[Mechanism of total flavonoids from Ampelopsis grossedentata against gouty arthritis based on multi-level interactive network and in vivo experimental validation].

The present study investigated the mechanism of total flavonoids from Ampelopsis grossedentata(AGTF) against gouty arthritis(GA) by network pharmacology and experimental validation. The main active ingredients and targets of AGTF, as well as disease targets, were screened out using relevant databases and literature data. The "protein-protein interaction"(PPI) network and "drug-ingredient-target-pathway" network were constructed, and the potential targets and mechanism of AGTF against GA were predicted. The hyperuricemia(HUA) combined with GA model was induced in rats. The gait behaviors of rats were scored, and ankle swelling degree was observed. The uric acid(UA) level and xanthine oxidase(XOD) activity in the rat serum were detected, and the levels of interleukin-1ß(IL-1ß), interleukin-6(IL-6), and tumor necrosis factor-α(TNF-α) were measured. The protein expression of toll-like receptor 4(TLR4), myeloid differentiation factor 88(MyD88), and nuclear factor-kappa B(NF-κB) in the synovial tissues of the rat ankle joint was determined by immunohistochemistry. Ten active ingredients of AGTF and 73 candidate targets of AGTF against GA were screened out by network pharmacology. Eighty-six signaling pathways were enriched, including TNF signaling pathway, NF-κB signaling pathway, TLR signaling pathway, Nod-like receptor signaling pathway, and purine metabolism signaling pathway, which were closely related to AGTF against GA. Animal experimental results showed that AGTF could effectively improve the abnormal gait behaviors of GA rats, relieve ankle inflammation, and reduce ankle joint swelling. In addition, AGTF could significantly reduce UA level, inhibit XOD activity, decrease TNF-α, IL-6, and IL-1ß content, and down-regulate the expression of TLR4, MyD88, and NF-κB in ankle synovial tissues(P<0.05, P<0.01). The results of network pharmacology and experimental validation are consistent, indicating that AGTF exerts its therapeutic effect on GA by regulating UA metabolism, improving abnormal UA level, reducing the release of inflammatory factors, and regulating immunity and the TLR4/MyD88/NF-κB inflammatory pathway.

Pseudolaric acid B exhibits anti-cancer activity on human hepatocellular carcinoma through inhibition of multiple carcinogenic signaling pathways.

BACKGROUND: Pseudolaric acid B (PAB), a diterpene acid isolated from the root bark of Pseudolarix kaempferi, exhibits a potent anti-cancer activity in a variety of tumor cells. PURPOSE: The present study was designed to evaluate the anti-cancer effects of PAB on hepatocellular carcinoma (HCC) cell lines in vitro, and to explore the underlying mechanism. METHODS: The anti-proliferative activity of PAB on HCC cells were assessed via sulforhodamine B staining, colony formation, cell cycle analysis, respectively. Apoptosis was detected using Annexin V/propidium iodide double staining and diamidino-phenyl-indole staining, respectively. Protein expression regulated by PAB treatment was tested by western blotting. RESULTS: The present results showed that PAB significantly inhibited the proliferation of HepG2, SK-Hep-1, and Huh-7 HCC cell lines in vitro with IC50 values of 1.58, 1.90, and 2.06 µM, respectively. Furthermore, PAB treatment repressed the colony formation in HepG2, SK-Hep-1, and Huh-7 HCC cell lines. Flow cytometry analysis revealed that PAB caused an obvious cell cycle arrest in G2/M phase and induced apoptosis with the induction of p21, Bax, cleaved-caspase-3, and cleaved-PARP in human HepG2 and SK-Hep-1 cells. Mechanistically, PAB treatment down-regulated the phosphorylation of STAT3, ERK1/2, and Akt. Moreover, abnormal GSK-3ß/ß-catenin signaling in HepG2 cells was remarkably suppressed by PAB treatment. Finally, proliferation markers including cyclin D1 and c-Myc, and anti-apoptosis proteins such as Bcl-2 and survivin were also down-regulated by PAB treatment in HepG2 cells. CONCLUSION: Taken together, our results suggest that PAB exerts anti-cancer activity in HCC cells through inhibition of STAT3, ERK1/2, Akt, and GSK-3ß/ß-catenin carcinogenic signaling pathways, and may be used as a phytomedicine in the treatment of HCC.

Astragalus polysaccharides exerts anti-infective activity by inducing human cathelicidin antimicrobial peptide LL-37 in respiratory epithelial cells.

