[High frequency electrical stimulation of sciatic nerve enhances skeletal muscle autophagy in mice].

The aim of the present study was to investigate the effects of exercise on skeletal muscle autophagy. Trains of high-frequency electrical stimulation (pulses frequency: 100 Hz) were used to stimulate sciatic nerve and consequently induce muscle contraction of the left hindlimb. The unstimulated right hindlimb muscles were taken as control. The mice were sacrificed immediately (0), 30 or 60 min after the electrical stimulation by cervical dislocation, and gastrocnemius muscles were rapidly dissected and freeze-clamped in liquid nitrogen. AMP-activated protein kinase (AMPK) and the autophagy marker protein LC3 were detected by Western blotting, and muscle atrophy related genes including atrogin-1, MuRF-1, Bnip3, Bnip3l and CathepsinL were detected by using real-time qPCR. The results showed that, at 0 min after the electrical stimulation, the activity of AMPK and LC3-II/I ratio were significantly increased in left gastrocnemius muscles, compared with those of the muscles in the right hindlimb. The levels of atrogin-1, MuRF-1, Bnip3, Bnip3l and CathepsinL mRNA expressions were up-regulated by electrical stimulation. Meanwhile, the activity of autophagy related protein, ULK1 was significantly enhanced by electrical stimulation. These results suggest that electrical stimulation of sciatic nerve may induce the skeletal muscle autophagy, and this may be regulated through AMPK/ULK1-mediated signaling pathway.

Roles of Wnt/beta-catenin signaling in adipogenic differentiation potential of adipose-derived mesenchymal stem cells.

Wnt/beta-catenin signaling pathway controls differentiation of various cells by regulating the expression of target genes. beta-Catenin plays a central role in Wnt/beta-catenin signaling pathway. To investigate the molecular mechanisms of fate determination in adipose-derived mesenchymal stem cells (AMSCs), we investigated effects of Wnt3a and beta-catenin, two key members of the Wnt/beta-catenin signaling, in adipogenic differentiation of porcine AMSCs. We demonstrated that Wnt3a protein can inhibit the adipogenic differentiation of porcine AMSCs in vitro culture. By stabilization of cytoplasmic beta-catenin with continuous treatment by LiCl, the adipogenic differentiation of AMSCs was also suppressed and the osteogenesis was stimulated. In contrast, a loss of beta-catenin in AMSCs enhanced the adipogenic differentiation and rescued LiCl-induced anti-adipogenesis. In addition, the mutual activation of CCAAT/enhancer-binding protein-alpha (C/EBPalpha) and peroxisome proliferator-activated receptor-gamma (PPARgamma) were repressed in the presence of Wnt3a or LiCl, but increased in the gene silencing of beta-catenin. Taken together, our study indicated that Wnt/beta-catenin signaling pathway inhibited the adipogenic differentiation potential and alter the cell fate from adipocytes to osteoblasts.

Functional proteomic analysis revels that the ethanol extract of Annona muricata L. induces liver cancer cell apoptosis through endoplasmic reticulum stress pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Annona muricata L. is used to treat cancer in some countries. Extracts of Annona muricata have been shown to cause apoptosis of various cancer cells in vitro, and inhibit tumor growth in vivo in animal models. However, the molecular mechanisms underlying its anti-cancer and apoptotic effects of the herb remain to be explored. AIM OF STUDY: The study investigated the molecular mechanisms underlying liver cancer cell apoptosis triggered by the ethanol extract of leaves of Annona muricata L. MATERIALS AND METHODS: Liver cancer HepG2 cells were used as experimental model. MTT assay was employed to evaluate cell viability. Flow cytometry and TUNEL assays were performed to confirm apoptosis. We employed functional proteomic analysis to delineate molecular pathways underlying apoptosis triggered by the herbal extract. RESULTS: We showed that the extract was able to reduce viability and trigger apoptosis of the cancer cells. Proteomic analysis identified 14 proteins associated with the extract-elicited apoptosis, which included the increased expression levels of HSP70, GRP94 and DPI-related protein 5. Western blot analysis confirmed that the extract did up-regulated the protein levels of HSP70 and GRP94. Results from bioinformatic annotation pulled out two molecular pathways for the extract, which, notably, included endoplasmic reticulum (ER) stress which was evidenced by the up-regulation of HSP70, GRP94 and PDI-related protein 5. Further examinations of typical protein signaling events in ER stress using western blot analysis have shown that the extract up-regulated the phorsphorelation of PERK and eIF2α as well as the expression level of Bip and CHOP. CONCLUSION: Our results indicate that the ethanol extract of leaves of Annona muricata L. causes apoptosis of liver cancer cells through ER stress pathway, which supports the ethnomedicinal use of this herb as an alternative or complementary therapy for cancer.

Overcome Cancer Cell Drug Resistance Using Natural Products.

Chemotherapy is one of the major treatment methods for cancer. However, failure in chemotherapy is not uncommon, mainly due to dose-limiting toxicity associated with drug resistance. Management of drug resistance is important towards successful chemotherapy. There are many reports in the Chinese literature that natural products can overcome cancer cell drug resistance, which deserve sharing with scientific and industrial communities. We summarized the reports into four categories: (1) in vitro studies using cell line models; (2) serum pharmacology; (3) in vivo studies using animal models; and (4) clinical studies. Fourteen single compounds were reported to have antidrug resistance activity for the first time. In vitro, compounds were able to overcome drug resistance at nontoxic or subtoxic concentrations, in a dose-dependent manner, by inhibiting drug transporters, cell detoxification capacity, or cell apoptosis sensitivity. Studies in vivo showed that single compounds, herbal extract, and formulas had potent antidrug resistance activities. Importantly, many single compounds, herbal extracts, and formulas have been used clinically to treat various diseases including cancer. The review provides comprehensive data on use of natural compounds to overcome cancer cell drug resistance in China, which may facilitate the therapeutic development of natural products for clinical management of cancer drug resistance.

Beta-catenin protein utilized by Tumour necrosis factor-alpha in porcine preadipocytes to suppress differentiation.

The Wnt/beta-catenin signaling pathway alters adipocyte differentiation by inhibiting adipogenic gene expression. beta-catenin plays a central role in the Wnt/beta-catenin signaling pathway. In this study, we revealed that tumour necrosis factor-alpha (TNF-alpha), a potential negative regulator of adipocyte differentiation, inhibits porcine adipogenesis through activation of the Wnt/beta-catenin signaling pathway. Under the optimal concentration of TNF-alpha, the intracellular beta-catenin protein was stabilized. Thus, the intracellular lipid accumulation of porcine preadipocyte was suppressed and the expression of important adipocyte marker genes, including peroxisome proliferator-activated receptor-gamma (PPARgamma) and CCAAT/enhancer binding protein-alpha (C/EBPalpha), were inhibited. However, a loss of beta-catenin in porcine preadipocytes enhanced the adipogenic differentiation and attenuated TNF-alpha induced anti-adipogenesis. Taken together, this study indicated that TNF-alpha inhibits adipogenesis through stabilization of beta-catenin protein in porcine preadipocytes.