Rôle du système kallicréine-kinine(s) dans les complications du diabète

The kallikrein-kinin(s) system (KKS) is a peptide system with various pathophysiological effects. Kinins exert their actions through activation of two different receptor subtypes: B1 receptor (B1R) and B2 receptor (B2R). The aim of my thesis was to study the role of this system in diabetic complications in mice. We used pharmacological approaches using new specific agonists of B1R or B2R, or a specific antagonist of B2R. In the first part, we showed that selective pharmacological activation of B1R or B2R overcomes the effect of diabetes on post-ischemic neovascularization and restores tissue perfusion through inflammation. In a second model, we showed that B2R agonist impairs wound repair in mice, inducing skin disorganization and epidermis thickening. Interestingly, B2R blockade improves skin wound healing in two mouse models of diabetes. In a last part, we showed that B2R activation increases mortality after transient cerebral ischemia. In diabetic mice, B1R activation has neuroprotective effects. Indeed, B1R agonist treatment decreases infarct size and improves neurological deficit at day 2 after transient cerebral ischemia. To conclude, KKS activation has contradictory effects depending on the organs studied. The study of new B1 or B2 kinin receptor agonists opens new therapeutic options in diabetic complications.Le système kallicréine-kinine(s) (SKK) est un système peptidique vasomoteur avec des effets multiples et complexes qui dépendent principalement du tissu et des pathologies étudiés. Les kinines activent deux types de récepteurs : le récepteur B1 (RB1) et le récepteur B2 (RB2). L’objectif de mon travail de thèse a été d’étudier les effets de nouveaux agonistes spécifiques des RB1 ou des RB2 dans certaines complications associées au diabète chez la souris. Dans une première partie, nous avons pu montrer que l’administration chronique d’un agoniste spécifique des RB1 ou des RB2 corrige entièrement le défaut de néovascularisation post-ischémique observé chez la souris diabétique notamment en augmentant l’inflammation. Dans un deuxième modèle, nous avons pu montrer que l’activation des RB2 entraîne une désorganisation des couches cutanées et un retard important de cicatrisation chez la souris. En revanche, l’administration d’un antagoniste des RB2 accélère de façon significative la cicatrisation dans deux modèles de diabète. La dernière partie de ce travail a permis de montrer que l’administration chronique des RB2 augmente la mortalité des animaux ayant subit une ischémie cérébrale. Chez la souris diabétique, l’activation des RB1 a des effets neuroprotecteurs puisqu’elle réduit la taille de l'infarctus et les déficits neurologiques après une ischémie cérébrale transitoire. Pour conclure, nous avons pu montrer au cours de ces travaux que l’activation du SKK a des effets contradictoires en fonction des organes étudiés. Ainsi, l’étude de ces nouveaux agonistes des récepteurs des kinines ouvre des nouvelles voies thérapeutiques dans les complications du diabète

0052: Role of kinins in diabetic wound healing

The diabetic foot is associated with pain, decrease in patient's quality of life, considerable costs, and amputation. In this study, we determined the role of KKS, via activation of bradykinin receptors (B1R or B2R), in a mouse model of diabetic wound healing. Diabetic or nondiabetic mice are wounded with an 8-mm punch biopsy and then are treated or not with specific B1R or B2R agonists (720nmol/kg.d-1) and/or B2R antagonist (Icatibant, 500μg/kg.dg/d-1). The wound-healing surface was daily followed up. At 11 days, the scar were analysed by histology (Masson's trichrome staining) and B1R and B2R expression were assessed (RT-qPCR). Effects of the agonists on cells (fibroblasts and keratinocytes) migration and proliferation were also analysed. In diabetic condition, mRNA of B1R and B2R was increased in skin (p<0.01). B1R activation had no effect on wound closure in our model. In contrast, B2R activation dramatically delayed wound healing in diabetic (p<0.001) or nondiabetic (p<0.01) mice. Histological analysis of scar showed significant skin disorganization and epidermis thickening with B2R agonist (p<0.05). In vitro, B2R agonist induced an increase of keratinocyte proliferation (+46% after 48h, p<0.01) and a stimulation of keratinocyte migration (+30% after 24h, p<0.05). These effects was associated with ERK phosphorylation which occurs downstream of EGFR activation (p<0.05). B2R agonist had no effect on fibroblast migration but decreased fibroblast proliferation (–33% after 48h, p<0.05). Co-treatment with Icatibant abrogated in vivo and in vitro effects observed with B2R agonist. Moreover, Icatibant alone hastened wound healing and decrease the epidermis thickening induced by diabetes. In conclusion, KKS, through the B2R but not the B1R, plays a critical role in proliferation and remodelling phases of skin wound healing in mice. While more studies are needed, Icatibant could be used to correct the diabetic wound healing defect

