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.

Objective To observe the effects of amyloid-β(Aβ)receptor PirB on mouse astrocyte proliferation and reactive astrogliosis in vitro.Methods Mouse primary astrocytes were cultured,and divided into control group,Aβ group,Aβ+0.2 μmol/L PEP group,Aβ+0.4 μmol/L PEP group,Aβ+Fluspirilene group,Aβ+GFP-LV group,and Aβ+mPirB-LV group.The mouse astrocytes were treated with soluble PirB extracellular peptide PEP or PirB inhibitor Fluspirilene,respectively,to inhibit endogenous PirB receptor,or overexpressed PirB gene via lentivirus transfection and then treated with Aβ1-42 oligomers.The proliferation of astrocytes was observed by RTCA and EdU methods,and the mRNA expression levels of S-100 calcium-binding protein B(S-100β),Vimentin,Nestin and amyloid precursor protein(APP)associated with reactive astrogliosis of astrocytes were observed by real-time PCR,and the expression level of glial fibrillary acid protein(GFAP)was detected by Western-blotting.Results The results of RTCA monitoring showed that normalized cell index(NCI)values of each group decreased sharply after treatment,and then increased gradually and tended to be stable.The results of EdU staining showed that the proliferative activity of astrocytes was significantly enhanced in the Aβ group(P<0.05)compared with control group;Compared with Aβ group,cell proliferation activity in Aβ + 0.2 μmol/L PEP group,Aβ+0.4 μmol/L PEP group and Aβ+Fluspirilene group were significantly decreased(P<0.01 or P<0.001).The results of real-time PCR showed that compared with control group,mRNA expressions of GFAP,S-100β,Vimentin,Nestin,APP and PirB in Aβ group were significantly increased(P<0.05);Compared with Aβ group,mRNA expressions of GFAP,S-100β,Vimentin,Nestin,APP and PirB in Aβ+0.4 μmol/L PEP group were significantly decreased(P<0.01);Compared with Aβ+GFP-LV group,mRNA expressions of GFAP,S-100β,Vimentin,Nestin,APP and PirB in Aβ +mPirB-LV group were significantly increased(P<0.05).The results of Western blotting showed that compared with control group,the expression of GFAP in Aβ group was significantly increased(P<0.05);Compared with Aβ group,the expression of GFAP in Aβ+0.4 μmol/L PEP group was significantly decreased(P<0.05).Conclusions PirB is an upstream molecule which could regulate astrocyte proliferation and reactive astrogliosis,and inhibiting PirB receptor in astrocytes may be a potential treatment for Alzheimer's disease.

【Objective】 To explore the feasibility of en-bloc resection of bladder tumors by flexible cystoscope combined with laparoscopic instruments through urethra and to provide reference for the clinical application of this technique. 【Methods】 Self-designed and processed transurethral single-hole PORT and Olympus electronic cystoscope were used as observation mirror; Φ1.8 mm soft grasper, tissue scissors, electric hook, and ultrasonic scalpel were used as instruments; the porcine bladder was used as a model.The PORT was placed through the urethra, and the cystoscope was inserted to observe the inner wall of the bladder and the condition of the mucosa.After the lesion site was identified in the bladder cavity, the soft grasper was inserted to pull the mucosa to be removed, which was then fixed with tension at the target position to maintain a satisfactory feild of view.The surgeon held the cystoscope in the left hand, and operated the laparoscopic instruments into the bladder cavity through the PORT with the right hand.Observing with the cystoscope and lifting and pulling the mucosa with the grasper, the surgeon simulated the cutting and pushing actions to realize the en-bloc resection of the lesioned mucosa. 【Results】 The mucosa at 4 different locations were successfully resected on 2 in vitro porcine bladder models. 【Conclusion】 The in vitro experiments show that the combination of flexible electronic cystoscope and laparoscopic instruments achieves synergistic effects in en-bloc resection of bladder tumor by transurethral single-hole laparoscope without additional iatrogenic bladder injury caused by percutaneous bladder incision.This method is feasible in the treatment of bladder tumors, and has the potential of clinical application after further optimization.

