Hypoxia-Inducible Factor-1α Regulates High Phosphate-Induced Vascular Calcification via Type III Sodium-Dependent Phosphate Cotransporter 1.

Vascular calcification (VC) has a high incidence in patients with chronic kidney disease, which is a worldwide public health problem and presents a heavy burden to society. Hypoxia-inducible factor (HIF)-1α, the active subunit of HIF-1, has been reported to play a vital role in high phosphate-induced VC. However, the underlying mechanism is still undetermined, and effective treatment is unavailable. In the present study, human aortic smooth muscle cells (HASMCs) were cultured under normal or high phosphate media conditions. HIF-1α small interfering RNA and overexpression plasmids were employed to regulate HIF-1α expression. Phosphonoformic acid was employed to restrain the function of type III sodium-dependent phosphate cotransporter 1 (Pit-1). The expression levels of HIF-1α, Pit-1, runt-related transcription factor 2 (Runx2), and smooth muscle 22 alpha (SM22α) were evaluated, and the calcium contents were also examined. Cell growth was assessed using an MTT assay. High phosphate stimulation caused an upregulation in HIF-1α and Pit-1 expression levels and induced calcium depositions in HASMCs. Upregulation of Runx2 expression accompanied by downregulation of SM22α expression was observed in the high phosphate group. Following the suppression of HIF-1α expression, there was a concomitant attenuation in Pit-1 expression, calcium deposition, the alteration of phenotypic transition marker genes, and vice versa. The most serious calcium deposition was noted in HASMCs cultured under high phosphate conditions which were pretreated with a HIF-1α overexpression plasmid. However, when the biological functions of Pit-1 were restrained, the putative serious calcium deposition was not formed even in HASMCs transfected with a HIF-1α overexpression plasmid. The findings confirmed that HIF-1α regulated Pit-1 expression and exerted its pro-calcifying effect through Pit-1, which identified HIF-1α and Pit-1 as therapeutic targets for high phosphate-induced VC.

Paraquat-induced ferroptosis suppression via NRF2 expression regulation.

Paraquat (PQ) is an environmentally friendly and efficient herbicide, but PQ misuse or intentional self-use can cause death through multiple organ damage and can cause acute lung injury. Existing clinical treatments alleviate symptoms but do not significantly improve the mortality rate. Ferroptosis is a type of necrosis that presents in a manner very similar to the cell damage induced by high doses of PQ, but the role of ferroptosis in paraquat-induced lung injury remains unclear. In this study, we aimed to explore the role of ferroptosis in PQ-induced A549 cell injury and identify the potential mechanisms and critical sites of protection against PQ-induced A549 injury by ferroptosis inhibitors. We found that the ferroptosis inhibitors Ferr-1 and Lip-1 inhibit ferroptosis by attenuating oxidative stress through the upregulation of NRF2 gene expression. The protective role of the ferroptosis inhibitor Dfo was most evident in paraquat-induced cell injury. Dfo inhibited ferroptosis by iron chelation and promoted NRF2 protein level reduction. NRF2 attenuated PQ-induced ferroptosis in A549 cells, mainly through the upregulation of SLC40A1 to encourage the movement of iron to the extracellular side to alleviate iron overload, and the upregulation of SLC7A11 to promote the expression of GPX4 to inhibit lipid peroxidation.

ETI signaling nodes are involved in resistance of Hawaii 7996 to Ralstonia solanacearum-induced bacterial wilt disease in tomato.

Bacterial wilt caused by the soil-borne pathogen Ralstonia solanacearum is a destructive disease of tomato. Tomato cultivar Hawaii 7996 is well-known for its stable resistance against R. solanacearum. However, the resistance mechanism of Hawaii 7996 has not yet been revealed. Here, we showed that Hawaii 7996 activated root cell death response and exhibited stronger defense gene induction than the susceptible cultivar Moneymaker after R. solanacearum GMI1000 infection. By employing virus-induced gene silencing (VIGS) and CRISPR/Cas9 technologies, we found that SlNRG1-silenced and SlADR1-silenced/knockout mutant tomato partially or completely lost resistance to bacterial wilt, indicating that helper NLRs SlADR1 and SlNRG1, the key nodes of effector-triggered immunity (ETI) pathways, are required for Hawaii 7996 resistance. In addition, while SlNDR1 was dispensable for the resistance of Hawaii 7996 to R. solanacearum, SlEDS1, SlSAG101a/b, and SlPAD4 were essential for the immune signaling pathways in Hawaii 7996. Overall, our results suggested that robust resistance of Hawaii 7996 to R. solanacearum relied on the involvement of multiple conserved key nodes of the ETI signaling pathways. This study sheds light on the molecular mechanisms underlying tomato resistance to R. solanacearum and will accelerate the breeding of tomatoes resilient to diseases.