A new view of macula densa cell microanatomy

Although macula densa (MD) cells are chief regulatory cells in the nephron with unique microanatomical features, they have been difficult to study in full detail due to their inaccessibility and limitations in earlier microscopy techniques. The present study used a new mouse model with a comprehensive imaging approach to visualize so far unexplored microanatomical features of MD cells, their regulation, and functional relevance. MD-GFP mice with conditional and partial induction of green fluorescent protein (GFP) expression, which specifically and intensely illuminated only single MD cells, were used with fluorescence microscopy of fixed tissue and live MD cells in vitro and in vivo with complementary electron microscopy of the rat, rabbit, and human kidney. An elaborate network of major and minor cell processes, here named maculapodia, were found at the cell base, projecting toward other MD cells and the glomerular vascular pole. The extent of maculapodia showed upregulation by low dietary salt intake and the female sex. Time-lapse imaging of maculapodia revealed highly dynamic features including rapid outgrowth and an extensive vesicular transport system. Electron microscopy of rat, rabbit, and human kidneys and three-dimensional volume reconstruction in optically cleared whole-mount MD-GFP mouse kidneys further confirmed the presence and projections of maculapodia into the extraglomerular mesangium and afferent and efferent arterioles. The newly identified dynamic and secretory features of MD cells suggest the presence of novel functional and molecular pathways of cell-to-cell communication in the juxtaglomerular apparatus between MD cells and between MD and other target cells.NEW & NOTEWORTHY This study illuminated a physiologically regulated dense network of basal cell major and minor processes (maculapodia) in macula densa (MD) cells. The newly identified dynamic and secretory features of these microanatomical structures suggest the presence of novel functional and molecular pathways of cell-to-cell communication in the juxtaglomerular apparatus between MD and other target cells. Detailed characterization of the function and molecular details of MD cell intercellular communications and their role in physiology and disease warrant further studies

Kallikrein-kinin(s) system and diabetic complications

Le système kallicréine-kinine(s) (SKK) est un système peptidique vasomoteur avec des effets multiples et complexes qui dépendent principalement du tissu et des pathologies étudiés. Les kinines activent deux types de récepteurs : le récepteur B1 (RB1) et le récepteur B2 (RB2). L’objectif de mon travail de thèse a été d’étudier les effets de nouveaux agonistes spécifiques des RB1 ou des RB2 dans certaines complications associées au diabète chez la souris. Dans une première partie, nous avons pu montrer que l’administration chronique d’un agoniste spécifique des RB1 ou des RB2 corrige entièrement le défaut de néovascularisation post-ischémique observé chez la souris diabétique notamment en augmentant l’inflammation. Dans un deuxième modèle, nous avons pu montrer que l’activation des RB2 entraîne une désorganisation des couches cutanées et un retard important de cicatrisation chez la souris. En revanche, l’administration d’un antagoniste des RB2 accélère de façon significative la cicatrisation dans deux modèles de diabète. La dernière partie de ce travail a permis de montrer que l’administration chronique des RB2 augmente la mortalité des animaux ayant subit une ischémie cérébrale. Chez la souris diabétique, l’activation des RB1 a des effets neuroprotecteurs puisqu’elle réduit la taille de l'infarctus et les déficits neurologiques après une ischémie cérébrale transitoire. Pour conclure, nous avons pu montrer au cours de ces travaux que l’activation du SKK a des effets contradictoires en fonction des organes étudiés. Ainsi, l’étude de ces nouveaux agonistes des récepteurs des kinines ouvre des nouvelles voies thérapeutiques dans les complications du diabète.The kallikrein-kinin(s) system (KKS) is a peptide system with various pathophysiological effects. Kinins exert their actions through activation of two different receptor subtypes: B1 receptor (B1R) and B2 receptor (B2R). The aim of my thesis was to study the role of this system in diabetic complications in mice. We used pharmacological approaches using new specific agonists of B1R or B2R, or a specific antagonist of B2R. In the first part, we showed that selective pharmacological activation of B1R or B2R overcomes the effect of diabetes on post-ischemic neovascularization and restores tissue perfusion through inflammation. In a second model, we showed that B2R agonist impairs wound repair in mice, inducing skin disorganization and epidermis thickening. Interestingly, B2R blockade improves skin wound healing in two mouse models of diabetes. In a last part, we showed that B2R activation increases mortality after transient cerebral ischemia. In diabetic mice, B1R activation has neuroprotective effects. Indeed, B1R agonist treatment decreases infarct size and improves neurological deficit at day 2 after transient cerebral ischemia. To conclude, KKS activation has contradictory effects depending on the organs studied. The study of new B1 or B2 kinin receptor agonists opens new therapeutic options in diabetic complications

