Effect of Electroacupuncture in "Zusanli" and "Kunlun" Acupoints on TLR4 Signaling Pathway of Adjuvant Arthritis Rats.

BACKGROUND: Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease. Previous study suggested that toll-like receptor (TLR) signaling pathway contributes to the development and progression of RA. In recent years, acupuncture has become one of the most vital treatments of arthralgia. But little is known about the mechanisms of improving RA by acupuncture. STUDY QUESTION: The study studied the effect of electroacupuncture in "Zusanli" and "Kunlun" acupoints on the expression of TLR4, myeloid differentiation factor 88 (MYD88), and NF-κB in adjuvant arthritis rats to clarify the molecular mechanism of acupuncture of RA. STUDY DESIGN: A rat model of adjuvant arthritis was established with injection of 0.1 mL Freund complete adjuvant in the right hindlimb footpad. We next punctured the Zusanli and Kunlun acupoints with 0.25 × 40-mm acupuncture needles to 5-mm depth. Then, we performed electroacupuncture treatment for 28 days with frequency of 2 Hz and intensity of 2 mA, once a day and 30 minutes each time. MEASURES AND OUTCOMES: Arthritis index and paw swelling were measured every week. FQ-PCR and western blot were used to detect the expression of TLR4, MYD88, and NF-κB. RESULTS: Paw swelling of rats injected with Freund complete adjuvant was more serious than that of the normal rats, which illustrated the successful establishment of adjuvant arthritis rat model. After treatment for 14 days, the paw swelling and joint symptoms score decreased, paw tissue inflammation eased in the rats of treatment group compared with the model group during the same period. After treatment for 28 days, the expression of TLR4, MYD88, and NF-κB in the ankle bone tissues decreased at both mRNA and protein levels. CONCLUSIONS: Stimulation with electric needle in Zusanli and Kunlun acupoints can reduce the expression of TLR4, MYD88, and NF-κB, which play an important role in treatment of adjuvant arthritis.

[Effect of ligustrazine on reverse cholesterol transport in foam cells].

OBJECTIVE: To discuss the intervention effect of ligustrazine on ox-LDL-induced foam cells from the perspective of reverse cholesterol transport. METHOD: RAW264.7 cultured in vitro was induced with 20 mg x L(-1) ox-LDL to establish the foam cell model, and intervened with ligustrazine. The lipid accumulation in cells was observed by the oil red O dyeing. The changes in total cholesterol and cholesterol ester in the cells were detected with the colorimetric method. The fluorescent quantitative PCR and Western blot were used to detect the mRNA expressions of PPARgamma, LXRalpha and ABCA1. RESULT: Ligustrazine could reduce total cholesterol and cholesterol ester in foam cells, inhibit the lipid accumulation, and increase the mRNA and protein expressions of PPARgamma, LXRalpha and ABCA1. CONCLUSION: Ligustrazine can promote the reverse cholesterol transport by increasing the gene expressions of PPARgamma, LXRalpha and ABCA1.

[Effect of cold and cool herbs on liver mitochondria proteome of rats with heat symptom].

In the 1960s, modern science began involving the essence of heat syndrome, but there have still no in-depth systematic studies on pathological mechanisms of heat syndrome and action mechanisms of cold and cool herbs. In this study, the animal model with heat syndrome was set up by feeding herbs with hot property, and then cold and cool herbs was applied in the experimental therapy. The two-dimensional electrophoresis and mass spectrometry technologies were adopted to compare the liver mitochondria proteome of the rats of the heat syndrome model and the ones treated with cold and cool herbs, so as to discover specificity-related proteins after heat syndrome and treatment with cold and cool herbs.

Carpesium abrotanoides (L.) Root as a Potential Source of Natural Anticancer Compounds: Targeting Glucose Metabolism and PKM2/HIF-1α Axis of Breast Cancer Cells.

Carpesium abrotanoides L. (CA) is widely used as a medicinal plant in Asia. The biological activities of the extract from the roots of Carpesium abrotanoides L. (PCA) and its major components were analyzed in this study. PCA was separated and identified with mass spectrometry. Furthermore, we sought to elucidate the anticancer activity of PCA and its mechanisms. PCA exerted its anti-breast cancer activity through inhibiting the expression of glycolysis-related genes, such as glucose transporter 1, lactate dehydrogenase A, and hexokinase 2. Moreover, PCA downregulated the expression of pyruvate kinase M2 and altered its cellular translocation. We also demonstrated PCA is an inhibitor of the PKM2/hypoxia-inducible factor-1α axis, indicating that PCA is potentially useful as an anti-breast cancer agent. PRACTICAL APPLICATION: In this study, the extract from roots of Carpesium abrotanoides Linn. (PCA) was shown to have a noticeable anticancer effect against breast cancer in vitro, and PCA exerts the anticancer activity by regulating glucose metabolism and PKM2 expression. These findings indicate that PCA is a promising agent with practical applications in the development of functional food containing Carpesium abrotanoides L. root extracts.

