Low-dose nifedipine rescues impaired endothelial progenitor cell-mediated angiogenesis in diabetic mice.

It is of great clinical significance to develop potential novel strategies to prevent diabetic cardiovascular complications. Endothelial progenitor cell (EPC) dysfunction is a key contributor to diabetic vascular complications. In the present study we evaluated whether low-dose nifedipine could rescue impaired EPC-mediated angiogenesis and prevent cardiovascular complications in diabetic mice. Diabetes was induced in mice by five consecutive injections of streptozotocin (STZ, 60 mg·kg-1·d-1, i.p.). Diabetic mice were treated with low-dose nifedipine (1.5 mg·kg-1·d-1, i.g.) for six weeks. Then, circulating EPCs in the peripheral blood were quantified, and bone marrow-derived EPCs (BM-EPCs) were prepared. We showed that administration of low-dose nifedipine significantly increased circulating EPCs, improved BM-EPCs function, promoted angiogenesis, and reduced the cerebral ischemic injury in diabetic mice. Furthermore, we found that low-dose nifedipine significantly increased endothelial nitric oxide synthase (eNOS) expression and intracellular NO levels, and decreased the levels of intracellular O2.- and thrombospondin-1/2 (TSP-1/2, a potent angiogenesis inhibitor) in BM-EPCs of diabetic mice. In cultured BM-EPCs, co-treatment with nifedipine (0.1, 1 µM) dose-dependently protected against high-glucose-induced impairment of migration, and suppressed high-glucose-induced TSP-1 secretion and superoxide overproduction. In mice with middle cerebral artery occlusion, intravenous injection of diabetic BM-EPCs treated with nifedipine displayed a greater ability to promote local angiogenesis and reduce cerebral ischemic injury compared to injection of diabetic BM-EPCs treated with vehicle, and the donor-derived BM-EPCs homed to the recipient ischemic brain. In conclusion, low-dose nifedipine can enhance EPCs' angiogenic potential and protect against cerebral ischemic injury in diabetic mice. It is implied that chronic treatment with low-dose nifedipine may be a safe and economic manner to prevent ischemic diseases (including stroke) in diabetes.

Sodium dichloroacetate improves migration ability by suppressing LPS-induced inflammation in HTR-8/SVneo cells via the TLR4/NF-κB pathway.

Objectives: Inadequate cytotrophoblast migration and invasion are speculated to result in preeclampsia, which is a pro-inflammatory condition. Sodium dichloroacetate (DCA) exerts anti-inflammatory actions. Thus,we sought to investigate the effect of DCA on the migration function of the lipopolysaccharide (LPS)-stimulated human-trophoblast-derived cell line (HTR-8/SVneo). Materials and Methods: HTR-8/SVneo cells were treated with LPS to suppress cell migration. Cell migration was examined by both scratch wound healing assay and transwell migration assay. Western blotting was used to analyze the expression levels of toll-like receptor-4 (TLR4), nuclear factor-κB (NF-κB), TNF-α, IL-1ß, and IL-6 in the cells. Results: DCA reversed LPS-induced inhibition of migration in HTR-8/SVneo cells. Furthermore, DCA significantly suppressed LPS-induced activation of TLR4, phosphorylation of NF-κB (p65), translocation of p65 into the nucleus, and the production of pro-inflammatory cytokines (TNF-α, IL-1ß, and IL-6). Treatment with inhibitors of TLR4 signal transduction (CLI095 or MD2-TLR-4-IN-1) reduced LPS-induced overexpression of pro-inflammatory cytokines, and a synergistic effect was found between TLR4 inhibitors and DCA in HTR-8/SVneo cells. Conclusion: DCA improved trophoblast cell migration function by suppressing LPS-induced inflammation, at least in part, via the TLR4/NF-κB signaling pathway. This result indicates that DCA might be a potential therapeutic candidate for human pregnancy-related complications associated with trophoblast disorder.

Berberine hydrochloride inhibits migration ability via increasing inducible NO synthase and peroxynitrite in HTR-8/SVneo cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Inadequate trophoblasts migration and invasion is considered as an initial event resulting in preeclampsia, which is closely related to oxidative stress. Berberine hydrochloride (BBR), extracted from the traditional medicinal plant Coptis chinensis Franch., exerts a diversity of pharmacological effects, and the crude drug has been widely taken by most Chinese women to treat nausea and vomit during pregnancy. But there is no research regarding its effects on trophoblast cell function. AIM OF THE STUDY: This study aimed to investigate the effect of BBR on human-trophoblast-derived cell line (HTR-8/SVneo) migration ability and its mechanism. MATERIALS AND METHODS: Cell viability was detected by CCK-8 assay. The effect of BBR on cells migration function was examined by scratch wound healing assay and transwell migration assay. Intracellular nitric oxide (NO), superoxide (O2-) and peroxynitrite (ONOO-) levels were measured by flow cytometry. The expression levels of inducible NO synthase (iNOS), eNOS, p-eNOS, MnSOD, CuZnSOD, Rac1, NOX1, TLR4, nuclear factor-κB (NF-κB), p-NFκB, pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) in cells were analyzed by Western blotting. Uric acid sodium salt (UA), the scavenger of ONOO-, PEG-SOD (a specific superoxide scavenger), L-NAME (a NOS inhibitor) and antioxidants (Vit E and DFO) were further used to characterize the pathway of BBR action. RESULTS: 5 µM BBR decreased both the migration distance and the number of migrated cells without affecting cells viability in HTR-8/SVneo cells after 24 h treatment. BBR could increase the level of NO in HTR-8/SVneo cells, and the over-production of NO might be attributable to iNOS, but not eNOS. BBR could increase intracellular O2- levels, and the over-production of O2- is closely related with Rac1 in HTR-8/SVneo cells. The excessive production of NO and O2- further react to form ONOO-, and the increased ONOO- level induced by BBR was blunted by UA. Moreover, UA improved the impaired migration function caused by BBR in HTR-8/SVneo cells. The depressed migration function stimulated by BBR in HTR-8/SVneo cells was diminished by PEG-SOD and L-NAME. Furthermore, BBR increased the expression of IL-6 in HTR-8/SVneo cells, and antioxidants (Vit E and DFO) could decrease the expression of IL-6 and iNOS induced by BBR. CONCLUSIONS: BBR inhibits the cell migration ability through increasing inducible NO synthase and peroxynitrite in HTR-8/SVneo cells, indicating that BBR and traditional Chinese medicines containing a high proportion of BBR should be used with caution in pregnant women.