Membrane proteins are integral components of cellular membranes, accounting for approximately 30% of the mammalian proteome and serving as targets for 60% of FDA-approved drugs. They are critical to both physiological functions and disease mechanisms. Their functional protein-protein interactions form the basis for many physiological processes, such as signal transduction, material transport, and cell communication. Membrane protein interactions are characterized by membrane environment dependence, spatial asymmetry, weak interaction strength, high dynamics, and a variety of interaction sites. Therefore, in situ analysis is essential for revealing the structural basis and kinetics of these proteins. This paper introduces currently available in situ analytical techniques for studying membrane protein interactions and evaluates the characteristics of each. These techniques are divided into two categories: label-based techniques (e.g., co-immunoprecipitation, proximity ligation assay, bimolecular fluorescence complementation, resonance energy transfer, and proximity labeling) and label-free techniques (e.g., cryo-electron tomography, in situ cross-linking mass spectrometry, Raman spectroscopy, electron paramagnetic resonance, nuclear magnetic resonance, and structure prediction tools). Each technique is critically assessed in terms of its historical development, strengths, and limitations. Based on the authors’ relevant research, the paper further discusses the key issues and trends in the application of these techniques, providing valuable references for the field of membrane protein research. Label-based techniques rely on molecular tags or antibodies to detect proximity or interactions, offering high specificity and adaptability for dynamic studies. For instance, proximity ligation assay combines the specificity of antibodies with the sensitivity of PCR amplification, while proximity labeling enables spatial mapping of interactomes. Conversely, label-free techniques, such as cryo-electron tomography, provide near-native structural insights, and Raman spectroscopy directly probes molecular interactions without perturbing the membrane environment. Despite advancements, these methods face several universal challenges: (1) indirect detection, relying on proximity or tagged proxies rather than direct interaction measurement; (2) limited capacity for continuous dynamic monitoring in live cells; and (3) potential artificial influences introduced by labeling or sample preparation, which may alter native conformations. Emerging trends emphasize the multimodal integration of complementary techniques to overcome individual limitations. For example, combining in situ cross-linking mass spectrometry with proximity labeling enhances both spatial resolution and interaction coverage, enabling high-throughput subcellular interactome mapping. Similarly, coupling fluorescence resonance energy transfer with nuclear magnetic resonance and artificial intelligence (AI) simulations integrates dynamic structural data, atomic-level details, and predictive modeling for holistic insights. Advances in AI, exemplified by AlphaFold’s ability to predict interaction interfaces, further augment experimental data, accelerating structure-function analyses. Future developments in cryo-electron microscopy, super-resolution imaging, and machine learning are poised to refine spatiotemporal resolution and scalability. In conclusion, in situ analysis of membrane protein interactions remains indispensable for deciphering their roles in health and disease. While current technologies have significantly advanced our understanding, persistent gaps highlight the need for innovative, integrative approaches. By synergizing experimental and computational tools, researchers can achieve multiscale, real-time, and perturbation-free analyses, ultimately unraveling the dynamic complexity of membrane protein networks and driving therapeutic discovery.

ObjectiveThis study employed the scoping review method to systematically retrieve and analyze the basic information and clinical research evidence of expensive Chinese patent medicines （CPMs）， aiming to provide a basis for future related research and clinical applications. MethodsEight Chinese and English databases were systematically searched for the clinical research evidence on expensive CPMs. ResultsEleven expensive CPMs （Angong Niuhuang Wan， Jufang Zhibao Wan， Suhexiang Wan， Pien Tze Huang， Niuhuang Qingxin Wan， Qinggong Shoutao Wan， Compound Realgar Natural Indigo Tablets， Xihuang Wan， Dingkun Wan， Babao Wan， and Guilingji Capsules） were selected. A total of 365 related studies were included in this review， comprising 331 clinical studies （of which 291 were randomized controlled trials）， 30 systematic reviews and Meta-analyses， 3 expert consensus， and 1 rapid health technology assessment. Among the 11 CPMs， 2（Angong Niuhuang Wan and Jufang Zhibao Wan） had a daily price over 500 yuan. The famous and precious Chinese medicinal materials involved included Moschus （frequency of 7）， Bovisc Alculus （7）， and Borneol （5）. The dosage forms included pills， capsules， oral liquid， tablets， and lozenges. The diseases treated by these CPMs mainly included malignant tumors， cerebrovascular diseases， gynecological diseases， and hepatobiliary system diseases. The sample sizes of the clinical studies were mainly concentrated within the range of 51-100 cases， and the main control form was CPM + basic Western medicine treatment vs. basic Western medicine treatment. The 331 clinical studies reported a total of 44 adverse events occurred， of which 36 were determined to be adverse reactions. ConclusionThe scarcity of raw materials leads to the high prices of expensive CPMs. The difficulty of conducting clinical research and the critical and severe cases treated lead to a lack of clinical research evidence with large sample sizes. The uneven distribution of existing studies， incomplete information on medicine package， and non-standard clinical research designs remain to be addressed in the future.