Astragalus polysaccharides (APS), one of the major active components in Astragalus membranaceus, is an effective immunomodulator used in the treatment of immunological diseases in China. However, the anti-infective action and mechanism of APS is not fully known. In the present study, we found that APS induced the expression of human cathelicidin antimicrobial peptide LL-37, a key host anti-infective molecule, in both mRNA and protein levels in respiratory epithelial cells HBE16 and A549. Furthermore, the lysate and supernatant from APS-treated HBE16 cells both exhibited an obvious antibacterial action, which was partially neutralizated by LL-37 monoclonal antibody. In addition, APS also significantly elevated the phosphorylation of p38 MAPK and JNK and caused the degradation of IκBα. Specific inhibitors of p38 MAPK, JNK, or NF-κB obviously abolished APS-induced LL-37 synthesis and antibacterial activity, respectively. Taken together, our results confirmed the enhancement of APS on LL-37 induction and antibacterial action in respiratory epithelial cells, which may be attributed to activation of p38 MAPK/JNK and NF-κB pathways. Furthermore, these results also supported the clinical application of APS in the treatment of infectious diseases.

The anti-arthritic activity of total glycosides from Pterocephalus hookeri, a traditional Tibetan herbal medicine.

CONTEXT: Pterocephalus hookeri (C. B. Clarke) Hock., a traditional Tibetan herbal medicine rich in glycosides, has been used to treat several diseases including rheumatoid arthritis. OBJECTIVE: To evaluate the anti-arthritic activity of total glycosides from P. hookeri, and its possible mechanisms of action. MATERIALS AND METHODS: Anti-arthritic activity of total glycosides from P. hookeri (oral administration for 30 days at 14-56 mg/kg) was evaluated using paw swelling, arthritis scores and histopathological measurement in adjuvant-induced arthritis (AA) Sprague-Dawley rats. The NF-κB p65 expression in synovial tissues, and serum superoxide dismutase (SOD) activity, malondialdehyde (MDA) and nitric oxide (NO) levels was measured in AA rats, respectively. Further assessment of anti-inflammatory and analgesic activities of these glycosides were carried out using inflammation and hyperalgesia models induced by xylene, carrageenan, agar and acetic acid, respectively. RESULTS: Total glycosides (56 mg/kg) decreased the paw swelling (38.0%, p < 0.01), arthritis scores (25.3%, p < 0.01) and synovial inflammation in AA rats. The glycosides significantly (p < 0.05-0.01) attenuated the inflammation induced by xylene, carrageenan, acetic acid and agar, increased the pain threshold in acetic acid-induced writhing in mice and mechanical stimuli-induced hyperalgia in AA rats. The glycosides (14, 28, 56 mg/kg) also suppressed the NF-κB p65 expression (33.1-78.2%, p < 0.05-0.01), reduced MDA (21.3-35.9%, p < 0.01) and NO (20.3-32.4%, p < 0.05-0.01) levels, respectively, enhanced the SOD activity (7.8%, p < 0.05) at 56 mg/kg in AA rats. DISCUSSION AND CONCLUSION: Our findings confirmed the anti-arthritic property of the total glycosides from P. hookeri, which may be attributed to its inhibition on NF-κB signalling and oxidative stress.

Anti-HBV Activities of Three Compounds Extracted and Purified from Herpetospermum Seeds.

The goal of this research was to evaluate the anti-hepatitis B virus (HBV) activities of three compounds extracted and purified from Herpetospermum seeds (HS) on HepG2.2.15 cells. Herpetin (HPT), herpetone (HPO), and herpetfluorenone (HPF) were isolated from HS and identified using HR-ESI-MS and NMR. Different concentrations of the drugs were added to the HepG2.2.15 cells. Cell toxicity was observed with an MTT assay, cell culture supernatants were collected, and HBsAg and HBeAg were detected by ELISA. The content of HBV DNA was determined via quantitative polymerase chain reaction (PCR) with fluorescent probes. The 50% toxicity concentration (TC50) of HPF was 531.48 µg/mL, suggesting that this species is less toxic than HPT and HPO. HPT and HPF showed more potent antiviral activities than HPO. The 50% inhibition concentration (IC50) values of HPF on HBsAg and HBeAg were 176.99 and 134.53 µg/mL, respectively, and the corresponding therapeutic index (TI) values were 2.66 and 3.49, respectively. HPT and HPF were shown to significantly reduce the level of HBV DNA in the HepG2.2.15 culture medium compared to the negative control. This initial investigation of the anti-HBV constituents of HS yielded three compounds that revealed a synergistic effect of multiple components in the ethnopharmacological use of HS.