ETHNOPHARMACOLOGICAL RELEVANCE:Grape seed proanthocyanidin extract (GSPE), derived from Vitis vinifera L., has been used in traditional European and Mediterranean medicine to treat inflammation, vascular diseases, and wounds. Historically employed for "detoxification" and "hard lumps" (possibly tumors), its effects are now attributed to GSPE's antioxidant and antiproliferative properties. Modern studies suggest these mechanisms may underlie its potential anticancer applications, including against bladder cancer. AIM OF THE STUDY:To validate the traditional antitumor claims of grape seeds by investigating grape seed proanthocyanidin extract (GSPE)'s efficacy against bladder cancer (BCa) in vitro and in vivo, with a focus on its metabolic regulatory mechanisms. METHODS:We employed an integrated multi-omics approach combining in vitro assays (including MTT, colony formation, migration, and cell cycle/apoptosis analyses), in vivo orthotopic BCa models, and untargeted metabolomics using ultrahigh-performance liquid chromatography-Q Exactive™ mass spectrometry (UHPLC-Q-Exactive MS). Bioinformatics analysis was performed to examine inosine 5'-monophosphate dehydrogenase 1/2 (IMPDH1/2) expression in BCa tissues and evaluate their prognostic significance. Subsequently, IMPDH1 and IMPDH2 expression levels were quantified by qPCR and Western blot, and their enzymatic activity was assessed in vitro. Molecular docking was then performed to predict potential interactions between GSPE and IMPDH1/2. RESULTS:GSPE exhibited potent dose-dependent anti-tumor effects in BCa cells, suppressing proliferation, colony formation, and migration through induction of G0/G1 arrest and apoptotic cell death. In orthotopic BCa mouse models, GSPE administration achieved 82.66% tumor growth inhibition (p < 0.001) without inducing systemic toxicity or bladder irritation, as confirmed by normal serum biochemistry and histopathological analysis. Untargeted metabolomic profiling identified 307 significantly altered metabolites, with purine/pyrimidine metabolism showing the most profound dysregulation. Bioinformatics analysis revealed that elevated IMPDH1/2 expression in BCa tissues correlated with poor patient prognosis. Mechanistically, GSPE both dose-dependently downregulated IMPDH1/2 expression (at mRNA and protein levels), directly inhibited their enzymatic activity, and bound to IMPDH1/2 active sites with high affinity. CONCLUSION:Our findings demonstrate that GSPE serves as a novel IMPDH1/2-targeted therapeutic agent capable of disrupting nucleotide metabolism in BCa through dual inhibition of both enzymatic function and gene expression. While the exact active form(s) in vivo require further pharmacokinetic investigation, the potent anti-tumor efficacy and exceptional safety profile of intravesical GSPE support its significant clinical potential as a phytotherapeutic agent for BCa treatment.

Ribavirin is a common antiviral drug, especially for patients with hepatitis C. Our recent studies demonstrated that ribavirin showed anti-tumor activity in colorectal cancer and hepatocellular carcinoma, but its effects on lung cancer remains unclear. This study aimed to evaluate the anti-tumor activity of ribavirin against lung cancer and elucidate the underlying mechanism. We established orthotopic mouse model of lung cancer (LLC and GLC-82) and employed an ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS)-based metabolomics approach. We found that ribavirin significantly inhibited the proliferation and colony formation of lung cancer cells. Tumor sizes of orthotopic lung cancer in ribavirin-treated groups were also significantly lower than those in control groups. Metabolomics analysis revealed that ribavirin mainly affected 5 metabolic pathways in orthotopic lung tumor models, taurine and hypotaurine metabolism, nicotinate and nicotinamide metabolism, linoleic acid metabolism, arginine biosynthesis and arachidonic acid metabolism. Furthermore, we identified 5 upregulated metabolites including β-nicotinamide adenine dinucleotide (NAD+), nicotinamide (NAM), taurine, ornithine and citrulline, and 7 downregulated metabolites including 1-methylnicotinamide (MNAM), S-adenosyl-l-homocysteine (SAH), N1-Methyl-2-pyridone-5-carboxamide (2PY), homocysteine (Hcy), linoleic acid, arachidonic acid (AA) and argininosuccinic acid in ribavirin-treated groups. These results provide new insight into the anti-tumor mechanism of ribavirin for lung cancer.

