PSTPIP2 protects against alcoholic liver injury and invokes STAT3-mediated suppression of apoptosis.

Alcoholic liver injury (ALI) stands as a prevalent affliction within the spectrum of complex liver diseases. Prolonged and excessive alcohol consumption can pave the way for liver fibrosis, cirrhosis, and even hepatocellular carcinoma. Recent findings have unveiled the protective role of proline serine-threonine phosphatase interacting protein 2 (PSTPIP2) in combating liver ailments. However, the role of PSTPIP2 in ALI remains mostly unknown. This study aimed to determine the expression profile of PSTPIP2 in ALI and to uncover the mechanism through which PSTPIP2 affects the survival and apoptosis of hepatocytes in ALI, using both ethyl alcohol (EtOH)-fed mice and an EtOH-induced AML-12 cell model. We observed a consistent decrease in PSTPIP2 expression both in vivo and in vitro. Functionally, we assessed the impact of PSTPIP2 overexpression on ALI by administering adeno-associated virus 9 (AAV9)-PSTPIP2 into mice. The results demonstrated that augmenting PSTPIP2 expression significantly shielded against liver parenchymal distortion and curbed caspase-dependent hepatocyte apoptosis in EtOH-induced ALI mice. Furthermore, enforcing PSTPIP2 expression reduced hepatocyte apoptosis in a stable PSTPIP2-overexpressing AML-12 cell line established through lentivirus-PSTPIP2 transfection in vitro. Mechanistically, this study also identified signal transducer and activator of transcription 3 (STAT3) as a direct signaling pathway regulated by PSTPIP2 in ALI. In conclusion, our findings provide compelling evidence that PSTPIP2 has a regulatory role in hepatocyte apoptosis via the STAT3 pathway in ALI, suggesting PSTPIP2 as a promising therapeutic target for ALI.

Ginsenoside Rb1 for overcoming cisplatin-insensitivity of A549/DDP cells in vitro and vivo through the dual-inhibition on two efflux pumps of ABCB1 and PTCH1.

BACKGROUND: The multi-drug resistance is an inherent weakness in the chemotherapeutics of non-small cell lung cancer occurring frequently all over the world. Clinically, ginseng and Chinese medicinal prescriptions including ginseng usually used as anti-tumor adjuncts due to its characteristic of qi-invigorating, which could improve the curative effect of chemotherapy drugs and reduce their toxic side effects. Triterpenoid saponins are the crucial active ingredients in Panax ginseng, and Ginsenoside Rb1 is of the highest quantities. However, the research on the tumor drug-resistance reversal effect and mechanism of ginsenoside Rb1 is still not clear. PURPOSE: This study aimed to systematically estimate the reversal activity of Ginsenoside Rb1 on cisplatin-insensitivity of A549/DDP cells and to reveal its prospective molecular mechanism. METHODS: MTT assay were conducted to evaluate the reversal activity on cisplatin-insensitivity of A549/DDP cells of Ginsenoside Rb1in vitro, and the behavior was also studied by establishing a subcutaneous transplanted tumor model of A549/DDP in BALB/c-nu mice. In addition, P-gp ATPase activity assay, cisplatin accumulation assay, Annexin V-FITC apoptosis assay, real-time qPCR analysis and western blotting analysis were used to clarify the potential mechanism. RESULTS: Ginsenoside Rb1 could effectively reverse the cisplatin-resistance of A549/DDP in vitro and vivo. And after the co-treatment of Ginsenoside Rb1 plus cisplatin, the accumulation of cisplatin increased in A549/DDP cells, which was accompanied with the down-regulation of the mRNA and protein expression levels of ABCB1, SHH, PTCH1 and GLI2. Besides, the apoptosis-inducing ability of cisplatin improved by the relative regulation on the protein expression level of Bax and Bcl-2. Far more importantly, the changes of CYP3A4 mRNA and protein levels were not significant. CONCLUSION: Ginsenoside Rb1 could increase the concentration of intracellular cisplatin and improve the insensitivity for cisplatin on A549/DDP cells. Even better, there was perhaps no unpredictable CYP3A4-mediated pharmacokinetic interactions after the combination of Ginsenoside Rb1 plus cisplatin. Ginsenoside Rb1 was a probable reversal agent for the cisplatin-insensitivity of A549/DDP cells, with a bifunction of inhibiting the efflux of two drug pumps (P-gp and PTCH1) by targeting ABCB1 and Hedgehog (Hh) pathway. In general, this research laid the groundwork for the development of a new reversal agent for the cisplatin-insensitivity of NSCLC.