AVC et nouvelles stratégies thérapeutiques (effet d un activateur des canaux potassiques, le SKA-31, sur un modèle de souris double transgéniques hypertendu développant des AVC spontanés)

MONTPELLIER-BU Pharmacie (341722105) / SudocSudocFranceF

Neuroprotective effect of kinin B1 receptor activation in acute cerebral ischemia in diabetic mice

Abstract Activation of the kallikrein-kinin system enhances cardiac and renal tolerance to ischemia. Here we investigated the effects of selective agonists of kinin B1 or B2 receptor (R) in brain ischemia-reperfusion in diabetic and non-diabetic mice. The role of endogenous kinins was assessed in tissue kallikrein deficient mice (TK−/−). Mice underwent 60min-middle cerebral artery occlusion (MCAO), eight weeks after type 1-diabetes induction. Treatment with B1R-, B2R-agonist or saline was started at reperfusion. Neurological deficit (ND), infarct size (IS), brain water content (BWC) were measured at day 0, 1 and 2 after injury. MCAO induced exaggerated ND, mortality and IS in diabetic mice. B2R-agonist increased ND and mortality to 60% and 80% in non-diabetic and diabetic mice respectively, by mechanisms involving hemodynamic failure and renal insufficiency. TK−/− mice displayed reduced ND and IS compared to wild-type littermate, consistent with suppression of B2R activity. B1R mRNA level increased in ischemic brain but B1R-agonist had no effect on ND, mortality or IS in non-diabetic mice. In contrast, in diabetic mice, B1R-agonist tested at two doses significantly reduced ND by 42–52% and IS by 66–71%, without effect on BWC or renal function. This suggests potential therapeutic interest of B1R agonism for cerebral protection in diabetes

Re-epithelialization of pathological cutaneous wounds is improved by local mineralocorticoid receptor antagonism

International audienceImpaired cutaneous wound healing is a social burden. It occurs as a consequence of glucocorticoid treatment and in several pathologies. Glucocorticoids (GC) bind not only to the glucocorticoid receptor (GR), but also to the mineralocorticoid receptor (MR), both expressed by keratinocytes. In addition to its beneficial effects through the GR, GC exposure may lead to inappropriate MR occupancy. We hypothesized that dermatological use of MR antagonists (MRA) may be beneficial by overcoming the negative impact of GC treatment on pathological wounds. The potent GC clobetasol, applied as an ointment to mouse skin, or added to cultured human skin explants, induced delayed wound closure and outgrowth of epidermis with reduced proliferation of keratinocytes. Delayed wound re-epithelialization was rescued by local MRA application. Normal skin was unaffected by MRA. The benefit of MR blockade is explained by the increased expression of MR in clobetasol-treated mouse skin. Blockade of the epithelial sodium channel by phenamil also rescued cultured human skin explants from GC- impaired growth of the epidermis. MRA application over post-biopsy wounds of clobetasol-treated skin zones of healthy volunteers (from the SPIREPI clinical trial) also accelerated wound closure. In conclusion, we propose repositioning MRA for cutaneous application to improve delayed wound closure occurring in pathology

Essential role and therapeutic targeting of the glomerular endothelial glycocalyx in lupus nephritis

Lupus nephritis (LN) is a major organ complication and cause of morbidity and mortality in patients with systemic lupus erythematosus (SLE). There is an unmet medical need for developing more efficient and specific, mechanism-based therapies, which depends on improved understanding of the underlying LN pathogenesis. Here we present direct visual evidence from high-power intravital imaging of the local kidney tissue microenvironment in mouse models showing that activated memory T cells originated in immune organs and the LN-specific robust accumulation of the glomerular endothelial glycocalyx played central roles in LN development. The glomerular homing of T cells was mediated via the direct binding of their CD44 to the hyaluronic acid (HA) component of the endothelial glycocalyx, and glycocalyx-degrading enzymes efficiently disrupted homing. Short-course treatment with either hyaluronidase or heparinase III provided long-term organ protection as evidenced by vastly improved albuminuria and survival rate. This glycocalyx/HA/memory T cell interaction is present in multiple SLE-affected organs and may be therapeutically targeted for SLE complications, including LN