P53/Rb inhibition induces metastatic adrenocortical carcinomas in a preclinical transgenic model.

Adrenocortical carcinoma (ACC) is a rare cancer with poor prognosis. Pan-genomic analyses identified p53/Rb and WNT/ß-catenin signaling pathways as main contributors to the disease. However, isolated ß-catenin constitutive activation failed to induce malignant progression in mouse adrenocortical tumors. Therefore, there still was a need for a relevant animal model to study ACC pathogenesis and to test new therapeutic approaches. Here, we have developed a transgenic mice model with adrenocortical specific expression of SV40 large T-antigen (AdTAg mice), to test the oncogenic potential of p53/Rb inhibition in the adrenal gland. All AdTAg mice develop large adrenal carcinomas that eventually metastasize to the liver and lungs, resulting in decreased overall survival. Consistent with ACC in patients, adrenal tumors in AdTAg mice autonomously produce large amounts of glucocorticoids and spontaneously activate WNT/ß-catenin signaling pathway during malignant progression. We show that this activation is associated with downregulation of secreted frizzled related proteins (Sfrp) and Znrf3 that act as inhibitors of the WNT signaling. We also show that mTORC1 pathway activation is an early event during neoplasia expansion and further demonstrate that mTORC1 pathway is activated in ACC patients. Preclinical inhibition of mTORC1 activity induces a marked reduction in tumor size, associated with induction of apoptosis and inhibition of proliferation that results in normalization of corticosterone plasma levels in AdTAg mice. Altogether, these data establish AdTAg mice as the first preclinical model for metastatic ACC.

All-trans-retinoic acid regulates aquaporin-3 expression and related cellular membrane permeability in the human amniotic environment.

INTRODUCTION: The aquaporins (AQP1, 3, 8, 9 and 11) are known to be expressed, and involved in the transport of water and small molecules through fetal membranes. To exert these crucial functions, these AQPs have to be finely regulated. All-trans-retinoic acid (atRA) was previously found to regulate some genes in this environment, raising the question of whether these AQPs were regulated by atRA. METHODS: Explants, and primary and established amniotic cells were cultured to determine which AQP were transcriptionally modified by atRA, using the qRT-PCR strategy. Immunohistochemistry and glycerol uptake tests were used to determine the impact of atRA on AQP protein expression and function. Specific agonists of retinoic acid receptors were used to identify the molecular mechanisms of AQP promoter activation. A classical gene AQP promoter study was also used to identify DR5 retinoic acid receptor elements (RAREs). RESULTS: Beyond these AQPs, only one specific atRA-dependent increase in AQP3 transcripts and proteins level was established in amnion (not in chorion) and in related primary and established cells. We found three DR5-RAREs essential for inducing this transcriptional AQP3 through RARα. This transactivation of the AQP3 coding gene was functionally related to an increase of AQP3 permeability tests by a glycerol uptake assay. DISCUSSION: Our data support an atRA regulatory model of AQP3 expression leading to an increased cellular permeability in the epithelial amniotic environment. We cast new light on AF regulation in healthy pregnancy, and advance new hypotheses for obstetrical complications linked to impairment of the retinoic signaling pathway.

Long-term intermittent glutamine supplementation repairs intestinal damage (structure and functional mass) with advanced age: assessment with plasma citrulline in a rodent model.

