mTORC1 and mTORC2 Co-Protect against Cadmium-Induced Renal Tubular Epithelial Cell Apoptosis and Acute Kidney Injury by Regulating Protein Kinase B.

The potential threat of cadmium (Cd)-induced acute kidney injury (AKI) is increasing. In this study, our primary goal was to investigate the individual roles played by mTOR complexes, specifically mTORC1 and mTORC2, in Cd-induced apoptosis in mouse kidney cells. We constructed a mouse model with specific deletion of Raptor/Rictor renal cells. Inhibitors and activators of mTORC1 or mTORC2 were also applied. The effects of protein kinase B (AKT) activation and autophagy were studied. Both mTORC1 and mTORC2 were found to mediate the antiapoptotic mechanism of renal cells by regulating the AKT activity. Inhibition of mTORC1 or mTORC2 exacerbated Cd-induced kidney cell apoptosis, suggesting that both proteins exert antiapoptotic effects under Cd exposure. We further found that the AKT activation plays a key role in mTORC1/TORC2-mediated antiapoptosis, protecting Cd-exposed kidney cells from apoptosis. We also found that mTOR activators inhibited excessive autophagy, alleviated apoptosis, and promoted cell survival. These findings provide new insights into the regulatory mechanisms of mTOR in renal diseases and provide a theoretical basis for the development of novel therapeutic strategies to treat renal injury.

Cadmium exposure exacerbates kidney damage by inhibiting autophagy in diabetic rats.

The incidence of diabetes mellitus (DM) is gradually increasing, making it a widespread global health concern. Cadmium (Cd) is a common toxic heavy metal in the environment, and cadmium exposure may be associated with diabetic nephropathy (DN). However, the mechanism of Cd-induced DN remains unclear. In this study, we aimed to determine the effect of cadmium on diabetic kidney injury and the underlying mechanism in diabetic rats and a renal tubular epithelial cell line (NRK-52E cells). Our results could provide novel insights on the nephrotoxic mechanism of cadmium. HE, PAS, and Masson staining were used to observe pathological renal injury. COL-I, COL-IV, CTSB, and CTSD protein levels were detected by immunohistochemistry and western blotting. Immunofluorescence was used to detect the fluorescence intensity of p62 and LC3 proteins in kidney tissue. TEM was used to observe the ultrastructure of mitochondria and number of autophagosomes. After cadmium exposure, DM rats showed a dramatic decrease in body weight compared to the unexposed DM group. Relative kidney weight showed a contrasting trend after cadmium exposure. Urinary microalbumin/creatinine significantly increased in normal and DM rats after cadmium exposure. However, the trend was clearer in the DM groups than in the control groups. Endogenous creatinine clearance exhibited a contrasting trend. After cadmium exposure in DM rats, MDA content significantly increased and GSH, CAT, SOD, and GSH-PX activation reduced compared to normal controls. Pathological damage was more pronounced, and the expression of autophagy related proteins and apoptosis and fibrosis proteins was significantly higher in vivo and vitro in the cadmium-exposed groups than in unexposed controls. Further, lysosomal protein levels were lower, and ROS content and autophagosome count significantly higher in the cadmium exposed groups compared to the unexposed controls. Therefore, Cadmium exposure aggravates diabetic kidney injury via autophagy inhibition.

N-acetylcysteine delayed cadmium-induced chronic kidney injury by activating the sirtuin 1-P53 signaling pathway.

With the development of modern industrial civilization, cadmium (Cd), a known nephrotoxic metal, has become a growing public safety issue due to its ability to induce various types of kidney disease. Maladaptive proximal tubule repair is a significant cause of Cd-induced chronic kidney disease (CKD), which is characterized by premature senescence and pro-fibrosis. Previously, we demonstrated that cadmium causes DNA damage and cycle arrest in renal tubular epithelial cells, which may be relevant to premature senescence regulated by sirtuin 1 (SIRT1). In this study, in vivo and in vitro studies were conducted to elucidate the role of SIRT1-mediated premature renal senescence in Cd-induced CKD. As oxidative stress is a significant cause of aging, we evaluated whether N-acetylcysteine (NAC) would inhibit Cd-induced premature aging and dysfunction in rat renal tubular epithelial cells. Cadmium induced premature renal senescence and fibrosis, and NAC inhibited premature renal senescence and fibrosis through the SIRT1-P53 pathway and delayed CKD progression. Overall, the results suggested that the SIRT1-P53 pathway mediates oxidative stress, premature renal senescence, and renal fibrosis during cadmium exposure, which may be a potential therapeutic target for Cd-induced CKD.

Nuclear factor-kappaB mediates the survival of rat kidney cells after cadmium exposure via promoting autophagy and inhibiting apoptosis.

