Loss of Nix in Pdx1-deficient mice prevents apoptotic and necrotic β cell death and diabetes

Mutations in pancreatic duodenal homeobox (PDX1) are linked to human type 2 diabetes and maturity-onset diabetes of the young type 4. Consistent with this, Pdx1-haploinsufficient mice develop diabetes. Both apoptosis and necrosis of β cells are mechanistically implicated in diabetes in these mice, but a molecular link between Pdx1 and these 2 forms of cell death has not been defined. In this study, we introduced an shRNA into mouse insulinoma MIN6 cells to deplete Pdx1 and found that expression of proapoptotic genes, including NIP3-like protein X (Nix), was increased. Forced Nix expression in MIN6 and pancreatic islet β cells induced programmed cell death by simultaneously activating apoptotic and mitochondrial permeability transition-dependent necrotic pathways. Preventing Nix upregulation during Pdx1 suppression abrogated apoptotic and necrotic β cell death in vitro. In Pdx1-haploinsufficient mice, Nix ablation normalized pancreatic islet architecture, β cell mass, and insulin secretion and eliminated reactive hyperglycemia after glucose challenge. These results establish Nix as a critical mediator of β cell apoptosis and programmed necrosis in Pdx1-deficient diabetes.     

Maternal diet–induced microRNAs and mTOR underlie β cell dysfunction in offspring

A maternal diet that is low in protein increases the susceptibility of offspring to type 2 diabetes by inducing long-term alterations in β cell mass and function. Nutrients and growth factor signaling converge through mTOR, suggesting that this pathway participates in β cell programming during fetal development. Here, we revealed that newborns of dams exposed to low-protein diet (LP0.5) throughout pregnancy exhibited decreased insulin levels, a lower β cell fraction, and reduced mTOR signaling. Adult offspring of LP0.5-exposed mothers exhibited glucose intolerance as a result of an insulin secretory defect and not β cell mass reduction. The β cell insulin secretory defect was distal to glucose-dependent Ca²⁺ influx and resulted from reduced proinsulin biosynthesis and insulin content. Islets from offspring of LP0.5-fed dams exhibited reduced mTOR and increased expression of a subset of microRNAs, and blockade of microRNA-199a-3p and -342 in these islets restored mTOR and insulin secretion to normal. Finally, transient β cell activation of mTORC1 signaling in offspring during the last week of pregnancy of mothers fed a LP0.5 rescued the defect in the neonatal β cell fraction and metabolic abnormalities in the adult. Together, these findings indicate that a maternal low-protein diet alters microRNA and mTOR expression in the offspring, influencing insulin secretion and glucose homeostasis.     

Growth hormone receptor regulates β cell hyperplasia and glucose-stimulated insulin secretion in obese mice

Insulin, growth hormone (GH), and insulin-like growth factor-1 (IGF-1) play key roles in the regulation of β cell growth and function. Although β cells express the GH receptor, the direct effects of GH on β cells remain largely unknown. Here we have employed a rat insulin II promoter-driven (RIP-driven) Cre recombinase to disrupt the GH receptor in β cells (βGHRKO). βGHRKO mice fed a standard chow diet exhibited impaired glucose-stimulated insulin secretion but had no changes in β cell mass. When challenged with a high-fat diet, βGHRKO mice showed evidence of a β cell secretory defect, with further deterioration of glucose homeostasis indicated by their altered glucose tolerance and blunted glucose-stimulated insulin secretion. Interestingly, βGHRKO mice were impaired in β cell hyperplasia in response to a high-fat diet, with decreased β cell proliferation and overall reduced β cell mass. Therefore, GH receptor plays critical roles in glucose-stimulated insulin secretion and β cell compensation in response to a high-fat diet.     

PPARβ/δ affects pancreatic β cell mass and insulin secretion in mice

PPARβ/δ protects against obesity by reducing dyslipidemia and insulin resistance via effects in muscle, adipose tissue, and liver. However, its function in pancreas remains ill defined. To gain insight into its hypothesized role in β cell function, we specifically deleted Pparb/d in the epithelial compartment of the mouse pancreas. Mutant animals presented increased numbers of islets and, more importantly, enhanced insulin secretion, causing hyperinsulinemia. Gene expression profiling of pancreatic β cells indicated a broad repressive function of PPARβ/δ affecting the vesicular and granular compartment as well as the actin cytoskeleton. Analyses of insulin release from isolated PPARβ/δ-deficient islets revealed an accelerated second phase of glucose-stimulated insulin secretion. These effects in PPARβ/δ-deficient islets correlated with increased filamentous actin (F-actin) disassembly and an elevation in protein kinase D activity that altered Golgi organization. Taken together, these results provide evidence for a repressive role for PPARβ/δ in β cell mass and insulin exocytosis, and shed a new light on PPARβ/δ metabolic action.     

