Jean-Louis Beneytout

Induction of antiproliferative effect by diosgenin through activation of p53, release of apoptosis-inducing factor (AIF) and modulation of caspase-3 activity in different human cancer cells

ABSTRACTPreviously, we demonstrated that a plant steroid, diosgenin, altered cell cycle distribution and induced apoptosis in the human osteosarcoma 1547 cell line. The objective of this study was to investigate if the antiproliferative effect of diosgenin was similar for different human cancer cell lines such as laryngocarcinoma HEp-2 and melanoma M4Beu cells. Moreover, this work essentially focused on the mitochondrial pathway. We found that diosgenin had an important and similar antiproliferative effect on different types of cancer cells. In addition, our new results show that diosgenin-induced apoptosis is caspase-3 dependent with a fall of mitochondrial membrane potential, nuclear localization of AIF and poly (ADP-ribose) polymerase cleavage. Diosgenin treatment also induces p53 activation and cell cycle arrest in the different cell lines studied.

Citral inhibits cell proliferation and induces apoptosis and cell cycle arrest in MCF‐7 cells

Many natural components of plants extract are studied for their beneficial effects on health and particularly on carcinogenesis chemoprevention. In this study, we investigated the effect of citral (3,7‐dimethyl‐2,6‐octadienal), a key component of essential oils extracted from several herbal plants, on the proliferation rate, cell cycle distribution, and apoptosis of the human breast cancer cell line MCF‐7. The effects of this compound were also tested on cyclo‐oxygenase activity. Citral treatment caused inhibition of MCF‐7 cell growth (IC50‐48 h: 18 × 10−5 m), with a cycle arrest in G2/M phase and apoptosis induction. Moreover, we observed a decrease in prostaglandin E2 synthesis 48 h after citral treatment. These findings suggest that citral has a potential chemopreventive effect.

Aromatase in synovial cells from postmenopausal women.

Peripheral aromatization of androgens exerts estrogenic actions in many tissues. In this study, osteoarthritis synoviocytes were examined to clarify the possible action of adrenal androgen on synovial cell. Synoviocytes from postmenopausal women are able to express aromatase mRNA. By sequence analysis, the PCR fragment (485 bp) was determined to be 100% identical to that of human placental aromatase cDNA. Moreover, this study demonstrates that adrenal androgen, androstenedione, is converted to estrone (E(1)) and estradiol (E(2)) in synoviocytes by aromatase which is positively regulated by glucocorticoids such as dexamethasone. E(2) production reduced significantly IL-6 secretion. These data provide preliminary evidence that in situ estrogen production by synoviocytes may have a role in OA susceptibility. However the role of E(2) in OA is not clear and remains to be determined.

The P2Y2/Src/p38/COX-2 pathway is involved in the resistance to ursolic acid-induced apoptosis in colorectal and prostate cancer cells

One of the hallmarks of cancer is resistance to apoptosis. Elucidating the mechanisms of how cancer cells evade or delay apoptosis should lead to novel therapeutic strategies. Previously, we showed that HT-29 colorectal cancer cells undergoing apoptosis overexpressed cyclooxygenase-2 (COX-2), in a p38 dependent pathway, to delay ursolic acid-induced apoptosis. Here, we focused on elucidating the upstream signaling pathways regulating this resistance mechanism. The role of ATP as an extracellular signaling molecule took a long time to be accepted. In recent years, ATP and its analogs, via the activation of specific purinergic receptors, have been implicated in many biological processes including cell proliferation, differentiation and apoptosis. In the present report, we have demonstrated a novel role involving purinergic receptors and particularly the P2Y(2) receptor in resistance to ursolic acid-induced apoptosis in both colorectal HT-29 and prostate DU145 cancer cells. We found that ursolic acid induced an increase in intracellular ATP and P2Y(2) transcript levels. Upon activation, P2Y(2) activated Src which in turn phosphorylated p38 leading to COX-2 overexpression which induced resistance to apoptosis in both HT-29 and DU145 cells. Furthermore, Ca(2+)-independent PLA(2) (iPLA(2)) and Ca(2+)-dependent secretory PLA(2) (sPLA(2)) were responsible for arachidonic acid release, the substrate of COX-2. Our findings document that apoptosis triggering was dependent on protein kinase C (PKC) activation in both cell lines after ursolic acid treatment.

HT-29 colorectal cancer cells undergoing apoptosis overexpress COX-2 to delay ursolic acid-induced cell death

Colorectal cancer is one of the most common cancer types and the third leading cause of cancer-related death in the western world. Generally, colorectal cancers are resistant to anticancer drugs. Several lines of evidence support a critical role for cyclooxygenase-2 (COX-2) during colorectal tumorigenesis and its role in chemoresistance. In this study, we focused our interest on the role played by COX-2 in apoptosis induced in HT-29 human colorectal cancer cells by ursolic acid (UA), a triterpenoid found in a large variety of plants. We showed that UA-induced apoptosis and that COX-2 was overexpressed only in apoptotic cells. We demonstrated that this overexpression was mediated by the p38 MAP kinase pathway as inhibiting its activation using a p38-specific inhibitor, SB 203580, abrogated COX-2 expression. Inhibiting COX-2 expression either by using a p38-specific inhibitor or COX-2-specific siRNA increased apoptosis. These results demonstrated that COX-2 was involved in a resistance mechanism to UA-induced apoptosis in HT-29 cells. Cells undergoing apoptosis were able to trigger a resistance mechanism by overexpressing a protein such as COX-2 to delay their death. Furthermore, we demonstrated that this resistance mechanism was independent of PGE(2) production as the addition of the specific COX-2 activity inhibitor, NS-398, did not affect apoptosis in UA-treated cells.