Cadmium (Cd) is a heavy metal pollutant in the environment, and the kidney is one of the target organs after Cd exposure. Previous studies have shown that apoptosis and autophagy disorders are the main mechanisms of Cd-induced nephrotoxicity in rats. As a transcription factor that balances cell survival and death, nuclear factor-kappaB (NF-κB) protein plays dual regulatory effects on apoptosis and autophagy in multiple renal diseases. However, the regulatory mechanisms of NF-κB in Cd-induced kidney injury remain unclear. Therefore, the normal rat kidney cell line (NRK-52E cells) was applied to investigate the above questions in this study. Here, we found that Cd promotes the nuclear translocation and activation of NF-κB in a concentration-dependent manner, and activated NF-κB mediates NRK-52E cells survival after Cd exposure. Next, our study elaborated the mechanisms of NF-κB in antagonizing Cd-induced renal cytotoxicity. Inhibition of NF-κB by inhibitor BAY 11-7082 (BAY) and NF-κB p65 siRNA (siNF-κB p65) exacerbate Cd-induced apoptosis and autophagy inhibition, and then aggravate Cd-induced NRK-52E cells injury. Activation of NF-κB by activator phorbol-12-myristate-13-acetate (PMA) alleviates Cd-induced apoptosis and autophagy inhibition, and then attenuates Cd-induced NRK-52E cells injury. In conclusion, Cd exposure promotes the activation of NF-κB, and activated NF-κB mediates the survival of NRK-52E cells after Cd exposure via promoting autophagy and inhibiting apoptosis.

Honokiol Antagonizes Cadmium-Induced Nephrotoxicity in Quail by Alleviating Autophagy Dysfunction, Apoptosis and Mitochondrial UPR Inhibition with Its Antioxidant Properties.

Japanese quail is a highly economically valuable bird due to its commercial production for meat and eggs. Although studies have reported Cadmium (Cd) is a ubiquitous heavy metal that can cause injury to various organs, the molecular mechanisms of Cd on quail kidney injury remain largely unknown. It has been reported that Honokiol (HKL), a highly functional antioxidant, can protect cells against oxidative stress effectively. This study was conducted to investigate the effects of Cd on quail kidneys injury and the protective effect of HKL on Cd-induced nephrotoxicity. A total of 40 Japanese quails were randomly divided into four groups: the control group, Cd treatment group, Co-treatment group and HKL treatment group. The results showed that Cd resulted in significant changes in growth performance, kidney histopathology and kidney biochemical status, antioxidant enzymes and oxidative stress parameters, and ultrastructure of renal tubular epithelial cells, compared with controls. Cd increased the expression of autophagy-related and apoptosis-related genes, but decreased expression of lysosomal function-related and UPRmt-related genes. The co-treatment group ameliorated Cd-induced nephrotoxicity by alleviating oxidative stress, inhibiting apoptosis, repairing autophagy dysfunction and UPRmt disorder. In conclusion, dietary supplementation of HKL showed beneficial effects on Japanese quail kidney injury caused by Cd.

Melatonin improves mitochondrial function by preventing mitochondrial fission in cadmium-induced rat proximal tubular cell injury via SIRT1-PGC-1α pathway activation.

Melatonin is an indoleamine produced in the pineal gland and has many physiological roles. There is increasing evidence that melatonin ameliorates cadmium (Cd)-induced nephrotoxicity. The potential protective impact of melatonin against Cd-induced nephrotoxicity and the mechanisms behind this protection are unknown. The relevance of mitochondrial dynamics in Cd-induced nephrotoxicity and the putative mechanism of melatonin-mediated protection were examined in this study. We show that melatonin prevents Cd-induced nephrotoxicity by inhibiting dynamin-related protein 1 (Drp1)- and mitochondrial fission protein 1 (Fis1)-mediated mitochondrial fission. Melatonin treatment attenuated cytotoxicity, suppressed oxidative stress, restored mitochondrial membrane potential, and increased mitochondrial mass in response to Cd exposure. Consistent with this finding, melatonin treatment increased Cd-inhibited sirtuin 1 (SIRT1) and peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) expression and inhibited Drp1- and Fis1-mediated mitochondrial fission. Like melatonin, SIRT1 overexpression via resveratrol attenuated Drp1- and Fis1-mediated mitochondrial fission and other Cd-induced mitochondrial oxidative injuries effectively. Melatonin has significant pharmacological potential for protecting against Cd-induced nephrotoxicity by preventing excessive mitochondrial fission.

Paeonol protects renal tubular cells against cadmium-induced cytotoxicity via alleviating oxidative stress, inhibiting inflammatory responses and restoring autophagy.