The von Hippel–Lindau Chuvash mutation promotes pulmonary hypertension and fibrosis in mice

Mutation of the von Hippel–Lindau (VHL) tumor suppressor protein at codon 200 (R200W) is associated with a disease known as Chuvash polycythemia. In addition to polycythemia, Chuvash patients have pulmonary hypertension and increased respiratory rates, although the pathophysiological basis of these symptoms is unclear. Here we sought to address this issue by studying mice homozygous for the R200W Vhl mutation (Vhl^R/R mice) as a model for Chuvash disease. These mice developed pulmonary hypertension independently of polycythemia and enhanced normoxic respiration similar to Chuvash patients, further validating Vhl^R/R mice as a model for Chuvash disease. Lungs from Vhl^R/R mice exhibited pulmonary vascular remodeling, hemorrhage, edema, and macrophage infiltration, and lungs from older mice also exhibited fibrosis. HIF-2α activity was increased in lungs from Vhl^R/R mice, and heterozygosity for Hif2a, but not Hif1a, genetically suppressed both the polycythemia and pulmonary hypertension in the Vhl^R/R mice. Furthermore, Hif2a heterozygosity resulted in partial protection against vascular remodeling, hemorrhage, and edema, but not inflammation, in Vhl^R/R lungs, suggesting a selective role for HIF-2α in the pulmonary pathology and thereby providing insight into the mechanisms underlying pulmonary hypertension. These findings strongly support a dependency of the Chuvash phenotype on HIF-2α and suggest potential treatments for Chuvash patients.     

PGC-1α promotes recovery after acute kidney injury during systemic inflammation in mice

Sepsis-associated acute kidney injury (AKI) is a common and morbid condition that is distinguishable from typical ischemic renal injury by its paucity of tubular cell death. The mechanisms underlying renal dysfunction in individuals with sepsis-associated AKI are therefore less clear. Here we have shown that endotoxemia reduces oxygen delivery to the kidney, without changing tissue oxygen levels, suggesting reduced oxygen consumption by the kidney cells. Tubular mitochondria were swollen, and their function was impaired. Expression profiling showed that oxidative phosphorylation genes were selectively suppressed during sepsis-associated AKI and reactivated when global function was normalized. PPARγ coactivator-1α (PGC-1α), a major regulator of mitochondrial biogenesis and metabolism, not only followed this pattern but was proportionally suppressed with the degree of renal impairment. Furthermore, tubular cells had reduced PGC-1α expression and oxygen consumption in response to TNF-α; however, excess PGC-1α reversed the latter effect. Both global and tubule-specific PGC-1α-knockout mice had normal basal renal function but suffered persistent injury following endotoxemia. Our results demonstrate what we believe to be a novel mechanism for sepsis-associated AKI and suggest that PGC-1α induction may be necessary for recovery from this disorder, identifying a potential new target for future therapeutic studies.     

Evidence that vitamin D3 promotes mast cell–dependent reduction of chronic UVB-induced skin pathology in mice

