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Smoking and HIV-1 infection are risk factors for COPD, which is among the most common comorbid conditions in people living with HIV-1. HIV-1 infection leads to persistent expansion of CD8+ T cells, and CD8+ T cell-mediated inflammation has been implicated in COPD pathogenesis. In this study, we investigated the effects of HIV-1 infection and smoking on T cell dynamics in patients at risk of COPD. Bronchoalveolar lavage (BAL), endobronchial brushings and blood from HIV-1 infected and uninfected non-smokers and smokers were analyzed by flow cytometry, and lungs were imaged by computed tomography. Chemokines were measured in BAL fluid, and CD8+ T cell chemotaxis in the presence of cigarette smoke extract was assessed in vitro. HIV-1 infection increased CD8+ T cells in the BAL, but this increase was abrogated by smoking. Smokers had reduced BAL levels of the T cell-recruiting chemokines CXCL10 and CCL5, and cigarette smoke extract inhibited CXCL10 and CCL5 production by macrophages and CD8+ T cell transmigration in vitro. In contrast to the BAL, CD8+ T cells in endobronchial brushings were increased in HIV-1 infected smokers, driven by an accumulation of effector memory T cells in the airway mucosa and an increase in tissue resident memory T cells. Mucosal CD8+ T cell numbers inversely correlated with lung aeration, suggesting an association with inflammation and remodeling. HIV-1 infection and smoking lead to retention of CD8+ T cells within the airway mucosa.

Neuroblastoma is the most common extracranial solid tumor of childhood. While survival rates are high for localized disease, treatment response remains poor for a subset of patients with large tumors or disseminated disease. Thus, there remains much room for improvement in treatment strategies for this disease. Using in vitro and in vivo systems, we present glycogen synthase kinase-3β (GSK-3β) inhibition as a potential mechanism to treat neuroblastoma. Using the specific GSK-3β inhibitor SB415286, we demonstrate that GSK-3β inhibition decreases the viability of Neuro-2A cells, as determined by cell proliferation assay and clonogenic survival. Moreover, we show that GSK-3β inhibition induces apoptosis in neuroblastoma cells, as determined by Annexin V staining and confirmed with DAPI staining. Using flow cytometry, we are able to demonstrate that SB415286 induces the accumulation of cells in the G2/M phase of the cell cycle. Finally, we show that these in vitro results translate into delayed tumor growth in vivo using a heterotopic tumor model in nude mice treated with SB415286. These findings suggest that GSK-3β is a potential molecular target for the treatment of neuroblastoma.

Purpose Development of new treatments is critical to effective protection against radiation-induced injury. We investigate the potential of developing small-molecule inhibitors of glycogen synthase kinase 3β (GSK-3β)—SB216763 or SB415286—as radioprotective agents to attenuate intestinal injury. Methods and Materials A survival study was done by use of C57BL/6J mice to evaluate the radioprotective effect of GSK-3β inhibitors. Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay and immunohistochemical staining for Bax and Bcl-2 were used to assess apoptosis in the small intestines of the treated mice. A clonogenic survival study, apoptosis assays (staining with annexin V or 4′,6-diamidino-2-phenylindole), and immunoblot analysis of β-catenin, Bcl-2, Bax, and caspase 3 were done by use of Rat intestinal epithelial cell line IEC-6 cells. Results Pretreatment with SB415286 significantly improved survival of mice irradiated with 8 and 12 Gy. Mice pretreated with SB216763 or SB415286 showed a significant reduction in TUNEL- and Bax-positive cells and an increase in Bcl-2–positive cells in intestinal crypts at 4 and/or 12 h after radiation with 4 and/or 8 Gy compared with radiation alone. Pretreatment of irradiated IEC-6 cells with GSK-3β inhibitors significantly increased clonogenic survival compared with cells treated with radiation alone. This increase was due to the attenuation of radiation-induced apoptosis, as shown by annexin V and 4′,6-diamidino-2-phenylindole assays, as well as immunoblot analysis of Bcl-2, Bax, and caspase 3. Conclusions Glycogen synthase kinase 3β small-molecule inhibitors protect mouse intestine from radiation-induced damage in cell culture and in vivo and improve survival of mice. Molecular mechanisms of this protection involve attenuated radiation-induced apoptosis regulated by Bcl-2, Bax, and caspase 3. Therefore GSK-3β inhibitors reduce deleterious consequences of intestinal irradiation and thereby improve quality of life during radiation therapy.