Vascular calcification is a highly regulated process of ectopic calcification in cardiovascular system while no effective intervention can be clinically performed up to date.As vascular calcification undergoes a common regulatory mechanism within bone formation,bone morphogenetic protein 7(BMP-7)main-tains contractile phenotype of vascular smooth muscle cells and further inhibits vascular calcification via promoting the process of osteoblast differentiation,reducing ectopic calcification pressure by increasing bone formation and reducing bone resorption.This work systematically reviews the role of BMP-7 in vascular calcifi-cation and the possible mechanism,and their current clinical application as well.The current proceedings may help develope early diagnostic strategy and therapeutic treatment with BMP-7 as a new molecular marker and potential drug target.The expec-tation could achieve early prevention and intervention of vascular calcification and improve poor prognosis on patients.

Aim To explore the mechanism of modified Wuzi Yanzong pill to reduce testicular toxicity of Tripterygium wilfordii by means of network pharmacolo-gy.Methods The effective chemical composition and target of modified Wuzi Yanzong Pill to reduce testicu-lar toxicity of Tripterygium wilfordii were analyzed by using multiple databases to understand the mechanism of action.Results The first five compounds of modi-fied Wuzi Yanzong pill are quercetin,sucrose,kaempferol,galactose and ellagic acid,respectively.The first five targets were TP53,SRC,EP300,MAPK3,STAT3;KEGG pathway enrichment analysis mainly concentrated in cancer pathway,lipid and atherosclero-sis,hepatitis B,apoptosis,diabetes complications AGE-RAGE signaling pathway,PI3K-Akt signaling pathway and other signaling pathways.Conclusions The ac-tive components of modified Wuzi Yanzong pill can re-duce the testicular toxicity of Tripterygium wilfordii mainly by interfering with TP53,SRC,EP300,MAPK3,STAT3 and other targets,regulating apoptosis,PI3K-Akt signaling pathway,diabetes complications AGE-RAGE signaling pathway,lipid metabolism and other pathways.

Objective:To standardize the bacterial endotoxin testing criteria for zoledronic acid injection and estab-lish a detection method using recombinant factor C(rFC).Methods:The gel-clot method(BET)was utilized to test 13 batches of zoledronic acid injection from national supervision and random inspection.Interference tests were conducted on zoledronic acid injections from three manufacturers at different concentrations(500,100,50,25 μg·mL-1)using rFC test kits from two manufacturers.Results:Detection was performed for the specification of 100 mL∶5 mg and other specifications according to＜0.50 EU per 1 mL and＜5.0 EU per 1 mg,respectively,and all results met the criteria.The recovery rate for 25 μg·mL-1 using rFC kits from both manufacturers ranged between 50％and 200％.Validation of rFC.Methods:Eight batches of zoledronic acid injection were validated at 25 μg·mL-1,five batches of zoledronic acid injection(100 mL∶5 mg)were validated at 0.5 EU·mL-1,and all recovery rates were between 50％and 200％.Conclusion:The bacterial endotoxin testing method for zoledronic acid injection can be established as follows:for large volume injection products with 100 mL or more,each 1 mL should contain less than 0.50 EU of endotoxin(following the Chinese Pharmacopoeia 2020,general chapter 1143).For other specifications,each 1 mg of zoledronic acid should contain less than 10.0 EU of endotoxin.The rFC test kit method for bacterial endotoxins involved diluting the sample with water for bacterial endotoxin testing to contain 25 μg of zoledronic acid per 1 mL,or performing 1∶1 dilution for large volume injections(100 mL∶5.0 mg specifi-cation),with recovery rate between 50％and 200％as per the kit instructions.

