Objective To investigate the role and potential mechanism of secreted phosphoprotein 1 (SPP1)⁺ macrophages in the formation of chronic renal allograft fibrosis. Methods The expression features of SPP1⁺ macrophages in renal allografts of chronic allograft dysfunction (CAD) patients were analyzed based on single-cell transcriptome data of renal tissues from patients with CAD. Transcription factor VIPER analysis and DoRothEA transcription factor activity analysis were performed on the single-cell transcriptome data. Renal tissue samples were collected from kidney transplant recipients, including the CAD group (n=5) and the non-renal allograft fibrosis group (CTL group, n=5). A mouse model of chronic allograft rejection was established and divided into the allogeneic kidney transplantation group (CAD group, n=3) and the syngeneic kidney transplantation group (SYN group, n=3). Hematoxylin-eosin staining was used to detect renal tissue injury in mice, and Masson staining was used to detect renal tissue fibrosis. Immunofluorescence staining was performed to detect SPP1 expression in renal tissues of transplant recipients and mouse renal allografts. Bone marrow-derived macrophages (BMDMs) were extracted from mice and subjected to hypoxia stimulation. The expression of hypoxia-inducible factor (HIF)-1α and SPP1 was detected by Western blot, and SPP1 expression was detected by flow cytometry. BMDMs were transfected with HIF-1α overexpression plasmid and HIF-1α small interfering RNA (siRNA) followed by hypoxia intervention, and the expression of HIF-1α and SPP1 was detected by Western blot. Mouse aortic endothelial cells (MAECs) were co-cultured with the supernatant of BMDMs, and the expression of endothelial-mesenchymal transition (EndMT)-related markers was detected by Western blot and immunofluorescence. Results Single-cell transcriptome analysis showed that the proportion of SPP1⁺ macrophages in renal allograft tissues was significantly higher in the CAD group than in the CTL group (P<0.05). The renal injury score and the percentage of interstitial fibrotic area in the CAD group were significantly higher than those in the SYN group (both P<0.05). Immunofluorescence staining showed that the proportion of SPP1⁺ macrophages was increased in the CAD group compared with the CTL group, and also increased in the CAD group compared with the SYN group (both P<0.05). VIPER analysis and DoRothEA transcription factor activity analysis revealed activation of the hypoxia pathway and upregulated expression of transcription factors such as HIF-1α in SPP1⁺ macrophages. SPP1 expression was elevated in BMDMs under hypoxic conditions. Knockdown of HIF-1α inhibited hypoxia-induced SPP1 protein expression, whereas overexpression of HIF-1α upregulated SPP1 protein levels. After co-culture of hypoxia-induced BMDMs with MAECs, the expression levels of EndMT-related markers were increased. Conclusions SPP1⁺ macrophages differentiated under hypoxia are significantly infiltrated in the formation of chronic renal allograft fibrosis, and may promote renal allograft fibrosis by inducing EndMT in renal vascular endothelial cells.

ObjectiveTo characterize the seasonal patterns of influenza in Kunming City, Yunnan Province before, during, and after the COVID-19 pandemic, and provide scientific evidence for optimizing influenza prevention and control strategies. MethodsInfluenza-like illness (ILI) and etiological surveillance data for influenza from the 14^th week of 2010 to the 13^th week of 2024 in Kunming City of Yunnan Province were collected. Harmonic regression models were constructed to analyze the epidemic characteristics and seasonal patterns of influenza before (2010/2011‒2019/2020 influenza seasons), during (2020/2021‒2022/2023 influenza seasons), and after (2023/2024 influenza season) the COVID-19 pandemic. ResultsBefore the COVID-19 pandemic, influenza in Kunming City mainly exhibited an annual cyclic pattern without a significant semi-annual periodicity, peaking from December to February of the next year, with an epidemic duration of 20‒30 weeks. During the pandemic, influenza seasonality shifted, with an increase in semi-annual periodicity and an approximate one month delay in annual peaks. However, after the pandemic, the annual amplitude of influenza increased compared with that before the pandemic, and the epidemic duration extended by about one month. Although the annual peak largely reverted to the pre-pandemic levels, the annual peaks for different influenza subtypes/lineages had not fully recovered. ConclusionInfluenza seasonality in Kunming City underwent substantial alterations following the COVID-19 pandemic and has not yet fully reverted to pre-pandemic levels. Continuous surveillance on different subtypes/lineages of influenza viruses remains essential, and prevention and control strategies should be adjusted and optimized in a timely manner based on current epidemic trends.

