In protein engineering, while computational models are increasingly used to predict mutation effects, their evaluations primarily rely on high-throughput deep mutational scanning (DMS) experiments that use surrogate readouts, which may not adequately capture the complex biochemical properties of interest. Many proteins and their functions cannot be assessed through high-throughput methods due to technical limitations or the nature of the desired properties, and this is particularly true for the real industrial application scenario. Therefore, the desired testing datasets, will be small-size (∼10-100) experimental data for each protein, and involve as many proteins as possible and as many properties as possible, which is, however, lacking. Here, we present VenusMutHub, a comprehensive benchmark study using 905 small-scale experimental datasets curated from published literature and public databases, spanning 527 proteins across diverse functional properties including stability, activity, binding affinity, and selectivity. These datasets feature direct biochemical measurements rather than surrogate readouts, providing a more rigorous assessment of model performance in predicting mutations that affect specific molecular functions. We evaluate 23 computational models across various methodological paradigms, such as sequence-based, structure-informed and evolutionary approaches. This benchmark provides practical guidance for selecting appropriate prediction methods in protein engineering applications where accurate prediction of specific functional properties is crucial.

Objective To investigate the effects of scribble planar cell polarity protein(SCRIB)on proliferation,apoptosis,and autophagy of glioblastoma(GBM)and elucidate its potential underlying mechanisms.Methods The expression level of SCRIB in GBM tissue was queried through the Biomarker Exploration of Solid Tumors(BEST)database.Lentivirus-mediated shRNA interference was employed to downregulate SCRIB expression in human glioblastoma cell lines U87 and U251,which were divided into negative control group(mock)and SCRIB shRNA interference groups(kd1 and kd2).SCRIB expression levels were detected using Western blotting(WB)and quantitative polymerase chain reaction(qPCR).EdU incorporation and cell apoptosis rates were detected by flow cytometry(FCM).CCK-8 assay was used to detect the proliferation vitality of U87 and U251 cells,and WB was used to detect the expression of proliferation-related proteins.Immunofluorescence(IF)staining was conducted to detect the expression of autophagy-related proteins LC3 and p62,followed by quantitative analysis across multiple fields.WB was also used to detect the expression levels of LC3,p62,and proteins in the JAK-STAT3 signaling pathway.Results Compared with that of normal tissues,SCRIB mRNA expression level was significantly upregulated in GBM tissues(P＜0.05).FCM results showed that EdU incorporation rates were significantly reduced(P＜0.001)while cell apoptosis rates were markedly increased(P＜0.001)in U87 and U251 cells with SCRIB knockdown.CCK-8 results indicated that compared with the mock group,the proliferation vitality of U87 and U251 cells in the SCRIB knockdown group was significantly downregulated(P＜0.001).IF staining showed that LC3 fluorescence aggregation was significantly enhanced(P＜0.001),while p62 fluorescence aggregation was significantly reduced(P＜0.001)in the SCRIB knockdown group.WB results showed that compared with the mock group,the protein expression levels of p27,LC3,p-JAK2 and p-STAT3 were upregulated,while C-Myc,Cyclin D1,MCM,PCNA and p62 were downregulated,with statistically significant differences(P＜0.05).Conclusion Downregulation of SCRIB may induce autophagy and apoptosis in glioblastoma cells by inhibiting the JAK-STAT3 signaling pathway,thereby suppressing cell proliferation.

