Down-regulation of dihydrofolate reductase inhibits the growth of endothelial EA.hy926 cell through induction of G1 cell cycle arrest via up-regulating p53 and p21(waf1/cip1) expression.

Folic acid supplementation may meliorate cardiovascular disease risk by improving vascular endothelial structure and function. However, the underlying mechanisms are still lack of a global understanding. To be used, folic acid must be converted to 7,8-dihydrofolate by dihydrofolate reductase to generate one-carbon derivatives serving as important cellular cofactors in the synthesis of nucleotides and amino acids required for cell growth. Therefore, this study explored the effect of dihydrofolate reductase knockdown on endothelial EA.hy926 cell growth and the mechanism involved. We found that down-regulation of dihydrofolate reductase inhibited EA.hy926 cell proliferation, and induced G1 phase arrest. Meanwhile, the expression of regulators necessary for G1/S phase transition, such as cyclin-dependent kinases CDK2, CDK4 and CDK6, were remarkably down-regulated; by contrast, the cell cycle inhibitors p21(waf/cip1), p27(Kip1) and p53 were significantly up-regulated after dihydrofolate reductase knockdown. Furthermore, supplementation of 5-methyltetrahydrofolate to the dihydrofolate reductase knockdown cells could weaken the inhibitory effect of dihydrofolate reductase knockdown on cell proliferation, simultaneously, inducing the expression of p53 and p21(waf/cip1) falling back moderately. Our findings suggest that attenuating dihydrofolate reductase may cause imbalanced expression of cell cycle regulators, especially up-regulation of p53-p21(waf/cip1) pathway, leading to G1 cell cycle arrest, thereby inhibiting the growth of endothelial EA.hy926 cells.

hERG1/Kv11.1 activation stimulates transcription of p21waf/cip in breast cancer cells via a calcineurin-dependent mechanism.

The function of Kv11.1 is emerging in breast cancer biology, as a growing body of evidence indicates that the hERG1/Kv11.1 potassium channel is aberrantly expressed in several cancer types including breast cancers.The biological effects of Kv11.1 channel blockers and their associated side effects are very well known but the potential use of Kv11.1 activators as an anticancer strategy are still unexplored. In our previous work, we have established that stimulation of the Kv11.1 potassium channel activates a senescent-like program that is characterized by a significant increase in tumor suppressor protein levels, such as p21waf/cip and p16INK4A. In this study we investigated the mechanism linking Kv11.1 stimulation to augmentation of p21waf/cip protein level. We have demonstrated that the Kv11.1 channel activator NS1643 activates a calcineurin-dependent transcription of p21waf/cip and that this event is fundamental for the inhibitory effect of NS1643 on cell proliferation. Our results reveal a novel mechanism by which stimulation of Kv11.1 channel leads to transcription of a potent tumor suppressor and suggest a potential therapeutic use for Kv11.1 channel activators.

Resveratrol and curcumin synergistically induces apoptosis in cigarette smoke condensate transformed breast epithelial cells through a p21(Waf1/Cip1) mediated inhibition of Hh-Gli signaling.

Combination therapy using two or more small molecule inhibitors of aberrant signaling cascade in aggressive breast cancers is a promising therapeutic strategy over traditional monotherapeutic approaches. Here, we have studied the synergistic mechanism of resveratrol and curcumin induced apoptosis using in vitro (cigarette smoke condensate mediated transformed breast epithelial cell, MCF-10A-Tr) and in vivo (tumor xenograft mice) model system. Resveratrol exposure increased the intracellular uptake of curcumin in a dose dependent manner and caused apoptosis in MCF-10A-Tr cells. Approximately, ten fold lower IC50 value was noted in cells treated with the combination of resveratrol (3µM) and curcumin (3µM) in comparison to 30µM of resveratrol or curcumin alone. Resveratrol+curcumin combination caused apoptosis by increasing Bax/Bcl-xL ratio, Cytochrome C release, cleaved product of PARP and caspase 3 in cells. Interestingly, this combination unaltered the protein expressions of WNT-TCF and Notch signaling components, ß-catenin and cleaved notch-1 val1744, respectively. Furthermore, the combination also significantly decreased the intermediates of Hedgehog-Gli cascade including SMO, SHH, Gli-1, c-MYC, Cyclin-D1, etc. and increased the level of p21(Waf/Cip1) in vitro and in vivo. A significant reduction of Gli- promoter activity was noted in combinational drug treated cells in comparison to individual drug treatment. Un-alteration of the expressions of the above proteins and Gli1 promoter activity in p21(Waf/Cip1) knockout cells suggests this combination caused apoptosis through p21(Waf/Cip1). Thus, our findings revealed resveratrol and curcumin synergistically caused apoptosis in cigarette smoke induced breast cancer cells through p2(Waf/Cip1) mediated inhibition of Hedgehog-Gli cascade.