To explore the impacts of TGR5 on liver lipid metabolism and bile acid synthesis in dairy cows with fatty liver.Liver tissues of healthy cows and cows with fatty liver were collected through puncture technique.The protein and mRNA expressions of lipid synthesis-related factors ACC1,FAS,SREBF1,lipid oxidation factor CPT1A,and bile acid synthesis-related factors CYP8B1,CYP7B1,CYP27A1 were detected by Western blot and fluorescent quantitative PCR.Moreover,the mRNA levels of CYP7B1 were determined.Primary hepatocytes of 1-day-old calves were extracted and cultured in vitro,and four treatment groups were established,namely Control,NEFA,INT-777,and the INT-777+NEFA group.The concentration of NEFA group was 1.2 mmol/L,the con-centration of INT-777 group was 1 μmol/L,and the concentration of INT-777+NEFA group was 1.2 mmol/L NEFA and 1 μmol/L INT-777 simultaneously.After 12 h of stimulation,cells were collected,and the protein and mRNA levels of ACC1,FAS,SREBF1,CPT1A,CYP8B1,CYP7A1,CYP27A1,and the mRNA levels of CYP7B1 were detected by Western blot and fluorescent quanti-tative PCR.The content of lipid droplets and TG in the cells were detected by flow cytometry and kit.The results demonstrated that compared with healthy cows,the protein and mRNA expressions of ACC1,FAS,SREBF1,CYP8B1,and CYP7A1 in the liver tissues of fatty liver cows were upreg-ulated,while the protein and mRNA levels of CPT1 A,CYP27A1,TGR5,and the mRNA levels of CYP7B1 were downregulated.In vitro experiments revealed that compared with the Control group,the protein and mRNA levels of ACC1,FAS,SREBF1,CYP8B1,and CYP7A1 in the NEFA group were upregulated,and the protein and mRNA levels of CPT1A,CYP27A1,and TGR5,as well as the mRNA level of CYP7B1,were downregulated.Compared with the NEFA group,the protein and mRNA levels of ACC1,FAS,SREBF1,CYP8B1,CYP7A1 were downregulated in the INT-777+NEFA group,while the protein and mRNA levels of CPT1A CYP27A1,and TGR5 as well as the mRNA level of CYP7B1,were upregulated.The results of flow cytometry and the kit indicated that the lipid droplets and TG content in the NEFA group were upregulated compared with the Control group,while the lipid droplets and TG content in the INT-777+NEFA group were downregulated compared with the NEFA group.The above results suggested that the addition of TGR5 agonist promoted the expression of TGR5 and ameliorated the abnormal lipid metabolism and bile acid synthesis in the liver of dairy cows with fatty liver.

The purpose of this study was to investigate the effect of medium-chain fatty acids(MC-FAs)caprylic acid(C8∶0)on lipid metabolism of calf hepatocytes.Primary calf hepatocytes were extracted and cultured,and 1.2 mmol/L nonesterified fatty acids(NEFAs)were added to the hep-atocytes to construct a model of hepatic lipid deposition in primary calf hepatocytes,Five process-ing groups have been set up:Control group(Ctrl),NEFA added group(NEFA),C8∶0 1.2 mmol/L treatment group(C8∶0 1.2),NEFA+C8∶0 0.2 mmol/L treatment group(NEFA+C8∶00.2),C8∶0 0.2 mmol/L treatment group(C8∶0 0.2).Stimulate calf liver cells for 12 hours,and the levels of triglyceride(TG),lipid oxidation(MDA),hydrogen peroxide(H2O2)and total SOD activity were detected by biochemical kit,and FAS,a protein related to lipid synthesis,was detec-ted by Western blot.The results showed that compared with the control group,the concentrations of TG,MDA and H2O2 in NEFA group increased significantly(P＜0.01),and the activity of SOD decreased significantly(P＜0.05).The protein expression levels of FAS,ACC1,DGAT2 and SREBP-1C were significantly up-regulated(P＜0.01),while the expression level of CPT1A was significantly down-regulated(P＜0.01).Compared with the NEFA group,the protein expression levels of SREBP-1C and DGAT2 in the NEFA+C8∶0(concentration 0.2 mmol/L)group de-creased significantly(P＜0.05),and the protein expression level of fatty acid β-oxidation related molecule CPT1A was slightly higher than that in the NEFA group,but there was no statistical sig-nificance(P＞0.05),and the MDA level in hepatocytes decreased significantly(P＜0.05).In a word,the results of this study show that C8∶0 has antioxidant effect,which can effectively reduce the liver injury caused by oxidative stress,regulate the expression of liver fat gene,and then pro-tect liver injury.