Objective To investigate the effects of scribble planar cell polarity protein(SCRIB)on proliferation,apoptosis,and autophagy of glioblastoma(GBM)and elucidate its potential underlying mechanisms.Methods The expression level of SCRIB in GBM tissue was queried through the Biomarker Exploration of Solid Tumors(BEST)database.Lentivirus-mediated shRNA interference was employed to downregulate SCRIB expression in human glioblastoma cell lines U87 and U251,which were divided into negative control group(mock)and SCRIB shRNA interference groups(kd1 and kd2).SCRIB expression levels were detected using Western blotting(WB)and quantitative polymerase chain reaction(qPCR).EdU incorporation and cell apoptosis rates were detected by flow cytometry(FCM).CCK-8 assay was used to detect the proliferation vitality of U87 and U251 cells,and WB was used to detect the expression of proliferation-related proteins.Immunofluorescence(IF)staining was conducted to detect the expression of autophagy-related proteins LC3 and p62,followed by quantitative analysis across multiple fields.WB was also used to detect the expression levels of LC3,p62,and proteins in the JAK-STAT3 signaling pathway.Results Compared with that of normal tissues,SCRIB mRNA expression level was significantly upregulated in GBM tissues(P＜0.05).FCM results showed that EdU incorporation rates were significantly reduced(P＜0.001)while cell apoptosis rates were markedly increased(P＜0.001)in U87 and U251 cells with SCRIB knockdown.CCK-8 results indicated that compared with the mock group,the proliferation vitality of U87 and U251 cells in the SCRIB knockdown group was significantly downregulated(P＜0.001).IF staining showed that LC3 fluorescence aggregation was significantly enhanced(P＜0.001),while p62 fluorescence aggregation was significantly reduced(P＜0.001)in the SCRIB knockdown group.WB results showed that compared with the mock group,the protein expression levels of p27,LC3,p-JAK2 and p-STAT3 were upregulated,while C-Myc,Cyclin D1,MCM,PCNA and p62 were downregulated,with statistically significant differences(P＜0.05).Conclusion Downregulation of SCRIB may induce autophagy and apoptosis in glioblastoma cells by inhibiting the JAK-STAT3 signaling pathway,thereby suppressing cell proliferation.

The aim of this study was to investigate the effects of high-intensity interval training(HIIT)on lipopolysaccharide(LPS)-induced septic myocardial injury in mice and the roles of NLRP3 inflammasome and macrophage M1 polarization in the process.C57BL/6 male mice were randomly divided into 4 groups:control(CON)group,LPS(L)group,HIIT+saline injection(E)group,and HIIT+LPS(EL)group.Six weeks of HIIT intervention was followed by intraperitoneal injection of LPS,and cardiac function indexes were measured by echocardiography 12 hours post the injection.Hematoxylin-eosin(HE)staining was used to evaluate the morphology and pathological characteristics of myocardium for assessing myocardial damage score;enzyme-linked immunosorbent assay(ELISA)was used to test the content of myocardial damage indicators(AST,CK-MB,LDH);RT-PCR was used to detect the relative mRNA levels of NLRP3 inflammasome(NLRP3,Caspase-1),atrial natriuretic peptide(ANP),brain natriuretic peptide(BNP),myeloperoxidase(MPO)and macrophage M1-associated inflammasome factors(IL-1β,TNF-α,IL-6);Western blot was applied to measure the protein expression of inducible nitric oxide synthase(iNOS)and apoptosis-associated speck-like protein containing a caspase recruitment domain(ASC)in cardiac tissues;immunofluorescence staining was used to detect the protein expression of NLRP3 inflammasome,ASC,IL-18 and iNOS.Compared with the CON group,mice in the LPS group showed obvious decrease in body weight,a significant decrease in EF and FS,a significant increase in LVESD and LVEDD,obvious pathological damage in myocardial tissue,a significant increase in myocardial damage fraction,a significant increase in serum myocardial damage indexes,and a significant increase in the expression levels of BNP,MPO,NLRP3 inflammasome,iNOS,IL-1β,IL-6 and TNF-α.HIIT treatment could reverse these changes mentioned above in model mice.In conclusion,6 weeks of HIIT inhibits the activation of LPS-induced NLRP3 inflammasome and suppressed macrophage M1-type polarization,thereby combating septic myocardial injury.