Objective To explore the clinical efficacy and safety of early surgical treatment for spinal thoracolumbar fracture without nerve injury.Methods The clinical data of 80 patients with spinal thoracolumbar fracture without nerve injury who were admitted to the department of spinal surgery in our hospital were retrospectively analyzed.According to the different operation timing,those who underwent surgery within 72 hours after fracture were included in the early operation group(n=41),and those who underwent surgery 72 hours to 2 weeks after fracture were included in the elective operation group(n=39).All operations were performed through the Wiltse approach for short-segment pedicle screw fixation on the injured vertebra.The operation time,intraoperative blood loss,hospital stay and incidence of complication of the two groups were compared.The visual analogue scale(VAS)scores,Oswestry disability index(ODI),compression rate of the anterior edge height of the injured vertebra,and the Cobb angle in the sagittal position of the injured vertebra before surgery,1 week after surgery and 1 year after surgery were compared between the two groups.The improvement rates of the anterior edge height compression and the Cobb angle in the sagittal position of the injured vertebra 1 week and 1 year after surgery were compared between the two groups.Results There was no significant difference in the operation time,intraoperative blood loss or total incidence of complications between the two groups(P>0.05).The hospital stay in the early operation group was shorter than that in the elective operation group,and the difference was statistically significant(P<0.05).The VAS scores and ODI 1 week and 1 year after surgery of the two groups were better than those before surgery,and the differences were statistically significant(P<0.05).There was no significant difference in the VAS scores or ODI at each time point before and after surgery between the two groups(P>0.05).The compression rate of the anterior edge height and Cobb angle in the sagittal position of the injured vertebra 1 week and 1 year after surgery in the two groups were lower/smaller than those before surgery,with statistically significant differences(P<0.05).There was no statistically significant difference in the compression rate of the anterior edge height or Cobb angle before surgery in the sagittal position of the injured vertebrae between the two groups(P>0.05).The compression rate of the anterior edge height and Cobb angle in the sagittal position of the injured vertebra 1 week and 1 year after surgery in the early operation group were lower/smaller than those in the elective operation group,and the differences were statistically significant(P<0.05).The improvement rates of the anterior edge height compression and the Cobb angle in the sagittal position of the injured vertebra 1 week and 1 year after surgery in the early operation group were better than those in the elective operation group,and the differences were statistically significant(P<0.05).Conclusion Early surgical treatment for spinal thoracolumbar fracture without nerve damage is safe,it can significantly shorten hospitalization time,obtain good fracture reduction quality and definite therapeutic effects.However,a comprehensive preoperative evaluation of the patients' condition is necessary to ensure surgical safety.