Berberine Ameliorates Abnormal Lipid Metabolism via the Adenosine Monophosphate-Activated Protein Kinase/Sirtuin 1 Pathway in Alcohol-Related Liver Disease.

Alcoholic fatty liver disease (AFLD) is an early stage of alcohol-related liver disease characterized by abnormal lipid metabolism in hepatocytes. To date, to our knowledge, there have been no effective strategies for preventing or treating alcohol-related liver disease besides alcohol abstinence. Berberine (BBR) is the main bioactive ingredient extracted from traditional Chinese medicines, such as Coptis and Scutellaria, which protect liver function and relieve liver steatosis. However, the potential role of BBR in AFLD remains unclear. Therefore, this study investigated the protective effects of BBR against Gao-binge model-induced AFLD in 6- to 8-week-old C57BL/6J male mice in vivo and ethyl alcohol (EtOH)-induced alpha mouse liver 12 (AML-12) cells in vitro. The results showed that BBR (200 mg/kg) attenuated alcoholic liver injury and suppressed lipid accumulation and metabolism disorders in vivo. Consistently, BBR effectively inhibited the expression of sterol regulatory element-binding transcription factor 1C, sterol regulatory element-binding transcription factor 2, fatty acid synthase, and 3-hydroxy-3-methylglutaryl-CoenzymeA reductase in EtOH-stimulated AML-12 cells in vitro and promoted the expression of sirtuin 1 (SIRT1) in EtOH-fed mice and EtOH-treated AML-12 cells. Furthermore, SIRT1 silencing attenuated the hepatic steatosis alleviation potential of BBR treatment. Mechanistically, molecular docking revealed the binding effect of BBR and adenosine monophosphate-activated protein kinase (AMPK). The results of further studies showed that a decrease in AMPK activity was accompanied by a significant inhibition of SIRT1 expression. SIRT1 silencing attenuated the protective effect of BBR, whereas the inhibition of its expression had no apparent effect on AMPK phosphorylation, suggesting that SIRT1 acts downstream of AMPK in AFLD. Collectively, BBR ameliorated abnormal lipid metabolism and alleviated EtOH-induced liver injury via the AMPK/SIRT1 pathway in AFLD mice.

BRD4 promotes hepatic stellate cells activation and hepatic fibrosis via mediating P300/H3K27ac/PLK1 axis.

Hepatic fibrosis (HF) is a reversible wound-healing response characterized by excessive extracellular matrix (ECM) deposition and secondary to persistent chronic injury. Bromodomain protein 4 (BRD4) commonly functions as a "reader" to regulate epigenetic modifications involved in various biological and pathological events, but the mechanism of HF remains unclear. In this study, we established a CCl4-induced HF model and spontaneous recovery model in mice and found aberrant BRD4 expression, which was consistent with the results in human hepatic stellate cells (HSCs)- LX2 cells in vitro. Subsequently, we found that distriction and inhibition of BRD4 restrained TGFß-induced trans-differentiation of LX2 cells into activated, proliferative myofibroblasts and accelerated apoptosis, and BRD4 overexpression blocked MDI-induced LX2 cells inactivation and promoted the proliferation and inhibited apoptosis of inactivated cells. Additionally, adeno-associated virus serotype 8-loaded short hairpin RNA-mediated BRD4 knockdown in mice significantly attenuated CCl4-induced fibrotic responses including HSCs activation and collagen deposition. Mechanistically, BRD4 deficiency inhibited PLK1 expression in activated LX2 cells, and ChIP and Co-IP assays revealed that BRD4 regulation of PLK1 was dependent on P300-mediated acetylation modification for H3K27 on the PLK1 promoter. In conclusion, BRD4 deficiency in the liver alleviates CCl4-induced HF in mice, and BRD4 participates in the activation and reversal of HSCs through positively regulating the P300/H3K27ac/PLK1 axis, providing a potential insight for HF therapy.