Objective:To investigate HER2 mRNA expression in breast cancer with HER2 immunohistochemistry (IHC) 0 and to analyze the feasibility of distinguishing between the tumor with HER2 μltra-low expression and the one without expression of HER2 (no staining by IHC) by HER2 mRNA level preliminarily.Methods:HER2 mRNA was analyzed by reverse transcription digital PCR in 41 cases of formalin-fixed paraffin-embedded surgical tissue samples of invasive breast cancer obtained between January 2020 and March 2023 at Peking Union Medical College Hospital. The cohort included 21 HER2 IHC 1+ and 20 IHC 0 (12 ultra-low and 8 non-expression of HER2). HER2 mRNA expression level was quantitatively evaluated by the FAM (HER2)/VIC (reference gene) ratio.Results:The expression of HER2 mRNA for the cases with 1+, ultra-low, and non-expression of HER2 by IHC was 0.30 to 1.78 (average 0.90, median 0.82), 0.55 to 1.51 (average 0.93, median 0.90) and 0.22 to 0.78 (average 0.41, median 0.36), respectively. For the mean and median HER2 mRNA levels, there was no significant difference between HER2 IHC 1+ and HER2 ultra-low expression diseases ( P=0.757). A remarkable difference in HER2 gene expression was found between the tumors with 1+ and non-expression of HER2 by IHC ( P=0.002). And, HER2 ultra-low cases contained statistically higher levels of HER2 mRNA compared with non-expression of HER2 subgroup by IHC ( P=0.001). Conclusions:Based on HER2 mRNA, HER2 non-expression and HER2 weak expression (including HER2 IHC 1+ and ultra-low) belong to two different types of the tumor and the disease with HER2 IHC 1+ and HER2 ultra-low expression may be the same. It is necessary to further test the performance of HER2 mRNA detection for stratifying the HER2 weak expression subgroup and to determine the threshold.

Objective:To investigate the clinical and pathological characteristics of breast cancer with HER2 low expression.Methods:The data from 3 422 patients with invasive breast cancer which archived in Peking Union Medical College Hospital between April 2019 and July 2022 were retrospectively reviewed. Among them, 136 patients were treated with neoadjuvant chemotherapy. The tumor size, histological type, tumor differentiation, lymph node metastasis, Ki-67 index, the status of estrogen receptor, progesterone receptor and HER2 as well as pathological complete response (pCR) rate were collected.Results:The HER2 status of 3 286 patients without neoadjuvant therapy, 616 (616/3 286, 18.7%) score 0, 1 047 (1 047/3 286, 31.9%) score 1+, 1 099 (1 099/3 286,33.4%) score 2+ and 524 (524/3 286,15.9%) score 3+ by immunohistochemistry (IHC). Among the 1 070 IHC 2+ cases, 161 were classified as HER2 positive by reflex fluorescence in situ hybridization (FISH) assay. In our cohort, 1 956 cases of HER2-low (IHC 1+ and IHC 2+/FISH-) breast cancer were identified. Compared to the HER2 IHC 0 group, HER2-low tumors more frequently occurred in patients with hormone receptor (HR) positive ( P<0.001), Ki-67 index below 35% ( P<0.001), well or moderate differentiation ( P<0.001) and over the age of 50 ( P=0.008). However, there were no significant differences in histological type, tumor size, and lymph node metastasis between HER2-low and HER2 IHC 0 group. For patients who had neoadjuvant therapy, the pCR rate in the patients with HER2-low was lower than those with HER2 IHC 0 (13.3%, 23.9%), but there was no significant difference. Although HER2-low breast cancers showed a slightly lower pCR rate than HER2 IHC 0 tumors, no remarkable difference was observed between tumors with HER2-low and HER2 IHC 0 regardless of hormone receptor status. Conclusions:The clinicopathological features of HER2-low breast cancers are different from those with HER2 IHC 0. It is necessary to accurately distinguish HER2-low breast cancer from HER2 IHC 0 and to reveal whether HER2-low tumor is a distinct biological entity.

Objective To study the efficacy of breathing training in the patients with chronic non-spe-cific low back pain (CNLBP).Methods The databases of Pubmed,Embase,Web of Science,Cochrane Librar-y,CNKI,China Biomedical Literature Database,Wanfang Database and VIP Database were searched for obtai-ning relevant trials of respiratory-related training for treating CNLBP.The retrieval time limit was from the database establishment to October 2022.The quality assessment was performed by using the Cochrane Manual for Systematic Reviews,and the data analysis was performed by using StataSE15.1 software.Results Nine-teen articles were finally obtained,involving in 1011 cases.The meta analysis showed that the visual analogue scale (VAS) score or pain numerical scale (NRS) score in the experiment group were lower than those in the control group (MD=-1.19,95％CI:-1.51 to-0.87,P<0.05),and the Oswestry dysfunction index (ODI) score was lower than that in the control group (MD=-0.64,95％CI:-0.91 to-0.38,P<0.05). For different types of the patients,the improvement effects of VAS score for different types of patients with CNLBP from high to low were in turn postpartum patients (MD=-1.89,95％CI:-2.51 to-1.27,P<0.05),athletes (MD=-1.46,95％CI:-1.79 to-1.13,P<0.05) and general population (MD=-1.01,95％CI:-1.40 to-0.61,P<0.05).Conclusion Thebreathing training has the improvement effect for pain,dysfunction,proprioception and posture control in various populations with CNLBP.

