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Oxidative stress alters mitochondrial bioenergetics and modifies pancreatic cell death independently of cyclophilin D, resulting in an apoptosis-to-necrosis shift.

ABSTRACT: Mitochondrial dysfunction lies at the core of acute pancreatitis (AP). Diverse AP stimuli induce Ca²⁺-dependent formation of the mitochondrial permeability transition pore (MPTP), a solute channel modulated by cyclophilin D (CypD), the formation of which causes ATP depletion and necrosis. Oxidative stress reportedly triggers MPTP formation and is elevated in clinical AP, but how reactive oxygen species influence cell death is unclear. Here, we assessed potential MPTP involvement in oxidant-induced effects on pancreatic acinar cell bioenergetics and fate. H₂O₂ application promoted acinar cell apoptosis at low concentrations (1-10 ?m), whereas higher levels (0.5-1 mm) elicited rapid necrosis. H₂O₂ also decreased the mitochondrial NADH/FAD⁺ redox ratio and ??_m in a concentration-dependent manner (10 ?m to 1 mm H₂O₂), with maximal effects at 500 ?m H₂O₂ H₂O₂ decreased the basal O₂ consumption rate of acinar cells, with no alteration of ATP turnover at <50 ?m H₂O₂ However, higher H₂O₂ levels (?50 ?m) diminished spare respiratory capacity and ATP turnover, and bioenergetic collapse, ATP depletion, and cell death ensued. Menadione exerted detrimental bioenergetic effects similar to those of H₂O₂, which were inhibited by the antioxidant N-acetylcysteine. Oxidant-induced bioenergetic changes, loss of ??_m, and cell death were not ameliorated by genetic deletion of CypD or by its acute inhibition with cyclosporine A. These results indicate that oxidative stress alters mitochondrial bioenergetics and modifies pancreatic acinar cell death. A shift from apoptosis to necrosis appears to be associated with decreased mitochondrial spare respiratory capacity and ATP production, effects that are independent of CypD-sensitive MPTP formation.

SUBMITTER: Armstrong JA

PROVIDER: S-EPMC5971444 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Oxidative stress alters mitochondrial bioenergetics and modifies pancreatic cell death independently of cyclophilin D, resulting in an apoptosis-to-necrosis shift.

Armstrong Jane A JA Cash Nicole J NJ Ouyang Yulin Y Morton Jack C JC Chvanov Michael M Latawiec Diane D Awais Muhammad M Tepikin Alexei V AV Sutton Robert R Criddle David N DN

The Journal of biological chemistry 20180406 21

Mitochondrial dysfunction lies at the core of acute pancreatitis (AP). Diverse AP stimuli induce Ca<sup>2+</sup>-dependent formation of the mitochondrial permeability transition pore (MPTP), a solute channel modulated by cyclophilin D (CypD), the formation of which causes ATP depletion and necrosis. Oxidative stress reportedly triggers MPTP formation and is elevated in clinical AP, but how reactive oxygen species influence cell death is unclear. Here, we assessed potential MPTP involvement in ox ...[more]

PMID: 29626097

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Project description:Linker of nucleoskeleton and cytoskeleton (LINC) complexes connect the nucleus to the cytoskeleton in eukaryotic cells. We previously reported that the overexpression of SUN2, an inner nuclear membrane protein and LINC complex component, inhibits HIV infection between the steps of reverse transcription and nuclear import in a capsid-specific manner. We also reported that SUN2 silencing does not modulate HIV infection in several cell lines. Silencing of SUN2 was recently reported to decrease HIV infection of CD4 T cells, an effect which was suggested to result from modulation of cyclophilin A (CypA)-dependent steps of HIV infection. We confirm here that HIV infection of primary CD4 T cells is compromised in the absence of endogenous SUN2, and we extend these findings to additional viral strains. However, we find that CypA is not required for the decreased infection observed in SUN2-silenced cells and, conversely, that endogenous SUN2 is not required for the well-documented positive modulation of HIV infection by CypA. In contrast, CD4 T cells lacking SUN2 exhibit a considerable defect in proliferative capacity and display reduced levels of activation markers and decreased viability. Additionally, SUN2-silenced CD4 T cells that become infected support reduced levels of viral protein expression. Our results demonstrate that SUN2 is required for the optimal activation and proliferation of primary CD4 T cells and suggest that the disruption of these processes explains the contribution of endogenous SUN2 to HIV infection in primary lymphocytes.IMPORTANCE Linker of nucleoskeleton and cytoskeleton (LINC) complexes connect the nucleus to the cytoskeleton. We previously reported that the overexpression of the LINC complex protein SUN2 inhibits HIV infection by targeting the viral capsid and blocking infection before the virus enters the nucleus. A recent report showed that the depletion of endogenous SUN2 in primary CD4 T cells results in decreased HIV infection and that this involves cyclophilin A (CypA), a host protein that interacts with the capsid of HIV to promote infection. We confirm that HIV infection is reduced in CD4 T cells lacking SUN2, but we find no role for CypA. Instead, SUN2 silencing results in CD4 T cells with decreased viability and much lower proliferation rates. Our results show that SUN2 is required for optimal CD4 T cell activation and proliferation and explain the reduced level of HIV infection in the absence of SUN2.

Project description:Pathological stimuli, such as bacterial activity, dental bleaching, and nonpolymerized resin monomers, can cause death of dental pulp cells (DPCs) through oxidative stress- (OS-) induced mitochondrial dysfunction. However, the crucial molecular mechanisms that mediate such a phenomenon remain largely unknown. OS is characterized by the overproduction of reactive oxygen species (ROS), e.g., H2O2, O2 -, and ·OH. Mitochondria are a major source of ROS and the principal attack target of ROS. Cyclophilin D (CypD), as the only crucial protein for mitochondrial permeability transition pore (mPTP) induction, facilitates the opening of mPTP and causes mitochondrial dysfunction, leading to cell death. In the present study, we hypothesized that CypD-mediated mitochondrial molecular pathways were closely involved in the process of OS-induced death of human DPCs (HDPCs). We tested the phenotypic and molecular changes of HDPCs in a well-established OS model-H2O2 treatment. We showed that H2O2 dramatically reduced the viability and increased the death of HDPCs in a time- and dose-dependent manner by performing MTT, flow cytometry, and TUNEL assays and quantifying the expression changes of Bax and Bcl-2 proteins. H2O2 also induced mitochondrial dysfunction, as reflected by the increased mitochondrial ROS, reduced ATP production, and activation of mPTP (decreased mitochondrial membrane potential and enhanced intracellular Ca2+ level). An antioxidant (N-acetyl-L-cysteine) effectively preserved mitochondrial function and significantly attenuated H2O2-induced cytotoxicity and death. Moreover, H2O2 treatment markedly upregulated the CypD protein level in HDPCs. Notably, genetic or pharmacological blockade of CypD significantly attenuated H2O2-induced mitochondrial dysfunction and cell death. These findings provided novel insights into the role of a CypD-dependent mitochondrial pathway in the H2O2-induced death in HDPCs, indicating that CypD may be a potential therapeutic target to prevent OS-mediated injury in dental pulp.

Dataset Information

Oxidative stress alters mitochondrial bioenergetics and modifies pancreatic cell death independently of cyclophilin D, resulting in an apoptosis-to-necrosis shift.

Publications

Oxidative stress alters mitochondrial bioenergetics and modifies pancreatic cell death independently of cyclophilin D, resulting in an apoptosis-to-necrosis shift.

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