Proteomic analysis of hepatocellular carcinoma HepG2 cells treated with platycodin D.

Platycodin D (PD), a triterpenoid saponin isolated from Platycodonis Radix, is a famous Chinese herbal medicine that has been shown to have anti-proliferative effects in several cancer cell lines. The aim of this study was to determine the changes in cellular proteins after the treatment of hepatocellular carcinoma HepG2 cells with PD using proteomics approaches. The cell viability was determined using the MTT assay. The proteome was analyzed by two-dimensional difference gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Western blot analysis was used to confirm the expression of changed proteins. Our results showed that PD inhibited the proliferation of HepG2 cells in concentration- and time-dependent manners. Sixteen proteins were identified to be up-regulated in PD-treated HepG2 cells, including ATP5H, OXCT1, KRT9, CCDC40, ERP29, RCN1, ZNF175, HNRNPH1, HSP27, PA2G4, PHB, BANF1, TPM3, ECH1, LGALS1, and MYL6. Three proteins (i.e., RPS12, EMG1, and KRT1) decreased in HepG2 cells after treatment with PD. The changes in HSP27 and PHB were further confirmed by Western blotting. In conclusion, our results shed new lights on the mechanisms of action for the anti-cancer activity of PD.

Proteomic analysis of liver mitochondria from rats with nonalcoholic steatohepatitis.

AIM: To explore mitochondrial dysfunction in nonalcoholic steatohepatitis (NASH) by analyzing the proteome of liver mitochondria from a NASH model. METHODS: The NASH rat model was established by feeding rats a fat-rich diet for 24 wk and was confirmed using hematoxylin and eosin staining of liver tissue and by changes in the levels of serum alanine transaminase, aspartate aminotransferase, triglyceride, total cholesterol and other markers. Liver mitochondria from each group were isolated using differential centrifugation. The mitochondrial samples were lyzed, purified and further analyzed using two-dimensional electrophoresis combined with matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry. Bioinformatic analyses of assigned gene ontology and biological pathway was used to study functional enrichments in the abundant proteomic data. RESULTS: Eight up-regulated and sixteen down-regulated proteins were identified that showed greater than 1.5-fold differences between the controls and the NASH group. These dysregulated proteins were predicted to be involved in different metabolic processes including fatty acid ß-oxidation processes, lipid metabolic processes, cell-cycle arrest, cell polarity maintenance, and adenosine triphosphate/sex hormone metabolic processes. Novel proteins that may be involved in NASH pathogenesis including the trifunctional enzyme Hadha, thyroxine, prohibitin, aldehyde dehydrogenase ALDH1L2, UDP-glucuronosyltransferase 2B31, and carbamoyl-phosphate synthase were identified using bioinformatics tools. The decreased expression of Hadha in NASH liver was verified by Western blotting, which was used as a complementary technique to confirm the proteomic results. CONCLUSION: This novel report on the liver mitochondrial proteome of a NASH model may provide a reservoir of information on the pathogenesis and treatment of NASH.

Dihydroartemisinin-induced inhibition of proliferation in BEL-7402 cells: an analysis of the mitochondrial proteome.

Artemisinin, the active ingredient of the Chinese medicinal herb Artemisia annua L., and its derivatives (ARTs) are currently widely used as anti-malarial drugs around the world. In this study, we found that dihydroartemisinin (DHA), one of the main active metabolites of ARTs, inhibited the proliferation of human hepatocarcinoma BEL-7402 cells in a concentration-dependent manner. To interpret the mechanisms involved, an analysis of the mitochondrial proteome was performed employing two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Seven mitochondrial proteins including fumarate hydratase, 60 kDa heat shock protein, enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, two subunits of ATP synthase and NADPH:adrenodoxin oxidoreductase were identified to be differentially expressed between the control and DHA-treated groups. Our results indicate that the imbalance of energy metabolism induced by DHA may contribute, at least in part, to its anti-cancer potential in BEL-7402 cells.