AIM: To investigate the potential inhibitory effect of pterostilbene on the endothelial-to-mesenchymal transition(EndMT)induced by high glucose conditions in human retinal microvascular endothelial cells(HRMECs).METHODS: The optimal concentration of pterostilbene for treating HRMECs was determined using the CCK-8 assay, with 12.5 and 25 μmol/L concentrations selected for subsequent experiments. Four experimental groups were established: control group, high glucose group, high glucose combined with 12.5 μmol/L pterostilbene treatment group, and high glucose combined with 25 μmol/L pterostilbene treatment group. The expression levels of HDAC7 and EndMT-associated markers were detected via Western blot analysis. Cell migration ability was assessed using Transwell migration assays and scratch wound healing tests, while vasculogenic capability was evaluated through tube formation assays.RESULTS: The CCK-8 assay revealed that pterostilbene at a concentration of 22.07 μmol/L inhibited 50% of cell viability in HRMECs. Western blot analysis demonstrated that compared with the control group, the expression levels of HDAC7, ZEB1, Vimentin, and Snail were significantly upregulated in HRMECs cultured in high glucose(all P<0.01), while the expressions of VE-cadherin and CD31 were significantly reduced(all P<0.01). Compared to the high glucose group, the treatment with 12.5 and 25 μmol/L pterostilbene significantly reduced the expression of HDAC7, ZEB1, Vimentin, and Snail under high glucose conditions(all P<0.01). Notably, 25 μmol/L pterostilbene enhanced the expression of VE-cadherin and CD31(all P<0.01). Scratch wound healing tests revealed that HRMECs treated with high glucose exhibited a significantly increased cell migration rate compared to the control group(P<0.05), while the application of 25 μmol/L pterostilbene significantly suppressed HRMECs migration under high glucose conditions(P<0.01). Transwell migration assays demonstrated that the cell migration rate in the high glucose group was significantly higher than that in the control group(P<0.01), with cell migration rate markedly reduced following treatment with both of 12.5 and 25 μmol/L pterostilbene(all P<0.01). The tube formation assay revealed that the ability of HRMECs to form tubular structures was significantly enhanced under high glucose conditions(P<0.01), and both 12.5 and 25 μmol/L of pterostilbene effectively inhibited this effect(all P<0.01).CONCLUSION: Pterostilbene can inhibit HDAC7 expression, suppress EndMT-mediated migration of HRMECs, and impair tube formation under high-glucose conditions.

AIM: To investigate the potential inhibitory effect of pterostilbene on the endothelial-to-mesenchymal transition(EndMT)induced by high glucose conditions in human retinal microvascular endothelial cells(HRMECs).METHODS: The optimal concentration of pterostilbene for treating HRMECs was determined using the CCK-8 assay, with 12.5 and 25 μmol/L concentrations selected for subsequent experiments. Four experimental groups were established: control group, high glucose group, high glucose combined with 12.5 μmol/L pterostilbene treatment group, and high glucose combined with 25 μmol/L pterostilbene treatment group. The expression levels of HDAC7 and EndMT-associated markers were detected via Western blot analysis. Cell migration ability was assessed using Transwell migration assays and scratch wound healing tests, while vasculogenic capability was evaluated through tube formation assays.RESULTS: The CCK-8 assay revealed that pterostilbene at a concentration of 22.07 μmol/L inhibited 50% of cell viability in HRMECs. Western blot analysis demonstrated that compared with the control group, the expression levels of HDAC7, ZEB1, Vimentin, and Snail were significantly upregulated in HRMECs cultured in high glucose(all P<0.01), while the expressions of VE-cadherin and CD31 were significantly reduced(all P<0.01). Compared to the high glucose group, the treatment with 12.5 and 25 μmol/L pterostilbene significantly reduced the expression of HDAC7, ZEB1, Vimentin, and Snail under high glucose conditions(all P<0.01). Notably, 25 μmol/L pterostilbene enhanced the expression of VE-cadherin and CD31(all P<0.01). Scratch wound healing tests revealed that HRMECs treated with high glucose exhibited a significantly increased cell migration rate compared to the control group(P<0.05), while the application of 25 μmol/L pterostilbene significantly suppressed HRMECs migration under high glucose conditions(P<0.01). Transwell migration assays demonstrated that the cell migration rate in the high glucose group was significantly higher than that in the control group(P<0.01), with cell migration rate markedly reduced following treatment with both of 12.5 and 25 μmol/L pterostilbene(all P<0.01). The tube formation assay revealed that the ability of HRMECs to form tubular structures was significantly enhanced under high glucose conditions(P<0.01), and both 12.5 and 25 μmol/L of pterostilbene effectively inhibited this effect(all P<0.01).CONCLUSION: Pterostilbene can inhibit HDAC7 expression, suppress EndMT-mediated migration of HRMECs, and impair tube formation under high-glucose conditions.

