We investigated the potential correlation between miR-223 and NAcHT, LRR, and PYd domain-containing protein 3 (NLRP3) in the context of renal ischemia-reperfusion injury (RIRI), which is a leading cause of acute renal failure with significant mortality rates. Additionally, miR-223 has been implicated in renal inflammation, further highlighting its relevance to this study. C57BL/6 male mice were used as RIRI models. After successful modeling, pathological examinations and serum creatinine and miR-223 levels were tested. Pro-inflammatory cytokine (IL-1β, IL-6, IL-8, NLPR3, TLR4) expression was detected in mice by western blot (kidney tissue) and enzyme-linked immunosorbent assay (serum). HK-2 cells were used for in vitro experiments. A hypoxia/reoxygenation (H/R) model was used, and miR-223 and pro-inflammatory cytokine levels were detected using PCR and western blot assays, respectively. A dual-luciferase reporter assay was conducted to confirm the binding of miR-223 to NLPR3. Next, NLRP3 was knocked down to determine whether the anti-inflammatory function of miR-223 is dependent on NLRP3. MiR-223 expression was lower in RIRI mice than in the sham operation group. The level of miR-223 negatively correlated with serum creatinine levels and the severity of tubule injury. Increased proinflammatory cytokine levels in RIRI mice were observed. In vitro, miR-223 alleviated the inflammatory response in H/R treated cells by inhibiting proinflammatory cytokines. Dual-luciferase reporter and western blot assays confirmed the binding of miR-223 to NLRP3. NLRP3 knockdown reversed the anti-inflammatory effects of miR-223 in HK-2 cells. MiR-223 plays an anti-inflammatory role in RIRI by targeting NLRP3 to repress pro-inflammatory factors.

BACKGROUND: Diabetic nephropathy (DN) is a common complication of diabetes mellitus, and Prolyl 4-Hydroxylase Subunit Beta (P4HB) expression is increased in high glucose (HG)-induced renal tubular epithelial cells (TECs). But it\'s role in HG-induced TECs remains to be elucidated.
METHODS: The HK-2 cells were induced using HG and transfected with SiRNA-P4HB. DCFH-DA staining was utilized for the detection of cellular levels of ROS. WB and immunofluorescence were utilized to detect the expression of P4HB, epithelial-mesenchymal transition (EMT), fibrosis, and TGFβ/SMAD3-related proteins in HK-2 cells. Online databases were utilized for predicting the interaction target of P4HB, and immunoprecipitation (IP) experiments were employed to validate the binding of P4HB with the target. SiRNA and overexpression vectors of target gene were used to verify the mechanism of action of P4HB.
RESULTS: HG induced an increase in the expression of P4HB and TGFβ, p-SMAD3, and ROS in HK-2 cells. Furthermore, HG downregulated the expression of E-cadherin and upregulated the expression of N-cadherin, Vimentin, α-SMA, Fibronectin, Collagen IV, SNAIL, and SLUG in HK-2 cells. Interfering with P4HB significantly reversed the expression of these proteins. Database predictions and IP experiments showed that P4HB interacts with PRMT1, and the expression of PRMT1 was increased in HG-induced HK-2 cells. Interfering with PRMT1 inhibited the changes in expression of EMT and fibrosis related proteins induced by HG. However, overexpression of PRMT1 weakened the regulatory effect of P4HB interference on the EMT, fibrosis, and TGFβ/SMAD3-related proteins in HK-2 cells.
CONCLUSIONS: P4HB regulated the TGFβ/SMAD3 signaling pathway through PRMT1 and thus participates in HG-induced EMT and fibrosis in HK-2 cells.

Oxidative damage to human retinal pigment epithelial (RPE) cells is the main cause of age-related macular degeneration (AMD), in our previous work, we showed that ghrelin has an antioxidative effect on human lens epithelium (HLE) cells, however, the studies of using ghrelin in treating the degenerative diseases of the retina have rarely been reported. In this article, we assessed the effect of ghrelin on preventing oxidative stress induced by hydrogen peroxide (H2O2) in ARPE-19 cells and its mechanism. We observed that pretreatment with ghrelin protected ARPE-19 cells from H2O2-induced cell oxidative injuries and apoptosis responses. Furthermore, an oxidative stress-induced mouse model of AMD was established via injection of sodium iodate (NaIO3) to tail veins, and treatment with ghrelin preserved retinal function, and protected photoreceptors.