Cadmium (Cd) is a widespread environmental pollutant with highly toxic to kidney. Paeonol (Pae) is a natural flavonoid isolated from Moutan Cortex that exhibits antioxidant and anti-inflammatory properties. Herein, we investigated the protective effects of Pae against Cd-induced nephrotoxicity and elucidated its underlying mechanisms. First, we screened the optimum treatment conditions for Pae and confirmed its ability to protect NRK-52E cells against Cd-induced cytotoxicity. The results showed that Pae alleviated Cd-induced oxidative stress by scavenging excessive reactive oxygen species. Furthermore, Pae suppressed Cd-induced inflammatory responses by inhibiting the activation of nuclear factor-κB and the transcriptions of pro-inflammatory cytokines including cytokines including interleukin (IL)-1ß, IL-6, monocyte chemotactic protein (MCP)-1 and tumour necrosis factor (TNF)-α. Pae alleviated Cd-induced autophagy inhibition, which was partly attributable to alleviation of oxidative stress. Meanwhile, Pae improved the structural integrity and degradation function of lysosomes, indicating that Pae can also target lysosomes to restore Cd-inhibited autophagic flux. Collectively, Pae alleviated Cd-induced renal cytotoxicity by alleviating oxidative stress, inhibiting inflammatory responses and restoring autophagy, implicating that Pae may serve as new candidate drug to treat Cd-induced nephrotoxicity.

Epigenetic regulator BRD4 is involved in cadmium-induced acute kidney injury via contributing to lysosomal dysfunction, autophagy blockade and oxidative stress.

Cadmium (Cd) is a known nephrotoxic heavy metal and proximal tubules are the major target of Cd-induced acute kidney injury (AKI). We previously demonstrated that lysosomal dysfunction and dysregulated autophagy contribute to Cd-induced AKI. Recent studies have revealed that bromodomain-containing protein 4 (BRD4) is a transcriptional repressor of autophagy and lysosomal function. Hence, in vivo and in vitro studies were performed to clarify the role of BRD4 in Cd-induced AKI. Firstly, Cd has no effect on BRD4 expression levels, but increases H4K16 acetylation. Resultantly, Cd promotes the recruitment of BRD4 to lysosomal gene promoter regions to make it as a transcriptional regulator. Pharmacological and genetic inhibition of BRD4 alleviates Cd-inhibited lysosomal gene transcript levels and lysosomal function, leading to the alleviation of Cd-induced autophagy inhibition. Moreover, inhibition of BRD4 relieves Cd-induced oxidative stress and concurrent cytotoxicity, which is counteracted by the inhibition of autophagy via Atg5 knockdown, indicating that alleviation of oxidative stress by BRD4 inhibition is ascribed to its restoration of autophagic flux. Collectively, these results demonstrate that BRD4 acts as a transcriptional repressor to mediate lysosomal dysfunction, autophagy blockade and oxidative stress during Cd exposure, which may be a potential therapeutic target for Cd-induced AKI.

The epigenetic regulator BRD4 is involved in cadmium-triggered inflammatory response in rat kidney.

Cadmium (Cd) has been described as a potential inflammatory inducer, while increasing evidence shows that inappropriate inflammation is a contributing factor to kidney injury. Hence, research on Cd-triggered inflammatory response is of great significance for elucidating the mechanism of Cd-induced nephrotoxicity. Bromodomain-containing 4 (BRD4) is an important epigenetic regulator involved in the development of many inflammatory diseases, but its regulatory roles in Cd-triggered inflammatory response remain to be clarified. Here, we found that treatment with Cd in Sprague-Dawley rats (2 mg/kg bw, i.p., 5 consecutive days) and in rat kidney cell line (NRK-52E, 0-10 µM, 12 h) induced the transcription of inflammatory cytokines, which could be reduced by JQ1 (BRD4 inhibitor, 25 mg/kg bw, i.p., 3 consecutive days in vivo; 0.5 µM, 12 h in vitro) or BRD4 small interfering RNA (siRNA, in vitro), suggesting that BRD4 participates in Cd-triggered inflammatory response. Next, our study clarified the roles of BRD4 in Cd-triggered inflammatory response. The inhibition of BRD4 decreased Cd-promoted NF-κB nuclear translocation and activation in vivo and in vitro. Cd increased the acetylation level of RelA K310 and enhanced BRD4 binding to acetylated NF-κB RelA in vivo and in vitro, which were abrogated by inhibiting BRD4. In summary, our study suggests that BRD4 is involved in Cd-triggered transcription of inflammatory cytokines by mediating the activation of NF-κB signaling pathway and increasing itself binding to acetylated NF-κB RelA in rat kidney, therefore, BRD4 could be a potential therapeutic target for Cd-induced renal diseases.