Objective To develop and validate an automated organoid image segmentation approach based on deep learning for addressing the issues of high misidentification rate,blurred boundary and poor generalization in current organoid segmentation,thereby facilitating researchers to monitor and analyze organoid growth more efficiently.Methods The wavelet scattering coefficient matrix and capsule convolution module were integrated into the U-Net architecture to construct the organoid image segmentation model OrgCapsU-Net which was trained and evaluated on 3 organoid image datasets from different tissue sources.Results Compared with current mainstream segmentation algorithms,OrgCapsU-Net could better distinguish organoid and impurity,and lead to smoother segmentation boundaries,achieving superior performance across 4 evaluation metrics on 3 datasets.Conclusion OrgCapsU-Net delivers excellent segmentation performance and can be applied to organoids from various tissue sources,showing strong potential for applications in the in vitro model establishment,high-throughput drug screening,and personalized medicine.

To explore the impacts of TGR5 on liver lipid metabolism and bile acid synthesis in dairy cows with fatty liver.Liver tissues of healthy cows and cows with fatty liver were collected through puncture technique.The protein and mRNA expressions of lipid synthesis-related factors ACC1,FAS,SREBF1,lipid oxidation factor CPT1A,and bile acid synthesis-related factors CYP8B1,CYP7B1,CYP27A1 were detected by Western blot and fluorescent quantitative PCR.Moreover,the mRNA levels of CYP7B1 were determined.Primary hepatocytes of 1-day-old calves were extracted and cultured in vitro,and four treatment groups were established,namely Control,NEFA,INT-777,and the INT-777+NEFA group.The concentration of NEFA group was 1.2 mmol/L,the con-centration of INT-777 group was 1 μmol/L,and the concentration of INT-777+NEFA group was 1.2 mmol/L NEFA and 1 μmol/L INT-777 simultaneously.After 12 h of stimulation,cells were collected,and the protein and mRNA levels of ACC1,FAS,SREBF1,CPT1A,CYP8B1,CYP7A1,CYP27A1,and the mRNA levels of CYP7B1 were detected by Western blot and fluorescent quanti-tative PCR.The content of lipid droplets and TG in the cells were detected by flow cytometry and kit.The results demonstrated that compared with healthy cows,the protein and mRNA expressions of ACC1,FAS,SREBF1,CYP8B1,and CYP7A1 in the liver tissues of fatty liver cows were upreg-ulated,while the protein and mRNA levels of CPT1 A,CYP27A1,TGR5,and the mRNA levels of CYP7B1 were downregulated.In vitro experiments revealed that compared with the Control group,the protein and mRNA levels of ACC1,FAS,SREBF1,CYP8B1,and CYP7A1 in the NEFA group were upregulated,and the protein and mRNA levels of CPT1A,CYP27A1,and TGR5,as well as the mRNA level of CYP7B1,were downregulated.Compared with the NEFA group,the protein and mRNA levels of ACC1,FAS,SREBF1,CYP8B1,CYP7A1 were downregulated in the INT-777+NEFA group,while the protein and mRNA levels of CPT1A CYP27A1,and TGR5 as well as the mRNA level of CYP7B1,were upregulated.The results of flow cytometry and the kit indicated that the lipid droplets and TG content in the NEFA group were upregulated compared with the Control group,while the lipid droplets and TG content in the INT-777+NEFA group were downregulated compared with the NEFA group.The above results suggested that the addition of TGR5 agonist promoted the expression of TGR5 and ameliorated the abnormal lipid metabolism and bile acid synthesis in the liver of dairy cows with fatty liver.

The purpose of this study was to investigate the effect of medium-chain fatty acids(MC-FAs)caprylic acid(C8∶0)on lipid metabolism of calf hepatocytes.Primary calf hepatocytes were extracted and cultured,and 1.2 mmol/L nonesterified fatty acids(NEFAs)were added to the hep-atocytes to construct a model of hepatic lipid deposition in primary calf hepatocytes,Five process-ing groups have been set up:Control group(Ctrl),NEFA added group(NEFA),C8∶0 1.2 mmol/L treatment group(C8∶0 1.2),NEFA+C8∶0 0.2 mmol/L treatment group(NEFA+C8∶00.2),C8∶0 0.2 mmol/L treatment group(C8∶0 0.2).Stimulate calf liver cells for 12 hours,and the levels of triglyceride(TG),lipid oxidation(MDA),hydrogen peroxide(H2O2)and total SOD activity were detected by biochemical kit,and FAS,a protein related to lipid synthesis,was detec-ted by Western blot.The results showed that compared with the control group,the concentrations of TG,MDA and H2O2 in NEFA group increased significantly(P＜0.01),and the activity of SOD decreased significantly(P＜0.05).The protein expression levels of FAS,ACC1,DGAT2 and SREBP-1C were significantly up-regulated(P＜0.01),while the expression level of CPT1A was significantly down-regulated(P＜0.01).Compared with the NEFA group,the protein expression levels of SREBP-1C and DGAT2 in the NEFA+C8∶0(concentration 0.2 mmol/L)group de-creased significantly(P＜0.05),and the protein expression level of fatty acid β-oxidation related molecule CPT1A was slightly higher than that in the NEFA group,but there was no statistical sig-nificance(P＞0.05),and the MDA level in hepatocytes decreased significantly(P＜0.05).In a word,the results of this study show that C8∶0 has antioxidant effect,which can effectively reduce the liver injury caused by oxidative stress,regulate the expression of liver fat gene,and then pro-tect liver injury.