Supplemental Table 1-4. Supplemental table 1 and 2 were the design for drug treatment of rats and mice respectively; supplemental table 3 and 4 were the body weight changes of rats and mice during the experimental period; supplemental table 5-13 were the RAW DATA of DMH+DSS and Met-M+VD3-M groups.

目的 探讨蛇葡萄素(AMP)对人膀胱癌J82细胞增殖和凋亡的影响及相关作用机制.方法 配置不同浓度(40.00、44.74、50.04、55.96、62.59、70.00μg/ml)的AMP作用于人膀胱癌J82细胞,分别培养48、72 h,使用噻唑蓝检测不同浓度的AMP对人膀胱癌J82细胞增殖能力的影响,流式细胞术检测不同浓度(40.00、50.04、62.59μg/ml)的AMP对人膀胱癌J82细胞凋亡能力的影响,透射电镜观察细胞形态,蛋白质印迹法(Western blot)检测磷酸化的B细胞淋巴瘤/白血病-2(p-Bcl-2)、p-Bad的相对表达量.结果 不同浓度AMP处理人膀胱癌J82细胞48、72 h,抑制率(IR)随AMP浓度升高、作用时间的延长逐渐升高,具有明显的浓度依赖性和时间依赖性.AMP处理48 h的半数抑制浓度(IC50)明显高于AMP处理72 h细胞,差异有统计学意义(P﹤0.01).40.00、50.04、62.59μg/ml AMP组和阳性对照组人膀胱癌J82细胞的总凋亡率均高于阴性对照组,40.00、50.04、62.59μg/ml AMP组人膀胱癌J82细胞的总凋亡率均高于阳性对照组,差异均有统计学意义(P﹤0.05).随着AMP浓度的增加,J82细胞存活细胞率逐渐降低,早期凋亡率先上升后下降,晚期凋亡率和总凋亡率均逐渐升高.40μg/ml的AMP作用于人膀胱癌J82细胞48 h后,透射电镜下可见凋亡细胞体积变小,细胞质浓缩,出现凋亡小体.作用24、48 h,70μg/ml的AMP处理的人膀胱癌J82细胞凋亡抑制蛋白p-Bad、p-Bcl-2相对表达量均低于处理12 h时(P﹤0.05).结论 AMP可有效抑制人膀胱癌J82细胞的增殖,促进其凋亡,诱导机制可能与其能够下调抗细胞凋亡蛋白p-Bad、p-Bcl-2的表达有关.

Type I co-activator-associated arginine methyltransferase 1 (CARM1) and type II protein arginine methyltransferase 5 (PRMT5) are highly expressed in multiple cancers including liver cancer and their overexpression contributes to poor prognosis, thus making them promising therapeutic targets. Here, we evaluated anti-tumor activity of ribavirin in hepatocellular carcinoma (HCC). We found that ribavirin significantly inhibited the proliferation of HCC cells in a time- and dose-dependent manner. Furthermore, ribavirin suppressed the growth of subcutaneous and orthotopic xenograft of HCC in mice, decreased vascular endothelial growth factor (VEGF) and peritoneal permeability to reduce ascites production, and prolonged the survival of mice in HCC ascites tumor models. Mechanistically, ribavirin potently down-regulated global protein expression of CARM1 and PRMT5, and concurrently decreased accumulation of H3R17me2a and H3R8me2s/H4R3me2s. However, ribavirin did not affect the activity and mRNA levels of both CARM1 and PRMT5 in vivo and in vitro HCC cells. In addition, our ChIP results shown that ribavirin inhibited CARM1 which in turn decreased the H3R17me2a, binds to the eukaryotic translation initiation factor 4E (eIF4E) and VEGF promoter region, and reduced the relative mRNA expression level of eIF4E and VEGF in HCC cells. Our findings suggested a potential therapeutic strategy for patients with HCC through inhibition of the abnormal activation/expression of both CARM1 and PRMT5.