Mast cell production of interleukin-10 (IL-10) can limit the skin pathology induced by chronic low-dose ultraviolet (UV)-B irradiation. Although the mechanism that promotes mast cell IL-10 production in this setting is unknown, exposure of the skin to UVB irradiation induces increased production of the immune modifying agent 1α,25-dihydroxyvitamin D₃ (1α,25[OH]₂D₃). We now show that 1α,25(OH)₂D₃ can up-regulate IL-10 mRNA expression and induce IL-10 secretion in mouse mast cells in vitro. To investigate the roles of 1α,25(OH)₂D₃ and mast cell vitamin D receptor (VDR) expression in chronically UVB-irradiated skin in vivo, we engrafted the skin of genetically mast cell–deficient WBB6F₁-Kit^W/W-v mice with bone marrow–derived cultured mast cells derived from C57BL/6 wild-type or VDR^−/− mice. Optimal mast cell–dependent suppression of the inflammation, local production of proinflammatory cytokines, epidermal hyperplasia, and epidermal ulceration associated with chronic UVB irradiation of the skin in Kit^W/W-v mice required expression of VDR by the adoptively transferred mast cells. Our findings suggest that 1α,25(OH)₂D₃/VDR-dependent induction of IL-10 production by cutaneous mast cells can contribute to the mast cell’s ability to suppress inflammation and skin pathology at sites of chronic UVB irradiation.In humans and mice, chronic UVB irradiation (wavelengths; 290–320 nm) induces cutaneous inflammation, photoaging, and gene mutations that can lead to the development of malignancies at affected sites (Kligman, 1996; Melnikova and Ananthaswamy, 2005). Although factors that promote UV-induced innate responses have been investigated intensively (Melnikova and Ananthaswamy, 2005), less is known about regulatory mechanisms that can limit UVB-induced inflammation.Although mast cells were once thought to function primarily as proinflammatory effector cells that can contribute to either allergic reactions (Galli et al., 2005) or inflammation caused by innate responses to exogenous environmental agents such as UV irradiation (Metz et al., 2006), recent evidence indicates that, in certain settings, mast cells can also limit inflammation and tissue injury, including that induced by UVB irradiation (Grimbaldeston et al., 2007; Galli et al., 2008). Mast cells are well positioned to participate in cutaneous immune responses, as skin mast cells are located predominately in the dermis near blood and lymphatic vessels and nerves. Although the majority of UVB rays do not penetrate to the dermis (Noonan and De Fabo, 1992), the UVB-dependent activation of dermal mast cells is thought to be achieved indirectly, e.g., via nerve growth factor derived from the epidermis (Hart et al., 2002; Townley et al., 2002) or by sensory C fibers (which innervate the epidermis and extend into the dermis) that have been activated by cis-urocanic acid (Hart et al., 1999, 2002; Khalil et al., 2001). Such indirect mechanisms of mast cell activation contribute to the UVB-induced systemic immunosuppression of acquired immune responses in mice subjected to a single acute dose (12 kJ/m²) of UVB irradiation (Hart et al., 1998, 2002). However, it is not clear how mast cells are activated when the skin is exposed to multiple cycles of UVB irradiation.We investigated whether vitamin D₃ (or cholecalciferol), which has important effects in immunity (van Etten and Mathieu, 2005; Moro et al., 2008), might influence mast cell function during chronic low-dose UVB irradiation. Vitamin D₃ is synthesized in the epidermis upon UVB-induced photochemical conversion of 7-dehydrocholesterol to previtamin D₃, which then undergoes spontaneous thermal isomerization to vitamin D₃ (Lehmann, 2005; Moro et al., 2008). Vitamin D₃ is converted into its biologically active metabolite, 1α,25-dihydroxyvitamin D₃ (1α,25[OH]₂D₃) via the 25-hydroxylation of vitamin D₃ by the cytochrome P450 proteins CYP2DII, CYP2D25, CYP3A4, or CYP2R1, followed by the hydroxylation of 25(OH)D₃ to 1α,25(OH)₂D₃ by CYP27B1 (Lehmann, 2005; Bouillon et al., 2008; Moro et al., 2008).Recently, we reported evidence that mast cell production of the antiinflammatory cytokine IL-10 can limit the inflammation and tissue damage associated with chronic low-dose UVB irradiation of mouse skin (Grimbaldeston et al., 2007). UVB irradiation can increase production of vitamin D₃, and mouse mast cells can express receptors for vitamin D₃ (i.e., vitamin D receptor [VDR]; Baroni et al., 2007). We therefore investigated whether 1α,25(OH)₂D₃ can activate mast cells to produce IL-10 and to what extent effects mediated via mast cell VDRs contribute to the mast cell’s ability to curtail the skin pathology associated with chronic low-dose UVB irradiation.     

Plasma exchange—A useful adjunct therapy to red cell exchange in patients with sickle cell disease and multiorgan dysfunction

Background

Urgent red cell exchange (RBCx) is indicated for many complications of sickle cell disease (SCD), including acute chest syndrome, stroke, and hepatic/splenic sequestration. Many who receive RBCx remain hospitalized and develop further complications, including multiple organ dysfunction syndrome (MODS), a leading cause of death in intensive care units. Therapeutic plasma exchange (TPE) has been advocated as an effective treatment of MODS, but its role in SCD compared with RBCx alone is not well studied.

Methods

We identified all ICU encounters from 2013 to 2019 involving RBCx procedures for MODS or SCD crisis that progressed to MODS, a total of 12 encounters. Data regarding hospital length of stay (LOS), survival, number of TPE procedures following RBCx, and procedure characteristics were collected. Surrogate laboratory markers of end-organ damage and disease severity scores were recorded at the time of admission, post-RBCx, post-TPE, and at discharge.

Results

Eight encounters involved RBCx followed by TPE (TPE group) while four involved RBCx alone (RBCx group). The TPE group had a higher SOFA score at ICU admission (9.5 vs. 7.0), greater predicted mortality, and a statistical trend toward higher disease severity scores following RBCx relative to the RBCx group (p = 0.10). The TPE group showed a significantly greater decrease in SOFA score between RBCx and discharge (p = 0.04). No significant difference in mortality or hospital LOS was observed between the groups.

Conclusion

The findings suggest TPE may be considered as an adjunct treatment for patients with acute complications of SCD that progress to MODS, especially in cases where there is no significant improvement following RBCx.     

The RAGE axis in systemic inflammation, acute lung injury and myocardial dysfunction: an important therapeutic target?

Background  

The sepsis syndromes, frequently complicated by pulmonary and cardiac dysfunction, remain a major cause of death amongst the critically ill. Targeted therapies aimed at ameliorating the systemic inflammation that characterises the sepsis syndromes have largely yielded disappointing results in clinical trials. Whilst there are many potential reasons for lack of success of clinical trials, one possibility is that the pathways targeted, to date, are only modifiable very early in the course of the illness. More recent approaches have therefore attempted to identify pathways that could offer a wider therapeutic window, such as the receptor for advanced glycation end-products (RAGE) and its ligands.