Background At present, the treatment of silicosis is still limited, and no method is available to cure the disease. miRNAs are involved in the process of fibrosis at the transcriptional level by directly degrading target gene mRNA or inhibiting its translation. However, how miR-130a-3p regulates silicosis fibrosis has not been fully elucidated yet. Objective To investigate whether miR-130a-3p promotes epithelial-mesenchymal transition (EMT) by inhibiting peroxisome proliferators-activated receptors gamma (PPAR-γ), thereby pro-moting the process of silicotic fibrosis. To identify effective new targets for the treatment of silicotic fibrosis. Methods (1) Animal experiments: C57BL/6J mice were intratracheally injected with a one-time dose of 10 mg silica suspension (dissolved in 100 μL saline) as positive lung exposure. A silicosis model group was established 28 d after the exposure. A control group was injected with the same amount of normal saline into the trachea. Hematoxylin-eosin staining and Sirius red staining were used to observe the pathological changes and collagen deposition in lung tissues respectively. Realtime fluorescence-based quantitative polymerase chain reaction (RT-qPCR) was used to assay the expression of miR-130a-3p and PPAR-γ mRNA in lung tissues. Western blotting was used to detect the protein expression of PPAR-γ, transforming growth factor (TGF)-β1, E-cadherin, α-smooth muscle actin (α-SMA), and Collagen Ⅰ in lung tissues. (2) Cells experiments: Mouse lung epithelial cells (MLE-12) were induced with 5 µg·L⁻¹ TGF-β1 for different time (0, 12, 24, 48 h). RT-qPCR was used to detect the expression of miR-130a-3p and PPAR-γ mRNA in cells. The binding relationship between miR-130a-3p and PPAR-γ mRNA was verified by dual luciferase reporter gene assay. MLE-12 cells were stimulated by 5 µg·L⁻¹ TGF-β1 after transfection of miR-130a-3p inhibitor, and Western blotting was used to measure the protein expression of PPAR-γ, E-cadherin, and α-SMA in the TGF-β1-induced cells. Results In the silicosis model group, the alveolar septum was widened and the pulmonary nodules were formed. The Sirius red staining collagen deposition in pulmonary nodules indicated that a silicosis fibrosis model was successfully established. The expressions of TGF-β1, α-SMA, and Collagen Ⅰ proteins were increased, and the expressions of E-cadherin and PPAR-γ proteins were decreased in lung tissues of the silicosis group, compared with the control group (P<0.05 or P<0.01). The expression of miR-130a-3p was increased and the expression of PPAR-γ mRNA was decreased in lung tissues of the silicosis model (P<0.01). The expression of miR-130a-3p was significantly increased, while the expression of PPAR-γ mRNA was decreased in the TGF-β1 induced MLE-12 cells (P<0.05 or P<0.01). The dual luciferase reporter assay showed a direct relationship between miR-130a-3p and PPAR-γ mRNA in MLE-12 cells. The transfection of miR-130a-3p inhibitor in the TGF-β1 induced MLE-12 cells inhibited the decrease of PPAR-γ and E-cadherin proteins, and the increase of α-SMA protein in the MLE-12 cells induced by TGF-β1 (P<0.05 or P<0.01). Conclusion miR-130a-3p promotes the development of silicosis fibrosis by targeting PPAR-γ to increase pulmonary EMT.