Objective To develop and validate an automated organoid image segmentation approach based on deep learning for addressing the issues of high misidentification rate,blurred boundary and poor generalization in current organoid segmentation,thereby facilitating researchers to monitor and analyze organoid growth more efficiently.Methods The wavelet scattering coefficient matrix and capsule convolution module were integrated into the U-Net architecture to construct the organoid image segmentation model OrgCapsU-Net which was trained and evaluated on 3 organoid image datasets from different tissue sources.Results Compared with current mainstream segmentation algorithms,OrgCapsU-Net could better distinguish organoid and impurity,and lead to smoother segmentation boundaries,achieving superior performance across 4 evaluation metrics on 3 datasets.Conclusion OrgCapsU-Net delivers excellent segmentation performance and can be applied to organoids from various tissue sources,showing strong potential for applications in the in vitro model establishment,high-throughput drug screening,and personalized medicine.

Developing universal CARs with improved flexible targeting and controllable activities is urgently needed. While several studies have suggested the potential of CD16a in tandem with monoclonal antibodies to construct universal CAR-T cells, the weak affinity between them is one of the limiting factors for efficacy. Herein, we systematically investigated the impact of Fcγ receptor (FcγR) affinity on CAR-T cells properties by constructing universal CARs using Fcγ receptors with different affinities for IgG1 antibodies, namely CD16a, CD32a, and CD64. We demonstrated that the activities of these universal CAR-T cells on tumor cells could be redirected and regulated by IgG1 antibodies. In xenografted mice, 64CAR chimeric Jurkat cells with the highest affinity showed significant antitumor effects in combination with herceptin in the HER2 low expression U251 MG model. However, in the CD20 high expression Raji model, 64CAR caused excessive activation of CAR-T cells, which resulted in cytokine release syndrome (CRS) and the decline of antitumor activity, and 32CAR with a moderate affinity brought the best efficacy. Our work extended the knowledge about FcγR-based universal CAR-T cells and suggested that only the FcγRCAR with an appropriate affinity can offer the optimal antitumor advantages of CAR-T cells.

Objective:To observe the postoperative complications and revision rates of robot arm-assisted unicompartment arthroplasty by means of a meta-analysis.Methods:Relevant databases including Cochrane Library, PubMed, EMBASE, Wanfang, VIP, CNKI, and Web of Science were searched by computer for high-quality studies on complications and revision rates after robot arm-assisted unicompartment arthroplasty in both English and Chinese from the database establishment date to March 2021. The quality of the studies retrieved was evaluated. Relevant data including postoperative complications, infection, pain, prosthesis loosening, and revision were extracted for a meta-analysis using STATA 15.0 software.Results:A total of 16 studies were included, including one randomized controlled study, 6 case-control studies and 9 cohort studies. By the methodological index for non-randomized studies (MINORS), 7 studies scored 14 points, 3 studies 13 points, one study 12 points, 4 studies 11 points, and one study 10 points. Meta analysis showed that the total rate of complications was 2% (95% CI: 1%to 4%) . Three studies used NAVIO robot, 7 studies MAKO robot, one study NAVIO and MAKO robots, and one study Acrobot robot. Since just one study used Acrobot robot, only MAKO and NAVIO robots were included for the subgroup analysis which showed that the postoperative complication rates for NAVIO and MAKO robots were 4.0% (95% CI: -2% to 10%) and 3% (95% CI: 1% to 5%) , respectively. The incidence of postoperative pain was 0.2% (95% CI: 0.1% to 0.3%), the incidence of postoperative infection 0.5% (95% CI: 0.3% to 0.8%), the incidence of postoperative prosthesis loosening 0.5% (95% CI: 0.3% to 0.8%), and the revision rate 2% (95% CI: 1% to 2%). According to the subgroup analysis of NAVIO and MAKO robots, their revision rates were 4% (95% CI: 2% to 7%) and 2% (95% CI: 1% to 2%), respectively. Conclusion:The clinical efficacy of robot arm-assisted unicompartment arthroplasty is good, for the complications in the patients are limited and the long-term survival rate of the prosthesis is excellent.