Apoptotic effect and mechanisms of AHPN on human skin malignant melanoma cell A375

Objective： To study apoptotic effects of synthetic retinoic acid 6-[3-（1-adamantyl）-4-hydroxyphenyl]-2-naphthalene carboxylic acid（AHPN） on human skin malignant melanoma A375 cells in comparison with the natural ligand all-trans-retinoic acid（ATRA） in vitro and the mechanisms related to the actions of AHPN. Methods：MTT assay was used to determine the anti-proliferative effects of AHPN and ATRA on A375 cells. Flow cytometry was performed to investigate the influence of AHPN and ATRA on cell cycle and cell apoptosis. In addition, transfection and luciferase activity assays were employed to explore the mechanisms of how AHPN executes its proapoptotic function. Results：Firstly, AHPN promoted apoptosis and GI arrest in A375 cells compared with ATRA. Secondly, the activity of NF-K B in A375 cells treated with AHPN increased 2-3 times compared with solvent DMSO treatment. Conclusion：AHPN, in comparison with ATRA, is a more effective alternative for therapy of malignant melanoma. The potentially proapoptotic function of AHPN requires activation of NF-K B.

Comparison between synthetic retinoid CD437 and acitretin inhibiting melanoma A375 cell in vitro

Objective： To investigate the effects of synthetic retinoid CD437 and acitretin on cell proliferation, apoptosis, cycle arrest and Bax/ Bcl-2 protein expression of melanoma A375 cell, Methods：MTT assay was used to determine the anti-proliferative effects of CD437 and acitretin on melanoma A375 cell, Flow cytometry was performed to investigate the influence of CD437 and acitretin on cell cycle and cell apoptosis. SABC immunocytochemistry was employed for detection of Bax/bcl-2 protein expressions. Results：10^-5 mol/L CD437 was more effective than acitretin in inhibiting proliferation and inducing apoptosis of A375 cell after 24 h treatment, growth inhibiting ratio and apoptosis ratio（58.6%vs43.25% and 28.03%vs17.13%, P 〈 0.05 respectively）. CD437 promoted G0/G1 arrest in melanoma A375 cell, however acitretin could not. CD437 and acitretin could up-regulate the expression of Bax protein and downregulate the expression of bcl-2 protein（P 〈 0.05）. Conclusion：CD437 is more effective than acitretin in inhibiting proliferation and inducing apoptosis and cycle arrest on A375 cell, CD437 may have more potentialities than acitretin for subsidiary treatment of melanoma. Mitochondrial apoptosis pathway is partially involved in two drugs inducing apoptosis on A375 cell.

Effects of histamine on growth and apoptosis of human melanoma cells A375

Objective: To investigate the effects of histamine on growth and apoptosis of human melanoma cells A375. Methods: The effect of histamine on growth of A375 cells in vitro was examined by MTT assay and Trypan blue exclusion assay. Cell cycle analysis, early apoptosis analysis by double staining with Annexin V-FITC and PI, and active caspase-3 analysis by staining FITC-conjugated monoclonal rabbit anti-active caspase-3 antibody were made by flow cytometer. StreptAvidin-Biotin Complex (SABC) immunocytochemical assays were adopted to detect Bax/Bcl-2 protein expressions.Results: Histamine inhibited proliferation of A375 cells in a dose- and time-dependent manner, and altered cell cycle distribution of A375 cells revealing an increase in G0/G1-phase population, a decrease in S-phase population and the inhibition of G1/S switching. Histamine induced apoptosis of A375 cells (P<0.05), elevated the cells population with detectable active caspase-3 (P<0.05), increased the number of cells forming Bax and decreased the number of cells forming Bcl-2 significantly (P<0.05). Conclusion: That histamine inhibits cell cycle progress of A375 cells is one of the possible mechanisms of proliferation arrest of A375 cells elicited by histamine. Histamine mediates apoptosis in A375 cells that may be caspase-dependent through mitochondria routine. Histamine with high concentration inhibits growth of A375 cells in vitro by interfering proliferation and inducing apoptosis of cells.

