Programmed Cell Death Induced by (?)-8,9-Dehydroneopeltolide in Human Promyelocytic Leukemia HL-60 Cells under Energy Stress Conditions

(+)-Neopeltolide is a marine macrolide natural product that exhibits potent antiproliferative activity against several human cancer cell lines. Previous study has established that this natural product primarily targets the complex III of the mitochondrial electron transport chain. However, the biochemical mode-of-actions of neopeltolide have not been investigated in detail. Here we report that (−)-8,9-dehydroneopeltolide (8,9-DNP), a more accessible synthetic analogue, shows potent cytotoxicity against human promyelocytic leukemia HL-60 cells preferentially under energy stress conditions. Nuclear morphology analysis, as well as DNA ladder assay, indicated that 8,9-DNP induced significant nuclear condensation/fragmentation and DNA fragmentation, and these events could be suppressed by preincubating the cells with a pan-caspase inhibitor, N-benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (zVAD). Immunoblot analysis demonstrated the release of cytochrome c from the mitochondria and the cleavage of full-length caspase-3 and poly(ADP-ribose) polymerase (PARP). These results indicated that 8,9-DNP induced caspase-dependent apoptotic programmed cell death under energy stress conditions. It was also found that 8,9-DNP induced non-apoptotic cell death in the presence/absence of zVAD under energy stress conditions. Immunoblot analysis showed the intracytosolic release of apoptosis-inducing factor (AIF), although it did not further translocate to the nucleus. It appears most likely that, in the presence of zVAD, 8,9-DNP triggered necrotic cell death as a result of severe intracellular ATP depletion.     

Interannual variability of net ecosystem production for a broadleaf deciduous forest in Sapporo, northern Japan

To estimate net ecosystem production (NEP), ecosystem respiration (R _E), and gross primary production (GPP), and to elucidate the interannual variability of NEP in a cool temperate broadleaf deciduous forest in Sapporo, northern Japan, we measured net ecosystem exchange (NEE) using an eddy covariance technique with a closed-path infrared gas analyzer from 2000 to 2003. NEP, R _E, and GPP were derived from NEE, and data gaps were filled using empirical regression models with meteorological variables such as photosynthetic active radiation and soil temperature. In general, NEP was positive (CO₂ uptake) from May to September, either positive or negative in October, and negative (CO₂ release) from November to the following April. NEP rapidly increased during leaf expansion in May and reached its maximum in June or July. The four-year averages (±?standard deviation) of annual NEP, GPP, and R _E were 443?±?45, 1,374?±?39, and 931?±?11?g?C?m^?2?year^?1, respectively. The lower annual NEP and GPP in 2000 may have been caused by lower solar radiation in the foliated season. During the foliated season, monthly GPP varied from year to year more than monthly R _E. Variations in the amount of incoming solar radiation may have caused the interannual variations in the monthly GPP. Additionally, in May, the timing of leaf expansion had a large impact on GPP. Variations in GPP affected the interannual variation in NEP at our site. Thus, interannual variation in NEP was affected by the incoming solar radiation and the timing of leaf expansion.     

Endocrine status and development of porcine testicular tissues in host mice

Clarification of the endocrine status of host mice provides us with basic knowledge with which we can manipulate the growth and function of xenografted testicular tissues. We investigated the hormonal profiles of castrated mice grafted with porcine immature testicular tissues from 30 to 210 or more days after grafting (day 0=castration and grafting). The serum follicle stimulating hormone (FSH) concentrations of the host mice declined (P<0.05) from day 60 compared with those of the castrated, ungrafted mice. The serum inhibin and testosterone levels were higher (P<0.05) than those in the castrated, ungrafted mice from days 30 and 90 days, respectively. The inhibin levels further increased (P<0.05) from day 90, during which time the levels were higher (P<0.05) than those in the intact male mice. In the grafts, formation of lumens occurred in the seminiferous cords on day 90 and spermatozoa appeared in the lumens from day 120. However, spermatogenesis in the grafts did not reach the qualitatively normal levels observed in adult boars. The intensity of the immune reaction to inhibin alpha subunits in the Sertoli cells of the grafts decreased with differentiation of the seminiferous tubules. The present findings indicate that a feedback loop was established between the mouse hypothalamo-pituitary axis and the grafted porcine tissues from 60 days post-grafting. The results also indicate that the serum inhibin levels in the host mice remained high even after the appearance of lumens in the seminiferous tubules of the grafted tissues; this is strikingly different to the situation in normal male animals, in which the serum inhibin levels decline at around the time of tubular differentiation. The lack of efferent ducts in the tubules of the grafted tissues probably caused the accumulation of inhibin to be released into the lumens, resulting in high concentrations of circulating inhibin. These high levels of inhibin may directly affect spermatogenic activity and suppress FSH secretion.