BACKGROUND: Sepsis-induced pulmonary injury (SPI) is a common complication of sepsis with a high rate of mortality. N4-acetylcytidine (ac4C) is mediated by the ac4C \"writer\", N-acetyltransferase (NAT)10, to regulate the stabilization of mRNA. This study aimed to investigate the role of NAT10 in SPI and the underlying mechanism.
METHODS: Twenty-three acute respiratory distress syndrome (ARDS) patients and 27 non-ARDS volunteers were recruited. A sepsis rat model was established. Reverse transcription-quantitative polymerase chain reaction was used to detect the expression of NAT10 and transferrin receptor (TFRC). Cell viability was detected by cell counting kit-8. The levels of Fe2+, glutathione, and malondialdehyde were assessed by commercial kits. Lipid reactive oxygen species production was measured by flow cytometric analysis. Western blot was used to detect ferroptosis-related protein levels. Haematoxylin & eosin staining was performed to observe the pulmonary pathological symptoms.
RESULTS: The results showed that NAT10 was increased in ARDS patients and lipopolysaccharide-treated human lung microvascular endothelial cell line-5a (HULEC-5a) cells. NAT10 inhibition increased cell viability and decreased ferroptosis in HULEC-5a cells. TFRC was a downstream regulatory target of NAT10-mediated ac4C acetylation. Overexpression of TFRC decreased cell viability and promoted ferroptosis. In in vivo study, NAT10 inhibition alleviated SPI.
CONCLUSIONS: NAT10-mediated ac4C acetylation of TFRC aggravated SPI through promoting ferroptosis.

The interplay between crystals and epithelial cells forms the cornerstone of kidney stone development, communication between epithelial cells and macrophages emerging as a pivotal role in this process. We conducted next-generation sequencing on the secreted exosomes of TCMK-1 cells treated with calcium oxalate monohydrate (OX_EXO) or controls (NC_EXO), and on the macrophage cell line RAW264.7 stimulated with OX_EXO or NC_EXO, followed by validation of differentially expressed target proteins and miRNAs through Western blot and PCR. UPSET plots were employed to identify genes co-targeted by exosomal miRNAs. Various bioinformatic analyses were employed to predict potential mechanisms of the dysregulated genes. We integrated sequencing data from the GEO database, and validated findings using clinical patient urine and kidney tissues. We identified 665 differentially expressed exosomal miRNAs between OX_EXO and NC_EXO. Among the top 10 down-regulated miRNAs, the most targeted genes were AAK1 and NUFIP2, whereas PLCB1 was significantly targeted among the top 10 up-regulated miRNAs. In clinical specimens, we confirmed the differential expressions of five homologous miRNAs, as well as CNOT3, CNCNA1C, APEX1, and TMEM199. In conclusion, treatment of TCMK-1 cells with calcium oxalate significantly alerted the expression profile of exosomal miRNAs, subsequently influencing gene expression in macrophages, thereby modulating the processes of kidney stone formation.

Aneuploidy is a hallmark of human cancer, yet the molecular mechanisms to cope with aneuploidy-induced cellular stresses remain largely unknown. Here, we induce chromosome mis-segregation in non-transformed RPE1-hTERT cells and derive multiple stable clones with various degrees of aneuploidy. We perform a systematic genomic, transcriptomic and proteomic profiling of 6 isogenic clones, using whole-exome DNA, mRNA and miRNA sequencing, as well as proteomics. Concomitantly, we functionally interrogate their cellular vulnerabilities, using genome-wide CRISPR/Cas9 and large-scale drug screens. Aneuploid clones activate the DNA damage response and are more resistant to further DNA damage induction. Aneuploid cells also exhibit elevated RAF/MEK/ERK pathway activity and are more sensitive to clinically-relevant drugs targeting this pathway, and in particular to CRAF inhibition. Importantly, CRAF and MEK inhibition sensitize aneuploid cells to DNA damage-inducing chemotherapies and to PARP inhibitors. We validate these results in human cancer cell lines. Moreover, resistance of cancer patients to olaparib is associated with high levels of RAF/MEK/ERK signaling, specifically in highly-aneuploid tumors. Overall, our study provides a comprehensive resource for genetically-matched karyotypically-stable cells of various aneuploidy states, and reveals a therapeutically-relevant cellular dependency of aneuploid cells.

Oxidative stress is a major pathogenic factor in many intestinal diseases, such as inflammatory bowel disease (IBD) and colorectal cancer (CRC). The Nrf2 signaling pathway and mitophagy can reduce reactive oxygen species (ROS) and alleviate oxidative stress, but their relationship is unclear. Hydroxytyrosol (HT), a polyphenolic compound abundant in olive oil, has strong antioxidant activity and may help treat these diseases. We used pigs as a model to investigate HT\'s effect on intestinal oxidative damage and its mechanisms. Diquat (DQ) induced oxidative stress and impaired intestinal barrier function, which HT mitigated. Mechanistic studies in IPEC-J2 cells showed that HT protected against oxidative damage by activating the PI3K/Akt-Nrf2 signaling pathway and promoting mitophagy. Our study highlighted the synergistic relationship between Nrf2 and mitophagy in mediating HT\'s antioxidant effects. Inhibition studies confirmed that disrupting either pathway compromised HT\'s protective effects. Maintaining redox balance through Nrf2 and mitophagy is important for eliminating excess ROS. Nrf2 increases antioxidant enzymes to clear existing ROS, while mitophagy removes damaged mitochondria and reduces ROS generation. This study demonstrates that these pathways collaboratively modulate the antioxidant effects of HT, with neither being dispensable. Targeting Nrf2 and mitophagy could be a promising strategy for treating oxidative stress-related intestinal diseases, with HT as a potential treatment.

