Role of MAP kinases in nitric oxide induced muscle-derived adult stem cell apoptosis.

Apoptosis in heart failure has been intensively investigated in vitro and in vivo. Stem cells have therapeutic value in the direct treatment of diseases, including cardiovascular disease. The main drawback of stem cell therapy is their poor survival in the diseased tissues. Since intracellular mitogen-activated protein kinases (MAPKs) actively participate in the regulation of cell survival and of proapoptotic signals, the ability to manipulate the mechanisms of MAPKs activation in myogenic stem cells might increase the survival of transplanted stem cells. Our results clearly demonstrate sustained activation of all three MAPKs, ERK, JNK and p38 in myogenic stem cells after exposure to the NO inducer, NOC-18. Inhibition of MAPKs phosphorylation by specific inhibitors revealed the anti-apoptotic role of MAPKs in myogenic stem cells.

On the combination of photodynamic therapy with ionizing radiation.

Ehrlich ascites carcinoma growth and cell damage have been examined after photodynamic therapy (PDT), radiotherapy (RT) and combined treatment. Haematoporphyrin dimethyl ether (HPde) is used as a photosensitizer for PDT and tested as a radiosensitizer for RT. For PDT a non-coherent light source (370 < lambda < 680 nm) equipped with filters is used. gamma-Irradiation consists of 60Co irradiation at a dose of 2 Gy. Both PDT and RT induce a significant delay and inhibition in tumour growth (33 and 38%, respectively). Nevertheless cell damage after these treatments is different: after PDT the cell membrane integrity is damaged and no serious chromosomal aberrations are observed; whereas after gamma-irradiation there is no cell membrane integrity damage, but more significant DNA injuries are observed. It seems evident that HPde is able to act as a photosensitizer as well as a radiosensitizer. Combining PDT and RT produces an additive effect, not dependent on the sequence in which the two treatments are given, when a 1 h time window is used.

Prooxidant character of flavonoid cytotoxicity: structure-activity relationships.

The action of flavonoids on bovine leukemia virus-transformed lamb fibroblasts (line FLK) and HL-60 cells was accompanied by lipid peroxidation, their toxicity was partly prevented by iron chelator desferrioxamine and antioxidant N,N'-diphenyl-p-phenylene diamine. This pointed out to the involvement of oxidative stress in flavonoid cytotoxicity. The concentration of compound for 50% survival of FLK cells (cL50) did not show correlation with polarographic oxidation half-peak potential (Ep/2) and/or partition coefficient (log P) of flavonoids; however, their toxicity to HL-60 cells was described by equation log cL50 (microM) = 3.0161 + 1.1099 Ep/2 (V) - 0.3369 log P. The toxicity of quercetin was partly prevented by nontoxic concentrations of other flavonoids examined, thus pointing out to potential neutralization of quercetin cytotoxicity by intake of flavonoid mixtures.

Proto-oncogene expression in bovine peripheral blood leukemic lymphocytes during their spontaneous proliferation, differentiation and apoptosis in vitro.

The expression of various proto-oncogenes in primary culture of lymphocytes from peripheral blood of bovine with chronic lymphocytic leukemia (CLL) was studied. Cellular proto-oncogenes encode proteins that propagate growth, differentiation or apoptosis signals from cell membrane to nucleus. The proliferation and differentiation of normal eukaryotic cells are precisely controlled. Tumor cells usually are characterized both by the continuous growth signal and by the block of cell differentiation. We have previously reported that along with spontaneous proliferation, bovine CLL lymphocytes continuously differentiate and enter apoptosis in vitro. CLL cells with an autocrine growth mechanism and at the same time undergoing spontaneous differentiation and apoptosis in vitro provide a new model system to investigate the possible involvement of various proto-oncogenes in the regulation of cellular proliferation, differentiation and apoptosis. Northern blot analysis revealed simultaneous expression of a number of proto-oncogenes in CLL cells. Transcripts of c-fos, c-myc, c-myb, A-raf, c-raf1, hck, IL-2 receptor alpha-chain (IL-2R alpha) were found in lymphocytes at the peak of their proliferative activity in culture. Kinetics studies demonstrated that CLL cells constitutively express transcripts of so-called immediate response nuclear proto-oncogenes c-myc, c-fos as well as cytoplasmic proto-oncogenes hck and c-raf1, i.e., genes coding for tyrosine and serine-threonine protein kinases, respectively. Expression level did not change significantly during all stages of CLL cells in culture. The results show that continuous expression of c-myc mRNA does not prevent CLL cell differentiation and may be associated with apoptotic cell death.

Toxicity of daunorubicin and naphthoquinones to HL-60 cells: an involvement of oxidative stress.

Incubation of HL-60 cells with anthracycline daunorubicin caused an appearance of viable apoptotic, nonviable apoptotic, necrotic (nonviable nonapoptotic) and chromatin-free (late apoptotic) cells. Both necrotic and apoptotic cell responses were partly prevented by antioxidant N,N'-diphenyl-p-phenylene diamine (DPPD) and iron-chelating agent, desferrioxamine, suggesting an involvement of activated oxygen species. The comparison of cytotoxicity of daunorubicin and of 5-hydroxy- and 5,8-dihydroxy-1,4-naphthoquinones revealed that at equitoxic concentrations, hydroxynaphthoquinones induced a larger number of necrotic cells in comparison to daunorubicin, a process being partly prevented by DPPD and desferrioxamine. However, we have found that cytotoxicity of daunorubicin was markedly higher in comparison with a series of naphtho- and benzoquinones, where an increase of cytotoxicity upon increase in single-electron reduction potential of quinones (E(1)7) was observed, pointing out to redox cycling as to the main factor of cytotoxicity. This discrepancy could be explained by additional factor(s) of daunorubicin cytotoxicity, e.g., DNA-intercalation, or selective accumulation of daunorubicin in cell nucleus.

Phenomena of spontaneous proliferation, differentiation, and apoptosis in primary culture of leukemic lymphocytes.

The proliferative capacity of lymphocytes from peripheral blood of bovine with chronic lymphocytic leukemia (CLL) in vitro was investigated. We have shown earlier that CLL cells spontaneously proliferate in serum-free medium in the absence of added growth factors and mitogenic stimulation; autocrine growth factors provide the growth-initiating signal for CLL cells. The results of the present study showed that bovine serum albumin or fetal calf serum greatly enhanced the number of CLL cells incorporating [3H]thymidine. Although some CLL cells proceeded through more than one cell cycle, proliferation of CLL cells in culture was temporary. On the other hand, it was shown that CLL cells differentiated spontaneously in culture. This differentiation was characterized by the appearance of plasmacytoid cells possessing cytoplasmic immunoglobulins that coincided with the cessation of cell proliferation. Moreover, together with spontaneous proliferation and differentiation, the phenomenon of programmed cell death (apoptosis) was found, as was evidenced by the appearance of apoptotic bodies as well as DNA fragmentation. The findings indicate that the loss of proliferative potential of CLL cells in culture may be a consequence of their differentiation and/or apoptosis in vitro. CLL cells, with an autocrine growth mechanism, spontaneous differentiation, and apoptosis in vitro, provide a new model system for studies of the relationship between cellular proto-oncogene expression and inhibition of growth and/or induction of differentiation.