ObjectiveBased on fluorescence lifetime imaging technology, a novel method for viscosity detection is proposed and the capability of different weighting of fluorescence lifetimes in distinguishing the viscosity of glycerol-water mixtures is evaluated, aiming to enhance the accuracy and reliability of viscosity differentiation. MethodsThis approach incorporates the principles of electronic weighting, introducing both amplitude-weighted average fluorescence lifetime (τ_m) and intensity-weighted average fluorescence lifetime (τ_i). Viscosity changes in glycerol-water mixtures are detected through τ_m and τ_i. τ_m Reflects the relationship between fluorescence signal amplitude and time, while τ_i focuses on the time-varying characteristics of fluorescence signal intensity. ResultsThe results of both τ_m and τ_i mutually corroborate each other, not only enhancing the reliability in detecting viscosity changes in glycerol-water mixtures but also revealing their unique roles in the detection process. Although τ_m plays a crucial role in capturing changes in fluorescence signal amplitude, τ_i exhibits higher accuracy in viscosity detection when considering the time-varying characteristics of fluorescence signal intensity. It is particularly noteworthy that, due to τ_i’s greater sensitivity, microenvironment viscosity detection can be directly analyzed using τ_i. This provides a more convenient approach for real-time, highly sensitive microfluidic viscosity monitoring. Therefore, through the comprehensive utilization of τ_m and τ_i, a more thorough and accurate understanding of the viscosity information in glycerol-water mixtures can be obtained, and specific parameters can be selected for in-depth analysis based on specific needs. ConclusionThe combination of amplitude weighting and intensity weighting allows for a more sensitive identification of subtle changes in viscosity under different conditions. The innovation of this method lies in its simultaneous consideration of multiple parameters, enhancing sensitivity and distinguishability to variations in viscosity. Therefore, this weighted-dependent fluorescence lifetime imaging technique not only introduces a novel approach for viscosity detection in glycerol-water mixtures but also provides a powerful analytical tool for various fields, including microfluidics, rheology, and research on novel functional materials.

AIM To explore the effects of Zishui Qinggan Decoction on the mouse model of depression induced by chronic restraint stress(CRS)via ERK/GSK3β/CREB/BDNF signaling pathway.METHODS Except for those of the blank group,the mice of other groups were induced into depression models by CRS,and divided into the model group,the fluoxetine hydrochloride group(10 mg/kg)and the low,medium and high dose Zishui Qinggan Decoction groups(8.835,17.670 and 35.340 g/kg)for the corresponding drug intervention and simultanous CRS treatment.The mice had their sugar water preference experiment and behavior experiment on the 7th and 14th day after administration;the observation of the hippocampal morphological changes by HE staining,the detection of the superoxide dismutase(SOD)activity and malondialdehyde(MDA)level in serum by kits,the detection of levels of serum 5-hydroxytryptamine(5-HT),tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)by ELISA,the detection of the hippocampal mRNA expressions of BDNF,TNF-α and IL-1β by RT-qPCR method,and the detection of the hippocampal protein expressions of ERK1/2,p-ERK1/2,GSK3β,p-GSK3β,CREB and BDNF by Western blot method 14 days after administration.RESULTS Compared with the model group,after 14 days of administration,both fluoxetine hydrochloride group and medium-dose Zishui Qinggan Decoction group displayed increased preference rate of sugar water(P<0.01),shortened immobility time of tail suspension and forced swimming(P<0.01),improved hippocampal damage of nerve cells,decreased levels of serum MDA,TNF-α and IL-1β(P<0.05,P<0.01),increased SOD activity and 5-HT level(P<0.05,P<0.01),decreased hippocampal mRNA expressions of TNF-α and IL-1β(P<0.01),and decreased expressions of BDNF mRNA and p-ERK1/2,p-GSK3β,CREB and BDNF proteins(P<0.05,P<0.01).CONCLUSION Zishui Qinggan Decoction can improve the depression-like behaviors in mice exposed to CRS,and its mechanism may be related to the regulation of hippocampal ERK/GSK3β/CREB/BDNF signaling pathway.