Effect of Guizhi Fuling Bolus on expression of Survivin、P21~(waf/cip) in tumor tissue of tumor-bearing mice / 中华中医药杂志

Objective:To Study the molecular mechanism of Guizhi Fuling Bolus(GFW) controlling the tumor growth.Methods:Each of 30 mice received 0.2ml S180 tumor cell suspension subcutaneously in the right axilla.The mice were randomly divided into model group,GFW group and Cyclophosphane(CY) group and with 10 mice in each group.Another ten mice without cancer were used as control.The model group received intragastric normal saline,0.2ml?10g-1?d-1;the GFW group was given intragastric administration,0.2ml?10g-1?d-1 whilst the CY group received intraperitoneal injection of CY at 20mg?kg-1?d-1.The medicines were given to the mice in each group after they were inoculated,once a day for 10 days.The mice were sacrificed and the tumor removed,the S180 mice sarcoma model was used to detect the inhibiting effects of GFW.The protein expressions of P21waf/cip and mRNA expression of Survivin were evaluated by immunohistochemistry and in situ hybridization.Results:For S180 sarcoma,the inhibiting ratio of GFW was 38.93%.Survivin mRNA was markedly declined in the GFW group when compared with the model group(P

The molecular mechanisms for mouse's neoplasm apoptosis triggered by lotus GFW / 中国免疫学杂志

Objective:To study on GFW-inducing tumor apoptosis and to find the experimenting basis for utilizing GFW in cancei theiapy.Methods:S180 mouse sarooma model was used to detect inhibiting effects of GFW on tumor grouth in viro,Morphological changes were observed by electron microscopy and flow cytometer was used for determination apoptosis.The protein expression of p21 and the mRNA expression of Survivin were evaluated by immunohistochemistry and in situ hybridization.Results:For S180 sarcoma ,the inhibiting rate of GFW was 38.93%.Apoptosis was 17.79% detected by flow cytomety.Typical apoptotic cells and apoptotic bodies were observed through electron microscope. Chromatin was found gathering as aggregates in nucleus or under nucleus membrane and the nuclear body formed at the same time. Survivin mRNA decreased markedly in the GFW group when compared with the model group(P

Guizhi Fuling Wan's effect on the expression of P21~(waf/cip) in bearing tumor mice / 西安交通大学学报(医学版)

Objective To study the effect of Guizhi Fuling Wan(GFW) on tumor apoptosis and apply the experimenting basis of GFW's development and utilization.Methods The S180 mice sarcoma model was used to detect the inhibitory effects of GFW,observe the ultrastructural change by electron microscopy,and the flow cytometer was used for apoptosis determination.The protein expression of P21waf/cip and the mRNA expression of Survivin were evaluated by immunohistochemistry and in situ hybridization.Results For S180 sarcoma,the inhibition ratio of GFW was 38.93%.The apoptosis was 17.79% by the flow cytometer.Some typical apoptotic cells and apoptotic bodies were observed through electron microscope.Chromatin gathered at the side of cell,the nucleus turned into the nucleus band and nucleus projected.Survivin mRNA markedly declined in the GFW group when compared with that in the model group(P