Serca2 deletion in the mouse adult Bmi1 + compartment induces a lethal phenotype involving a severe gastric dysfunction

BACKGROUND Cytoplasmic calcium ions (Ca2+) are well-known intracellular signalling molecules, being involved in the integration of a variety of metabolic, proliferative and survival signals. They have been demonstrated an important role in embryonic and most adult stem cells (ASC). In the gut, it has been long recognized that Ca2+ handling is critical for the generation of pacemaker activity by Cajal interstitial cells. More recently, some studies in have shown that cytosolic Ca2+ plays a central role in intestinal stem cell (ISC) regulation, while this connection remains undefined in mammalian ISC populations. AIM To evaluate whether cytosolic Ca2+ could play an equivalent role in murine ISC population as in Drosophila melanogaster , and compare this effect to the lymphohematopoietic compartment. METHODS To this aim, we disrupted the expression of the calcium transporter Serca2, one of the crucial players in Ca2+ storage, within murine ISC population. Murine ISC compartment was selected using Bmi1 , a well-established marker for reserve ISCs crucial in regeneration upon injury, and Serca2 conditional deletion was evaluated in the gut epithelium using the Serca2Bmi1homo mouse strain. Main results were compared with the Serca2villinhomo strain. Metabolic parameters were evaluated, including food and water intake, body weight evolution, glucose response, serum biochemical markers and fecal fat content. Histological analysis along the gastric tract was used to assess Bmi1 + ISC proliferative state. Intestinal crypts were purified and analyzed for proliferation, cell death and gene expression. Total and segmented intestine was analyzed for differential gene expression. Due to the severe phenotype found, the impact of Serca2 conditional deletion was later evaluated in the lymphohematopoietic system, to eliminate a generalized effect. RESULTS Conditional deletion of Serca2 in Bmi1 + adult ASC (DSerca2Bmi1homo mice) resulted, unexpectedly, in a lethal phenotype. Important weight loss suggested severe malabsorption/malnutrition as the most probable cause of deaths. Results were further confirmed in DSerca2Villin mice, that developed a fulminant phenotype during the first week after tamoxifen-induction. Blood biochemistry showed a reduced glucose uptake, while feces content showed an increased total amount of fat in DSerca2Villin mice. Histological analysis along gastric tract revealed major extension of Bmi1 + cell compartment in DSerca2Bmi1homo mice. In the gut, Bmi1 + ISC population showed a substantial increase (3-fold) in their proliferative rate upon Serca2 deletion. Differential gene expression did not identify major changes in intestinal crypts, but key calcium regulatory players such as Orai-1 , Pmca-1 and Pmca-4 expression were affected, as well as endoplasmic reticulum stress response genes. Finally, important thymic alterations were characterized in DSerca2Bmi1 mice, plausibly associated to the malnourished state of the animals. CONCLUSION As in Drosophila melanogaster , cytosolic Ca2+ is a key element of murine ISC regulation, affecting their proliferation rate and the proper differentiation equilibrium. Severe lethality provoked by Serca2 deletion in both gut Bmi1 + and Villin + compartments raises the possibility that dysregulation of cytosolic Ca2+ could play some role in some gastric pathologies.