Down-regulation of GPR137 expression inhibits proliferation of colon cancer cells

G protein-coupled receptors （GPRs） are highly related to oncogenesis and cancer metastasis. G protein-coupled re- ceptor 137 （GPR137） was initially reported as a novel orphan GPR about 10 years ago. Some orphan GPRs have been implicated in human cancers. The aim of this study is to investigate the role of GPR137 in human colon cancer. Expression levels of GRP137 were analyzed in different colon cancer cell lines by quantitative polymerase chain re- action and western blot analysis. Lentivirus-mediated short hairpin RNA was specifically designed to knock down GPR137 expression in colon cancer cells. Cell viability was measured by methylthiazoletetrazolium and colony forma- tion assays. In addition, cell cycle characteristic was investi- gated by flow cytometry. GRP137 expression was observed in aH seven colon cancer cell lines at different levels. The mRNA and protein levels of GPR137 were down-regulated in both HCTll6 and RKO cells after lentivirus infection. Lentivirus-mediated silencing of GPR137 reduced the proliferation rate and colonies numbers. Knockdown of GPR137 in both cell lines led to cell cycle arrest in the G0/G1 phase. These results indicated that GPR137 plays an important role in colon cancer cell proliferation. A better understanding of GPR137＇s effects on signal transduction pathways in colon cancer cells may provide insights into the novel gene therapy of colon cancer.     

Cloning,molecular characterization,and spatial and developmental expression analysis of GPR41 and GPR43 genes in New Zealand rabbits

Short-chain fatty acids (SCFAs) play a regulatory role in various physiological processes in mammals and act as endogenous ligands for the G protein-coupled receptors (GPR) 41 and 43. The role of GPR41 and GPR43 in mediating SCFA signaling in the rabbit remains unclear. The present study was to investigate the sequence of the GPR41 and GPR43 messenger RNA (mRNA) and their expression pattern in different tissues and developmental stages in New Zealand rabbit. Comparison of genomic sequences in GenBank using the Basic Local Alignment Search Tool program suggested that the New Zealand rabbit GPR41 mRNA has high similarities with the human (84%), bovine (84%) and Capra hircus (84%) genes. Similarly, GPR43 mRNA has high similarity with the rat (84%) and mouse (84%) genes. Real-time PCR results indicated that GPR41 and GPR43 mRNA were expressed throughout rabbit’s whole development and were expressed in several tissues. G protein-coupled receptor 41 and GPR43 mRNA were most highly expressed in pancreas (P<0.05) and s.c. adipose tissue (P<0.05), respectively. The expression levels of GPR41 mRNA was down-regulated in duodenum, cecum (P<0.05) and pancreas and up-regulated in jejunum, ileum, adipose tissue and spleen during growth. G protein-coupled receptor 43GPR43 mRNA was highly expressed in the duodenum, jejunum, ileum, colon, cecum and lung at 15^th day (P<0.05), whereas the expression levels in the pancreas and spleen increased later after birth, with the highest expression at 60^th day (P<0.05).     

Synthesis and activity of tumor-homing peptide iRGD and histone deacetylase inhibitor valproic acid conjugate

In this Letter, we present a concise strategy to prepare a conjugate of the tumor homing peptide iRGD and histone deacetylase inhibitor valproic acid, VPA–GFLG-iRGD. The conjugate VPA–GFLG-iRGD and a mixture of VPA and GFLG-iRGD have shown similar cytotoxicity against DU-145 prostate cancer cells. However, the treatment of DU-145 cells with conjugate VPA–GFLG-iRGD resulted in a decreased percentage of cells in the G2 phase, whereas the exposure of a mixture of VPA and GFLG-iRGD led to an increased percentage of cells in the G2 phase. We also found that GFLG-iRGD possessed cytotoxicity at the tested concentrations.     

组蛋白去乙酰化酶抑制剂调控干细胞分化与体细胞重编程的研究进展

表观遗传调控,如组蛋白乙酰化修饰,是决定干细胞分化方向的重要机制。组蛋白去乙酰化酶抑制剂(HDACi)通过影响不同亚类的组蛋白去乙酰化酶(HDAC)活性,提高组蛋白乙酰化水平,调控基因表达,从而影响胚胎干细胞自我更新,以及沿神经元、心肌和造血等细胞谱系的定向分化。HDACi类小分子化合物在体细胞重编程中也有广泛的应用,可替代致癌因子c-Myc和Klf4,促进体细胞克隆。研究显示,HDACi的效应与药物剂量、细胞类型和细胞分化状态密切相关。本文主要阐述了HDACi在干细胞分化和体细胞重编程中的应用进展,并对所涉及的分子通路进行讨论,有助于揭示干细胞定向分化的关键分子机制,优化干细胞定向分化诱导策略,对干细胞诱导分化具有重要的理论和实用价值。     

