Background: Plasmodium falciparumcan cause a diffuse encephalopathy known as cerebral malaria (CM), a major contributor to malaria associated mortality. Despite treatment, mortality due to CM can be as high as 30% while 10% of survivors of the disease may experience short- and long-term neurological complications. The pathogenesis of CM and other forms of severe malaria is multi-factorial and appear to involve cytokine and chemokine homeostasis, inflammation and vascular injury/repair. Identification of prognostic markers that can predict CM severity will enable development of better intervention.Methods: Postmortem serum and cerebrospinal fluid (CSF) samples were obtained within 2–4 hours of death in Ghanaian children dying of CM, severe malarial anemia (SMA), and non-malarial (NM) causes. Serum and CSF levels of 36 different biomarkers (IL-1β, IL-1ra, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12 (p70), IL-13, IL-15, IL-17, Eotaxin, FGF basic protein, CRP, G-CSF, GM-CSF, IFN-γ, TNF-α, IP-10, MCP-1 (MCAF), MIP-1α, MIP-1β, RANTES, SDF-1α, CXCL11 (I-TAC), Fas-ligand [Fas-L], soluble Fas [sFas], sTNF-R1 (p55), sTNF-R2 (p75), MMP-9, TGF-β1, PDGF bb and VEGF) were measured and the results compared between the 3 groups.Results: After Bonferroni adjustment for other biomarkers, IP-10 was the only serum biomarker independently associated with CM mortality when compared to SMA and NM deaths. Eight CSF biomarkers (IL-1ra, IL-8, IP-10, PDGFbb, MIP-1β, Fas-L, sTNF-R1, and sTNF-R2) were significantly elevated in CM mortality group when compared to SMA and NM deaths. Additionally, CSF IP-10/PDGFbb median ratio was statistically significantly higher in the CM group compared to SMA and NM groups.Conclusion: The parasite-induced local cerebral dysregulation in the production of IP-10, 1L-8, MIP-1β, PDGFbb, IL-1ra, Fas-L, sTNF-R1, and sTNF-R2 may be involved in CM neuropathology, and their immunoassay may have potential utility in predicting mortality in CM.

The HIV-1 Nef protein is known to be secreted, and our group has shown that Nef is secreted from nef-transfected and HIV-1-infected cells in small exosome-like vesicles (d. 40–100 nm). The role of secreted Nef remains to be fully characterized. Thus, it is important to characterize the nature of and the mechanisms regulating Nef secretion. We hypothesized that specific structural domains on the Nef protein interact with components of the endosomal trafficking machinery, sorting Nef into multivesicular bodies (MVB) and packaging it in exosome-like vesicles. To identify those domains, a series of mutants spanning the entire nef sequence were made and cloned into the expression vector pQB1, which expresses the mutants as Nef-GFP fusion proteins. These constructs were used in transient transfection assays to identify sequences necessary for secretion of the Nef-GFP fusion protein. N-terminal domains were identified as critical for Nef-induced vesicle secretion: (1) a basic cluster of four arginine residues (aa 17, 19, 21, 22), (2) the phosphofurin acidic cluster sequence (PACS; Glu62–65), and (3) a previously uncharacterized domain spanning amino acid residues 66–70 (VGFPV), which we named the secretion modification region (SMR). Additional amino acids P25, 29GVG31, and T44 were identified in HIV-1 Nef as regulating its secretion. These residues have not been associated with other reported Nef functions. The myristoylation domain, ubiquitination lysine residues, and the C-terminal portion of Nef (aa 71–206) had no effect on secretion. A minimal HIV-1 Nef sequence, comprising the identified motifs, was sufficient for Nef-induced vesicle secretion.