Treatment with paraquat affects the expression of ferroptosis-related genes.

OBJECTIVE: We aimed to explore the mechanisms underlying paraquat (PQ)-induced damage using cell lines (NCTC1469, TC-1, TCMK-1) and bioinformatic analysis of the GSE153959 dataset. Assessment of changes in the expression of ferroptosis-related genes in cellular damage due to paraquat poisoning and the important value of these genes in the pathogenesis. METHODS: Data were retrieved from the Gene Expression Omnibus database. The differentially expressed genes (DEGs) related to ferroptosis were identified by Venn plots and analyzed for enrichment. Proteins encoded by these DEGs were studied for interactions. qRT-PCR and western blotting analyses of cultured cells were used to determine the expression of ferroptosis-related DEGs and their corresponding protein levels. RESULTS: We identified 25 DEGs primarily involved in epidermal growth factor receptor signaling, apoptotic signaling pathways, endoplasmic reticulum (ER) stress, and ferroptosis. From these, we uncovered eight ferroptosis-related DEGs, four of which were involved in ER response and regulators of ferroptosis-Chac1 (ChaC glutathione specific gamma-glutamylcyclotransferase 1), Atf3 (activating transcription factor 3), Tfrc (transferrin receptor), and Slc7a11 (solute carrier family 7 member 11). Significant changes in mRNA and protein levels of CHAC1, ATF3, TFRC, and SLC7A11 were confirmed in PQ-exposed cells. CONCLUSION: ER stress and ferroptosis are critical for PQ-induced cell damage. CHAC1, ATF3, TFRC, and SLC7A11 are essential molecules implicated in PQ-induced ferroptosis that may serve as therapeutic targets for the amelioration of PQ poisoning.

Duality of immune recognition by tomato and virulence activity of the Ralstonia solanacearum exo-polygalacturonase PehC.

Ralstonia solanacearum is a devastating soil-borne bacterial pathogen capable of infecting many plant species, including tomato (Solanum lycopersicum). However, the perception of Ralstonia by the tomato immune system and the pathogen's counter-defense strategy remain largely unknown. Here, we show that PehC, a specific exo-polygalacturonase secreted by Ralstonia, acts as an elicitor that triggers typical immune responses in tomato and other Solanaceous plants. The elicitor activity of PehC depends on its N-terminal epitope, and not on its polygalacturonase activity. The recognition of PehC specifically occurs in tomato roots and relies on unknown receptor-like kinase(s). Moreover, PehC hydrolyzes plant pectin-derived oligogalacturonic acids (OGs), a type of damage-associated molecular pattern (DAMP), which leads to the release of galacturonic acid (GalA), thereby dampening DAMP-triggered immunity (DTI). Ralstonia depends on PehC for its growth and early infection and can utilize GalA as a carbon source in the xylem. Our findings demonstrate the specialized and dual functions of Ralstonia PehC, which enhance virulence by degrading DAMPs to evade DTI and produce nutrients, a strategy used by pathogens to attenuate plant immunity. Solanaceous plants have evolved to recognize PehC and induce immune responses, which highlights the significance of PehC. Overall, this study provides insight into the arms race between plants and pathogens.

A bioartificial transgenic porcine whole liver expressing human proteins alleviates acute liver failure in pigs.

BACKGROUND: Preventing heterologous protein influx in patients is important when using xenogeneic bioartificial livers (BALs) to treat liver failure. The development of transgenic porcine livers synthesizing human proteins is a promising approach in this regard. Here, we evaluated the safety and efficacy of a transgenic porcine liver synthesizing human albumin (hALB) and coagulation factor VII (hFVII) within a bioartificial system. METHODS: Tibetan miniature pigs were randomly subjected to different interventions after surgery-induced partially ischemic liver failure. Group A (n = 4) was subjected to basic treatment; group B (n = 4) was to standard medical treatment and wild-type porcine BAL perfusion, and group C (n = 2) was to standard medical treatment and transgenic BAL perfusion. Biochemical parameters, coagulation status, survival time, and pathological changes were determined. Expressions of hALB and hFVII were detected using immunohistochemistry and enzyme-linked immunosorbent assays. RESULTS: The survival time in group A was 9.75 ± 1.26 days; this was shorter than that in both perfused groups, in which all animals reached an endpoint of 12 days (P = 0.006). Ammonia, bilirubin, and lactate levels were significantly decreased, whereas albumin and fibrinogen levels were increased after perfusion (all P < 0.05). hALB and hFVII were detected in transgenic BAL-perfused pig serum and ex vivo in the liver tissues. CONCLUSIONS: The humanized transgenic pig livers could synthesize and secrete hALB and hFVII ex vivo in a whole organ-based bioartificial system, while maintaining their metabolism, detoxification, transformation, and excretion functions, which were comparable to those observed in wild-type porcine livers. Therefore, the use of transgenic bioartificial whole livers is expected to become a new approach in treating acute liver failure.