Increased cyclooxygenase-2 and thromboxane synthase expression is implicated in diosgenin-induced megakaryocytic differentiation in human erythroleukemia cells.

Differentiation induction as a therapeutic strategy has, so far, the greatest impact in hematopoietic malignancies, most notably leukemia. Diosgenin is a very interesting natural product because, depending on the specific dose used, its biological effect is very different in HEL (human erythroleukemia) cells. For example, at 10 microM, diosgenin induced megakaryocytic differentiation, in contrast to 40 microM diosgenin, which induced apoptosis in HEL cells previously demonstrated using sedimentation field-flow fractionation (SdFFF). The goal of this work focused on the correlation between cyclooxygenase-2 (COX-2) and thromboxane synthase (TxS) and megakaryocytic differentiation induced by diosgenin in HEL cells. Furthermore, the technique of SdFFF, having been validated in our models, was used in this new study as an analytical tool that provided us with more or less enriched differentiated cell fractions that could then be used for further analyses of enzyme protein expression and activity for the first time. In our study, we showed the implication of COX-2 and TxS in diosgenin-induced megakaryocytic differentiation in HEL cells. Furthermore, we showed that the analytical technique of SdFFF may be used as a tool to confirm our results as a function of the degree of cell differentiation.

Antiproliferative effect of extracts from Aristolochia baetica and Origanum compactum on human breast cancer cell line MCF-7.

Aristolochia baetica L. (Aristolochiaceae) and Origanum compactum Benth. (Lamiaceae) are native plants of Morocco used in traditional medicine. In order to systematically evaluate their potential activity on human breast cancer, four different polarity extracts from each plant were assessed in vitro for their antiproliferative effect on MCF-7 cells. As a result, several extracts of those plants showed potent cell proliferation inhibition on MCF-7 cells. Chloroform extract of A. baetica (IC50: 216.06 ± 15 μg/mL) and ethyl acetate of O. compactum (IC50: 279.51 ± 16 μg/mL) were the most active. Thin layer chromatography examination of the bioactive extracts of A. baetica and O. compactum showed the presence of aristolochic acid and betulinic acid, respectively. These results call for further studies of these extracts.

Cytotoxicity of arachidonic acid and of its lipoxygenase metabolite 15-hydroperoxyeicosatetraenoic acid on human breast cancer MCF-7 cells in culture

Monolayer cultures of MCF-7 human breast cancer cell line were treated with arachidonic acid (AA) and 15-L-(s)-hydroperoxyeicosatetraenoic acid (15-L-(s)-HPETE) in a concentration range of 10(-5)-10(-11) M and their relative cytotoxic potential was determined. Both compounds had a time and dose-dependent cytotoxic effect on MCF-7 cells. 15-L-(S)-HPETE, the 15-lipoxygenase product of AA was the most effective.

Evidence for an arachidonic acid 5-, 8- and 15-lipoxygenase in Lupinus albus seeds

Abstract A lipoxygenase preparation was obtained from dried Lupinus albus seeds and was shown to differ from previously characterized soybean lipoxygenases in the positional specificity and pH characteristics of the dioxygenation reaction. The L. albus enzyme converted arachidonic acid into three HPETEs: 15-HPETE was the major product (60%) besides 5-HPETE (34%), and there were small amounts of 8-HPETE (6%). The enzyme had a pH optimum at 6.0 and was inactive at pH 9.O. The molecular weight of the enzyme was estimated at 71000, and pI was 5.35. The enzyme activity was inhibited strongly in the presence of lipoxygenase inhibitors, such as esculetin or BW 755 C.

Low dose leflunomide activates PI3K/Akt signalling in erythroleukemia cells and reduces apoptosis induced by anticancer agents

Rheumatoid arthritis (RA) is characterized by persistent joint synovial tissue inflammation. Leflunomide is an immunomodulatory agent that has been approved for treatment of active RA. In the past few years, uses other than RA treatment have appeared. Leflunomide has been reported to show antitumor potential through inhibition of cancer cell proliferation. We thus tested the antiproliferative potential of leflunomide on HEL and K562 erythroleukemia cells. The findings summarized in this report demonstrate for the first time that low dose leflunomide prolonged survival and reduced apoptosis induced by several anticancer agents in erythroleukemia cells. We showed that in treated cells, leflunomide reduced the signalling pathways involved in promoting apoptosis by reducing p38 MAPK and JNK basal activity. On the other hand, leflunomide transiently activated the ERK signalling pathway and induced a sustained activation of Akt. We also showed that leflunomide reduced caspase-3 activity and DNA fragmentation induced by anticancer agents. By using an inhibitory strategy, we showed that inhibition of Akt activation but not ERK abolished the protective effect of leflunomide. Thus our findings suggested that leflunomide reduced apoptosis induced by anticancer agents through PI3K/Akt signalling activation.