Apoptotic and antiproliferative effects of silk protein sericin conjugated-AgNO3 nanoparticles in human breast cancer cells

Bombyx mori silk sericin is a globular-like protein that is used as an antioxidant, antibacterial, and antitumor agent. In this current research, we isolated sericin by degumming process and formation of sericin-AgNO₃ NPs confirmed by UV–vis spectra, SEM, EDX, FTIR, and XRD patterns. The sericin and sericin-AgNO₃ NPs mediated changes in human breast cancer cells were determined. The antiproliferative activity of sericin-AgNO₃ NPs was analyzed by MTT dye reduction assay. Alterations at molecular levels were investigated by qRT-PCR, while apoptotic effects were studied by nuclear DNA staining. After 72 h treatment, sericin and sericin-AgNO₃ NPs showed significant antiproliferative effects in MDA-MB-231 (26 %) and MCF-7 (41 %) cells. Expression modification showed prominent stimulation of cell cycle arrest and stress related genes such as cyclin-dependent kinase inhibitors (CDKN1A, CDKN1B), and GADD family genes. RT-PCR results of the GADD family include GADD45A, B, G, 34, 153 and cyclin-dependent kinase inhibitors (CDKN1A, 1B) showed pronounced induction of 3.1 to 19.8-folds in MCF-7 cell line while induction in MDA-MB-231 cell line was 2.5 to 34.3-folds. Nuclear DAPI staining showed significant induction of apoptosis and nuclear fragmentation in MDA-MB-231 cells at a concentration of 1 mg/mL for both sericin and sericin-AgNO₃ NPs. Meanwhile, in case of MCF-7 cells, after treatment with sericin and sericin-AgNO₃ NPs (1 mg/mL), the cells changed into a round shape and lost their original spindle outlook in dose-dependent manners. We concluded that sericin-AgNO₃ NPs have significant antiproliferative, apoptosis, and genetic profiling effects in both breast cancer cell lines at the highest concentration.     

Design,synthesis and biological evaluation of pyrimidine derivatives as novel CDK2 inhibitors that induce apoptosis and cell cycle arrest in breast cancer cells

Cyclin-dependent kinase 2 (CDK2) plays a key role in eukaryotic cell cycle progression which could facilitate the transition from G1 to S phase. The dysregulation of CDK2 is closely related to many cancers. CDK2 is utilized as one of the most studied kinase targets in oncology. In this article, 24 benzamide derivatives were designed, synthesized and investigated for the inhibition activity against CDK2. Our results revealed that the compound 25 is a potent CDK2 inhibitor exhibiting a broad spectrum anti-proliferative activity against several human breast cancer cells. Additionally, compound 25 could block cell cycle at G0 or G1 and induce significant apoptosis in MDA-MB-468 cells. These findings highlight a rationale for further development of CDK2 inhibitors to treat human breast cancer.     

TGF-beta2 inhibits AKT activation and FGF-2-induced corneal endothelial cell proliferation

The corneal endothelial cells form a boundary layer between anterior chamber and cornea. This single cell layer is important to maintain cornea transparency by eliciting net fluid transport into the anterior chamber. Injuries of the corneal endothelial layer in humans lead to corneal swelling and translucence. This hindrance is thought to be due to limited proliferative capacity of the endothelial layer. Fibroblast growth factor 2 (FGF-2) and transforming growth factor-beta 2 (TGF-beta2) are both found in aqueous humor, and these two cytokines promote and inhibit cell growth, respectively. The intracellular signaling mechanisms by which TGF-beta2 suppresses the mitogenic response to FGF-2, however, remain unclear. We have addressed this question by investigating potential crosstalk between FGF-2-induced and TGF-beta2-regulated intracellular signaling events in cultured bovine corneal endothelial (BCE) cells. We found that TGF-beta2 and FGF-2 oppositely affect BCE cell proliferation and TGF-beta2 can override the stimulating effects of FGF-2 by increasing COX-2 expression in these cells. Consistent with these findings, overexpression of COX-2 significantly reduced FGF-2-induced cell proliferation whereas a COX-2 specific inhibitor NS398 reversed the effect of TGF-beta2 on FGF-2-induced cell proliferation. The COX-2 product prostaglandin E2 (PGE-2) blocks FGF-2-induced cell proliferation. Whereas FGF-2 stimulates cell proliferation by activating the AKT pathway, TGF-beta2 and PGE-2 both inhibit this pathway. In accordance with the effect of PGE-2, cAMP also inhibits FGF-2-induced AKT activation. These findings suggest that the mitogenic response to FGF-2 in vivo in the corneal endothelial layer may be inhibited by TGF-beta2-induced suppression of the PI3-kinase/AKT signaling pathway.     