Two new dammarane-type ginsenosides from Panax ginseng.

Two new dammarane-type ginsenosides elucidated as 3-O-[ß-D-glucopyranosyl-(1→6)-ß-D-glucopyranosyl-(1→2)-ß-D-glucopyranosyl]-20-O-[α-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl]-20(S)-protopanaxadiol, named Ginsenoside Rb6(2); (Z)-6-O-ß-D-glucopyranosyl-dammar-20(22), 24-diene-3ß, 6α, 12ß-triol, named (20Z)-Ginsenoside Rh4 (4); along with five known ginsenosides(1, 3, 5, 6, 7) were isolated from the radix and rhizome of wild ginseng (Panax ginseng). The structures were determined by chemical and comprehensive spectroscopic analysis, including 1D, 2D NMR, HR-ESI-MS and hydrolysis reaction.

Two new dammarane-type saponins from radix and rhizomes of Panax ginseng C. A. Meyer.

Two new dammarane-type ginsenosides elucidated as 6-O-[α-D-glucopyranosyl-(1→3)-ß-D-glucopyranosyl]-dammar-24-ene-3ß, 6α, 12ß, 20S-tetraol, named 20(S)-Ginsenoside Re10 (4); 6-O-[α-L-rhamnopyranosyl-(1→2)-ß-D-glucopyranosyl]-20-O-[α-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl]-dammar-24-ene-3ß, 6α, 12ß, 20R-tetraol, named 20(R)-Ginsenoside Re11 (8); along with one steroidal saponin (1) and six known triterpenoid ginsenosides (2, 3, 5, 6, 7 and 9) were isolated from the radix and rhizomes of mountain-cultivated ginseng (Panax ginseng C. A. Meyer, family Araliaceae). Their structures were determined by comprehensive chemical and spectroscopic analysis. In addition, what's even more concerning is that protodioscin was isolated for the first time from Panax ginseng.

PLCL1 regulates fibroblast-like synoviocytes inflammation via NLRP3 inflammasomes in rheumatoid arthritis.

BACKGROUND: Phospholipase C-like 1 (PLCL1), a protein that lacks catalytic activity, has similar structures to the PLC family. The aim of this research was to find the function and underlying mechanisms of PLCL1 in fibroblast-like synoviocyte (FLS) of rheumatoid arthritis (RA). METHODS: In this study, we first analyzed the expression of PLCL1 in the synovial tissue of RA patients and K/BxN mice by immunohistochemical staining. Then silencing or overexpressing PLCL1 in FLS before stimulating by TNF-α. The levels of IL-6, IL-1ß and CXCL8 in FLS and supernatants were detected by Western Blot (WB), Real-Time Quantitative PCR and Enzyme Linked Immunosorbent Assay. We used INF39 to specifically inhibit the activation of NLRP3 inflammasomes, and detected the expression of NLRP3, Cleaved Caspase-1, IL-6 and IL-1ß in FLS by WB. RESULT: When PLCL1 was silenced, the level of IL-6, IL-1ß and CXCL8 were down-regulated. When PLCL1 was overexpressed, the level of IL-6, IL-1ß and CXCL8 were unregulated. The previous results demonstrated that the mechanism of PLCL1 regulating inflammation in FLS was related to NLRP3 inflammasomes. INF39 could counteract the release of inflammatory cytokines caused by overexpression of PLCL1. CONCLUSION: Result showed that the function of PLCL1 in RA FLS might be related to the NLRP3 inflammasomes. We finally confirmed our hypothesis with the NLRP3 inhibitor INF39. Our results suggested that PLCL1 might promote the inflammatory response of RA FLS by regulating the NLRP3 inflammasomes.