Objective To explore root cause of poor prognosis after successful endovascular treatment(EVT)in patients with acute ischemic stroke with large vascular occlusion(AIS-LVO)of anterior circulation.Methods Patients with AIS-LOV of anterior circulation who received successful EVT(postoperative modified thrombolysis incerebral infarction[mTICI]grade≥2b)were retrospectively and continuously collected in the Department of Neurology of Bozhou People's Hospital from January 2022 to March 2024.The baseline and clinical data of the patients were collected,including gender,age,vascular risk factors(hypertension,diabetes,coronary heart disease,hyperlipidemia,valvular heart disease,atrial fibrillation,smoking,and alcohol consumption),prior stroke or transient ischemic attack,baseline blood pressure,baseline National Institutes of Health Stroke scale(NIHSS)score,laboratory test indicators(pre-operative C-reactive protein and D-dimer,post-operative fasting blood glucose,lipid levels,homocysteine,etc).Meanwhile,the data of perioperative indicators was collected,including the time from onset to admission,the time from admission to puncture,the time from puncture to revascularization,the time from onset to puncture,the time from onset to revascularization,remedial measures(balloon dilation,stent placement,arterial thrombolysis)during the surgery or not,using tirofiban or not,postoperative complications(stroke-related pneumonia,stress ulcers,deep vein thrombosis,acute heart failure or renal failure,etc)or not.The patient's medical history and imaging data were collected,and these indicators were defined and collected,including Alberta stroke program early CT score(ASPECTS),location of occlusion(C1 segment of the internal carotid artery,C2 segment to C7 segment of the internal carotid artery,M1 segment of the middle cerebral artery),and the trial of org 10172 in acute stroke treatment(TOAST)classification and a postoperative transformation of cerebral infarction after ischemic stroke and symptomatic intracranial hemorrhage or not.According to the modified Rankin scale(mRS)score at 90 d after surgery,all patients were divided into poor prognosis group(mRS score≥ 3)and good prognosis group(mRS score≤2).The baseline and clinical data of two groups were compared using univariate analysis.Variables with P＜0.1 in the univariate analysis were selected as independent variables,and the poor prognosis was used as the dependent variable.Further,multivariate Logistic regression analysis was performed to identify the influencing factors of poor prognosis after EVT.Results Finally,a total of 192 patients with AIS-LVO of anterior circulation who received successful revascularization were included in this study.There were 101 male patients and 91 female patients.The poor prognosis group had 102 cases and the good prognosis group had 90 cases.Univariate analysis showed that the poor prognosis group had statistically significant differences with the good prognosis group in terms of age(Z=-3.088,P=0.002)and age distribution(x2=13.457,P=0.001),fasting blood glucose(Z=-3.347,P=0.001),baseline NIHSS score(Z=-4.469,P＜0.01),location of occlusion(x2=10.488,P=0.005),transformation of hemorrhage after ischemic stroke(x2=16.943,P＜0.01),and symptomatic intracranial hemorrhage(X2=25.449,P＜0.01),and the baseline ASPECTS of the poor prognosis group was significantly lower than that of the good prognosis group(Z=-4.547,P＜0.01).There were no significant differences in other baseline and clinical data(all P＞0.05).Further multivariate Logistic regression analysis showed that age＞80 years(OR,3.224,95％CI 1.033-10.058,P=0.044),baseline NIHSS score(OR,1.102,95％CI 1.013-1.199,P=0.023),baseline ASPECTS(OR,0.375,95％CI 0.212-0.665,P=0.001),and symptomatic intracranial hemorrhage(OR,7.127,95％CI 1.296-39.203,P=0.024)were independent influencing factors of poor prognosis.Conclusion The independent factors of 90 d poor prognosis after successful EVT in patients with AIS-LVO of anterior circulation are age＞80 years,baseline NIHSS score,baseline ASPECTS,and symptomatic intracranial hemorrhage.

Perilla frutescens,a short-day plant,is rich in biologically active substances and nutrients.Current research on Perilla frutescens focuses on agronomic traits such as yield and fatty acid accumula-tion,with limited exploration of the flowering process and floral organ development.The molecular regu-latory mechanisms underlying these aspects remain unclear.FLOWERING LOUC T(FT)is a florigen in Arabidopsis,plays critical roles in floral transition.PfFT3 is unannotated by genome but annotated by transcriptomics data to the FT-like subfamily.Its function in controlling flowering is yet to be explored.Here subcellular localization analysis showed that PfFT3 is localized in the nucleus and cytoplasm.The plant over-expression vector pCAMBIAI1303-PfFT3 was constructed and transformed into wild-type(Col-0)and mutant fd-2,fd-3,and ft-10 plants by agrobacterium-mediated inflorescence infiltration as a means of obtaining genetically stable and pure overexpression and backfill transgenic lines in Arabidopsis,respectively.Analysis of the results showed that overexpression of PfFT3 significantly promoted early flowering in Arabidopsis and rescued the late-flowering phenotype of the mutants fd-2,fd-3,and ft-10,and that expression of the exogenous PfFT3 promoted the expression of the downstream endogenous flow-ering genes AtSOC1,AtAP1,AtFUL,and AtLFY.This study demonstrates the positive role of PfFT3 in promoting flowering,providing a foundation for further investigation of PfPEBP function and advancing the breeding of early-flowering Perilla frrutescens cultivars.