Epidermal growth factor receptor tyrosine kinase inhibitors （EGFR-TKIs） are targeted drugs for the treatment of advanced non-small cell lung cancer （NSCLC）， but long-term use inevitably leads to drug resistance. Resistance to EGFR-TKIs can alter the tumor microenvironment， and patients with NSCLC resistant to EGFR-TKIs can regain the benefits of immune checkpoint inhibitors （ICIs）， but the changes in the tumor microenvironment are complex and the efficacy is unclear. This article reviews the clinical studies of ICIs in the treatment of EGFR-TKIs-resistant NSCLC， and finds that for patients with EGFR-TKIs-resistant NSCLC， the efficacy of ICIs as a single agent is unclear， and other relevant biomarkers need to be found to screen the beneficiary population. ICIs+EGFR-TKIs have potential toxicity and are not recommended for clinical use. There is controversy about the efficacy of ICIs+chemotherapy， and it is recommended to use it cautiously in clinical practice. ICIs+anti-vascular endothelial growth factor （VEGF） drug therapy has a synergistic effect， but may increase the incidence of adverse events. ICIs+chemotherapy+anti- VEGF drug have a synergistic effect and the incidence of adverse events is similar to that of chemotherapy. New ICIs such as lymphocyte activating gene 3 inhibitors are still in the clinical research stage or preclinical research stage， but they may be a new promising treatment.

Epidermal growth factor receptor tyrosine kinase inhibitors （EGFR-TKIs） are targeted drugs for the treatment of advanced non-small cell lung cancer （NSCLC）， but long-term use inevitably leads to drug resistance. Resistance to EGFR-TKIs can alter the tumor microenvironment， and patients with NSCLC resistant to EGFR-TKIs can regain the benefits of immune checkpoint inhibitors （ICIs）， but the changes in the tumor microenvironment are complex and the efficacy is unclear. This article reviews the clinical studies of ICIs in the treatment of EGFR-TKIs-resistant NSCLC， and finds that for patients with EGFR-TKIs-resistant NSCLC， the efficacy of ICIs as a single agent is unclear， and other relevant biomarkers need to be found to screen the beneficiary population. ICIs+EGFR-TKIs have potential toxicity and are not recommended for clinical use. There is controversy about the efficacy of ICIs+chemotherapy， and it is recommended to use it cautiously in clinical practice. ICIs+anti-vascular endothelial growth factor （VEGF） drug therapy has a synergistic effect， but may increase the incidence of adverse events. ICIs+chemotherapy+anti- VEGF drug have a synergistic effect and the incidence of adverse events is similar to that of chemotherapy. New ICIs such as lymphocyte activating gene 3 inhibitors are still in the clinical research stage or preclinical research stage， but they may be a new promising treatment.

Objective To establish a rat model of Alzheimer's disease (AD) expressing human triple mutant amyloid precursor protein (APP) in the hippocampus, and to provide a model for the study of disease mechanisms and drug development. Methods Twenty-four 12-week-old SPF-grade female SD rats were randomly divided into a blank control group, a virus control group and an experimental group, with eight rats in each group; among them, the experimental group received a stereotaxic injection of adeno-associated virus (AAV) carrying the human triple mutant APP and NanoLuc luciferase genes into the hippocampus. In vivo imaging was used to observe viral expression in the brains of rats in each group, the novel object recognition test was used to assess the recognition memory of the rats in each group, real-time fluorescent quantitative PCR was used to detect the expression level of the APP gene, HE staining was used to examine the brain histopathology, Nissl staining was used to assess the hippocampal lesions, and immunohistochemistry was used to detect the deposition of amyloid β-protein (Aβ). Results In vivo imaging showed that reporter fluorescence was detected in the brains of rats in both experimental and virus control groups. Fluorescence quantitative PCR showed that the expression level of the APP gene was significantly increased in the brains of rats in the experimental group (P<0.01). Novel object recognition test revealed that the recognition memory of rats in the experimental group was significantly reduced compared with that of the blank control group (P<0.01). Six months after recombinant AAV virus infection, HE staining and Nissl staining of brain tissues showed that the number of neurons and Nissl bodies in the CA1 region of the hippocampus in the experimental group was reduced and disorganized; immuno-histochemistry testing of the CA1 region of the hippocampus and the pyramidal cell layer of the experimental group revealed prominent brown deposits, indicating Aβ protein deposition. Conclusion The rat model successfully established by stereotaxic injection and AAV-mediated delivery of human triple mutant APP gene exhibits typical AD features, providing a valuable animal model for studying AD pathology and developing drug therapies targeting Aβ protein deposition.