OBJECTIVE: Glutamine is the preferred fuel for the rat small intestine and promotes the growth of intestinal mucosa, especially in the event of gut injury. Quantitatively, glutamine is one important precursor for intestinal citrulline release. The aim of this study was to determine whether the effect of glutamine on the increase in intestinal villus height is correlated with an increase in both gut mass and citrulline plasma level in very old rats. METHODS: We intermittently supplemented very old (27-mo) female rats with oral glutamine (20% of diet protein). Intestinal histomorphometric analysis of the small bowel was performed. Amino acids, in particular citrulline, were measured in the plasma, liver and jejunum. Markers of renal (creatinine, urea) and liver (alanine aminotransferase [ALT]) and aspartate aminotransferase (AST) functions were measured to evaluate renal and liver functions in relation to aging and to glutamine supplementation. Liver glutathione was also determined to evaluate cellular redox state. RESULTS: Glutamine supplementation maintains the body weight of very old rats, not by limiting sarcopenia but rather by increasing the organ mass of the splanchnic area. Total intestine mass was significantly higher in glutamine-supplemented rats than in controls (15%). Measurement of villus height and crypt depth demonstrated that the difference between villus and crypt was significantly improved in glutamine pre-treated rats compared to controls (~ 11%). Plasma citrulline also increased by 15% in glutamine-supplemented rats compared to controls. CONCLUSION: Citrulline appears as a biomarker of enterocyte mass in villous atrophy associated with advanced age. Non-invasive measurement of this metabolite may be useful in following the state of the gastrointestinal tract in very old people, whose numbers are increasing worldwide and the care of whom is a major public health issue. The gut may contribute to the malnutrition caused by malabsorption frequently observed in the elderly.

[Implications of retinoid pathway in human fetal membranes: study of target genes]. / Étude de la voie des rétinoïdes au sein des membranes fœtales humaines: mise en évidence de gènes cibles.

Retinoids (active derivatives of vitamin A) were already demonstrated to be important morphogenes and their implication at the placental and fetal level was already established. A new field of research is now developed in order to show their role on fetal membranes constituted by amnion and chorion. To describe the role of retinoids on these membranes, our studies were focused on target gene research. Firstly, all metabolism enzymes needed to vitamin A pathways were demonstrated to be present and active in signal transduction. Secondly, a bioinformatic analysis was performed to assess a list of potential target genes that could be classified in different biological pathways (inflammation, retinoids, hormones, vascularization, extracellular matrix and water homeostasis). Then, it was demonstrated that the gene coding for PLAT, implied in the degradation of extracellular matrix during programmed or premature rupture of membranes, is regulated by retinoids in a two steps mechanism. Finally, preliminary data showed that some aquaporins, which control water transport across membranes, are expressed and regulated by retinoids in the fetal membranes. A disregulation in pathologies like oligo or poly-hydramnios can be anticipated. Improvement of our knowledge about the retinoid implications is a key point in order to obtain a precise and complete documented cartography of the vitamin A (regulating) in amniotic membranes (regulated) that will permit the development of new diagnostic and therapeutic strategies.

Liver X Receptor activation downregulates AKT survival signaling in lipid rafts and induces apoptosis of prostate cancer cells.

Cholesterol is a structural component of lipid rafts within the plasma membrane. These domains, used as platforms for various signaling molecules, regulate cellular processes including cell survival. Cholesterol contents are tightly correlated with the structure and function of lipid rafts. Liver X receptors (LXRs) have a central role in the regulation of cholesterol homeostasis within the cell. Therefore, we investigated whether these nuclear receptors could modulate lipid raft signaling and consequently alter prostate cancer (PCa) cell survival. Treatment with the synthetic LXR agonist T0901317 downregulated the AKT survival pathway and thus induced apoptosis of LNCaP PCa cells in both xenografted nude mice and cell culture. The decrease in tumor cholesterol content resulted from the upregulation of ABCG1 and the subsequent increase in reverse cholesterol transport. RNA interference experiments showed that these effects were mediated by LXRs. Atomic force microscopy scanning of the inner plasma membrane sheet showed smaller and thinner lipid rafts after LXR stimulation, associated with the downregulation of AKT phosphorylation in these lipid rafts. Replenishment of cell membranes with exogenous cholesterol antagonized these effects, showing that cholesterol is a key modulator in this process. Altogether, pharmacological modulation of LXR activity could thus reduce prostate tumor growth by enhancing apoptosis in a lipid raft-dependent manner.

Hematopoietic progenitor cell mobilization and harvesting in children with malignancies: do the advantages of pegfilgrastim really translate into clinical benefit?

