Cyclosporine A Induces Apoptotic and Autophagic Cell Death in Rat Pituitary GH3 Cells

Abstract

Cyclosporine A (CsA) is a powerful immunosuppressive drug with side effects including the development of chronic nephrotoxicity. In this study, we investigated CsA treatment induced apoptotic and autophagic cell death in pituitary GH3 cells. CsA treatment (0.1 to 10 µM) decreased survival of GH3 cells in a dose-dependent manner. Cell viability decreased significantly with increasing CsA concentrations largely due to an increase in apoptosis, while cell death rates due to autophagy altered only slightly. Several molecular and morphological features correlated with cell death through these distinct pathways. At concentrations ranging from 1.0 to 10 µM, CsA induced a dose-dependent increase in expression of the autophagy markers LC3-I and LC3-II. Immunofluorescence staining revealed markedly increased levels of both LC3 and lysosomal-associated membrane protein 2 (Lamp2), indicating increases in autophagosomes. At the same CsA doses, apoptotic cell death was apparent as indicated by nuclear and DNA fragmentation and increased p53 expression. In apoptotic or autophagic cells, p-ERK levels were highest at 1.0 µM CsA compared to control or other doses. In contrast, Bax levels in both types of cell death were increased in a dose-dependent manner, while Bcl-2 levels showed dose-dependent augmentation in autophagy and were decreased in apoptosis. Manganese superoxide dismutase (Mn-SOD) showed a similar dose-dependent reduction in cells undergoing apoptosis, while levels of the intracellular calcium ion exchange maker calbindin-D9k were decreased in apoptosis (1.0 to 5 µM CsA), but unchanged in autophagy. In conclusion, these results suggest that CsA induction of apoptotic or autophagic cell death in rat pituitary GH3 cells depends on the relative expression of factors and correlates with Bcl-2 and Mn-SOD levels.

Autophagy and the Kidney: Implications for Ischemia-Reperfusion Injury and Therapy

“…Several diuretics (furosemide) and immunosuppressive drugs (rapamycin, tacrolimus, MMF, and corticosteroids) used in the setting of AKI and kidney transplantation, respectively, are autophagy modulators (Table 2), [49][50][51][52][53][54][55][56][57][58][59][60][61][62][63][64][65] which could partially explain their influences on renal IRI. Furosemide is often applied in an attempt to promote diuresis in AKI.…”

“…Interestingly, its long-term nephrotoxic effects have even been attributed to inhibition of the autophagic flux, leading to an accumulation of autophagosomes, 63 and this mechanism could account for cyclosporine Ainduced autophagy-dependent cell death observed in glioma and pituitary cell models in vitro. 64, 65 One final note is that corticosteroids also affect autophagy. Although some glucocorticoids (eg, dexamethasone) stimulate autophagy in vitro and in vivo, methylprednisolone, the most commonly used steroid in transplantation, suppresses autophagy in the rat spinal cord and thereby alleviates spinal cord injury, suggesting that methylprednisolone can counteract autophagydependent cell death.…”

Cell Death Conversion under Hypoxic Condition in Tumor Development and Therapy

“…Autophagic cell death is due to excessive autophagic degradation of intracellular contents, which is considered to be distinct from apoptosis and necrosis, lacking caspase activation, DNA fragmentation or inflammatory response [ 15 , 23 ]. Cyclosporine A (CsA), a powerful immunosuppressive drug, could induct autophagic cell death [ 24 ].…”

RecQL4 regulates autophagy and apoptosis in U2OS cells

“…General or selective autophagy is an important process in the maintenance of cellular metabolism and allows cells to survive under conditions of hypoxia, starvation, or nutrient deprivation. Several DNA repair-associated proteins, including XPA, CSA, and CSB, are located in the mitochondria and are involved in autophagy (Fang et al 2014;Kim et al 2014;Scheibye-Knudsen et al 2012). Specifically, XPA is involved in mitophagy via PARP-1 hyper activation and NAD+/SIRT1 reduction (Fang et al 2014).…”