Human immunodeficiency virus (HIV)-infected and viremic individuals exhibit elevated levels of plasma cytokines. Here we show that most cytokines are not in free form but appear associated with exosomes that are distinct from virions. Purified exosomes were analyzed to determine the levels of 21 cytokines and chemokines and compared with exosome-depleted plasma. Most cytokines were markedly enriched in exosomes from HIV-positive individuals relative to negative controls and to plasma. Moreover, exposure of naive peripheral blood mononuclear cells to exosomes purified from HIV-positive patients induced CD38 expression on naive and central memory CD4⁺ and CD8⁺ T cells, probably contributing to inflammation and viral propagation via bystander cell activation.

The HIV/AIDS data from the national surveillance systems of China and the United States from 1985 to 2014 were compared to characterize the HIV/AIDS epidemic in both countries. The current estimated national HIV prevalence rate in China and the United States are 0.0598% and 0.348%, respectively. In the United States, the annual number of new HIV infections has remained relatively stable (~50,000 each year) and has shown a downward trend in recent years. The Chinese national HIV prevalence is still low, and new HIV infections have been contained at a low level (50,000–100,000 each year). However, the epidemic has showed an increasing trend since 2012. By risk group, in both countries, men who have sex with men (MSM), heterosexual sex, and injection drug use (IDU) are the most common modes of transmission of new HIV infections. However, in the United States, MSM is the dominant transmission route, accounting for >60% of new infections; whereas in China, heterosexual sex has now become the dominant route, also accounting for >60% of new infections. A rapid increase in the proportion of HIV cases that were attributed to MSM and an obvious decrease in the proportion of HIV cases attributed to IDU in China in recent years imply that the China’s epidemic is still evolving, to some extent, copying what was experienced in the United States. By age group, the proportions of HIV cases that were attributed to the age group 25–59 were comparable between the two countries. However, the United States had a higher proportion of cases that were attributed to age groups 15–19 and 20–24 than China, indicating that youth account for more infections in the United States. One other fact worth noting: in China there is a significant increase in the number of HIV new infections in individuals over 50 years of age, which results in much higher proportion of cases that were attributed to age groups 60–64 and over 65 in China than those in the United States. By race/ethnicity, in the United States, Blacks/African Americans continue to experience the most severe HIV burden, followed by Hispanics/Latinos. In China, no official data on race/ethnicity disparities are currently available. Thus, region, risk group, age are important factors in the HIV epidemics in both countries.

Nef is secreted from infected cells in exosomes and is found in abundance in the sera of HIV-infected individuals. Secreted exosomal Nef (exNef) induces apoptosis in uninfected CD4(+) T cells and may be a key component of HIV pathogenesis. The exosomal pathway has been implicated in HIV-1 virus release, suggesting a possible link between these two viral processes. However, the underlying mechanisms and cellular components of exNef secretion have not been elucidated. We have previously described a Nef motif, the secretion modification region (SMR; amino acids 66 to 70), that is required for exNef secretion. In silico modeling data suggest that this motif can form a putative binding pocket. We hypothesized that the Nef SMR binds a cellular protein involved in protein trafficking and that inhibition of this interaction would abrogate exNef secretion. By using tandem mass spectrometry and coimmunoprecipitation with a novel SMR-based peptide (SMRwt) that blocks exNef secretion and HIV-1 virus release, we identified mortalin as an SMR-specific cellular protein. A second set of coimmunoprecipitation experiments with full-length Nef confirmed that mortalin interacts with Nef via Nef's SMR motif and that this interaction is disrupted by the SMRwt peptide. Overexpression and microRNA knockdown of mortalin revealed a positive correlation between exNef secretion levels and mortalin protein expression. Using antibody inhibition we demonstrated that the Nef/mortalin interaction is necessary for exNef secretion. Taken together, this work constitutes a significant step in understanding the underlying mechanism of exNef secretion, identifies a novel host-pathogen interaction, and introduces an HIV-derived peptide with antiviral properties.

• Extracellular vesicles’ (EVs) role in breast tumor microenvironment and pre-metastatic niche development. • Breast cancer EV-mediated transmission of pro-metastatic and drug-resistant phenotypes. • Precision medicine with EVs as biomarkers and delivery vehicles for drug and anticancer genetic material.