Estradiol and body weight during temporally targeted food restriction: Central pathways and peripheral metabolic factors.

We used temporally-targeted food restriction (TTFR), in which ovariectomized rats had chow only for 2 h/day, to test the hypothesis that estradiol benzoate (EB) suppresses feeding and decreases body weight during brief (4 day) TTFR, as it does during ad libitum feeding. All rats lost weight during TTFR, but the loss was greater with EB treatment. However, OIL and EB-treated rats ate comparable amounts of chow during TTFR. We next investigated central nervous system pathways and peripheral hormonal and metabolic changes that accompany the effects of TTFR to determine the mechanism for this effect. Immunolabeling for fos in the nucleus of the solitary tract, the terminal site of vagal afferents from the gastrointestinal tract, was increased when rats on TTFR had access to chow for 1 h on the test day, indicating neuronal activation associated with consumption of the meal. However, fos immunolabeling was not affected by EB treatment, nor were numbers of the α subtype of estrogen receptors. TTFR had the expected effects on carbohydrate and lipid metabolites and metabolic hormones, with only slight differences in plasma glucose, triglycerides, and free fatty acids attributable to EB treatment. Interestingly, plasma corticosterone levels were greater in EB-treated rats on TTFR, and increased further after eating. Given that corticosterone affects metabolism, these findings suggest that elevated corticosterone may explain the persistence of EB-induced differences in body weight during TTFR despite the lack of effect on food intake.

Disruption of thymopoiesis in ST6Gal I-deficient mice.

Thymocyte development is accompanied by sequential changes in cell surface glycosylation. For example, medullary thymocytes have increased levels of alpha2,3-linked sialic acid and a loss of asialo core 1 O-glycans as compared to cortical thymocytes. Some of these changes have been linked to fine tuning of the T cell receptor avidity. We analyzed ST6Gal I transcript abundance and levels of alpha2,6-linked sialic acid across thymocyte subsets. We found that ST6Gal I transcript levels increased following T cell receptor beta-selection suggesting that this sialyltransferase may influence the development of early thymocyte populations. Indeed, low levels of alpha2,6-linked sialic acid were found in the earliest T lineage cells, and then increased in T cell receptor beta-selected cells. To determine whether ST6Gal I influences T cell development, we analyzed ST6Gal I-deficient mice for disruptions in thymocyte populations. We found reduced thymic cellularity in the ST6Gal I-deficient mice starting in the early thymocyte compartments.

Effects of hepatic zonal oxygen levels on hepatocyte stress responses.

BACKGROUND: Hepatocytes spend their lifetimes in a gradient of oxygen, hormones, and enzymes. We used a three-dimensional Matrigel model to determine whether hepatocytes cultured at perivenous (zone 3) oxygen levels differed in susceptibility to anoxia-induced cell injury compared with hepatocytes cultured at periportal (zone 1) oxygen levels. MATERIALS AND METHODS: Hepatocytes were harvested from Sprague Dawley rats and cultured at 9% oxygen (hepatic zone 1) or 5% oxygen (hepatic zone 3) and stressed at 0% oxygen. Microscopy, real-time reverse transcriptase-polymerase chain reaction, and enzyme-linked immunosorbent assay were used to assess cell viability, mitochondrial potential, acute phase responses, and membrane blebbing. RESULTS: Hepatocytes cultured in Matrigel with HepatoZyme medium at zone 1 and zone 3 oxygen conditions were viable for 1 wk and showed acute phase responses as measured by interleukin-6-induced fibrinogen production. In response to 3 h anoxia, cells maintained at the perivenous oxygen level showed increased membrane blebbing and increased loss of mitochondrial membrane potential in comparison to the periportal oxygen cultured cells. Cells at perivenous oxygen also showed a reduced ability to recover following reoxygenation. CONCLUSIONS: Hepatocytes can remain viable and functional for extended periods in culture at low oxygen levels that mimic the hepatic perivenous environment, yet these cells are more susceptible to anoxia-induced damage than hepatocytes cultured at the periportal oxygen level. The small population of perivenous hepatocytes may be critical in determining the fate of the liver during ischemia/reperfusion since hepatocytes cultured at that concentration appear to be more labile in response to anoxia.

Attenuation of cytokine responsiveness during T cell development and differentiation.

Cytokines play critical roles during T cell development; however, it is unclear to what extent development is altered by the high levels of cytokines produced during immune responses. A potential mechanism to shield developing cells from cytokine influence is attenuation of cytokine signaling. Using intracellular staining and flow cytometry to detect cytokine-induced Stat phosphorylation, we analyzed the cytokine responsiveness of developmentally defined mouse T cells. We assessed CD4(-)CD8(-) (DN), CD4(+)CD8(+) (DP), CD4(+)CD8(-) (SP4), and CD4(-)CD8(+) (SP8) in the thymus, and CD4(+)CD44(lo) (naive), CD4(+)CD44(hi) (memory), CD8(+)CD44(lo) (naive), and CD8(+)CD44(hi) (memory) in the periphery for responsiveness to interleukin-2 (IL-2), IL-4, IL-6, IL-7, IL- 10, IL-15, interferon-alpha (IFN-alpha), and IFN-gamma. SP thymocytes responded to a wider range of cytokines than did the less mature DN and DP subpopulations. DP thymocytes were nonresponsive to all cytokines tested except for modest responses to IL-4 and IFN-alpha. Peripheral naive and memory T cells also displayed differential cytokine sensitivity. Memory T cells were less responsive to the proinflammatory cytokines IL-6 and IFN-gamma when compared with naive T cells, and the memory CD4(+) subset was less responsive to IL-4. In summary, developing thymocytes and memory T cells appear to be resistant to the influences of numerous cytokines produced during immune responses.