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We have established a novel in vitro co-culture system of human brain endothelial cells (HBEC), Plasmodium falciparum parasitised red blood cells (iRBC) and peripheral blood mononuclear cells (PBMC), in order to simulate the chief pathophysiological lesion in cerebral malaria (CM). This approach has revealed a previously unsuspected pro-inflammatory role of the endothelial cell through potentiating the production of interferon (IFN)-γ by PBMC and concurrent reduction of interleukin (IL)-10. The IFN-γ increased the expression of CXCL10 and intercellular adhesion molecule (ICAM)-1, both of which have been shown to be crucial in the pathogenesis of CM. There was a shift in the ratio of IL-10:IFN-γ protein from >1 to

Substance P (SP) is a prototypical neuropeptide with roles in pain and inflammation. Numerous mechanisms regulate endogenous SP levels, including the differential expression of SP mRNA and the controlled secretion of SP from neurons. Proteolysis has long been suspected to regulate extracellular SP concentrations but data in support of this hypothesis is scarce. Here, we provide evidence that proteolysis controls SP levels in the spinal cord. Using peptidomics to detect and quantify endogenous SP fragments, we identify the primary SP cleavage site as the C-terminal side of the ninth residue of SP. If blocking this pathway increases SP levels, then proteolysis controls SP concentration. We performed a targeted chemical screen using spinal cord lysates as a proxy for the endogenous metabolic environment and identified GM6001 (galardin, ilomastat) as a potent inhibitor of the SP(1-9)-producing activity present in the tissue. Administration of GM6001 to mice results in a greater-than-three-fold increase in the spinal cord levels of SP, which validates the hypothesis that proteolysis controls physiological SP levels.

We investigated the relationship between gut health, visceral fat dysfunction and metabolic disorders in diet-induced obesity. C57BL/6J mice were fed control or high saturated fat diet (HFD). Circulating glucose, insulin and inflammatory markers were measured. Proximal colon barrier function was assessed by measuring transepithelial resistance and mRNA expression of tight-junction proteins. Gut microbiota profile was determined by 16S rDNA pyrosequencing. Tumor necrosis factor (TNF)-α and interleukin (IL)-6 mRNA levels were measured in proximal colon, adipose tissue and liver using RT-qPCR. Adipose macrophage infiltration (F4/80⁺) was assessed using immunohistochemical staining. HFD mice had a higher insulin/glucose ratio (P = 0.020) and serum levels of serum amyloid A3 (131%; P = 0.008) but reduced circulating adiponectin (64%; P = 0.011). In proximal colon of HFD mice compared to mice fed the control diet, transepithelial resistance and mRNA expression of zona occludens 1 were reduced by 38% (P