Objective:To explore the application value of simultaneous monitoring of voriconazole(VRCZ)and voriconazole N-oxide(VNO)in efficacy and safety of VRCZ in the prevention and treatment of fungal infections in allogeneic hematopoietic stem cell transplantation(allo-HSCT)patients before engraftment(i.e.,days+1 to+30 after transplantation).Methods:The influencing factors of VRCZ,VNO concentration and MR(CVNO/CVRCz)and the difference of VRCZ in the prevention and treatment of fungal infection and liver and kidney injury were analyzed.The receiver operating characteristic curve(ROC)was used to analyze the differences(the corresponding to the maximum of the Youden index on the curve was set as the cut-off value)to confirm the critical value.Results:The factors affecting VRCZ concentration(CVRCZ),VNO concentration(CVNO)and MR were patient weight,VRCZ daily dose,and transplantation type(all P＜0.05).CVRCZ and CVNO in the effective group were higher than those in the ineffective group(P＜0.001),the opposite of MR(P＜0.001);the liver and renal injury group had lower MR than the normal group(P＜0.05).ROC showed that CVRCZ,CVNO and MR had important value in predicting VRCZ in the prevention and treatment of invasive fungal infections in allo-HSCT patients before engraftment,and their cutoff of concentrations were 0.95 μg/ml,1.35 μg/ml and 1.645,respectively(AUC:0.9677,0.7634,0.9564).CVRCZ and MR can assist in indicating liver[cutoff values:0.65 μg/ml,1.96(AUC:0.5971,0.6663)]and renal injury[cutoff values:0.95 μg/ml,1.705(AUC:0.6039,0.6164)].Conclusion:The great value of simultaneous monitoring of VRCZ,VNO and MR can predict in the efficacy and safety of VRCZ in allo-HSCT patients before engraftment.The prediction accuracy of CVRCZ was higher than that of MR,followed by that of CVNO.Increased CVRCZ and decreased MR increase the risk of liver and kidney injury.

Objective:To investigate the effects of Xiongcan Yishen Formula(XYF)on ferroptosis in mouse TM3 Leydig cells after oxidative stress injury(OSI)induced by H2O2.Methods:An oxidative stress injury model was established in mouse TM3 Leydig cells using H2O2 induction.The modeled TM3 cells were randomly divided into OSI group,XYF group,the ferroptosis inhibitor Ferrostatin-1(F-1)group,and F-1+XYF group,which were respectively intervened with blank serum,20%drug-containing serum,2μmol/L F-1,and2μmol/L F-1+20%drug-containing serum.A control group(normal TM3 cells+blank serum)was also set up.The morphology of cells in each group was observed,and the levels of testosterone,superoxide dismutase(SOD),reactive oxygen spe-cies(ROS),malondialdehyde(MDA),ferritin heavy chain 1(FTH1),solute carrier family 7 member 11(SLC7A11),glutathione(GSH),glutathione peroxidase 4(GPX4),fatty acid CoA ligase 4(FACL4),total iron ions,and ferrous ions were detected.Re-sults:Compared with the model group,the control group showed significantly decreased expression of ROS,MDA,FACL4,total iron,and ferrous ions(P<0.05),and significantly increased levels of testosterone,SOD,GSH,FTH1,SLC7A11,and GPX4(P<0.05).The male silkworm kidney-tonifying formula group significantly promoted testosterone secretion by TM3 cells and upregulated the expression of FTH1,SLC7A11,GPX4,GSH,and SOD in TM3 cells(P<0.05),while significantly downregulating ROS,MDA,FACL4,total iron ions,and ferrous ions(P<0.05).Conclusion:Following H2O2 exposure,oxidative stress can induce ferroptosis in mouse TM3 Leydig cells.XYF can antagonize OSI and ferroptosis in TM3 cells by activating the SLC7A11/GSH/GPX4 axis,which may underlie the mechanism of XYF in the treatment of male late-onset hypogonadism.