TGEV nucleocapsid protein induces cell cycle arrest and apoptosis through activation of p53 signaling

Our previous studies showed that TGEV infection could induce cell cycle arrest and apoptosis via activation of p53 signaling in cultured host cells. However, it is unclear which viral gene causes these effects. In this study, we investigated the effects of TGEV nucleocapsid (N) protein on PK-15 cells. We found that TGEV N protein suppressed cell proliferation by causing cell cycle arrest at the S and G2/M phases and apoptosis. Characterization of various cellular proteins that are involved in regulating cell cycle progression demonstrated that the expression of N gene resulted in an accumulation of p53 and p21, which suppressed cyclin B1, cdc2 and cdk2 expression. Moreover, the expression of TGEV N gene promoted translocation of Bax to mitochondria, which in turn caused the release of cytochrome c, followed by activation of caspase-3, resulting in cell apoptosis in the transfected PK-15 cells following cell cycle arrest. Further studies showed that p53 inhibitor attenuated TGEV N protein induced cell cycle arrest at S and G2/M phases and apoptosis through reversing the expression changes of cdc2, cdk2 and cyclin B1 and the translocation changes of Bax and cytochrome c induced by TGEV N protein. Taken together, these results demonstrated that TGEV N protein might play an important role in TGEV infection-induced p53 activation and cell cycle arrest at the S and G2/M phases and apoptosis occurrence.     

RPS27a promotes proliferation,regulates cell cycle progression and inhibits apoptosis of leukemia cells

Ribosomal protein S27a (RPS27a) could perform extra-ribosomal functions besides imparting a role in ribosome biogenesis and post-translational modifications of proteins. The high expression level of RPS27a was reported in solid tumors, and we found that the expression level of RPS27a was up-regulated in advanced-phase chronic myeloid leukemia (CML) and acute leukemia (AL) patients. In this study, we explored the function of RPS27a in leukemia cells by using CML cell line K562 cells and its imatinib resistant cell line K562/G01 cells. It was observed that the expression level of RPS27a was high in K562 cells and even higher in K562/G01 cells. Further analysis revealed that RPS27a knockdown by shRNA in both K562 and K562G01 cells inhibited the cell viability, induced cell cycle arrest at S and G2/M phases and increased cell apoptosis induced by imatinib. Combination of shRNA with imatinib treatment could lead to more cleaved PARP and cleaved caspase-3 expression in RPS27a knockdown cells. Further, it was found that phospho-ERK(p-ERK) and BCL-2 were down-regulated and P21 up-regulated in RPS27a knockdown cells. In conclusion, RPS27a promotes proliferation, regulates cell cycle progression and inhibits apoptosis of leukemia cells. It appears that drugs targeting RPS27a combining with tyrosine kinase inhibitor (TKI) might represent a novel therapy strategy in TKI resistant CML patients.     

Ivermectin inhibits the growth of glioma cells by inducing cell cycle arrest and apoptosis in vitro and in vivo

Glioma, the most predominant primary malignant brain tumor, remains uncured due to the absence of effective treatments. Hence, it is imperative to develop successful therapeutic agents. This study aimed to explore the antitumor effects and mechanisms of ivermectin (IVM) in glioma cells in vitro and in vivo. The effects of IVM on cell viability, cell cycle arrest, apoptosis rate, and morphological characteristics were determined respectively by MTT assay/colony formation assay, flow cytometry, and transmission electron microscope. In addition, the expression levels of cycle-related and apoptosis-associated proteins were individually examined by Western blot analysis. Moreover, cell proliferation and apoptosis analyses were carried out by TUNEL, Ki-67, cleaved caspase-3, and cleaved caspase-9 immunostaining assay. Our results demonstrated that IVM has a potential dosage-dependent inhibition effect on the apoptosis rate of glioma cells. Meanwhile, the results also revealed that IVM induced apoptosis by increasing caspase-3 and caspase-9 activity, upregulating the expressions of p53 and Bax, downregulating Bcl-2, activating cleaved caspase-3 and cleaved caspase-9, and blocking cell cycle in G0/G1 phase by downregulating levels of CDK2, CDK4, CDK6, cyclin D1, and cyclin E. These findings suggest that IVM has an inhibition effect on the proliferation of glioma cells by triggering cell cycle arrest and inducing cell apoptosis in vitro and in vivo, and probably represents promising agent for treating glioma.     

