Astragalus membranaceus-Carthamus tinctorius herb pair antagonizes parthanatos in cerebral ischemia/reperfusion injury via regulating PARP-1/TAX1BP1-mediated mitochondrial respiratory chain complex I

Abstract

Ethnopharmacological relevance: The combination of Astragalus membranaceus (Huang Qi in Chinese, HQ) and Carthamus tinctorius (Hong Hua in Chinese, HH) is commonly employed for treating ischemic stroke (IS). The heavily oxidative environment of cerebral ischemia/reperfusion injury (CI/RI) promotes activation of poly (ADP-ribose) polymerase-1 (PARP-1), which initiates parthanatos, a regulated cell death mode. Reactive oxygen species (ROS) bursting in mitochondrial respiratory chain complex I (Complex I) is a key cause of CI/RI. Nevertheless, the intrinsic mechanism of its involvement in Complex I in the parthanatos cascade remains obscure.

Aim of the study: This experiment aimed to investigate that HQ-HH antagonized parthanatos via regulating PARP-1/TAX1BP1-mediated Complex I to attenuate CI/RI.

Materials and methods: The HPLC fingerprint of HQ-HH was established, and the contents of 9 components were determined. The neuroprotective effect of HQ-HH in CI/RI was evaluated by rat middle cerebral artery occlusion/reperfusion (MCAO/R) and BV2 cell oxygen glucose deprivation/reoxygenation (OGD/R) models. Pathological changes in brain tissue of MCAO/R rats were observed using TTC staining, HE staining, and TEM. Complex I activity was measured in MCAO/R rats and OGD/R-treated BV2 cells. qRT-PCR and Western blot were performed to detect the expressions of related genes and proteins of parthanatos and Complex I as well as tax1 binding protein 1 (TAX1BP1). Immunofluorescence staining was employed to certify the nuclear translocation of apoptosis-inducing factor (AIF) in MCAO/R rats.

Results: The HPLC fingerprint of HQ-HH with 25 common peaks and the contents of 9 components were obtained. HQ-HH improved behavioral function and alleviated cerebral infarction in MCAO/R rats in a dose-dependent manner. HQ-HH alleviated parthanatos and exhibited the same repressive effect on PARP-1 transcription and translation as PJ34 (PARP-1 inhibitor). Moreover, the migration of TAX1BP1 to the mitochondria was restrained with HQ-HH treatment as a downstream of PARP-1, resulting in the inhibition of Complex I activity and less ROS production, accompanied by a decrease in mRNA and protein levels of ND1 and ND2. Subsequently, the nuclear translocation of AIF and the generation of poly(ADP-ribose) (PAR) polymers were suppressed.

Conclusions: HQ-HH mitigated CI/RI by regulating PARP-1/TAX1BP1 to inhibit the Complex I activity with less ROS production, further impeding nuclear translocation of AIF, and ultimately antagonizing parthanatos. By emphasizing the link between parthanatos and Complex I, we anticipate providing new empirical evidence for HQ-HH therapy of IS.

Keywords: Astragalus membranaceus-Carthamus tinctorius; Cerebral ischemia/reperfusion injury; Mitochondrial respiratory chain complex I; PARP-1; Parthanatos.