Ginsenoside Rd (GS-Rd), isolated from the Chinese traditional herbal medicine Panax ginseng, is used for the treatment of cardiovascular diseases, inflammation, different body pains, and trauma. Caspase-3 and Caspase-9 belong to cysteine aspartic acid specific protease (Caspase) family that plays an important role in apoptosis progression of cancers. In the present study, we investigated the anti-tumor effect of GS-Rd by MTT assay, colony formation assessment, flow cytometry, and Western blotting. Our results revealed that ginsenoside Rd significantly inhibits human gastric cancer (GC) growth and cell proliferation. Flow cytometer analysis showed that the GS-Rd could significantly induce apoptosis and arrest the G0/G1 phase in GC cells. Further, GS-Rd was found to increase the ratio of Bax/Bcl-2 and the expression of Caspase-3 and Caspase-9, respectively, and to decrease the expression of Cyclin D1. Taken together, our study suggests that GS-Rd significantly inhibits GC cell proliferation, induces cell apoptosis through increase the expression of Caspase-3, Caspase-9, and the ratio of Bax/Bcl-2. GS-Rd also induces cell cycle arrest at G0/G1 phase by down-regulation Cyclin D1. Thus, GS-Rd could serve as a lead to develop novel therapeutic agents to against human gastric cancer.

Ginsenoside Rd (GRd) is a protopanaxadiol-type ginsenoside found in ginseng whose antitumor activities have been reported. We previously reported the chemoprevention of vitamin D3 (VD3) and metformin (Met) in mice and rat models of colorectal carcinogenesis. Here we investigated the potential synergistic chemoprevention of GRd, VD3, and Met on colitis-associated colon neoplasia induced with 1,2-dimethylhydrazine dihydrochloride (DMH) via intraperitoneal injection, and dextran sodium sulfate (DSS) in the drinking water. Our results showed that combination of GRd, VD3, and Met treatment exhibit synergistic effects against the early colon cancer, as evidenced by reduced the numbers of aberrant crypt foci (ACF) and tumor in the colon of mice. The possible mechanism of their action was associated with downregulation S6P expression via the AMPK/mTOR pathway. These results suggest that the combination of GRd, VD3, and Met may prevent colon cancer associated with colitis.

Previous studies have demonstrated that ampelopsin (AMP), a type of flavonoid isolated from the stems and leaves of Ampelopsis grossedentata, exhibits anti-cancer activity in various types of cancer. Conversion of AMP into its sodium salt (AMP-Na) conferred enhanced solubility and stability to it. The present study aimed to evaluate the anti-cancer activity of AMP-Na in human lung adenocarcinoma cell lines and to investigate its mechanisms of action. Cell proliferation and viability were assessed by MTT and colony formation assays, and cell migration was determined using a scratch wound healing assay. The cell cycle distribution, apoptosis rate and tubulin immunofluorescence intensity were analyzed using flow cytometry, the cell ultra-microstructure was examined using transmission electron microscopy and the accumulation of tubulin was determined using laser confocal microscopy. The results demonstrated that AMP-Na significantly inhibited the proliferation, clonogenicity and migration of human lung adenocarcinoma cells. Furthermore, AMP-Na induced SPC-A-1 cell apoptosis, and promoted tubulin polymerization. The results suggested that the underlying mechanisms of AMP-Na may involve targeting of microtubules and tubulin polymerization to subsequently disrupt mitosis and induce cell cycle arrest at the S-phase.