Mitogen-activated protein kinase-dependent apoptosis in norcan-tharidin-treated A375-S2 cells is proceeded by the activation of protein kinase C

We have reported that norcantharidin (NCTD) induces human melanoma A375-S2cell apoptosis and that the activation of caspase and the mitochondrial pathway are involved in theapoptotic process. This study aimed at investigating the roles of mitogen-activated protein kinase(MAPK) and protein kinase C (PKC) in A375-S2 cell apoptosis induced by NCTD. We assessed theeffects of NCTD on cell growth inhibition using the 3-(4,5-dimethylthiazol-2-yl)-2 ,5-dipheyltetrazolium bromide ( MTT) assay, DNA fragmentation ( DNA agarose gel electrophoresis ) ,and MAPK protein levels (Western blot analysis) in A375-S2 cells. Photomicroscopic data were alsocollected. The NCTD inhibitory effect on A375-S2 cells was partially reversed by MAPK and PKCinhibitors. The expression of phosphorylated JNK and p38 also increased after the treatment withNCTD, and inhibitors of c-Jun NH2 - terminal kinase (JNK) and p38 ( SP600125 and SB203580,respectively) had significant inhibitory effects on the upregulation of phosphorylated JNK and p38expression. Simultaneously, the PKC inhibitor staurosporine blocked the upregulation ofphosphorylated JNK and phosphorylated p_(38), but had little effect on extracellularsignal-regulated kinase (ERK) expression. These results suggest that the activation of JNK andp_(38) MAPK promotes the process of NCTD-induced A375-S2 cell apoptosis and that PKC plays animportant regulation role in the activation of MAPKs.

Celecoxib in combination with retinoid CD437 inhibits melanoma A375 cell in vitro

This study aimed to investigate the effects of celecoxib,synthetic retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalenecarboxylicacid(CD437)and the combination of the two on cell proliferation,apoptosis,and cycle arrest of human malignant mela-noma A375 cells.3-(4,5-dimethylthiazol-2-yl)-2,5-di-phenyltetrazoliumbromide assay(MTT assay)was applied to determine the anti-proliferative effects of the drugs on human malignant melanoma A375 cells.Flow cytometry was performed to investigate the influence of the drugs on cell cycle and cell apoptosis.Both celecoxib and CD437 could inhibit the growth of human malignant melanoma A375 cells in a dose-dependent manner.Celecoxib at 80μmol/L inhibited proliferation,induced apoptosis and G2/M cell cycle arrest of human malignant melanoma A375 cells after treatment for 24 h[proliferation inhibiting rate:(50.2±2.51)%,apoptosis rate:(35.91±1.80)%].CD437 at 10μmol/L inhibited proliferation,induced apoptosis and G0/G1 cell cycle arrest of human malignant melanoma A375 cells after treatment for 24 h[proliferation inhibiting rate:(58.6±2.38)%,apoptosis rate:(28.03±0.77)%].Celecoxib in combination with CD437 could significantly enhance the effects of inhibiting proliferation and inducing apoptosis of human malignant melanoma A375 cells 24 h after treatment compared with the drug alone[proliferation inhibiting rate:(68.92±1.72)%,apop-tosis rate:(42.09±1.05)%,both P<0.05]and decrease the proportion of the S phase in the cell cycle.Celecoxib could inhibit the growth of human malignant melanoma A375 cells by inducing apoptosis and G2/M cycle arrest.CD437 could inhibit the growth of human malignant melanoma A375 cells by inducing apoptosis and G0/G1 cycle arrest.Celecoxib exhibited additive effects with CD437 on retarding the growth and inducing apoptosis of human malignant melanoma A375 cells.Celecoxib in combination with CD437 may become an effective method for prevention and treatment of human melanoma.