Objective To assess the value of acute gastrointestinal injury(AGI)and intestinal fatty acid-binding protein(I-FABP)in the prognosis of liver cirrhotic patients with sepsis.Methods Clinical data of 84 liver cirrhosis patients with sepsis who were admitted to the intensive care unit(ICU)of a hospital from September 2020 to March 2023 were analyzed retrospectively,and 41 patients with decompensated liver cirrhosis during the same period were selected as the control group.Serum I-FABP level in patients was determined with enzyme-linked immunosorbent assay(ELISA).Scores of the model of end-stage liver disease(MELD)and sequential organ failure assessment(SOFA)were calculated.AGI was evaluated based on medical records.30-day and 90-day survival was observed.Correlation among variables was analyzed by Spearman correlation.Risk factors for death in patients with liver cir-rhosis and sepsis was determined by multivariate Cox regression analysis.The optimal cut-off value was determined by receiver operating characteristic(ROC)curve,and the diagnostic efficacy was compared through the area under the ROC curve(AUC).Results Both AGI grading and I-FABP level in liver cirrhosis patients with sepsis were higher than those in the control group(both P＜0.05).I-FABP level was correlated with procalcitonin(PCT),MELD,and SOFA scores in patients with liver cirrhosis and sepsis(all P＜0.05).AGI grading was positively cor-related with SOFA score(P=0.038).The 30-day and 90-day mortality of patients in the liver cirrhosis with sepsis group were 25.0％(n=21)and 35.7％(n=30),respectively.Multivariate Cox regression analysis showed that baseline I-FABP and SOFA scores were independently correlated with 30-day and 90-day survival,and the I-FABP quartile showed good prognostic differentiation efficacy.ROC curve showed that I-FABP could significantly improve the predictive effect of SOFA score on the prognosis of patients.Conclusion AGI grading and I-FABP level in liver cirrhosis patients with sepsis are elevated significantly.Serum I-FABP is associated with the prognosis of patient and can improve the predictive efficacy of SOFA score for survival.

Ankylosing spondylitis (AS) combined with lower cervical fracture is often categorized into unstable fracture, with a high incidence of neurological injury and a high rate of disability and morbidity. As factors such as shoulder occlusion may affect the accuracy of X-ray imaging diagnosis, it is often easily misdiagnosed at the primary diagnosis. Non-operative treatment has complications such as bone nonunion and the possibility of secondary neurological damage, while the timing, access and choice of surgical treatment are still controversial. Currently, there are no clinical practice guidelines for the treatment of AS combined with lower cervical fracture with or without dislocation. To this end, the Spinal Trauma Group of Orthopedics Branch of Chinese Medical Doctor Association organized experts to formulate Clinical guidelines for the treatment of ankylosing spondylitis combined with lower cervical fracture in adults ( version 2024) in accordance with the principles of evidence-based medicine, scientificity and practicality, in which 11 recommendations were put forward in terms of the diagnosis, imaging evaluation, typing and treatment, etc, to provide guidance for the diagnosis and treatment of AS combined with lower cervical fracture.

Objective:To investigate the current situation of the use of transjugular intrahepatic portosystemic shunt (TIPS) for portal hypertension, which should aid the development of TIPS in China.Methods:The China Portal Hypertension Alliance (CHESS) initiated this study that comprehensively investigated the basic situation of TIPS for portal hypertension in China through network research. The survey included the following: the number of surgical cases, main indications, the development of Early-TIPS, TIPS for portal vein cavernous transformation, collateral circulation embolization, intraoperative portal pressure gradient measurement, commonly used stent types, conventional anticoagulation and time, postoperative follow-up, obstacles, and the application of domestic instruments.Results:According to the survey, a total of 13 527 TIPS operations were carried out in 545 hospitals participating in the survey in 2021, and 94.1% of the hospital had the habit of routine follow-up after TIPS. Most hospitals believed that the main indications of TIPS were the control of acute bleeding (42.6%) and the prevention of rebleeding (40.7%). 48.1% of the teams carried out early or priority TIPS, 53.0% of the teams carried out TIPS for the cavernous transformation of the portal vein, and 81.0% chose routine embolization of collateral circulation during operation. Most of them used coils and biological glue as embolic materials, and 78.5% of the team routinely performed intraoperative portal pressure gradient measurements. In selecting TIPS stents, 57.1% of the hospitals woulel choose Viator-specific stents, 57.2% woulel choose conventional anticoagulation after TIPS, and the duration of anticoagulation was between 3-6 months (55.4%). The limitation of TIPS surgery was mainly due to cost (72.3%) and insufficient understanding of doctors in related departments (77.4%). Most teams accepted the domestic instruments used in TIPS (92.7%).Conclusions:This survey shows that TIPS treatment is an essential part of treating portal hypertension in China. The total number of TIPS cases is far from that of patients with portal hypertension. In the future, it is still necessary to popularize TIPS technology and further standardize surgical indications, routine operations, and instrument application.