Genetic characterization and phylogenetic analysis of porcine epidemic diarrhea virus in Guangdong, China, between 2018 and 2019.

Porcine epidemic diarrhea virus (PEDV), a leading cause of piglet diarrhea outbreaks, poses a significant danger to the swine industry. The aim of this study was to investigate the epidemic characteristics of PEDV that was circulating in Guangdong province, one of China's major pig producing provinces. Clinical samples were collected from eight pig farms in Guangdong province between 2018 and 2019 and tested for the major porcine enteric pathogens, including PEDV, transmissible gastroenteritis virus (TGEV), Swine enteric coronavirus (SeCoV), Swine acute diarrhea syndrome coronavirus (SADS-CoV), porcine deltacoronavirus (PDCoV), and porcine rotavirus (RV). As a result, only PEDV and RV were detected at a rate of 47.0% (16/34) and 18.6% (8/34), respectively. Coinfectoin with PEDV and RV occurred at a rate of PEDV 12.5% (2/16). Subsequently, the full-length S gene sequences of 13 PEDV strains were obtained, and phylogenetic analysis suggested the presence of GII-c group PEDV strains in this region (non-S-INDEL). Two novel common amino acid insertions (55T/IG56 and 551L) and one novel glycosylation site (1199G+) were detected when the CV777 and ZJ08 vaccine strains were compared. Furthermore, intragroup recombination events in the S gene regions 51-548 and 2478-4208 were observed in the PEDV strains studied. In summary, the observations provide current information on the incidence of viral agents causing swine diarrhea in southern China and detailed the genetic characteristics and evolutionary history of the dominant PEDV field strains. Our findings will aid in the development of an updated vaccine for the prevention and control of PEDV variant strains.

Effect of cell cycle synchronization on cadmium-induced apoptosis and necrosis in NRK-52E cells.

Heavy metal pollution is a problem that cannot be ignored. Due to the prevalence of cadmium in the environment and its harmful effects on humans, cadmium pollution has become a research hotspot recently. The mechanism of cadmium-induced toxicity has also drawn much attention and most studies have been conducted using whole cells, but the toxicological mechanism of cadmium remains unclear. In this study, we aimed to obtain NRK-52E cells at different growth stages by various methods and analyze the differences in cadmium toxicity. The results show that the cadmium sensitivity of cells in each phase was different and the late apoptotic rate was increased significantly after 5 µM Cd treatment. In addition, cadmium easily induces apoptosis of G0- and S-phase cells, as well as necrosis of S- and M-phase cells, but has no significant effect on G1-phase cells. Overall, we first explored the differences in the effects of cadmium on NRK-52E cells at various growth phases. Besides, the findings of this study might provide a theoretical basis for further exploration of the toxicological mechanism of cadmium.Abbreviations Cd: cadmium; CDK: cyclin-dependent kinases; DAPI 2-(4-amidinophenyl)-1H-indole-6-carboxamidine; TBST: Tris-buffered saline with Tween-20; PI: propidium iodide; DMEM: Dulbecco's Modified Eagle Medium; BCA: bicinchoninic acid.

Trehalose prevents cadmium-induced hepatotoxicity by blocking Nrf2 pathway, restoring autophagy and inhibiting apoptosis.

Cadmium (Cd) is a ubiquitously distributed environmental pollutant that is highly toxic to liver. Trehalose (Tr), a novel autophagy activator, has been shown to exert cytoprotective effect in numerous pathological processes. However, it is yet to be established whether Tr affords protection against Cd-induced hepatotoxicity. Here, we aimed to investigate the protective effect of Tr on Cd-induced hepatic injury in rats. First, Cd-elevated serum hepatic enzymes and liver pathological changes were significantly ameliorated by Tr treatment. Also, Tr remarkably improved Cd-mediated oxidative stress and antioxidant status in serum, indicating its anti-oxidant action for the whole body. Cd-stimulated nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation and subsequent elevated expression of Nrf2-downstream targets in rat liver were significantly inhibited by Tr treatment. Simultaneously, Cd-elevated protein levels of hepatic antioxidant enzymes were markedly downregulated by administration with Tr. Moreover, Cd-induced autophagy inhibition in liver tissues was noticeably restored by Tr, evidenced by immunohistochemical analysis and immunoblot assays. Additionally, Tr treatment significantly mitigated Cd-induced apoptosis in hepatic tissues via inhibiting caspase-dependent apoptotic pathway. In conclusion, these observations demonstrate that Tr treatment alleviates Cd-induced liver injury by blocking Nrf2 pathway, restoring autophagy and inhibiting apoptosis.