The increased mitochondrial DNA damage leads to altered functional capacities of retinal pigment epithelial (RPE) cells. A previous study showed the increased autophagy in RPE cells caused by low concentrations of rotenone, a selective inhibitor of mitochondrial complex I. However, the mechanism by which autophagy regulates RPE cell death is still unclear. In the present study, we examined the mechanism underlying the regulation of RPE cell death through the inhibition of mitochondrial complex I. We report herein that rotenone induced mitotic catastrophe (MC) in RPE cells. We further observed an increased level of autophagy in the RPE cells undergoing MC (RPE-MC cells). Importantly, autophagy inhibition induced nonapoptotic cell death in RPE-MC cells. These findings indicate that autophagy has a pivotal role in the survival of RPE-MC cells. We next observed PINK1 accumulation in the mitochondrial membrane and parkin translocation into the mitochondria from the cytosol in the rotenone-treated RPE-MC cells, which indicates that increased mitophagy accompanies MC in ARPE-19 cells. Noticeably, the mitophagy also contributed to the cytoprotection of RPE-MC cells. Although there might be a significant gap in the roles of autophagy and mitophagy in the RPE cells in vivo, our in vitro study suggests that autophagy and mitophagy presumably prevent the RPE-MC cells from plunging into cell death, resulting in the prevention of RPE cell loss.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hematopoietic cytokine that stimulates the differentiation and function of hematopoietic cells. GM-CSF has been implicated in nervous system function. The goal of the present study was to understand the effects of hypoxia-induced GM-CSF on neural stem cells (NSCs) in a model of spinal cord injury (SCI). GM-CSF-overexpressing NSCs were engineered utilizing a hypoxia-inducible gene expression plasmid, including an Epo enhancer ahead of an SV promoter (EpoSV-GM-CSF). Cells were then subjected to hypoxia (pO 2 , 1%) or a hypoxia-mimicking reagent (CoCl 2 ) in vitro. The progression of time of GM-CSF expression was tracked in EpoSV-GM-CSF-transfected NSCs. Overexpression of GM-CSF in undifferentiated and differentiated NSCs created resistance to H 2 O 2 -induced apoptosis in hypoxia. NSCs transfected with EpoSV-GM-CSF or SV-GM-CSF were transplanted into rats after SCI to assess the effect of GM-CSF on NSC survival and restoration of function. Moreover, a significantly higher amount of surviving NSCs and neuronal differentiation was observed in the EpoSV-GM-CSF-treated group. Significant improvement in locomotor function was also found in this group. Thus, GM-CSF overexpression by the Epo enhancer in hypoxia was beneficial to transplanted NSC survival and to behavioral improvement, pointing toward a possible role for GM-CSF in the treatment of SCI.

Abstract. The Bcl-2 protein is known to exert not only antiapoptotic but also anti-autophagic activities. Numerous studies have demonstrated that etoposide, which is one of the most widely used cancer chemotherapy agents, induces apoptotic cell death. However, the exact molecular mechanism leading to cell death by etoposide remains to be resolved. This study aimed to dissect the mode of cell death induced by etoposide in Hep3B hepatoma cells. Furthermore, this study was conducted to examine whether etoposide overcomes the resistance conferred by Bcl-2 in Hep3B hepatoma cells. We observed that Hep3B cells treated with etoposide show not only apoptotic but autophagic phenotypes. Autophagy inhibition by 3-methyladenine (3MA) and caspase inhibition by zVAD-fmk effectively decreased autophagic and apoptotic phenotypes, respectively. However, either zVAD-fmk or 3MA only partially prevented cell death. These data indicate that etoposide concomitantly induces autophagic cell death and apoptosis in Hep3B cells. Importantly, etoposide can effectively induce cell death in Bcl-2-overexpressing Hep3B cells. Conversely, staurosporine, which exclusively induces apoptosis in Hep3B cells, did not efficiently induce cell death in Bcl-2-overexpressing Hep3B cells. Staurosporine-treated Hep3B cells also showed an autophagic phenotype. While autophagy is cell death-inducing in Hep3B cells treated with etoposide, it is cytoprotective in Hep3B cells treated with staurosporine. To this end, we observed that etoposide-induced mixed type of programmed cell death is associated with the dissociation of Bcl-2 from Beclin-1. Taken together, etoposide induces a mixed type of programmed cell death and overcomes the resistance conferred by Bcl-2 in Hep3B hepatoma cells.

ABSTRACT:Here we report the successful treatment of acute antibody-mediated rejection (AMR) with bortezomib. Bortezomib rescue treatment was administered after a 42-year-old woman failed to respond to steroid pulse and plasmapheresis with intravenous immunoglobulin (IVIG). The patient underwent a second renal transplantation with a deceased donor kidney. She was treated pre-operatively with rituximab (200 mg/body) and underwent plasmapheresis twice (day-1 and operation day) because ELISA screening revealed that her pre-operative peak panel reactive antibody (PRA) composition was 100% class I and 100% class II and 15 times of cross-match positive history during the waiting period for transplantation. The patients received induction therapy with Simulect (an IL-2-blocking agent). A 1-hour protocol biopsy revealed C4d-positivity and mild peritubular capillary inflammation. This was suggestive of early AMRassociated changes. After transplantation, the patient underwent plasmaphereses (nine times) with low-dose IVIG (2 mg/kg). Despite this treatment regimen, serum creatinine levels increased to 3.4 mg/dL on posttransplant day 15. A second graft biopsy was performed, which showed overt AMR with glomerulitis, peritubular capillary inflammation and no C4d deposition. On post-operative day (POD) 22, treatment with four doses of bortezomib (1.3 mg/m 2 ) was initiated with the patient's consent. On POD 55, renal function had recovered and serum creatinine was 1.5 mg/dL. In summary, bortezomib was administered as a rescue treatment for a patient who developed AMR that was refractory to a combination of plasmaphereses with low-dose IVIG and preemptive administration of rituximab.