Influence of A Continuous Very Low Dose of Gamma-Rays on Cell Proliferation, Apoptosis and Oxidative Stress.

We have previously shown a delay of death by lymphoma in SJL/J mice irradiated with continuous very low doses of ionizing radiation. In order to understand the mechanisms involved in this phenomenon, we have irradiated in vitro the Raw264.7 monocytic and the YAC-1 lymphoma cell lines at very low-dose rate of 4cGy.month(-1). We have observed a transient increase in production of both free radicals and nitric oxide with a transient adaptive response during at least two weeks after the beginning of the irradiation. The slight decrease of Ki67 proliferation index observed during the second and third weeks of YAC-1 cells culture under irradiation was not significant but consistent with the shift of the proliferation assay curves of YAC-1cells at these same durations of culture. These in vitro results were in good agreement with the slightly decrease under irradiation of Ki67 proliferative index evaluated on lymphomatous lymph nodes of SJL/J mice. A significant decrease of YAC-1 cells apoptotic rate under radiation appeared after 4 weeks of culture. Therefore very small doses of gamma-irradiation are able to modify the cellular response. The main observations did not last with increasing time under irradiation, suggesting a transient adaptation of cells or organisms to this level of irradiation.

Cell cycle and apoptotic effects of SAHA are regulated by the cellular microenvironment in HCT116 multicellular tumour spheroids.

Using multicellular tumour spheroids (MCTS) of HCT116 colon carcinoma cells, we analysed the effects of SAHA (suberoylanilide hydroxamic acid), a histone deacetylase inhibitor (HDACi). We found that, although SAHA-induced histone acetylation and ROS level upregulation occur throughout the spheroid, inhibition of cell cycle progression and induction of apoptosis are dependent on cell microenvironment. SAHA-induced growth inhibition of HCT116 MCTS results from the inhibition of cell cycle progression and induction of apoptosis. At a low concentration SAHA decreases Ki-67 and cyclin A positive cells and increases p21 positive cells in the outer layer while it induces a ROS-dependent apoptosis in the central zone of the spheroid. Coimmunostaining of p21 and apoptotic cells confirms that SAHA effects are different depending on the position of the cells within the spheroid. At a higher dose, SAHA induces mitotic defects and survivin downregulation in the outer layer of cells resulting in an additional cytotoxic effect in this part of the spheroid. Together these findings show that SAHA-induced cytostatic and cytotoxic effects occur in different cell populations, indicating that the cellular microenvironment is an important determinant in the regulation of the effects of SAHA treatment. Consequently, the MCTS model appears to be a valuable advanced tool for evaluating the effects of SAHA treatment in combination with other anticancer agents.

Human fetal chromaffin cells: a potential tool for cell pain therapy.

Transplantation of adrenal medulla cells has been proposed in the treatment of various conditions. Indeed, these cells possess a bipotentiality: neural and neuroendocrine, which could be exploited for brain repair or pain therapy. In a previous study, we characterized these human cells in vitro over 7-10 gestational weeks (GW) [Zhou, H., Aziza, J., Sol, J.C., Courtade-Saidi, M., Chatelin, S., Evra, C., Parant, O., Lazorthes, Y., and Jozan, S., 2006. Cell therapy of pain: Characterization of human fetal chromaffin cells at early adrenal medulla development. Exp. Neurol. 198, 370-381]. We report here our results on the extension to 23 GW. This developmental period can be split into three stages. During the first stage (7-10 GW), we observed in situ that extra-adrenal surrounding cells display the same morphology and phenotype as the intra-adrenal chromaffin cells. We also found that the intra-adrenal chromaffin cells could be committed in vitro towards an adrenergic phenotype using differentiating agents. During the second stage (11 to 15-16 GW), two types of cells (Type 1 and Type 2 cells) were identified morphologically both inside and outside the gland. Interestingly, we noted that the Type 2 cells stem from the Type 1 cells. However, during this developmental period only the intra-adrenal Type 2 cells will evolve towards an adrenergic phenotype. In the third stage (17-23 GW), we observed the ultimate location of the medulla gland. Both the in situ results and the in vitro experiments indicate that particular procedures need to be implemented prior transplantation of chromaffin cells. First, in order to obtain a large number of immature chromaffin cells, they must be isolated from the intra and extra-adrenal gland and should then be committed towards an adrenergic phenotype in vitro for subsequent use in pain therapy. This strategy is under investigation in our laboratory.

Histiocytic sarcoma in C57BL/6J female mice is associated with liver hematopoiesis: review of 41 cases.

Forty-one cases of histiocytic sarcoma (HS) in C57BL/6J mice were histopathologically studied with special regard to unexpected associated hematopoietic disorders. These cases were retrieved among C57BL/6J female mice used as control mice in a chronic low-dose irradiation experiment. Hematopoietic characteristics were analysed by comparison to 41 disease-free mice from the same cohort. Tumoral involvement of the liver was observed in all 41 HS-bearing mice, followed by infiltration of the spleen (61.8%), lung (32.4%), bone marrow (14.3%), uterus (12.2%), lymph node (9.8%), and kidney (2.4%). By comparative analysis, we were able to demonstrate a significant association of HS with liver hematopoiesis (89.5% in HS group vs 15% in control mice, p < 0.00001), and with central hematopoietic disorders involving the myelocytic cells (decreased in HS, p = 0.003) and erythrocytic cells (increased in HS, p = 0.001). Microscopic characteristics of these 41 cases and physiopathology of the newly described hematopoietic features in HS are further discussed.

All trans retinoic acid enhances CDDP-induced apoptosis: modulation of the CDDP effect on cell cycle progression.

We have previously shown that ATRA potentiates CDDP cytotoxicity in various ovarian carcinoma cell lines. In the present study, we found that the enhanced sensitivity to CDDP was due to an increase of CDDP-induced apoptosis in OVCCR1 and NIH-OVCAR-3 cells. In these cell lines, flow cytometric analysis indicated that CDDP induced an initial accumulation of cells in the S-phase, followed by an increase in the proportion of cells in G2/M phase. Pretreatment of OVCCR1 and NIH-OVCAR-3 cells with ATRA did not modify cell cycle parameters, but delayed S-phase exit of CDDP-treated cells. Bcl-2 over-expression inhibited both delay in S-phase exit and CDDP-induced apoptosis in ATRA-pretreated cells. The CDDP-induced S-phase accumulation of OVCCR1 cells resulted from an activation of CDK2/cyclin A activity. Our results indicate that ATRA-pretreatment modified the CDDP-induced regulation of CDK2 activity by the CDK inhibitors p21 and p27. Taken together, our findings suggest that ATRA potentiates the apoptosis induced by CDDP in ovarian carcinoma cells and that this action is sustained by modulation of the activity of CDK2/cyclin A.