Rôle des cellules ß - intercalaires dans le maintien de la balance du sodium et de la pression artérielle

Hypertension is one of the most common human diseases. Today, many studies support the hypothesis that hypertension necessarily involve abnormal transport of salt (NaCl) by the kidneys. The nephron, the functional unit of the kidney, is composed of several segments with different NaCl transport systems. Our team has recently demonstrated that β-intercalated cells of the collecting duct (CD) were able to reabsorb NaCl. This transport is possible through functional coupling of two exchangers : pendrin and NDCBE. The objective of this thesis is to characterize the transport and show that β-intercalated cells (β-IC) play an important role in maintaining of sodium balance and blood pressure. Through several transgenic mouse models, we have shown that deletion of NDCBE causes hypotension and induces a compensation by another transporter of NaCl : the NCC cotransporter. With another mouse model, knockout for the proton pump H+- ATPase specifically in the intercalated cells and necessary for the reabsorption of NaCI in the β-ICs, we have shown that the β-ICs were capable of modulating the sodium transport of the principal cell of the CD through the secretion of paracrine factors.Otherwise, in mice with a mutated form of the WNK4 kinase, known to cause the Gordon syndrome, a disease characterized by hypertension and hyperkalemia, the NaCl transport by β-ICs is overactive and this overactivation probably contributes to the establishment of the disease.L'hypertension est l'une des maladies humaines les plus courantes. Aujourd'hui, de nombreuses études confirment l'hypothèse selon laquelle l'hypertension implique nécessairement un transport anormal de sel (NaCl) par les reins.Le néphron, l'unité fonctionnelle du rein, est composé de plusieurs segments présentant différents systèmes de transport de NaCl. Notre équipe a récemment démontré que les cellules intercalaires du canal collecteur (CD) étaient capables de réabsorber du NaCl. Ce transport est possible grâce au couplage fonctionnel de deux échangeurs : la pendrine et NDCBE. L'objectif de cette thèse est de caractériser ce transport et de montrer que les cellules intercalaires ( IC) jouent un rôle important dans le maintien de la balance sodée et de la pression artérielle. Grâce à plusieurs modèles de souris transgéniques nous avons pu montrer que la délétion de NDCBE provoque une hypotension et la mise en place d'une compensation par un autre transporteur du néphron : le cotransporteur NCC. A partir d'un autre modèle murin knock-out pour la pompe à proton H+-ATPase spécifiquement dans les cellules intercalaires et nécessaire à la réabsorption de NaCl par les ICs, nous avons montré que les ICs étaient capables de moduler le transport de sodium des cellules principales du CD par la sécrétion de facteurs paracrines. Enfin, chez des souris possédant une mutation de kinase WNK4, connue pour provoquer le syndrôme de Gordon, une maladie caractérisée entre autre par une hypertension artérielle et une hyperkaliémie, le transport de NaCl par les ICs est suractivée et cette suractivation contribue probablement à la mise en place de la maladie

Detection of Circulating Tumor Cells in the Diagnostic Leukapheresis Product of Non-Small-Cell Lung Cancer Patients Comparing CellSearch® and ISET

Circulating tumor cells (CTCs) detected by CellSearch are prognostic in non-small-cell lung cancer (NSCLC), but rarely found. CTCs can be extracted from the blood together with mononuclear cell populations by diagnostic leukapheresis (DLA), therefore concentrating them. However, CellSearch can only process limited DLA volumes (≈2 mL). Therefore, we established a protocol to enumerate CTCs in DLA products with Isolation by SizE of Tumor cells (ISET), and compared CTC counts between CellSearch® and ISET. DLA was performed in NSCLC patients who started a new therapy. With an adapted protocol, ISET could process 10 mL of DLA. CellSearch detected CTCs in a volume equaling 2 × 108 leukocytes (mean 2 mL). CTC counts per mL were compared. Furthermore, the live cell protocol of ISET was tested in eight patients. ISET successfully processed all DLA products—16 with the fixed cell protocol and 8 with the live cell protocol. In total, 10–20 mL of DLA was processed. ISET detected CTCs in 88% (14/16), compared to 69% (11/16, p < 0.05) with CellSearch. ISET also detected higher number of CTCs (ISET median CTC/mL = 4, interquartile range [IQR] = 2–6, CellSearch median CTC/mL = 0.9, IQR = 0–1.8, p < 0.01). Cells positive for the epithelial cell adhesion molecule (EpCAM+) per mL were detected in similar counts by both methods. Eight patients were processed with the live cell protocol. All had EpCAM+, CD45−, CD235- cells isolated by fluorescence-activated cell sorting (FACS). Overall, ISET processed larger volumes and detected higher CTC counts compared to CellSearch. EpCAM+ CTCs were detected in comparable rates

The role of ß-intercalated cells in maintaining the sodium balance and blood pressure