Synthesis and characterization of new copper thiosemicarbazone complexes with an ONNS quadridentate system: cell growth inhibition, S-phase cell cycle arrest and proapoptotic activities on cisplatin-resistant neuroblastoma cells

Two new copper thiosemicarbazone complexes with an ONNS quadridentate system were synthesized and evaluated for anticancer activity on cisplatin-resistant neuroblastoma cells. Among these two copper complexes, the substituted 8-hydroxyquinoline-2-carboxaldehyde–4,4-dimethyl-3-thiosemicarbazide (CuHQDMTS) exhibited stronger cell growth inhibition activity than the unsubstituted copper 8-hydroxyquinoline-2-carboxaldehyde thiosemicarbazide complex (CuHQTS). Both CuHQTS and CuHQDMTS showed dose-dependent cell growth inhibition, cell cycle arrest and apoptosis induction activities on the SK-N-DZ neuroblastoma cells. Increased expression of p53 protein molecules was detected in the SK-N-DZ cells treated with CuHQTS. The data obtained in this study suggest that CuHQDMTS and CuHQTS hold potential as new, effective drugs for treatment of refractory neuroblastoma in children.     

Lipopolysaccharide inhibits GPR120 expression in macrophages via Toll‐like receptor 4 and p38 MAPK activation

Free fatty acid receptor G protein‐coupled receptor 120 (GPR120) is highly expressed in macrophages and was reported to inhibit lipopolysaccharide (LPS)‐stimulated cytokine expression. Under inflammation, macrophages exhibit striking functional changes, but changes in GPR120 expression and signaling are not known. In this study, the effects of LPS treatment on macrophage GPR120 expression and activation were investigated. The results showed that LPS inhibited GPR120 expression in mouse macrophage cell line Ana‐1 cells. Moreover, LPS treatment inhibited GPR120 expression in mouse alveolar macrophages both in vitro and in vivo. The inhibitory effect of LPS on GPR120 expression was blocked by Toll‐like receptor 4 (TLR4) inhibitor TAK242 and p38 mitogen‐activated protein kinase inhibitor LY222820, but not by ERK1/2 inhibitor U0126 and c‐Jun N‐terminal kinase inhibitor SP600125. LPS‐induced inhibition of GPR120 expression was not attenuated by GPR120 agonists TUG891 and GW9508. TUG891 inhibited the phagocytosis of alveolar macrophages, and LPS treatment counteracted the effects of TUG891 on phagocytosis. These results indicate that pretreatment with LPS inhibits GPR120 expression and activation in macrophages. It is suggested that LPS‐induced inhibition of GPR120 expression is a reaction enhancing the LPS‐induced pro‐inflammatory response of macrophages.     

Dihydrofolate reductase inhibition effect of 5-substituted pyrido[2,3-d]pyrimidines: Synthesis,antitumor activity and molecular modeling study

A new series of pyrido[2,3-d]pyrimidines 3–18 bearing substitution at C-5 position was synthesized. All compounds were tested for their in vitro antitumor activity against five human cancer cell lines namely; hepatocellular carcinoma (HePG2), breast carcinoma (MCF-7), human prostate carcinoma (PC3), colorectal carcinoma (HCT-116), and cervical carcinoma (Hela) using doxorubicin as a positive control. Compounds 3, 4, 9, 11, 13, 14, 15 and 17 exhibited the highest antitumor activity against the tested cell lines and were selected to screen their enzymatic inhibition against dihydrofolate reductase enzyme (DHFR) compared with the reference drug methotrexate (MTX), to explain the probable mechanism of action of the observed anticancer activity. Compound 11 displayed the highest inhibitory activity (IC₅₀ = 6.5 µM) among the tested compounds in comparison with MTX (IC₅₀ = 5.57 µM). Also, compounds 13 and 14 showed high inhibitory activity against DHFR with IC₅₀ values 7.1 and 8.7 µM, respectively. Comparative molecular modeling study was performed between DHFR inhibitors 11, 13 and 14 of the highest activity and 10 of the lowest activity among the eight inhibitors against MTX. Docking studies into the active site of DHFR domain showed good agreement with the obtained biological results. Finally, compound 11 was found to be best antitumor, DHFR inhibitor, and it induced the process of apoptosis at Pre-G phase and cell cycle arrest at G2/M phase in MCF-7 cells.     

