Endolysosome Iron Chelation Inhibits HIV-1 Protein-Induced Endolysosome De-Acidification-Induced Increases in Mitochondrial Fragmentation, Mitophagy, and Cell Death.

People with human immunodeficiency virus-1 (PLWH) experience high rates of HIV-1-associated neurocognitive disorders (HANDs); clinical symptoms range from being asymptomatic to experiencing HIV-associated dementia. Antiretroviral therapies have effectively prolonged the life expectancy related to PLWH; however, the prevalence of HANDs has increased. Implicated in the pathogenesis of HANDs are two HIV-1 proteins, transactivator of transcription (Tat) and gp120; both are neurotoxic and damage mitochondria. The thread-like morphological features of functional mitochondria become fragmented when levels of reactive oxygen species (ROS) increase, and ROS can be generated via Fenton-like chemistry in the presence of ferrous iron (Fe2+). Endolysosomes are central to iron trafficking in cells and contain readily releasable Fe2+ stores. However, it is unclear whether the endolysosome store is sufficient to account for insult-induced increases in levels of ROS, mitochondrial fragmentation, autophagy, and cell death. Using U87MG astrocytoma and SH-SY5Y neuroblastoma cells, we determined that chloroquine (CQ), Tat, and gp120 all (1) de-acidified endolysosomes, (2) decreased endolysosome numbers and increased endolysosome sizes, (3) increased mitochondrial numbers (fragmentation), (4) increased autophagosome numbers, (5) increased autolysosome numbers, (6) increased mitochondrial fragments within endolysosomes, and (7) increased cell death. These effects were all blocked by the endolysosome-specific iron chelator deferoxamine (DFO). Thus, the endolysosome de-acidification-induced release of endolysosome Fe2+ is sufficient to account for inter-organellar signaling events and cell biology consequences of HIV-1 proteins, including mitochondrial fragmentation, autophagy, and cell death.

HIV-1 Tat endocytosis and retention in endolysosomes affects HIV-1 Tat-induced LTR transactivation in astrocytes.

The presence of latent HIV-1 reservoirs in the periphery and brain represents a major obstacle to curing HIV-1 infection. As an essential protein for HIV-1 viral replication, HIV-1 Tat, mostly intracellular, has been implicated in latent HIV-1 infection. From HIV-1 infected cells, HIV-1 Tat is actively secreted and bystander cells uptake the released Tat whereupon it is endocytosed and internalized into endolysosomes. However, to activate the HIV-1 LTR promoter and increase HIV-1 replication, HIV-1 Tat must first escape from the endolysosomes and then enter the nucleus. Here, we tested the hypothesis that HIV-1 Tat can accumulate in endolysosomes and contribute to the activation of latent HIV-1 in astrocytes. Using U87MG astrocytoma cells expressing HIV-1 LTR-driven luciferase and primary human astrocytes we found that exogenous HIV-1 Tat enters endolysosomes, resides in endolysosomes for extended periods of time, and induces endolysosome de-acidification as well as enlargement. The weak base chloroquine promoted the release of HIV-1 Tat from endolysosomes and induced HIV-1 LTR transactivation. Similar results were observed by activating endolysosome Toll-like receptor 3 (TLR3) and TLR7/8. Conversely, pharmacological block of TLRs and knocking down expression levels of TLR3 and TLR7, but not TLR8, prevented endolysosome leakage and attenuated HIV-1 Tat-mediated HIV-1 LTR transactivation. Our findings suggest that HIV-1 Tat accumulation in endolysosomes may play an important role in controlling HIV-1 transactivation.

B7-H4(B7x)-Mediated Cross-talk between Glioma-Initiating Cells and Macrophages via the IL6/JAK/STAT3 Pathway Lead to Poor Prognosis in Glioma Patients.