Objective To compare the efficacy of TightRope loop plate and Endobutton plate in the treatment of acromioclavicular joint dislocation.Methods A retrospective analysis was conducted on 94 patients with acromioclavicular joint dislocation who were treated at this center from March 2021 to February 2023.They were divided into two groups based on different admission date.The Group E(n =47)received Endobutton plate treatment between March 2021 and February 2022,while the Group T(n =47)received TightRope loop plate treatment between March 2022 and February 2023.At the last follow-up,the perioperative indicators,Visual Analogue Scale(VAS),Constant-Murley shoulder joint function scores,and surgical complications were compared between the two groups.Results The surgical time,intraoperative bleeding,incision length,and VAS scores at 7 days after surgery in the Group T were shorter or lower than those in the Group E(P<0.05).There were no statistical differences in the incidence of perioperative nerve injury,internal fixation displacement,clavicle fracture,vascular injury,and infection between the two groups(P>0.05).The subjective and objective scores of Constant-Murley shoulder joint function in both groups at9 months after surgery showed significant improvement compared to preoperative scores(all P =0.000).There was no significant difference in the subjective and objective scores of Constant-Murley shoulder joint function between the two groups at 9 months after surgery(P>0.05).Conclusions The treatment of acromioclavicular joint dislocation with TightRope loop plate ot or Endobutton plate has a significant effect and can effectively improve shoulder joint function.Compared with Endobutton plate,use of TightRope loop plate has minor surgical trauma,less bleeding,and significantly reduced postoperative pain,being more conducive to early functional exercise for patients.

The prevention and treatment of cardiovascular diseases have always been a focus of attention in re-lated fields.This paper discusses the application and mechanism of exogenous and endogenous supplementation with β-hy-droxybutyrate in the prevention and treatment of cardiovascular diseases such as myocardial ischemia,myocardial infarc-tion,diabetic cardiomyopathy,hypertension,myocardial inflammation,hypertrophic cardiomyopathy and heart failure.Exogenous β-hydroxybutyrate supplementation can quickly and directly provide substitute energy for the heart and thus prevent and treat heart injury.The comprehensive effect of exogenous β-hydroxybutyrate supplementation is significantly better than that of endogenous β-hydroxybutyrate.Endogenous fatty acid,obtained through a ketogenic diet,needs to be oxidized to β-hydroxybutyrate in vivo for energy supply,which is greatly influenced by glucose and lipid metabolism under the conditions of the basic disease itself.At present,ketogenic diet is not recommended for use in the prevention and treat-ment of myocardial ischemia or myocardial infarction,and in the treatment of diabetic cardiomyopathy or hypertension.

Ketogenic diet can supplement blood ketone,provide energy for skeletal muscle,and affect the athletic performance and recovery of animals and humans after fatigue.In this paper,it was found that ketogenic diet transformed muscle fibers from Ⅱb to Ⅱa through axonal germination and nerve reinnervation,improved the quality and function of mitochondria of fast muscle and increased histone acetyltransferase to enhance skeletal muscle strength.Using ketone body to provide energy,it can reduce glycolysis and improve the ability of fatty acid oxidation in slow muscle to improve exercise endurance;It can reduce endoplasmic reticulum stress,oxidative stress and inflammatory reaction of skeletal muscle,protect the body from injury,reduce the consumption of muscle glycogen and the accumulation of lactic acid,reduce fatigue after exercise and promote recovery.It is considered that ketogenic diet has low negative effects on the body,can improve sports performance and accelerate fatigue recovery,and can be used as a diet scheme in competitive sports and rehabilitation training.