Objective To establish a rat model of Alzheimer's disease (AD) expressing human triple mutant amyloid precursor protein (APP) in the hippocampus, and to provide a model for the study of disease mechanisms and drug development. Methods Twenty-four 12-week-old SPF-grade female SD rats were randomly divided into a blank control group, a virus control group and an experimental group, with eight rats in each group; among them, the experimental group received a stereotaxic injection of adeno-associated virus (AAV) carrying the human triple mutant APP and NanoLuc luciferase genes into the hippocampus. In vivo imaging was used to observe viral expression in the brains of rats in each group, the novel object recognition test was used to assess the recognition memory of the rats in each group, real-time fluorescent quantitative PCR was used to detect the expression level of the APP gene, HE staining was used to examine the brain histopathology, Nissl staining was used to assess the hippocampal lesions, and immunohistochemistry was used to detect the deposition of amyloid β-protein (Aβ). Results In vivo imaging showed that reporter fluorescence was detected in the brains of rats in both experimental and virus control groups. Fluorescence quantitative PCR showed that the expression level of the APP gene was significantly increased in the brains of rats in the experimental group (P<0.01). Novel object recognition test revealed that the recognition memory of rats in the experimental group was significantly reduced compared with that of the blank control group (P<0.01). Six months after recombinant AAV virus infection, HE staining and Nissl staining of brain tissues showed that the number of neurons and Nissl bodies in the CA1 region of the hippocampus in the experimental group was reduced and disorganized; immuno-histochemistry testing of the CA1 region of the hippocampus and the pyramidal cell layer of the experimental group revealed prominent brown deposits, indicating Aβ protein deposition. Conclusion The rat model successfully established by stereotaxic injection and AAV-mediated delivery of human triple mutant APP gene exhibits typical AD features, providing a valuable animal model for studying AD pathology and developing drug therapies targeting Aβ protein deposition.

Objective To establish a dynamic model of lipopolysaccharide-induced acute lung injury in mice based on the NLRP3/Caspase-1/gasdermin D(GSDMD)pyroptosis pathway,and observe the result ing lung injury at different time points.We aimed to identify the optimal time for modelling according to the injury at different time points and the expression of pyroptosis pathway-related proteins,to lay the foundation for animal models for subsequent experiments.Methods Fifty-four 6～8 weeks old male SPF BALB/c mice were divided randomly into nine groups,including Con group and model groups at 1,3,6,12,18,24,48,and 72 h.Body weight and lung tissue were detected by general and pathological observations and semi-quantitative scoring,including lung index,lung water content,and wet and dry weight ratio.The white blood cell count and concentrations of tumor necrosis factor-α,interleukin(IL)-6,IL-1β,IL-18,and BCA protein were detected in bronchoalveolar lavage fluid(BALF).The classic pyroptosis pathway-related proteins NLRP3,pro-Caspase 1,Caspase 1,and GSDMD were detected by Western Blot.Results Body weight decreased in all experimental groups,with the most significant weight loss in the 24 and 48 h groups.Gross observation and pathological examination of lung tissue showed that the most severe lung injury occurred at 24～72 h,with significant differences between each group and the control group.The lung index,lung water content,and wet/dry weight ratio were also significantly increased at 24～72 h.White blood cells in BALF started to increase from 6 h after model initiation,48 h can reach a peak,72 h all keep increasing.IL-18 in BALF began to increase at 24 h and continued to increase at 72 h.The inflammatory factors tumor necrosis factor-α,IL-1β,IL-6 were highest at 6 h and significantly reduced at 48 h.Protein concentrations in BALF were significantly increased within 24,48,and 72 h compared with those in the control group.The pyroptosis pathway proteins NLRP3,pro-Caspase-1,Caspase-1,and GSDMD were significantly enhanced in each time series,and channel protein expression was significantly enhanced at 24～72 h compared with that in the Con group.Conclusions Comprehensive analysis of experimental indicators,inflammatory factors,and pathway proteins at different times showed that the mechanism of pyroptosis was closely related to the occurrence and progression of acute lung injury.Expression of the pyroptosis pathway was most obvious and lung injury was most serious at 24～48 h.This study provides a model reference and experimental basis for subsequent studies of the specific mechanism and intervention targets of acute lung injury.

