The HSP90 inhibitor, 17AAG, protects the intestinal stem cell niche and inhibits graft versus host disease development.

Graft versus host disease (GvHD), which is the primary complication of allogeneic bone marrow transplantation, can alter the intestinal barrier targeted by activated donor T-cells. Chemical inhibition of the stress protein HSP90 was demonstrated in vitro to inhibit T-cell activation and to modulate endoplasmic reticulum (ER) stress to which intestinal cells are highly susceptible. Since the HSP90 inhibitor 17-allylamino-demethoxygeldanamycin (17AAG) is developed in clinics, we explored here its ability to control intestinal acute GvHD in vivo in two mouse GvHD models (C57BL/6BALB/c and FVB/NLgr5-eGFP), ex vivo in intestine organoids and in vitro in intestinal epithelial cultures. We show that 17AAG decreases GvHD-associated mortality without impairing graft versus leukemia effect. While 17AAG effect in T-cell activation is just moderate at the dose used in vivo, we observe a striking intestinal integrity protection. At the intestine level, the drug promotes the splicing of the transcription factor X-box binding protein 1 (XBP1), which is a key component of the ER stress. This effect is associated with a decrease in intestinal damage and an increase in Lgr5(+) stem cells, Paneth cells and defensins production. The importance of XBP1 splicing control is further confirmed in cultured cells and organoids of primary intestinal epithelium where XBP1 is either shRNA depleted or inhibited with toyocamycin. In conclusion, 17AAG has a protective effect on the epithelial intestinal barrier in mouse models of acute GvHD. This compound deserves to be tested in the therapeutic control of acute GvHD.

MHC polymorphism in Caribbean African green monkeys.

African green monkeys (AGM) are among the most widely used nonhuman primate models used in various fields of medical research. One species of AGM that originated from West Africa, Chlorocebus sabaeus, was introduced three centuries ago in the Caribbean islands. We present here a systematic study of the major histocompatibility complex (MHC) polymorphism of Caribbean AGM which is currently frequently used as an animal model. We studied 54 animals originated from Barbados (N=25) or Saint Kitts (N=29). The MHC polymorphism was characterized by means of 17 MHC microsatellites spread across MHC and DRB genotyping by DGGE sequencing. We defined nine frequent MHC haplotypes of which two were found in the two insular populations suggesting either past exchanges between the two populations or a common origin of the founders of the two populations. By the analysis of a previously described EST library, we characterized 38 MHC cDNA sequences (17 class I and 21 class II). In conclusion, we characterized for the first time the MHC polymorphism of Barbados and Saint Kitts AGM. We found a restricted polymorphism due to a founding effect, which is responsible for a strong bottleneck. The poorness of MHC polymorphism observed in the Caribbean AGM populations is similar to that observed in the Mauritian cynomolgus macaque population.

AMP-activated protein kinase in the regulation of hepatic energy metabolism: from physiology to therapeutic perspectives.

As the liver is central in the maintenance of glucose homeostasis and energy storage, knowledge of the physiology as well as physiopathology of hepatic energy metabolism is a prerequisite to our understanding of whole-body metabolism. Hepatic fuel metabolism changes considerably depending on physiological circumstances (fed vs. fasted state). In consequence, hepatic carbohydrate, lipid and protein synthesis/utilization are tightly regulated according to needs. Fatty liver and hepatic insulin resistance (both frequently associated with the metabolic syndrome) or increased hepatic glucose production (as observed in type 2 diabetes) resulted from alterations in substrates oxidation/storage balance in the liver. Because AMP-activated protein kinase (AMPK) is considered as a cellular energy sensor, it is important to gain understanding of the mechanism by which hepatic AMPK coordinates hepatic energy metabolism. AMPK has been implicated as a key regulator of physiological energy dynamics by limiting anabolic pathways (to prevent further ATP consumption) and by facilitating catabolic pathways (to increase ATP generation). Activation of hepatic AMPK leads to increased fatty acid oxidation and simultaneously inhibition of hepatic lipogenesis, cholesterol synthesis and glucose production. In addition to a short-term effect on specific enzymes, AMPK also modulates the transcription of genes involved in lipogenesis and mitochondrial biogenesis. The identification of AMPK targets in hepatic metabolism should be useful in developing treatments to reverse metabolic abnormalities of type 2 diabetes and the metabolic syndrome.

Head-down tilt bed rest and immune responses.

Head-down tilt bed rest (HDT) is used as a model for studying the physiological changes occurring in weightlessness during spaceflight. In the present study, eight volunteers were subjected to a strict HDT of -6 degrees for 42 days. Blood samples were obtained 37 and 13 days before, at days 13, 34, and 41 during, and 12, 33, and 47 days after HDT. FACScan analysis was used to determine cell subpopulations. Plasma was used to quantify various circulating hormone levels. Whole blood and reconstituted blood were stimulated with various activators such as phytohaemagglutinin-P (PHA), PHA combined with phorbol-12-myristate 13-acetate (PMA), anti-CD2, anti-CD3, and lipopolysaccharide. Supernatants were collected and analysed for the interleukins IL-1beta, IL-2, IL-6, and IL-10, interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha). The total number of T lymphocytes and monocytes did not change significantly, whereas the number of polymorphonuclear cells increased during HDT. The percentage of CD2+ and CD3+ cells was increased at day 35 of HDT. The percentage and total number of natural killer cells (CD2+/CD3-/CD56+) was increased 12 days before and 14 days after HDT. TNF-alpha secretion did not change significantly during HDT. IL-2, IL-10 and IFN-gamma were increased at day 34 of HDT. IL-1beta levels were increased before and during HDT compared to post-HDT measurements. No significant changes were observed in plasma immunoglobulin, complement factors and other factors of the inflammatory system. Prolactin levels increased slightly but significantly at day 35 of HDT, thyreotropin and growth hormone levels remained virtually unchanged. Cortisol decreased slightly but significantly over the entire duration of the study. The changes observed during HDT do not indicate that the immune system is blunted, and these changes do not seem to correlate with the duration of HDT. Taken together these results show that a HDT does not reproduce the changes in immune responses observed after spaceflight.

The complete sequence of a 10.8kb fragment to the right of the chromosome III centromere of Saccharomyces cerevisiae.

The complete nucleotide sequence of the D10H fragment (10850 bp) was determined. The D10H fragment is located on the right arm of chromosome III near the centromere and contains the SUF2 gene. Six open reading frames (ORFs) larger than 300 bp were found. One of them is the CIT2 gene encoding the cytoplasmic citrate synthase. The others are new putative genes and show no significant similarity with any known gene. In addition two tRNA genes (Asn and Pro) and a solo delta element were identified. Two ORFs were disrupted; no peculiar phenotype was observed.