Effects of visfatin on proliferation and collagen synthesis in rat cardiac fibroblasts.

The proliferation of cardiac fibroblasts (CFs) and excessive extracellular matrix protein accumulation are the basic pathological processes of myocardial fibrosis. Visfatin is a novel adipokine involved in the regulation of inflammatory cytokines, however, the effects of visfatin on proliferation and collagen synthesis of CFs are unclear. The aim of this study was to determine whether visfatin has any effect on the proliferation and collagen synthesis in rat CFs. Incorporation of [ (3)H]-thymidine and [ (3)H]-proline were used for evaluating DNA and collagen synthesis. Flow cytometry techniques were adopted to analyze cell cycle. Enzyme-linked immunosorbent assay was used for measuring collagen type I and III production. RT-PCR and Western blot analysis were used for determining procollagen I and III mRNA expression and protein production. The inhibitors used for pathway determination were: wortmannin [phosphatiylinositol 3-kinase (PI3K)], SB203580 [p38 mitogen-activated protein kinase (MAPK)], PD98059 [extracellular signal-regulated kinase (ERK)1/2)], and JNK inhibitor II [c-Jun NH 2-terminal kinase (JNK)]. We demonstrated that visfatin significantly increased DNA and collagen synthesis in a dose- and time-dependent manner. Cell cycle analysis showed that visfatin increased S-stage percentage and proliferation index in a dose- and time-dependent manner. It was also found that visfatin upregulated collagen I and III production, procollagen I and III mRNA expression and protein production. These effects were diminished by SB203580, wortmannin, and PD98059, but not by JNK inhibitor II. These results suggest that visfatin promote CFs proliferation and collagen synthesis via p38MAPK, PI3K, and ERK 1/2 pathways rather than JNK pathways, which also indicate that visfatin might play a role in myocardial fibrosis.

Antiproliferative effects of D-polymannuronic sulfate on rat vascular smooth muscle cells and its related mechanisms.

AIM: To investigate the inhibitory effects of D-polymannuronic sulfate (DPS) on the proliferation of rat vascular smooth muscle cells (VSMC) induced by angiotensin II (Ang II) and its related mechanisms. METHODS: The effects of DPS on Ang II-induced proliferation of VSMC were evaluated by MTT assay. The intracellular free Ca2+ concentrations, protein contents, and cell cycle were analyzed by flow cytometry. RESULTS: DPS 0.001 - 100 mg/L blocked the cell cycle at the G0/G1-->S transit and prevented the cells from entering into the G2/M phase, and its inhibitory effects on an increase in intracellular free Ca2+ concentrations and the protein synthesis of VSMC were also observed. Also, the suppressing actions of DPS on intracellular Ca2+ were completely blocked by L-NAME, a nitric oxide synthase inhibitor, indicating that the counteracting effects on a rise in intracellular free Ca2+ contents by DPS might be mediated by participation of NO. CONCLUSION: DPS exerted an inhibitory effect on Ang II-induced proliferation of VSMC and its related mechanisms were considered to be related to its inhibition on the increment of intracellular Ca2+ concentrations, which subsequently suppressed the synthesis of DNA and protein of VSMC.

[Inhibitory effects of D-polymannuronic sulfate on proliferation of rat vascular smooth muscle cells and its related mechanisms of action in vitro].

AIM: To investigate the inhibitory effects of D-polymannuronic sulfate (DPS) on the proliferation of rat vascular smooth muscle cells (VSMC) induced by basic fibroblast growth factor (bFGF) or interleukin-1 (IL-1) and its related mechanisms. METHODS: Rat aortic smooth muscle cells pretreated with DPS in concentrations ranging from 0.001 microgram.mL-1 up to 100 micrograms.mL-1 were incubated at 37 degrees C for 24 h, followed by addition of bFGF (50 ng.mL-1) or IL-1 (50 U.mL-1) for another 24 h. The effects of DPS on the proliferation of VSMC were evaluated by MTT assays. VSMC were pretreated with DPS in concentrations ranging from 0.001 microgram.mL-1 up to 1 microgram.mL-1, followed by addition of L-NAME (0.1 microgram.mL-1) or bFGF (50 ng.mL-1) for 24 h. Supernatant nitric oxide (NO) was determined with NO assay kit, while supernatant angiotensin II (Ang II) and endothelin-1 (ET-1) were measured by radioimmunoassay. RESULTS: DPS exerted antiproliferative effects at concentrations ranging from 0.01 microgram.mL-1 to 10 micrograms.mL-1, and its maximal effect was observed at the concentration of 1 microgram.mL-1. Also, the suppressing actions of DPS on the proliferation of VSMC were diminished by increasing the concentrations of bFGF or IL-1. Furthermore, DPS increased NO synthesis and decreased Ang II and ET-1 contents released from VSMC in a concentration-dependent manner. CONCLUSION: DPS afforded the antiproliferative effects on bFGF- or IL-1-treated VSMC and its underlying mechanisms were associated with enhancement of NO synthesis and decrement of Ang II and ET-1 production/release in vitro.

Analysis of oligomannuronic acids and oligoguluronic acids by high-performance anion-exchange chromatography and electrospray ionization mass spectrometry.

Oligomannuronic acids and oligoguluronic acids were prepared by enzymatic hydrolysis of alginate with alginate lyases. The oligosaccharides generated up to degree of polymerization 16 were characterized by high-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection (PAD) and electrospray ionization mass spectrometry (ESI-MS). Acetate buffer linear gradients were used as mobile phases for separations of oligosaccharides. ESI-MS and HPAEC-PAD are very effective for the analysis and characterization of anionic oligosaccharides.

Antihypertensive effects of D-polymannuronic sulfate and its related mechanisms in renovascular hypertensive rats.

AIM: To investigate the antihypertensive effects of D-polymannuronic sulfate (DPS), a kind of sulfated polysaccharide, and the underlying mechanisms in renovascular hypertensive rats (RHR). METHODS: Used two-kidney one clip (Goldblatt, 2-K 1C) method to produce RHR model. DPS was given i.v. or ig for 5 wk with the initiation of establishment of RHR. Serum nitric oxide (NO) was determined with NO kit; plasma angiotensin II (Ang II) and endothelin-1 (ET-1) were measured by radioimmumoassays. RESULTS: In acute therapeutic experiments, DPS markedly reduced systolic blood pressure (SBP) and diastolic blood pressure (DBP) dose-dependently and decreased heart rate (HR) with reduction in arterial blood pressure. In the prophylactic experiments, DPS prevented the rise in SBP and DBP in a dose-dependent manner. The hypotensive potency of DPS 50 mg/kg is comparable to that of captopril (14 mg/kg). Moreover, DPS elevated serum NO contents and lowered plasma concentrations of Ang II and ET-1. CONCLUSION: The antihypertensive activities of DPS might be involved both in increasing the generation of nitric oxide and in decreasing the production of angiotensin II and endothelin-1 in vivo.