RESUMO
Mitochondrial stress releases mitochondrial DNA (mtDNA) into the cytosol, thereby triggering the type Ι interferon (IFN) response. Mitochondrial outer membrane permeabilization, which is required for mtDNA release, has been extensively studied in apoptotic cells, but little is known about its role in live cells. We found that oxidatively stressed mitochondria release short mtDNA fragments via pores formed by the voltage-dependent anion channel (VDAC) oligomers in the mitochondrial outer membrane. Furthermore, the positively charged residues in the N-terminal domain of VDAC1 interact with mtDNA, promoting VDAC1 oligomerization. The VDAC oligomerization inhibitor VBIT-4 decreases mtDNA release, IFN signaling, neutrophil extracellular traps, and disease severity in a mouse model of systemic lupus erythematosus. Thus, inhibiting VDAC oligomerization is a potential therapeutic approach for diseases associated with mtDNA release.
Assuntos
DNA Mitocondrial/metabolismo , Lúpus Eritematoso Sistêmico/metabolismo , Membranas Mitocondriais/metabolismo , Multimerização Proteica , Canais de Ânion Dependentes de Voltagem/metabolismo , Animais , Modelos Animais de Doenças , Endodesoxirribonucleases/genética , Humanos , Interferons/metabolismo , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Camundongos , Estresse Oxidativo , Domínios Proteicos , Multimerização Proteica/efeitos dos fármacos , Ratos , Canais de Ânion Dependentes de Voltagem/antagonistas & inibidores , Canais de Ânion Dependentes de Voltagem/genéticaRESUMO
It was previously shown that 14-3-3 η is overexpressed in the synovial fluid of patients with joint inflammation, which is often associated with growth failure. In this study, we investigated the role of 14-3-3 η in chondrogenesis using ATDC5 cells. Upon treatment with TNF-α, cells overexpressed 14-3-3 η with inhibition of chondrogenesis. Chondrogenesis was also inhibited by overexpression of 14-3-3 η without TNF-α treatment, whereas silencing of 14-3-3 η promoted chondrogenic differentiation. Further, G1 phase arrest was inhibited by overexpression of 14-3-3 η. In summary, we suggest that 14-3-3 η plays a regulatory role in chondrogenic differentiation.
Assuntos
Proteínas 14-3-3/fisiologia , Diferenciação Celular , Condrócitos/fisiologia , Condrogênese , Proteínas 14-3-3/genética , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular Tumoral , Condrócitos/citologia , Condrócitos/metabolismo , Condrogênese/efeitos dos fármacos , Condrogênese/genética , Fase G1/efeitos dos fármacos , Insulina/farmacologia , Camundongos , RNA Interferente Pequeno/genética , Fator de Necrose Tumoral alfa/farmacologia , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
Although generation of reactive oxygen species (ROS) by NADPH oxidases (Nox) is thought to be important for signal transduction in nonphagocytic cells, little is known of the role ROS plays in chondrogenesis. We therefore examined the possible contribution of ROS generation to chondrogenesis using both ATDC5 cells and primary chondrocytes derived from mouse embryos. The intracellular level of ROS was increased during the differentiation process, which was then blocked by treatment with the ROS scavenger N-acetylcysteine. Expression of Nox1 and Nox2 was increased upon differentiation of ATDC5 cells and primary mouse chondrocytes, whereas that of Nox4, which was relatively high initially, was decreased gradually during chondrogenesis. In developing limb, Nox1 and Nox2 were highly expressed in prehypertrophic and hypertrophic chondrocytes. However, Nox4 was highly expressed in proliferating chondrocytes and prehypertrophic chondrocytes. Depletion of Nox2 or Nox4 expression by RNA interference blocked both ROS generation and differentiation of ATDC5 cells, whereas depletion of Nox1 had no such effect. We also found that ATDC5 cells depleted of Nox2 or Nox4 underwent apoptosis. Further, inhibition of Akt phosphorylation along with subsequent activation of ERK was observed in the cells. Finally, depletion of Nox2 or Nox4 inhibited the accumulation of proteoglycan in primary chondrocytes. Taken together, our data suggest that ROS generated by Nox2 or Nox4 are essential for survival and differentiation in the early stage of chondrogenesis.