Cisplatin, a chemotherapeutic drug, can result in acute kidney injury (AKI). Currently, there are no effective prevention methods. An incomplete understanding of the pathogenesis of AKI is a major barrier to the development of effective therapies. Metabolism reprogramming shift to glycolysis was involved in AKI pathogenesis. Glycolysis results in the pyruvate production. The mitochondrial pyruvate carrier (MPC) conveys cytosol pyruvate into mitochondria, promoting the tricarboxylic acid cycle. In this current study, we found a reduction in MPC2 expression in mice and cultured HK2 cells with cisplatin-induced AKI. MPC2 overexpression attenuated cisplatin-mediated nephrotoxicity both in vitro and in vivo via restoring pyruvate metabolism and mitochondrial function. Knockdown of MPC2 reversed this effect. Furthermore, artemether, an MPC2 potential activator, could mitigate AKI via regulating MPC2-mediated pyruvate metabolism. Our findings revealed that MPC2-pyruvate metabolism axis was a promising strategy to alleviate AKI induced by cisplatin.

UNASSIGNED: The high infectivity of coronaviruses has led to increased interest in developing new strategies to prevent virus spread. Silver nanoparticles (AgNPs) and graphene oxide (GO) have attracted much attention in the antiviral field. We investigated the potential antiviral activity of GO and AgNPs combined in the nanocomposite GO-Ag against murine betacoronavirus MHV using an in vitro model.
UNASSIGNED: GO, AgNPs, and GO-Ag characterization (size distribution, zeta potential, TEM visualization, FT-IR, and EDX analysis) and XTT assay were performed. The antiviral activity of GO-Ag nanocomposites was evaluated by RT-qPCR and TCID50 assays. The results were compared with free AgNPs and pure GO. Cell growth and morphology of MHV-infected hepatocytes treated with GO-Ag composites were analyzed by JuLI™Br. Immunofluorescence was used to visualize the cell receptor used by MHV. Ultrastructural SEM analysis was performed to examine cell morphology after MHV infection and GO-Ag composite treatment.
UNASSIGNED: A significant reduction in virus titer was observed for all nanocomposites tested, ranging from 3.2 to 7.3 log10 TCID50. The highest titer reduction was obtained for GO 5 µg/mL - Ag 25 µg/mL in the post-treatment method. These results were confirmed by RT-qPCR analysis. The results indicate that GO-Ag nanocomposites exhibited better antiviral activity compared to AgNPs and GO. Moreover, the attachment of AgNPs to the GO flake platform reduced their cytotoxicity. In addition, the GO-Ag composite modulates the distribution of the Ceacam1 cell receptor and can modulate cell morphology.
UNASSIGNED: Graphene oxide sheets act as a stabilizing agent, inhibiting the accumulation of AgNPs and reducing their cellular toxicity. The GO-Ag composite can physically bind and inhibit murine betacoronavirus from entering cells. Furthermore, the constant presence of GO-Ag can inhibit MHV replication and significantly limit its extracellular release. In conclusion, GO-Ag shows promise as an antiviral coating on solid surfaces to minimize virus transmission and spread.

Diabetes-related bone loss represents a significant complication that persistently jeopardizes the bone health of individuals with diabetes. Primary cilia proteins have been reported to play a vital role in regulating osteoblast differentiation in diabetes-related bone loss. However, the specific contribution of KIAA0753, a primary cilia protein, in bone loss induced by diabetes remains unclear. In this investigation, we elucidated the pivotal role of KIAA0753 as a promoter of osteoblast differentiation in diabetes. RNA sequencing demonstrated a marked downregulation of KIAA0753 expression in pro-bone MC3T3 cells exposed to a high glucose environment. Diabetes mouse models further validated the downregulation of KIAA0753 protein in the femur. Diabetes was observed to inhibit osteoblast differentiation in vitro, evidenced by downregulating the protein expression of OCN, OPN and ALP, decreasing primary cilia biosynthesis, and suppressing the Hedgehog signalling pathway. Knocking down KIAA0753 using shRNA methods was found to shorten primary cilia. Conversely, overexpression KIAA0753 rescued these changes. Additional insights indicated that KIAA0753 effectively restored osteoblast differentiation by directly interacting with SHH, OCN and Gli2, thereby activating the Hedgehog signalling pathway and mitigating the ubiquitination of Gli2 in diabetes. In summary, we report a negative regulatory relationship between KIAA0753 and diabetes-related bone loss. The clarification of KIAA0753\'s role offers valuable insights into the intricate mechanisms underlying diabetic bone complications.