Protein arginine methyltransferase 5 (PRMT5), a type II PRMT, is highly expressed in several types of tumors including cervical cancer. Arginine methyltransferase inhibitor 1 (AMI-1) inhibits solid tumors by targeting PRMT5. However, the effect of AMI-1 on cervical cancer is still unknown. In this study, we provided the first evidence that AMI-1 reduced cervical cancer cell proliferation, colony formation and promoted cell apoptosis in vitro. Suppression of tumorigenicity was also confirmed in vivo. Mechanistic studies revealed that AMI-1 significantly reduced PRMT5 level in cells and mice xenografts model of cervical cancer. These results suggest that AMI-1 inhibits cervical cancer by type II PRMT5.

In medicine,the ability of the nervous system to repair nerve damage is often limited.In recent years,it has been found that adipose derived stem cells (ADSCs) have repair effects on various types of nerve injury and can be the seed cells for nerve injury repair.Not only being as stem cells,ADSCs also have some exclusive advantages.ADSCs belong to adult cells,which derived from the mesoderm,with multi-lineage differentiation potential,low immunogenicity,easy to gain,low risk after transplantation,thus make them become excellent immunogenicity,easy to gain,low risk after transplantation,thus make them become excellent seed cells for the nerve repair.The characteristics of adipose derived stem cells and their research progress and existing problems on nerve injury repair were reviewed.

Ciliary neurotrophic factor (CNTF) analogues were reported to ameliorate fatty liver in db/db or high-fat diet-fed mice. It is generally thought that CNTF exerts its actions centrally. The aim of this study was to investigate whether peripheral effects of CNTF analogues are involved in the therapeutic effect on high fat-induced hepatic steatosis. The rat model of fatty liver was induced by a high-fat diet (HFD) for 12 weeks. In the next 2 weeks, rats were fed the HFD along with subcutaneous injection of vehicle or mutant recombinant human CNTF (rhmCNTF 0.05-0.2 mg/kg per day). Steatotic HepG2 cells were induced by 50% fetal bovine serum (FBS) for 48 hours, and then treated with rhmCNTF for 24 hours. The results showed that after rhmCNTF treatment, hepatic triglyceride (TG) accumulation was attenuated both in vivo and in vitro. RhmCNTF increased protein expression of CPT-1 and PPARα, and decreased SREBP-1c, FAS and SCD-1 in steatotic HepG2 cells. But the production of nitric oxide and 8-isoPGF2α in steatotic HepG2 cells was not affected by rhmCNTF. These results suggest that rhmCNTF has a peripheral effect that alleviates fat-induced hepatic steatosis.

Ciliary neurotrophic factor (CNTF) and CNTF analogs were reported to have hepatoprotective effect and ameliorate hepatic steatosis in db/db or high-fat-diet-fed mice. Because hepatic steatosis and injury are also commonly induced by hepatotoxin, the aim of the present study is to clarify whether CNTF could alleviate hepatic steatosis and injury induced by carbon tetrachloride (CCl4). Unexpectedly, when combined with CCl4, CNTF aggravated hepatic steatosis and liver injury. The mechanism is associated with effects of CNTF that inhibited lipoprotein secretion and drastically impaired the ability of lipoproteins to act as transport vehicles for lipids from the liver to the circulation. While injected after CCl4 cessation, CNTF could improve liver function. These data suggest that CNTF could be a potential hepatoprotective agent against CCl4-induced hepatic injury after the cessation of CCl4 exposure. However, it is forbidden to combine recombinant mutant of human CNTF treatment with CCl4.

To evaluate the cytotoxic effects of ampelopsin sodium (Amp-Na) and carboplatin (CBP) used alone or in combination on human non-small cell lung cancer (NSCLC) cells SPC-A1 in vitro and its related mechanism.

