Mice harboring the human SLC30A8 R138X loss-of-function mutation have increased insulin secretory capacity.

SLC30A8 encodes a zinc transporter that is primarily expressed in the pancreatic islets of Langerhans. In ß-cells it transports zinc into insulin-containing secretory granules. Loss-of-function (LOF) mutations in SLC30A8 protect against type 2 diabetes in humans. In this study, we generated a knockin mouse model carrying one of the most common human LOF mutations for SLC30A8, R138X. The R138X mice had normal body weight, glucose tolerance, and pancreatic ß-cell mass. Interestingly, in hyperglycemic conditions induced by the insulin receptor antagonist S961, the R138X mice showed a 50% increase in insulin secretion. This effect was not associated with enhanced ß-cell proliferation or mass. Our data suggest that the SLC30A8 R138X LOF mutation may exert beneficial effects on glucose metabolism by increasing the capacity of ß-cells to secrete insulin under hyperglycemic conditions.

Inflammatory mediators reduce surface PrPc on human BMVEC resulting in decreased barrier integrity.

The cellular prion protein (PrPc) is a surface adhesion molecule expressed at junctions of various cell types including brain microvascular endothelial cells (BMVEC) that are important components of the blood-brain barrier (BBB). PrPc is involved in several physiological processes including regulation of epithelial cell barrier function and monocyte migration across BMVEC. BBB dysfunction and disruption are significant events in central nervous system (CNS) inflammatory processes including HIV neuropathogenesis. Tumor necrosis factor (TNF)-α and vascular endothelial growth factor (VEGF) are two inflammatory factors that have been implicated in the processes that affect BBB integrity. To examine the effect of inflammation on PrPc expression in BMVEC, we used these mediators and found that TNF-α and VEGF decrease surface PrPc on primary human BMVEC. We also showed that these factors decrease total PrPc protein as well as mRNA, indicating that they regulate expression of this protein by de novo synthesis. To determine the effect of PrPc loss from the surface of BMVEC on barrier integrity, we used small hairpin RNAs to knockdown PrPc. We found that the absence of PrPc from BMVEC causes increased permeability as determined by a fluorescein isothiocyanate (FITC)-dextran permeability assay. This suggests that cell surface PrPc is essential for endothelial monolayer integrity. To determine the mechanism by which PrPc downregulation leads to increased permeability of an endothelial monolayer, we examined changes in expression and localization of tight junction proteins, occludin and claudin-5, and found that decreased PrPc leads to decreased total and membrane-associated occludin and claudin-5. We propose that an additional mechanism by which inflammatory factors affect endothelial monolayer permeability is by decreasing cell-associated PrPc. This increase in permeability may have subsequent consequences that lead to CNS damage.

The Role of Shed PrPc in the Neuropathogenesis of HIV Infection.

HIV-1 enters the CNS soon after peripheral infection and causes chronic neuroinflammation and neuronal damage that leads to cognitive impairment in 40-70% of HIV-infected people. The nonpathogenic cellular isoform of the human prion protein (PrPc) is an adhesion molecule constitutively expressed in the CNS. Previously, our laboratory showed that shed PrPc (sPrPc) is increased in the cerebrospinal fluid of HIV-infected people with cognitive deficits as compared with infected people with no impairment. In this article, we demonstrate that CCL2 and TNF-α, inflammatory mediators that are elevated in the CNS of HIV-infected people, increase shedding of PrPc from human astrocytes by increasing the active form of the metalloprotease ADAM10. We show that the consequence of this shedding can be the production of inflammatory mediators, because treatment of astrocytes with rPrPc increased secretion of CCL2, CXCL-12, and IL-8. Supernatants from rPrPc-treated astrocytes containing factors produced in response to this treatment, but not rPrPc by itself, cause increased chemotaxis of both uninfected and HIV-infected human monocytes, suggesting a role for sPrPc in monocyte recruitment into the brain. Furthermore, we examined whether PrPc participates in glutamate uptake and found that rPrPc decreased uptake of this metabolite in astrocytes, which could lead to neurotoxicity and neuronal loss. Collectively, our data characterize mediators involved in PrPc shedding and the effect of this sPrPc on monocyte chemotaxis and glutamate uptake from astrocytes. We propose that shedding of PrPc could be a potential target for therapeutics to limit the cognitive impairment characteristic of neuroAIDS.

Astrocyte-shed extracellular vesicles regulate the peripheral leukocyte response to inflammatory brain lesions.

Brain injury induces a peripheral acute cytokine response that directs the transmigration of leukocytes into the brain. Because this brain-to-peripheral immune communication affects patient recovery, understanding its regulation is important. Using a mouse model of inflammatory brain injury, we set out to find a soluble mediator for this phenomenon. We found that extracellular vesicles (EVs) shed from astrocytes in response to intracerebral injection of interleukin-1ß (IL-1ß) rapidly entered into peripheral circulation and promoted the transmigration of leukocytes through modulation of the peripheral acute cytokine response. Bioinformatic analysis of the protein and microRNA cargo of EVs identified peroxisome proliferator-activated receptor α (PPARα) as a primary molecular target of astrocyte-shed EVs. We confirmed in mice that astrocytic EVs promoted the transmigration of leukocytes into the brain by inhibiting PPARα, resulting in the increase of nuclear factor κB (NF-κB) activity that triggered the production of cytokines in liver. These findings expand our understanding of the mechanisms regulating communication between the brain and peripheral immune system and identify astrocytic EVs as a molecular regulator of the immunological response to inflammatory brain damage.

Protease resistant protein cellular isoform (PrP(c)) as a biomarker: clues into the pathogenesis of HAND.

HIV infection and HIV neurocognitive impairment are major global health problems. The prevalence of HIV associated neurocognitive disorders (HAND) is increasing as people with HIV are living longer due to the success of antiretroviral therapies. Our laboratory identified the soluble form of (sPrP(c)), the cellular non-pathogenic isoform of the prion protein, as a biomarker of HAND. In this review we discuss the published data addressing PrP(c) biology in normal conditions and pathologies, as well as the mechanisms of sPrP(c) shedding and secretion. Lastly, we discuss our studies that demonstrated that sPrP(c) is a biomarker of neurocognitive impairment in the HIV infected population.