Chromatin Remodeling Induced by ARID1A Loss in Lung Cancer Promotes Glycolysis and Confers JQ1 Vulnerability.

ARID1A is a key mammalian SWI/SNF complex subunit that is mutated in 5% to 11% of lung cancers. Although recent studies have elucidated the mechanism underlying dysregulation of the switch/sucrose non-fermentable (SWI/SNF) complexes in cancers, the significance of ARID1A loss and its implications in lung cancers remain poorly defined. This study investigates how ARID1A loss affects initiation and progression of lung cancer. In genetically engineered mouse models bearing mutant Kras and a deficient Trp53 allele (KP), ARID1A loss (KPA) promoted lung tumorigenesis. Analysis of the transcriptome profiles of KP and KPA tumors suggested enhanced glycolysis following ARID1A loss, and expression of the glycolytic regulators Pgam1, pyruvate kinase M (Pkm), and Pgk1 was significantly increased in ARID1A-deficient lung tumors. Furthermore, ARID1A loss increased chromatin accessibility and enhanced hypoxia-inducible factor-1α (HIF1α) binding to the promoter regions of Pgam1, Pkm, and Pgk1. Loss of ARID1A in lung adenocarcinoma also resulted in loss of histone deacetylase 1 (HDAC1) recruitment, increasing acetylation of histone-4 lysine at the promoters of Pgam1, Pkm, and Pgk1, and subsequently enhancing BRD4-driven transcription of these genes. Metabolic analyses confirmed that glycolysis is enhanced in ARID1A-deficient tumors, and genetic or pharmacologic inhibition of glycolysis inhibited lung tumorigenesis in KPA mice. Treatment with the small molecule bromodomain and extraterminal protein (BET) inhibitor JQ1 compromised both initiation and progression of ARID1A-deficient lung adenocarcinoma. ARID1A negatively correlated with glycolysis-related genes in human lung adenocarcinoma. Overall, ARID1A loss leads to metabolic reprogramming that supports tumorigenesis but also confers a therapeutic vulnerability that could be harnessed to improve the treatment of ARID1A-deficient lung cancer. SIGNIFICANCE: This study links ARID1A loss with enhanced glycolysis in lung cancer and demonstrates the preclinical efficacy of BET inhibitor therapy as a strategy to combat tumor growth.

Hyperbaric Oxygen Therapy Represses the Warburg Effect and Epithelial-Mesenchymal Transition in Hypoxic NSCLC Cells via the HIF-1α/PFKP Axis.

BACKGROUND: Tumor cells initiate hypoxia-induced mechanisms to fuel cell proliferation, invasion, and metastasis, largely mediated by low O2-responsive Hypoxia-Inducible Factor 1 Alpha (HIF-1α). Therefore, hyperbaric oxygen therapy (HBO) is now being studied in cancer patients, but its impact upon non-small-cell lung cancer (NSCLC) cell metabolism remains uncharacterized. METHODS: We employed the NSCLC cell lines A549 and H1299 for in vitro studies. Glucose uptake, pyruvate, lactate, and adenosine triphosphate (ATP) assays were used to assess aerobic glycolysis (Warburg effect). A quantitative glycolytic flux model was used to analyze the flux contributions of HIF-1α-induced glucose metabolism genes. We used a Lewis lung carcinoma (LLC) murine model to measure lung tumorigenesis in C57BL/6J mice. RESULTS: HBO suppressed hypoxia-induced HIF-1α expression and downstream HIF-1α signaling in NSCLC cells. One HIF-1α-induced glucose metabolism gene-Phosphofructokinase, Platelet (PFKP)-most profoundly enhanced glycolytic flux under both low- and high-glucose conditions. HBO suppressed hypoxia-induced PFKP transactivation and gene expression via HIF-1α downregulation. HBO's suppression of the Warburg effect, suppression of hyperproliferation, and suppression of epithelial-to-mesenchymal transition (EMT) in hypoxic NSCLC cell lines is mediated by the HIF-1α/PFKP axis. In vivo, HBO therapy inhibited murine LLC lung tumor growth in a Pfkp-dependent manner. CONCLUSIONS: HBO's repression of the Warburg effect, repression of hyperproliferation, and repression of EMT in hypoxic NSCLC cells is dependent upon HIF-1α downregulation. HIF-1α's target gene PFKP functions as a central mediator of HBO's effects in hypoxic NSCLC cells and may represent a metabolic vulnerability in NSCLC tumors.