Our purpose was to assess success rates in children of achieving optimal hematopoietic progenitor cells (HPCs) harvest after mobilization with 300 microg/kg pegfilgrastim. Between January 2005 and January 2007, 26 children with solid malignancies who were referred for HPC collection were consecutively included. Hematopoietic progenitor cell mobilization consisted of one s.c. injection of 300 microg/kg body weight (BW) of pegfilgrastim. The success criterion was defined as at least 5 x 10(6) CD34+ cells/kg during the first standard apheresis (less than 3 blood volumes processed (BVP)). After 26 inclusions, the Bayesian analysis gave a mean estimated success rate of 60.7% (95% credibility interval: 42.0-78.0%). The first apheresis allowed the collection of 8.3 x 10(6) CD34+ cells/kg BW (range 0.6-37.8), with a median of 2.8 BVP (range 1.4-3.0). Overall, the median of CD34+ cells collected was 12.4 x 10(6)/kg (range 2.7-37.8). The cumulative dose of anthracyclin was the only variable associated with the total number of CD34+ collected cells (P<0.05). Mobilization was clinically well tolerated in 20 patients. No drug-related adverse events of grade > or =3 occurred. We conclude that a single injection of 300 microg/kg pegfilgrastim in the hematological steady state is an efficient and well-tolerated method of HPC mobilization in children with solid malignancies.

Expression of retinoid receptors during rabbit lung development.

Rabbit lung is often used to study the pathophysiology of some congenital anomalies affecting the lung because its development is very comparable with that of human. Retinoids and the molecular transducers of the retinoic signal play a crucial role in mammalian lung development. In the rabbit, the molecular retinoic pathway has so far been poorly studied. As a first step in elucidating this process, we aimed to identify the retinoic acid receptors (RARs) and retinoid X receptors (RXRs). We cloned a part of the nuclear receptors (RARs and RXRs), and we used reverse transcription - polymerase chain reaction (RT-PCR) and immunohistochemistry assays to demonstrate the presence of RAR (α,ß) and RXR (α, ß) at all stages of rabbit lung development. Our results initiate further analysis into the molecular and genetic functions of retinoids during normal and pathological rabbit lung development including the surgical model of congenital diaphragmatic hernia.

[Outcome of hyperlactatemia and lipodystrophy syndromes in patients infected with human immunodeficiency virus]. / Survenue des syndromes lipodystrophiques et hyperlactatémiques chez des patients infectés par le virus de l'immunodéficience humaine.

HIV-positive persons requiring a highly active antiretroviral therapy containing one or more nucleosidic reverse transcriptase inhibitors associated with or without protease inhibitors are exposed to metabolic side effects among which lipodystrophy and hyperlactemia, defined by blood lactates higher than 2,25 mmol/L. Hyperlactatemia had to be differentiated from lactic acidosis of type B (without hypoxemia, lactates higher than 5 mmol/L and arterial pH lower than 7,3), a rare but potentially fatal complication by multi-visceral failure. The accused INRT induce mitochondrial toxicity by inhibition of DNA gamma polymerase and deterioration of its DNA. Our exploratory study, troop of 282 patients, identified age and stavudine like statistically associated, which has occurred of this metabolic anomaly. The patients having profited of a therapeutic change with the profit from drugs minus hyperlactatogenic presented an obvious clinical and biological improvement; whereas similar switch of therapy occurred for the lipodystrophic patients presented any clinical improvement. Nevertheless, biological parameters (blood lactates, triglycerides, total cholesterol and LDL-cholesterol) were significantly decreased after this therapeutic switch occurred on the lipodystrophic patients. In conclusion, the measurement of the following biological parameters: glycemia, lactatemia, triglycerides, total cholesterol and LDL-cholesterol at patient VIH, in a simple and rigorous pre-analytical and analytical context, appears to us justified in the monitoring of metabolic disorders in treated HIV patients by INRT and/or IP.