Mitophagy has distinct functions, which can lead to either protection or damage of tissues. Though current evidence indicated that NaF triggers mitophagy, the role and regulation of mitophagy in sodium fluoride (NaF)-induced liver injury still remain unclear. Therefore, we exployed the cell and mouse models and confirmed that NaF treatment activates mitophagy. Knocking down PTEN-induced putative kinase protein 1 (PINK1) expression attenuated mitophagy and increased the degree of mitochondrial impairment, oxidative stress, and apoptosis in NaF-treated HepG2 cells. In vivo experiments indicated that PINK1 deficiency weakened NaF-induced mitophagy. Moreover, PINK1-deficient mices aggravated NaF-induced hepatic mitochondrial injury, oxidative stress, and inflammation in livers, evidenced by the increased number of abnormal mitochondria, decreased adenosine triphosphate (ATP) and glutathione (GSH) levels, elevated reactive oxygen species (ROS) and malondialdehyde (MDA) content, enhanced hepatic macrophage infiltration and inflammatory cytokine levels. Notably, NaF exposure activated Nrf2 signaling both in vitro and in vivo. Nrf2 siRNA transfection blocked the upregulation of PINK1 expression and the induction of mitophagy in NaF-treated HepG2 cells. Also, ML385 (Nrf2 inhibitor) partially blocked the upregulation of PINK1 expression caused by NaF in mice livers. To sum up, the present study provided the demonstration that Nrf2/PINK1-mediated mitophagy activation offers a hepatoprotective effect by inhibiting NaF-induced mitochondrial dysfunction, oxidative stress, and inflammation.

Marek’s disease (MD) caused by pathogenic Marek’s disease virus type 1 (MDV−1) is one of the most important neoplastic diseases of poultry. MDV−1-encoded unique Meq protein is the major oncoprotein and the availability of Meq-specific monoclonal antibodies (mAbs) is crucial for revealing MDV pathogenesis/oncogenesis. Using synthesized polypeptides from conserved hydrophilic regions of the Meq protein as immunogens, together with hybridoma technology and primary screening by cross immunofluorescence assay (IFA) on Meq-deleted MDV−1 viruses generated by CRISPR/Cas9-gene editing, a total of five positive hybridomas were generated. Four of these hybridomas, namely 2A9, 5A7, 7F9 and 8G11, were further confirmed to secrete specific antibodies against Meq as confirmed by the IFA staining of 293T cells overexpressing Meq. Confocal microscopic analysis of cells stained with these antibodies confirmed the nuclear localization of Meq in MDV-infected CEF cells and MDV-transformed MSB-1 cells. Furthermore, two mAb hybridoma clones, 2A9-B12 and 8G11-B2 derived from 2A9 and 8G11, respectively, displayed high specificity for Meq proteins of MDV−1 strains with diverse virulence. Our data presented here, using synthesized polypeptide immunization combined with cross IFA staining on CRISPR/Cas9 gene-edited viruses, has provided a new efficient approach for future generation of specific mAbs against viral proteins.

The avian immunosuppressive and neoplastic diseases are great threats to the poultry industry, causing huge economic losses worldwide. Most recently, the emerging hypervirulent variants of Marek’s disease virus (HV-MDV), partially co-infected with avian leukosis virus (ALV) and/or reticuloendotheliosis virus (REV), have been identified as the key driver of tumour outbreaks in vaccinated chicken flocks, but the role of chicken infectious anemia virus (CIAV) remains unclear. Herein, we have investigated the prevalence and co-infection of CIAV in 71 clinical tumour-bearing flocks collected from central China during 2021–2023, which has shown a CIAV positivity rate of 59.2% (42/71). Notably, the incidence of CIAV mono-infection increased significantly from 0% (0/29) in 2021 to 23.7% (9/38) in 2023, whereas CIAV + MDV co-infection decreased from 65.5% (19/29) to 31.6% (12/38). A total of 20 viral genomes of epidemic CIAV isolates from diverse sources were obtained, and the phylogenetic analysis, including 91 reference isolates were clustered into four major lineages (A–D), with clade C further subdivided into subclades C1 and C2. Clade C1 consisted predominantly of Asian isolates, with 88.5% (46/52) of the isolates originating from mainland China. Among the 20 new isolates, 17 were clustered in subclade C1, two in C2, and one in B. The VP1 gene phylogeny showed a topology largely consistent with that of the whole-genome analysis. Moreover, all newly characterized isolates contained glutamine (Q) at VP1 residue 394, a molecular marker associated with high pathogenicity. Collectively, our data suggest that prevalent HV-MDV variants together with CIAV co-infections are the primary drivers of the ongoing tumour outbreaks in Chinese poultry flocks. Notably, the significantly increased CIAV mono-infections, possibly resulting from an independently evolving lineage among circulating Chinese strains, are likely to pose a new challenge for future control of disease.

The avian immunosuppressive and neoplastic diseases are great threats to the poultry industry causing huge economic losses worldwide. Most recently, the emerging hypervirulent variants of Marek’s disease virus (HV-MDV), partially co-infected with avian leukosis virus (ALV) and/or reticuloendotheliosis virus (REV), have been identified as the key driver of tumour outbreaks in vaccinated chicken flocks, but the role of chicken infectious anaemia virus (CIAV) remains unclear. Herein, we have investigated the prevalence and co-infection of CIAV in 72 clinical tumour-bearing flocks collected from central China during 2020-2023, which has shown a CIAV positivity rate of 59.7% (43/72). Notably, the incidence of CIAV mono-infection increased significantly from 0% (0/30) in 2020-2021 to 26.2% (11/42) in 2022-2023, whereas CIAV+MDV co-infection decreased from 66.7% (20/30) to 28.6% (12/42). A total of 20 viral genomes of epidemic CIAV isolates from diverse sources were obtained, and the phylogenetic analysis including 91 reference isolates were clustered into four major lineages (A–D), with a clade C further subdivided into subclades C1 and C2. Clade C1 consisted predominantly of Asian isolates, with 88.5% (46/52) of the isolates originated from mainland China. Among the 20 new isolates, 17 were clustered in subclade C1, two in C2, and one in B. The VP1 gene phylogeny showed a topology largely consistent with that of the whole-genome analysis. Moreover, all newly characterized isolates contained glutamine (Q) at VP1 residue 394, a molecular marker associated with high pathogenicity. Collectively, our data suggest that prevalent HV-MDV variants together with CIAV co-infections are the primary drivers of the ongoing tumour outbreaks in Chinese poultry flocks. Furthermore, the increased pathogenicity observed in CIAV mono-infections, likely resulting from an independently evolving lineage among circulating Chinese strains, will pose a significant challenge for future control of disease.