7-O-(2- (Propylamino)-2-oxoethyl) hesperetin attenuates inflammation and protects against alcoholic liver injury by NLRP12.

Alcoholic liver disease (ALD) is a liver disease caused by long-term heavy drinking. Alcoholic liver injury is a part of alcoholic liver disease. A large number of studies have shown that alcohol metabolism and endotoxin / lipopolysaccharide (LPS) and cycles can cause massive activation of macrophages, leading alcoholic liver injury. Hesperetin is a dihydro-flavonoid extracted from the fruits of Citrus in Rutaceae. It has a variety of pharmacological activities, including antibacterial, anti-inflammatory, antioxidant and so on, but recent studies have shown that hesperetin derivatives have stronger anti-inflammatory effects than hesperetin. In order to improve the anti-inflammatory activity of hesperetin, our group used ethyl-bromoacetate to replace the hydroxyl group at the 7 position of hesperetin to obtain the hesperetin derivative 7-O-(2-(Propylamino)-2-oxoethyl) hesperetin (HD-4d). In this study, we found that HD-4d had hepatoprotective and anti-inflammatory effects on alcoholic liver injury in C57BL/6J mice, and it also had noticeable anti-inflammatory effects in EtOH and LPS-induced RAW264.7 cells. Besides, we found that HD-4d can reduce the expression of inflammatory factors by up-regulating NLRP12 in vivo and in vitro. We found that the expression of NLRP12 was significantly increased in EtOH and LPS-induced RAW264.7 cells compared with the control group. Moreover, the inhibitory effect of HD-4d on inflammation weakened considerably after silencing NLRP12 in RAW264.7 cells. However, when NLRP12 was overexpressed with plasmid pEX-3-NLRP12, the effect of HD-4d on alcohol and LPS induced inflammation was remarkably increased. In addition, further studies indicated that HD-4d inhibited the activation and phosphorylation of the p65 protein by up-regulating NLRP12. In conclusion, HD-4d activated NLRP12 to reduce liver injury and inflammatory response through the NF-кB pathway.

Role of the S100 protein family in rheumatoid arthritis.

Rheumatoid arthritis is a chronic systemic autoimmune disease characterized by synovial hyperplasia, inflammatory cell infiltration, and proliferation of inflammatory tissue (angiogranuloma). The destruction of joints and surrounding tissues eventually causes joint deformities and dysfunction or even loss. The S100 protein family is one of the biggest subtribes in the calcium-binding protein family and has more than 20 members. The overexpression of most S100 proteins in rheumatoid arthritis is closely related to its pathogenesis. This paper reviews the relationship between S100 proteins and the occurrence and development of rheumatoid arthritis. It will provide insights into the development of new clinical diagnostic markers and therapeutic targets for rheumatoid arthritis.

PLCL1 regulates fibroblast-like synoviocytes inflammation via NLRP3 inflammasomes in rheumatoid arthritis