L'hypertension est l'une des maladies humaines les plus courantes. Aujourd'hui, de nombreuses études confirment l'hypothèse selon laquelle l'hypertension implique nécessairement un transport anormal de sel (NaCl) par les reins.Le néphron, l'unité fonctionnelle du rein, est composé de plusieurs segments présentant différents systèmes de transport de NaCl. Notre équipe a récemment démontré que les cellules intercalaires du canal collecteur (CD) étaient capables de réabsorber du NaCl. Ce transport est possible grâce au couplage fonctionnel de deux échangeurs : la pendrine et NDCBE. L'objectif de cette thèse est de caractériser ce transport et de montrer que les cellules intercalaires ( IC) jouent un rôle important dans le maintien de la balance sodée et de la pression artérielle. Grâce à plusieurs modèles de souris transgéniques nous avons pu montrer que la délétion de NDCBE provoque une hypotension et la mise en place d'une compensation par un autre transporteur du néphron : le cotransporteur NCC. A partir d'un autre modèle murin knock-out pour la pompe à proton H+-ATPase spécifiquement dans les cellules intercalaires et nécessaire à la réabsorption de NaCl par les ICs, nous avons montré que les ICs étaient capables de moduler le transport de sodium des cellules principales du CD par la sécrétion de facteurs paracrines. Enfin, chez des souris possédant une mutation de kinase WNK4, connue pour provoquer le syndrôme de Gordon, une maladie caractérisée entre autre par une hypertension artérielle et une hyperkaliémie, le transport de NaCl par les ICs est suractivée et cette suractivation contribue probablement à la mise en place de la maladie.Hypertension is one of the most common human diseases. Today, many studies support the hypothesis that hypertension necessarily involve abnormal transport of salt (NaCl) by the kidneys. The nephron, the functional unit of the kidney, is composed of several segments with different NaCl transport systems. Our team has recently demonstrated that β-intercalated cells of the collecting duct (CD) were able to reabsorb NaCl. This transport is possible through functional coupling of two exchangers : pendrin and NDCBE. The objective of this thesis is to characterize the transport and show that β-intercalated cells (β-IC) play an important role in maintaining of sodium balance and blood pressure. Through several transgenic mouse models, we have shown that deletion of NDCBE causes hypotension and induces a compensation by another transporter of NaCl : the NCC cotransporter. With another mouse model, knockout for the proton pump H+- ATPase specifically in the intercalated cells and necessary for the reabsorption of NaCI in the β-ICs, we have shown that the β-ICs were capable of modulating the sodium transport of the principal cell of the CD through the secretion of paracrine factors.Otherwise, in mice with a mutated form of the WNK4 kinase, known to cause the Gordon syndrome, a disease characterized by hypertension and hyperkalemia, the NaCl transport by β-ICs is overactive and this overactivation probably contributes to the establishment of the disease

Double Knockout of the Na+-Driven Cl-/HCO3--Exchanger and Na+/Cl- Cotransporter Induces Hypokalemia and Volume Depletion

We recently described a novel thiazide-sensitive electroneutral NaCI transport mechanism resulting from the parallel operation of the Cl-/HCO3- exchanger pendrin and the Na+-driven Cl-/2HCO(3)(-) exchanger (NDCBE) in beta-intercalated cells of the collecting duct. Although a role for pendrin in maintaining Na+ balance, intravascular volume, and BP is well supported, there is no in vivo evidence for the role of NDCBE in maintaining Na+ balance. Here, we show that deletion of NDCBE in mice caused only subtle perturbations of Na homeostasis and provide evidence that the Na+/Cl- cotransporter (NCC) compensated for the inactivation of NDCBE. To unmaskthe role of NDCBE, we generated Ndcbe/Ncdclouble-knockout (dKO) mice. On a normal salt diet, dKO and single-knockout mice exhibited similar activation of the renin-angiotensin-aldosterone system, whereas only dKO mice displayed a lower blood K concentration. Furthermore, dKO mice displayed upregulation of the epithelial sodium channel (ENaC) and the Ca2+-activated K+ channel BKCa. During NaCI depletion, only dKO mice developed marked intravascular volume contraction, despite dramatically increased renin activity. Notably, the increase in aldosterone levels expected on NaCI depletion was attenuated in dKO mice, and single-knockout and dKO mice had similar blood K+ concentrations under this condition. In conclusion, NDCBE is necessary for maintaining sodium balance and intravascular volume during salt depletion or NCC inactivation in mice. Furthermore, NDCBE has an important role in the prevention of hypokalemia. Because NCC and NDCBE are both thiazide targets, the combined inhibition of NCC and the NDCBE/pendrin system may explain thiazide-induced hypokalemia in some patients

Renal β-intercalated cells maintain body fluid and electrolyte balance

Inactivation of the B1 proton pump subunit (ATP6V1B1) in intercalated cells (ICs) leads to type I distal renal tubular acidosis (dRTA), a disease associated with salt- and potassium-losing nephropathy. Here we show that mice deficient in ATP6V1B1 (Atp6v1b1-/- mice) displayed renal loss of NaCl, K+, and water, causing hypovolemia, hypokalemia, and polyuria. We demonstrated that NaCl loss originated from the cortical collecting duct, where activity of both the epithelial sodium channel (ENaC) and the pendrin/Na+-driven chloride/bicarbonate exchanger (pendrin/NDCBE) transport system was impaired. ENaC was appropriately increased in the medullary collecting duct, suggesting a localized inhibition in the cortex. We detected high urinary prostaglandin E2 (PGE2) and ATP levels in Atp6v1b1-/- mice. Inhibition of PGE2 synthesis in vivo restored ENaC protein levels specifically in the cortex. It also normalized protein levels of the large conductance calcium-activated potassium channel and the water channel aquaporin 2, and improved polyuria and hypokalemia in mutant mice. Furthermore, pharmacological inactivation of the proton pump in β-ICs induced release of PGE2 through activation of calcium-coupled purinergic receptors. In the present study, we identified ATP-triggered PGE2 paracrine signaling originating from β-ICs as a mechanism in the development of the hydroelectrolytic imbalance associated with dRTA. Our data indicate that in addition to principal cells, ICs are also critical in maintaining sodium balance and, hence, normal vascular volume and blood pressure