Vitamin E succinate inhibits human prostate cancer cell growth via modulating cell cycle regulatory machinery

Several epidemiological studies have demonstrated that vitamin E is a chemopreventative agent for prostate cancer. alpha-Tocopheryl succinate (VES), a derivative of vitamin E, effectively modulates prostate cancer cell growth. However, little is known about the mechanisms regarding this action. Here we show that VES causes human prostate cancer cell LNCaP arrest at G1 phase. This effect is accomplished through VES significantly decreasing expression of the cell cycle regulatory proteins cyclin D1, D3, and E, cdk2 and 4, but not cdk6. Furthermore, VES reduces cdk4 kinase activity, Rb phosphorylation, and cyclin E mRNA expression. Recently there is increasing interest in the protective effect of the VES and selenium combination on prostate cancer. Here we show that VES and selenium work through different mechanisms to exert their inhibitory effects on prostate cancer cells. Taken together, our studies suggest that VES-mediated prostate cancer cell G1/S arrest is a consequence of the regulation of multiple molecules of the cell cycle regulatory machinery.     

HDAC4 inhibits cell-cycle progression and protects neurons from cell death

HDAC4 is a Class II histone deacetylase (HDAC) that is highly expressed in the brain, but whose functional significance in the brain is not known. We show that forced expression of HDAC4 in cerebellar granule neurons protects them against low potassium-induced apoptosis. HDAC4 also protects HT22 neuroblastoma cells from death induced by oxidative stress. HDAC4-mediated neuroprotection does not require its HDAC catalytic domain and cannot be inhibited by chemical inhibitors of HDACs. Neuroprotection by HDAC4 also does not require the Raf-MEK-ERK or the PI-3 kinase-Akt signaling pathways and occurs despite the activation of c-jun, an event that is generally believed to condemn neurons to die. The protective action of HDAC4 occurs in the nucleus and is mediated by a region that contains the nuclear localization signal. HDAC4 inhibits the activity of cyclin-dependent kinase-1 (CDK1) and the progression of proliferating HEK293T and HT22 cells through the cell cycle. Mice-lacking HDAC4 have elevated CDK1 activity and display cerebellar abnormalities including a progressive loss of Purkinje neurons postnatally in posterior lobes. Surviving Purkinje neurons in these lobes have duplicated soma. Furthermore, large numbers of cells within these affected lobes incorporate BrdU, indicating cell-cycle progression. These abnormalities along with the ability of HDAC4 to inhibit CDK1 and cell-cycle progression in cultured cells suggest that neuroprotection by HDAC4 is mediated by preventing abortive cell-cycle progression.     

Methanolic extract of Citrullus colocynthis suppresses growth and proliferation of breast cancer cells through regulation of cell cycle

Breast cancer is a major cause of cancer related deaths in women worldwide. Available treatments pose serious limitations such as systemic toxicity, metastasis, tumor recurrence, off-target effects, and drug resistance. In recent years, phytochemicals such as secondary metabolites due to their effective anticancer potential at very low concentration have gained attention. Aim of the study was to evaluate anticancer potential of Citrullus colocynthis and its possible molecular targets on MCF-7, a human breast cancer cell line. Methanolic extract of leaves was prepared and fractionated by solvents (n-hexane, chloroform, ethyl acetate and n-butanol) with increasing polarity. Bioassays and gene expression regulation was conducted to evaluate the anticancer activity, proliferation rate and cell cycle regulation of breast cancer cells treated with extract and its fractions, separately. Results showed a significant anticancer activity of methanolic extract of C. colocynthis and two of its fractions prepared with chloroform and ethyl acetate. Bioassays depicted significant decrease in proliferation and growth potential along with cell cycle arrest of treated cells compared to control untreated cells. Expression regulation of genes further confirmed the cell cycle arrest through significant upregulation of cyclin-CDK inhibitors (p21 and p27) and cell cycle checkpoint regulators (HUS1, RAD1, ATM) followed by downregulation of downstream cell cycle progression genes (Cyclin A, Cyclin E, CDK2). It is concluded that C. colocynthis arrests cell cycle in human breast cancer cells through expression regulation of cyclin-CDK inhibitors and with further research can be proposed for therapeutic interventions.