Efficacy of Acetylshikonin in Preventing Obesity and Hepatic Steatosis in db/db Mice.

Zicao (Lithospermum erythrorhizon) has been used in clinics as a traditional Chinese medicine for thousands of years. Acetylshikonin (AS) is the main ingredient of Zicao, Xinjiang, China. The objective of this study was to investigate the anti-obesity and anti-nonalcoholic fatty liver disease (NAFLD) efficacy of AS in a model of spontaneous obese db/db mice. Mice were divided into Wild Type (WT) groups and db/db groups, which received no treatment or treatment with 100 mg/kg/day clenbuterol (CL) hydrochloride or 540 mg/kg/day AS by oral gavage for eight weeks. The results provided the evidence that AS prevented obesity and NAFLD including reduction in body weight, food efficiency ratio, serum triglyceride (TG) and free fatty acid (FFA) levels in db/db mice. Administration of AS markedly suppressed the levels of hepatic alanine aminotransferase (ALT), aspartate aminotransferase (AST) and pro-inflammatory cytokines in treated groups when compared with that of db/db groups. Further investigation of the lipid synthesis-related protein using Western blotting revealed that hepatic protein expression of sterol regulatory element-binding protein-1 (SREBP-1), fatty acid synthetase (FAS) and 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) were significantly downregulated by AS treatment. These findings suggest that AS exerts anti-obesity and anti-NAFLD effects through the regulation of lipid metabolism and anti-inflammatory effects.

Arnebin-1 promotes angiogenesis by inducing eNOS, VEGF and HIF-1α expression through the PI3K-dependent pathway.

Arnebin-1, a naphthoquinone derivative, plays a crucial role in the wound healing properties of Zicao (a traditional wound healing herbal medicine). It has been noted that Arnebin-1, in conjunction with vascular endothelial growth factor (VEGF), exerts a synergistic pro-angiogenic effect on human umbilical vein endothelial cells (HUVECs) and accelerates the healing process of diabetic wounds. However, the mechanisms responsible for the pro-angiogenic effect of arnebin­1 on HUVECs and its healing effect on diabetic wounds have not yet been fully elucidated. In this study, in an aim to elucidate these mechanisms of action of arnebin­1, we investigated the effects of arnebin­1 on the VEGF receptor 2 (VEGFR2) and the phosphoinositide 3-kinase (PI3K)­dependent signaling pathways in HUVECs treated with VEGF by western blot analysis. The pro­angiogenic effects of arnebin­1 on HUVECs, including its effects on proliferation and migration, were evaluated by MTT assay, Transwell assay and tube formation assay in vitro. The expression levels of hypoxia-inducible factor (HIF)­1α, endothelial nitric oxide synthase (eNOS) and VEGF were determined by western blot analysis in the HUVECs and wound tissues obtained from non­diabetic and diabetic rats. CD31 expression in the rat wounds was evaluated by immunofluorescence staining. We found that the activation of the VEGFR2 signaling pathway induced by VEGF was enhanced by arnebin­1. Arnebin­1 promoted endothelial cell proliferation, migration and tube formation through the PI3K­dependent pathway. Moreover, Arnebin­1 significantly increased the eNOS, VEGF and HIF­1α expression levels in the HUVECs and accelerated the healing of diabetic wounds through the PI3K­dependent signaling pathway. CD31 expression was markedly enhanced in the wounds of diabetic rats treated with arnebin­1 compared to the wounds of untreated diabetic rats. Therefore, the findings of the present study indicate that arnebin-1 promotes the wound healing process in diabetic rats by eliciting a pro-angiogenic response.

Arnebin-1 promotes the angiogenesis of human umbilical vein endothelial cells and accelerates the wound healing process in diabetic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Zicao is a traditional wound healing herbal medicine that has been used for several hundred years in China. A survey of the published literatures revealed that arnebin-1, one of the naphthoquinone derivatives, played the most important role in wound healing property of this plant. However, whether arnebin-1 affects angiogenesis in vitro and has an effect on wound healing process in diabetic rats remains enigmatic. To investigate the effect of arnebin-1 with or without VEGF on proliferation, migration and tube formation of HUVECs in vitro and the effect of its topical application in the form of ointment on wound healing in a cutaneous punch wound model of alloxan-induced diabetic rats in vivo. MATERIALS AND METHODS: The pro-angiogenic functions of arnebin-1 on HUVECs including proliferation, migration and angiogenesis were evaluated through MTT assay, wound healing assay, transwell assay and tube formation assay in vitro. Male Sprague-Dawley rats were injected intraperitoneally with alloxan to induce type І diabetic rats. Three wounds were created in each rat on the dorsal surface, and then divided to be basement treated, arnebin-1 ointment treated and untreated group correspondingly. The indicators including wound closure rate and histological evaluation were investigated on day 4 and 7 post-wounding. RESULTS: Without VEGF, arnebin-1 did not affect the proliferation of HUVECs significantly, but had a positive effect on cell migration and tube formation. However, in the presence of minimal VEGF, Arnebin-1 could increase the proliferation, enhance the migration and promote the tube formation of HUVECs significantly. The wound closure rate was increased significantly in arnebin-1 treated group compared to that of untreated and basement treated groups in diabetic rats, and the histological evaluation also showed well organized dermal layer, reduced number of macrophages, increased number of fibroblasts, remarkable degree of neovascularization and epithelization in arnebin-1 treated group. CONCLUSION: These findings suggest that arnebin-1 has a pro-angiogenic effect, and a synergetic effect with VEGF promotes the wound healing process in diabetic rats.

Scutellarin induced Ca(2+ ) release and blocked KCl-induced Ca(2+ ) influx in smooth muscle cells isolated from rat thoracic artery.

This study was designed to investigate the effect of scutellarin (1) on the modulation of intracellular Ca(2+ ) concentration in thoracic smooth muscle cells of rat. Single smooth muscle cells were obtained enzymatically. Fluo-3 AM was used to determine the alteration of intracellular-free Ca(2+ )([Ca(2+ )](i)) and the changes in fluorescence intensity under different agonists were recorded. Compound 1 induced Ca(2+ ) transients in the medium with and/or without Ca(2+ ). In the Ca(2+ )-free medium, after pretreatment of 1, thapsigargin failed to cause the elevation of [Ca(2+ )](i). However, 1 still caused the elevation of [Ca(2+ )](i) after pretreatment of thapsigargin. The infusion of 1 blocked KCl-induced Ca(2+ ) entry and this effect was hardly reversible. The results of present study suggested that 1 increased [Ca(2+ )](i) by blocking sarcoplasmic reticulum Ca(2+ )/ATPase and blocked voltage-dependent Ca(2+ ) channels in smooth muscle cells of the rat thoracic aortic artery.