Abstract Background: Phospholipase C-like 1 (PLCL1), a protein that lacks catalytic activity, has similar structures to the PLC family. The aim of this research was to find the function and underlying mechanisms of PLCL1 in fibroblast-like synoviocyte (FLS) of rheumatoid arthritis (RA). Methods: In this study, we first analyzed the expression of PLCL1 in the synovial tissue of RA patients and K/BxN mice by immunohistochemical staining. Then silencing or overexpressing PLCL1 in FLS before stimulating by TNF-α. The levels of IL-6, IL-1β and CXCL8 in FLS and supernatants were detected by Western Blot (WB), Real-Time Quantitative PCR and Enzyme Linked Immunosorbent Assay. We used INF39 to specifically inhibit the activation of NLRP3 inflammasomes, and detected the expression of NLRP3, Cleaved Caspase-1, IL-6 and IL-1β in FLS by WB. Result: When PLCL1 was silenced, the level of IL-6, IL-1β and CXCL8 were down-regulated. When PLCL1 was overexpressed, the level of IL-6, IL-1β and CXCL8 were unregulated. The previous results demonstrated that the mechanism of PLCL1 regulating inflammation in FLS was related to NLRP3 inflammasomes. INF39 could counteract the release of inflammatory cytokines caused by overexpression of PLCL1. Conclusion: Result showed that the function of PLCL1 in RA FLS might be related to the NLRP3 inflammasomes. We finally confirmed our hypothesis with the NLRP3 inhibitor INF39. Our results suggested that PLCL1 might promote the inflammatory response of RA FLS by regulating the NLRP3 inflammasomes.

Research progress on naturally-occurring and semi-synthetic ocotillol-type ginsenosides in the genus Panax L. (Araliaceae).

Ocotillol (OT)-type ginsenosides, one subtype of ginsenosides, consist of a dammarane skeleton and a tetrahydrofuran ring. Most naturally-occurring OT-type ginsenosides exist in Panax species, particularly in Panax quinquefolius, which may be attributed to the warm and humid climate of its native areas. Till now, merely 28 types of naturally-occurring OT-type ginsenosides have been isolated. In contrast, semi-synthesized OT-type ginsenosides are attracted considerable attentions. These ginsenosides can be obtained through oxidation and cyclization of side chains of dammarane-type ginsenosides, and other methods, which may change their physical and chemical properties and further improve their bioavailabilities. It is also notable that the pharmacological activities of ginsenosides are closely related to the stereoisomers caused by the configuration at C-20. Semi-synthesis of OT-type ginsenosides can facilitate our understanding of the biosynthesis, transformation and metabolism of OT-type ginsenosides in the body. This review will systematically summarize the research progress on naturally-occurring and semi-synthetic OT-type ginsenosides, which provides a theoretical basis for their bioactivity-guided research.

Cytotoxic triterpenoid saponins from the root of Anemone tomentosa (Maxim.) Pei.

Two new triterpenoid saponins, tomentoside T1 (1) and tomentoside T2 (2), together with eleven known compounds (3-13) were obtained from ethyl acetate extracts of Anemone tomentosa (Maxim.) Pei by bioactivity-guided isolation. Their structures were elucidated by chemical and spectral analysis, including 1D, 2D NMR data and HRESIMS. The cytotoxic activity of all isolated saponins was evaluated on MDA-MB-231 and PC-3 cell lines. Compounds 5 and 7 showed significant cytotoxicity against with MDA-MB-231 cell and PC-3 cell with IC50 values of 3.73-4.21 µM and 3.53-5.37 µM, respectively.

Simultaneous determination of 19 constituents in Cimicifugae Rhizoma by HPLC-DAD and screening for antioxidants through DPPH free radical scavenging assay.

Cimicifugae Rhizoma (sheng ma) is a well-known traditional Chinese medicine, which has been demonstrated to possess anti-inflammatory, antipyretic and analgesic activities. In the present study, a simple and efficient HPLC-DAD (high-performance liquid chromatography coupled with diode array detection) method was developed and validated for simultaneous quantification of 19 chemical components (including 16 phenolic acids, one coumarin and two alkaloids) in Cimicifugae Rhizoma. The result indicated that this method could effectively evaluate the quality of Cimicifugae Rhizoma and provide a valuable reference for further study. Additionally, the antioxidant activity of Cimicifugae Rhizoma was evaluated by DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging assay. The results showed that the content of phenolic acids and antioxidant activity exhibited significant correlation coefficients.