Objective To explore the prognostic value of the coronary angiography-derived index of microcirculatory resistance(caIMR)for major adverse cardiovascular events(MACE)in patients with acute ST-segment elevation myocardial infarction(STEMI)after primary percutaneous coronary intervention(PCI).Methods Between September 2019 and March 2022,541 patients diagnosed with STEMI at the Affiliated Hospital of Xuzhou Medical University were enrolled.The caIMR was calculated using the FlashAngio system(Suzhou Rainmed Medical Technology Co.,Ltd.).The patients were divided into MACE and non-MACE groups according to the occurrence of MACE during hospitalization or follow-up,with MACE defined as all-cause mortality,heart failure readmission,and unplanned revascularization.COX regression analysis,receiver operating characteristic(ROC)curves,and Kaplan-Meier survival curves were used to evaluate the prognostic value of caIMR for STEMI patients after primary PCI.Results During the 1-year follow-up,61 patients(11.28％)experienced MACE.The patients in the MACE group had higher caIMR values than those in the non-MACE group.Multivariate COX analysis showed that caIMR was an independent risk factor for MACE.ROC curve analysis showed that caIMR predicted MACE with an area under the curve of 0.688,and the optimal cutoff value was 25.3 U.caIMR significantly increased the discriminant and reclassification indexes when added to a model with clinical risk factors.The patients were further divided into a low caIMR group(caIMR＜25 U,n=377)and a high caIMR group(caIMR ≥25 U,n=164).Kaplan-Meier curve showed that patients with caIMR≥25 U had a worse prognosis.Conclusions caIMR is an independent risk factor for poor prognosis after PCI in patients with STEMI,and patients with caIMR≥25 U had a worse prognosis.

To investigate the potential therapeutic effects of the myelocytomatosis oncogene(c-Myc)using pharmacogenomic data from liv-er cancer cell lines and organoids.Methods:Drug sensitivity,functional genomics,and gene expression data from cell lines were collected to assess the correlation between the expression of c-Myc and drug sensitivity.Differential expression and prognostic relevance of c-Myc in a primary liver cancer cohort were analyzed.Immunohistochemistry(IHC)staining validated c-Myc expression in liver cancer patients and cor-related it with the drug sensitivity of organoids obtained from an organoid biobank.Three c-Myc inhibitors were screened on liver cancer cell lines and organoids to test tumor-killing effects in two-and three-dimensional levels.Bioinformatics analysis was conducted to explore the role of c-Myc in targeted drug resistance.Results:Bioinformatics analyses combined with IHC staining revealed a potential regulatory role of c-Myc in the drug sensitivity of liver cancer.The expression of c-Myc in cancerous tissues was significantly higher than in para-cancerous tis-sues,and c-Myc expression was higher in patients resistant to targeted drugs than in those sensitive to targeted drugs(P<0.05).Drug screening experiments showed that c-Myc inhibitors including THZ1,ABBV-744,and JQ1 reduced cell viability in both tumor cells and tumor organoids.The combination of lenvatinib and c-Myc inhibitors exhibited significant synergistic effects(all synergy scores>1)in the SNU-739 cell line.Analysis of organoids from an organoid biobank indicated a strong association between c-Myc and stemness-mediated targeted drug resistance(R=0.87).Conclusions:c-Myc showed increased expression in patients resistant to targeted therapy,suggesting it as a novel target for drugs in liver cancer.Inhibition of c-Myc effectively killed tumors in both two-and three-dimensional ex vivo models.