Ampelopsin is a well-known flavonoid which has variety of biological and pharmacological actions including anticancer effects and induction of apoptosis on the several cancer cell lines. The present study aimed to evaluate the role of ampelopsin sodium (Amp-Na) in the mitochondrial-mediated apoptosis of human lung adenocarcionma SPC-A-1 cells. The analysis of cell proliferation and ultrastructure were performed. Furthermore, to clarify its action mechanism by determining the mitochondrial membrane potential (Δψm), intracellular calcium (Ca2+) concentration, mitochondrial nitric oxide (NO) level and total ATPase activity. The results showed that Amp-Na markedly inhibited the SPC-A-1 cell proliferation and caused ultrastructural apoptosis feature in SPC-A-1 cells in a dose-dependent manner. Amp-Na led to a rapid and sustained Ca2+ elevation and Δψm reduction, and induced the mitochondrial NO production and decreased the total ATPase activity in SPC-A-1 cells. The results enhance the potential of Amp-Na as a therapeutic drug for treating lung cancer, and provide new information for mechanism of Amp-Na which induces mitochondrial-mediated apoptosis in tumor cells.

目的:研究蛇葡萄素钠单用及与卡铂联合用药对小鼠肺癌Lewis细胞的体外抗肿瘤作用,初步探讨蛇葡萄素钠抑瘤作用机制.方法:应用MTT比色法测定蛇葡萄素钠单用及与卡铂联合用药对小鼠肺癌Lewis细胞增殖的抑制作用;使用流式细胞仪检测蛇葡萄素钠单用及与卡铂合用诱导细胞凋亡作用.结果:MTT实验结果表明,蛇葡萄素钠单用在12.5、25、50、100μg/ml浓度范围内对小鼠肺癌Lewis细胞的增殖具有浓度依赖性的抑制作用;6.25～50 μg/ml的蛇葡萄素钠与卡铂3.12～ 25 μg/ml联合用药对卡铂抑制Lewis细胞的增殖作用具有协同效应;流式细胞仪结果表明,蛇葡萄素钠25、50、100 μg/ml与卡铂25 μg/ml联合应用时,凋亡率显著增高.结论:蛇葡萄素钠单用在一定的浓度范围内对Lewis细胞增殖具有浓度依赖性抑制作用,蛇葡萄素钠与卡铂合用可以增强卡铂的活性,有协同抑制作用,蛇葡萄素钠可能是通过诱导凋亡而发挥其抗肿瘤作用.

Objective: The close connection between high blood FFA and insulin resistance (IR) in obese individuals is well-known. The purpose of this study was to identify whether the blood FFA increased in obese-IR animals.Methods: Obese-IR animal models were established using high-fat diet (HFD) or HFD and streptozocin, and treated with drugs.Results: The serum FFA of obese-IR animals was not increased, even significantly lower than that of normal animals, and were not significantly decreased when insulin sensitivity and obesity-related indices were ameliorated after treatment.Conclusion: The results suggest that blood FFA are unlikely the link between obesity and insulin resistance. (C) 2015 Asian Oceanian Association for the Study of Obesity. Published by Elsevier Ltd. All rights reserved.

目的:研究蛇葡萄素钠对人肺腺癌SPC-A-1细胞、人正常胚肺MRC-5细胞的毒性作用。方法:应用MTT法检测细胞增殖,流式细胞仪检测细胞周期和细胞凋亡;透射电子显微镜（TEM）观察细胞超微结构的变化。结果:MTT法实验表明,蛇葡萄素钠、卡铂显著抑制SPC-A-1细胞的增殖,呈现浓度依赖性,IC50分别为57.18±9.42 ug/ml、33.56±11.2 ug/ml,但对MRC-5细胞增殖的抑制作用不明显,IC50分别大于5000 ug/ml及500ug/ml;流式细胞检测分析表明,25~100ug/ml的蛇葡萄素钠可使SPC-A-1细胞增殖阻滞于S期,G0/G1期细胞明显减少,且呈浓度依赖性,50、100 ug/ml的蛇葡萄素钠可使SPC-A-1细胞发生明显凋亡,而蛇葡萄素钠对MRC-5细胞周期及凋亡的影响不明显。蛇葡萄素钠浓度为50、100ug/ml时,透射电镜下可见SPC-A-1细胞表现出典型的凋亡特征,而MRC-5细胞变化不明显。结论:蛇葡萄素钠对肿瘤细胞具有较强的毒作用,对正常细胞毒性较小。

