transforming Growth Factor-?_1/Smad Pathway in the Collagen Synthesis of Cardiac Fibroblasts Induced by Chymase / 中华高血压杂志

Background Cardiac mast cell-derived chymase is involved in myocardial fibrosis,but the underlying mechanisms remain unclear. Objective To investigate the effect of chymase on the collagen synthesis and its relationship with transforming growth factor-?1 (TGF-?1)/Smad pathway in rat cardiac fibroblasts (CFs). Methods CFs,from neonatal SD rats,were isolated by trypsinization. The collagen synthesis of CFs was determined by 3H-proline incorporation. The protein expressions of TGF-?1,phosphorylated Smad2/3 (P-Smad2/3) and total Smad2/3 were determined by immunoblotting in CFs. Results Chymase (15,30 and 60 ?g/L) increased the 3H-proline incorporation in a concentration-dependent manner. 30 ?g/L chymase stimulation increased the protein expressions of TGF-?1 and P-Smad2/3 in a time-dependently,while little effect on Smad2/3 protein expression was found. The stimulatory effect of chymase on 3H-proline incorporation elicited by 30 ?g/L chymase was blocked in the presence of TGF-?1 antibody or staurosporine,a P-Smad2/3 inhibitor. Conclusion Chymase promotes collagen synthesis of rat CFs,TGF-?1/Smad might be involved into the signal pathway.

Inhibitory effect of cyclosporin A on growth and collagen synthesis of rat cardiac fibroblasts induced by arginine vasopressin / 药学学报

Aim To investigate the effects of cyclosporin A (CsA) on growth and collagen synthesis of cardiac fibroblasts (CFs) induced by arginine vasopressin (AVP). Methods CFs of neonatal Sprague-Dawley rats were isolated by trypsinization and cultured; growth-arrested CFs were stimulated with 1×10-7 mol·L-1 AVP in the presence or absence of CsA (0.05, 0.5 and 5 μmol·L-1). MTT and flow cytometry techniques were adopted to measure cell number and analyze cell cycle respectively. Collagen synthesis was determined by measurement of hydroxyproline content in culture supernatant with colorimetry. Calcineurin activity was estimated by chemiluminescence. Trypan blue staining to test the viability of CFs. Results 0.05, 0.5 and 5 μmol·L-1 CsA inhibited the increase of CFs number induced by 1×10-7 mol·L-1 AVP in a dose-dependent manner, with the inhibitory rates by 12%, 24% and 29%, respectively (P＜0.05). Furthermore, cell cycle analysis showed 0.5 μmol·L-1 CsA decreased the S stage percentage and proliferation index of CFs stimulated by AVP (P＜0.05). In culture medium, the hydroxyproline content induced by AVP decreased by 0.5 and 5 μmol·L-1 CsA (P＜0.05), with the inhibitory rates of 29% and 33%, respectively. CsA completely inhibited the increment of calcineurin activity induced by AVP (P＜0.01), but CsA itself had no effect on the baseline of calcineurin activity and CFs viability. Conclusion CsA inhibits proliferation and collagen synthesis of CFs by virtue of blocking calcineurin signaling pathway and might provide a novel target for prevention and treatment to cardiac fibrosis.

Polyclonal antibody production and expression of CREG protein in human vascular smooth muscle cells / 老年心脏病学杂志（英文版）

Objectives The cellular repressor of E1A-activated genes (CREG), a novel gene, was recently found to play a role in inhibiting cell growth and promoting cell differentiation. The purpose of this study was to obtain antibody against CREG protein and to study the expression of CREG protein in human internal thoracic artery cells (HITASY) which express different patterns of differentiation markers after serum withdrawal. Methods The open reading frame of CREG gene sequence was amplified by PCR and cloned into the pGEX-4T-1 vector. Glutathione-S-transferase (GST)-CREG fusion protein was expressed in E. Coli BL21 and purified from inclusion bodies by Sephacryl S-200 chromatography. Rabbits were immunized with the purified GST-CREG protein. Western blot examined with immunohistochemistry staining and the protein expression level was analyzed by Western blot in HITASY cells after serum removal. Results It was confirmed by using endonuclease digesting and DNA sequencing that the PCR product of CREG was correctly inserted into the vector. The GST-CREG protein was purified with gel filtration chromatography. Polyclonal antibody against GST-CREG was obtained from rabbits. CREG protein immunohistochemistry staining displayed a perinuclear distribution in the cytoplasm of HITASY cells. Results from Western blot suggested that comparing with the untreated cells upregulation of CREG polyclonal antibody against CREG was comfirmed. Using this antibody, the changes of CREG protein expression was observed in the process of phenotypic modulation of HITASY cells. These results provide basic understanding on the relationship of CREG gene with the cell phenotypic conversion.

Effects of TNF-? on changes of inducible nitric oxide synthase-nitric oxide system activity induced by arginine vasopressin in rat cardiac fibroblasts / 中国病理生理杂志

AIM: To explore the effects of tumor necrosis factor-? (TNF-?) on inducible nitric oxide synthase (iNOS)-nitric oxide (NO) system activity in arginine vasopressin (AVP)-induced rat cardiac fibroblasts (CFs). METHODS: CFs were isolated by trypsin digestion method. Nitrate reductase method, spectrophotometry and reverse transcription-polymerase chain reaction (RT-PCR) were used to detect NO contents, NOS activity and iNOS mRNA expression, respectively. RESULTS: AVP significantly increased iNOS mRNA expression, NOS activity and NO contents in CFs. Combined with AVP, TNF-? enhanced the effects of AVP on iNOS-NO system activity in a concentration-dependent manner. However, if the concentration of TNF-? was too high, the iNOS-NO system activity did not increase accordingly, but slightly decreased instead. CONCLUSION: TNF-? stimulates iNOS-NO system activity in coordination with AVP in CFs. The enhancement of NO contents inhibits ventricular remodeling induced by AVP and TNF-?. [

Effects of argipressin on proliferation and NO synthesis in rat cardiac fibroblasts / 解放军医学杂志

Objective To explore the effect of arginine vasopressin(AVP) on proliferation and nitric oxide(NO) synthesis in rat cardiac fibroblasts(CFs). Methods CFs were isolated by trypsin digestion method. The effect of AVP on proliferation and NO synthesis were measured by MTT technique and nitric acid reductase method respectively. Results The MTT OD values of CFs increased in a concentration-dependent manner with the rising of the concentration of AVP and in a time-dependent manner with the prolongation of the culture time. For example, the MTT OD values of 10 -6 mol/L AVP group were higher than those of the control, and the MTT OD values of CFs cultured for 36h were significantly higher than those cultured for 6h when intervened with 10 -7 mol/L AVP. The content of NO of CFs displayed the same change as the MTT OD values. The contents of NO in 10 -7 mol/L and 10 -6 mol/L AVP groups were statistically higher than those of the contral group, 10 -9 mol/L AVP group and 10 -8 mol/L AVP group. The NO contents of CFs cultured for 24h and 36h were significantly higher than those cultured for 6h and 12h. Conclusion AVP can stimulate the proliferation of CFs and the synthesis of NO in neonatal rat. The enhancement of NO contents would inhibit CFs proliferation and myocardial fibrosis induced by AVP.