PURPOSE: The objective of this study was to evaluate clinical significance and immunosuppressive mechanisms of B7-H4 (B7x/B7S1), a B7 family member, in glioma. EXPERIMENTAL DESIGN: B7-H4 levels in glioma tissue/cerebral spinal fluid (CSF) were compared between different grades of glioma patients. Survival data were analyzed with Kaplan-Meier to determine the prognostic value of B7-H4. Cytokines from CD133(+) cells to stimulate the expression of B7-H4 on human macrophages (Mφs) were investigated by FACS, neutralizing antibodies, and Transwell chemotaxis assay. shRNA, reporter vector, and chromatin immunoprecipitation were used to determine the binding of STAT3 to the B7-H4 promoter. The function of B7-H4(+) Mφs in vitro was evaluated through phagocytosis, T-cell proliferation/apoptosis, and cytokine production as well as in the xenografted model for in vivo analysis. RESULTS: We found that B7-H4 expression in tumors was associated with prognosis of human glioblastoma and correlated directly with malignant grades. Mechanistically, glioma initiating CD133(+) cells and Mφs/microglia cointeraction activated expression of B7-H4 via IL6 and IL10 in both tumor cells and microenvironment supporting cells. IL6-activated STAT3 bound to the promoter of B7-H4 gene and enhanced B7-H4 expression. Furthermore, CD133(+) cells mediated immunosuppression through B7-H4 expression on Mφs/microglia by silencing of B7-H4 expression on these cells, which led to increased microenvironment T-cell function and tumor regression in the xenograft glioma mouse model. CONCLUSIONS: We have identified B7-H4 activation on Mφs/microglia in the microenvironment of gliomas as an important immunosuppressive event blocking effective T-cell immune responses. Clin Cancer Res; 22(11); 2778-90. ©2016 AACR.

B7-H3 and B7-H1 expression in cerebral spinal fluid and tumor tissue correlates with the malignancy grade of glioma patients.

The B7 family consists of activating and inhibitory molecules that regulate immune responses. Recent research demonstrated the roles of soluble B7-H3 (sB7-H3) and soluble B7-H1 (sB7-H1) in the blood serum of various tumors; however, none of these studies investigated the expression of these proteins in the cerebral spinal fluid (CSF) and blood serum of patients with glioma. The aim of the present study was to investigate the expression of B7-H3 and B7-H1 in the CSF, blood serum and tissues of patients with glioma and their correlation with clinicopathological data. Between January 2012 and November 2012, samples were obtained from 78 patients with glioma, four CSF samples were obtained from patients with a moderate traumatic brain injury, four brain tissue samples were obtained from patients with a traumatic brain injury and 40 blood serum samples were obtained from healthy individuals. The expression of B7-H3 and B7-H1 in the CSF, blood serum and tumor samples of the patients with high-grade glioma was found to be higher than that in the patients with low-grade glioma. However, no significant differences in sB7-H3 and sB7-H1 expression were observed in the blood serum of the patients with glioma compared with the healthy control subjects. In addition, the expression of sB7-H3 and sB7-H1 in the CSF of the patients with glioma was higher than that in the CSF of the patients with a moderate traumatic brain injury. Furthermore, in the patients with glioma, B7-H3 and B7-H1 expression in the CSF and tumor tissue, although not in the blood serum, correlated with the glioma grade.

Magnetic resonance imaging characteristics of primary intraspinal peripheral primitive neuroectodermal tumour.

PURPOSE: The purpose of our study was to describe the salient magnetic resonance imaging (MRI) findings in primary intraspinal peripheral primitive neuroectodermal tumour (PNET). METHODS: A retrospective review of the clinical and MRI images of 7 pathologically proven cases of intraspinal peripheral PNETs was performed. The various parameters, such as vertebral level of involvement; tumour location, size, focality, and margin; signal intensity of the lesion; the presence of hemorrhage or calcification; any signal voids; assessment of the adjacent cord for cord compression; cord dilatation; the presence of paraspinal tissue mass; or vertebral or other bony changes, were analysed. RESULTS: All 7 patients had lesions in the thoracolumbar region. Three patients had extradural lesions, 4 had intradural extramedullary lesions, and none had intramedullary lesions. Six lesions were well circumscribed. Only 1 patient had multifocal involvement. All lesions were of hypointense or isointense signal on T1-weighted imaging, whereas all but one were hyperintense on T2-weighted imaging. Lesions enhanced heterogeneously except 1 intradural extramedullary lesion, which enhanced homogeneously. A paraspinal mass was noticed in 2 patients. Vertebral collapse was present in 1 patients. CONCLUSION: Intraspinal peripheral PNETs are rare spinal tumours. Although imaging characteristics are not specific, a focal circumscribed lesion in a young individual at the intramedullary, extramedullary intradural, or extradural spinal location that shows hypointense and hyperintense signal on T1- and T2-weighted images, respectively, requires PNET to be considered in the differentials.