Objective: The aim of the study was to investigate the therapeutic effect of administration route of ginsenoside Rd in the treatment of ulcerative colitis(UC). Methods:Colitis in rats was induced by colonic administration with 2,4,6-trinitrobenzene sulfonic acid(TNBS).Wistar rats were randomly divided into three groups:TNBS control group, ginsenoside Rd intragastric group(ig at 30mg/kg ginsenoside Rd)and intramuscular injection group(im at 30mg/kg ginsenoside Rd). Rats were administrated by ginsenoside Rd(30mg/kg)per day for 7days with different administration route. Macroscopic and microscopic damage scores and changes in weight/length ratio(mg/mm)of colon segments were analyzed. Results:1.Compared with the TNBS control group, Rd ig group and im group facilitated recovery of pathologic changes in the colon and decreased colonic weight/length ratio(P 0.01 or P 0.05). 2.Rd also notably reduced the colonic macroscopic and microscopic damage scores(P0.01). Conclusion:Rd exerts a beneficial effect in the TNBS-induced colitis in rats and compared with intramuscular injection, the administration of Rd by ig has a better effect on ulcerative colitis in rats.

OBJECTIVE:To study the inhibitory effects of ampelopsin sodium (AMP-Na) combined with carboplatin on the proliferation of Lewis cell of lung cancer. METHODS:6.25,12.5,25,50 and 100 μg/ml AMP-Na and 3.125,6.25,12.5,25 and 50 μg/ml carboplatin were used to culture the cells for 4 h,and the cell viability was determined and the inhibition rate and half in-hibition concentration(IC50)were calculated. 12.5,25,50 and 100 μg/ml AMP-Na and 12.5 μg/ml carboplatin were used to culture the cells for 12 h,and the flow cytometry was used to determine the expression of Caspase-3. RESULTS:AMP-Na with serial con-centrations combined with carboplatin with serial concentrations had obvious inhibitory effects on the cell proliferation. With the in-crease of mass concentration,the IC50 of carboplatin on the Lewis cells was gradually decreased;when the AMP-Na of 6.25-50μg/ml was combined with carboplatin of 3.125-25 μg/ml,it showed the strongest inhibitory effects on the Lewis cell proliferation. When cells were cultured with AMP-Na and carboplatin for 24 h,the expression of Caspase-3 increased significantly. CONCLU-SIONS:AMP-Na combined with carboplatin has synergistic inhibitory effect on the Lewis cell proliferation by a mechanism that may be related to the apoptosis induced by Caspase-3 activated by AMP-Na.

Protein arginine methyltransferases (PRMTs) plays critical roles in cancer. PRMT5 has been implicated in several types of tumors. However, the role of PRMT5 in cancer development remains to be fully elucidated. Here, we provide evidence that PRMT5 is overexpressed in colorectal cancer (CRC) cells and patient-derived primary tumors, correlated with increased cell growth and decreased overall patient survival. Arginine methyltransferase inhibitor 1 (AMI-1)strongly inhibited tumor growth, increased the ratio of Bax/Bcl-2, and induced apoptosis in mouse CRC xenograt model. AMI-1 also induced apoptosis and decreased the migratory activity in several CRC cells. In CRC xenografts AMI-1 significantly decreased symmetric dimethylation of histone 4 (H4R3me2s), a histone mark of type II PRMT5, but not the expression of H4R3me2a, a histone mark of type I PRMTs. These results suggest that the inhibition of PRMT5 contributes to the antitumor efficacy of AMI-1. Chromatin immunoprecipitation (ChIP) identified FGFR3 and eIF4E as two key genes regulated by PRMT5. PRMT5 knockdown reduced the levels of H4R3me2s and H3R8me2s methylation on FGFR3 and eIF4E promoters, leading to decreased expressions of FGFR3 and eIF4E. Collectively, our findings provide new evidence that PRMT5 plays an important role in CRC pathogenesis through epigenetically regulating arginine methylation of oncogenes such as eIF4E and FGFR3.