Objective To compare the effects of 20 mg qd and 10 mg bidadministration of iprrazole enteric-coated tablets on the control of gastric acid in healthy subjects.Methods A randomized,single-center,parallel controlled trial was designed to include 8 healthy subjects.Randomly divided into 2 groups,20 mg qd administration group:20 mg enteric-coated tablets of iprrazole in the morning;10 mg bid administration group:10 mg enteric-coated tablets of iprrazole in the morning and 10 mg in the evening.The pH values in the stomach of the subjects before and 24 h after administration were monitored by pH meter.The plasma concentration of iprazole after administration was determined by HPLC-MS/MS.The main pharmacokinetic parameters were calculated by Phoenix WinNonlin(V8.0)software.Results The PK parameters of iprrazole enteric-coated tablets and reference preparations in fasting group were as follows:The Cmax of 20 mg qd group and 10 mg bid group were(595.75±131.15)and(283.50±96.98)ng·mL-1;AUC0-t were(5 531.94±784.35)and(4 686.67±898.23)h·ng·mL-1;AUC0-∞ were(6 003.19±538.59)and(7 361.48±1 816.77)h·ng·mL-1,respectively.The mean time percentage of gastric pH＞3 after 20 mg qd and 10 mg bid were 82.64％and 61.92％,and the median gastric pH within 24 h were 6.25±1.49 and 3.53±2.05,respectively.The mean gastric pH values within 24 h were 5.71±1.36 and 4.23±1.45,respectively.The correlation analysis of pharmacokinetic/pharmacodynamics showed that there was no significant correlation between the peak concentration of drug in plasma and the inhibitory effect of acid.Conclusion Compared with the 20 mg qd group and the 10 mg bid group,the acid inhibition effect is better,the administration times are less,and the safety of the two administration regimes is good.

Objective:To study the effects of periodontitis on bone and tryptophan metabolism of gut microbiota in the context of estrogen deficiency.Methods:Thirty-two female C57BL6/J mice were randomly divided into four groups based on table of random numbers ( n=8 in each group): Sham group, in which mice were given sham surgery; Sham_Lig group, in which mice were given sham surgery and were induced to periodontitis by ligating the bilateral maxillary second molars with 5-0 silk threads at the fourth week; Ovx group, in which mice were given bilateral ovariectomy; Ovx_Lig group, in which mice were given bilateral ovariectomy and were induced to periodontitis at the fourth week. After 8 weeks of ligation, the mice of 4 groups were euthanized for collecting the samples of femur, tibia, mandible and skull. Those samples were scanned by micro-CT to measure the bone mineral density (BMD), bone volume versus total volume ratio (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th) and trabecular spacing (Tb.Sp). The cecum contents of 4 groups of mice were collected for gut microbiota 16S rRNA gene sequencing. The tryptophan and its metabolites in intestinal tracts were detected by liquid chromatography-mass spectrometry. Pearson correlation analysis was performed to analyze the correlation between the abundance of gut microbiota and the content of tryptophan and its metabolites. Results:Femur BMD [(82.23±3.97) mg/cm 3], BV/TV [(9.25±1.37)%] and Tb.Th [(70.95±5.70) μm] in Ovx_Lig group were significantly lower than Ovx group [(96.30±3.76) mg/cm 3 ( P=0.004); (14.45±1.55)% ( P=0.022) and (87.58±8.02) μm ( P<0.001), respectively]. The β-diversity analysis of gut microbiota based on Bray-Curtis distance showed that samples of Ovx_Lig group and Ovx group were obviously grouped. Linear discriminant analysis effect size (LEfSe) showed that Alistipes was the representative genus in Ovx_Lig group. The relative abundance of Alistipes in Ovx_Lig group [(0.42±0.14)%] were significantly higher than that in Ovx group [(0.17±0.05)%] ( t=4.45, P<0.001). Tryptophan metabolism analysis showed that the content of kynurenic acid [(531.12±158.60) ng/g] in Ovx_Lig group were significantly higher than that in Ovx group [(400.42±57.96) ng/g] ( t=2.19, P=0.046). And the content of indole-3-carbaldehyde [(383.37±144.06) ng/g] in Ovx_Lig group were significantly lower than Ovx group [(701.72±141.93) ng/g] ( t=4.45, P<0.001). Correlation analysis showed that relative abundance of Alistipes was positively correlated with kynurenic acid ( r=0.32, P=0.088), while negatively correlated with indole-3-carbaldehyde ( r=-0.32, P=0.088). Conclusions:Periodontitis can induce bone destruction of femur in estrogen-deficient mice, the mechanism of which may be related to Alistipes in gut and the tryptophan metabolites kynurenic acid and indole-3-carbaldehyde.