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.

BACKGROUND: Based on the China-VHD database, this study sought to develop and validate a Valvular Heart Disease- specific Age-adjusted Comorbidity Index (VHD-ACI) for predicting mortality risk in patients with VHD. METHODS & RESULTS: The China-VHD study was a nationwide, multi-centre multi-centre cohort study enrolling 13,917 patients with moderate or severe VHD across 46 medical centres in China between April-June 2018. After excluding cases with missing key variables, 11,459 patients were retained for final analysis. The primary endpoint was 2-year all-cause mortality, with 941 deaths (10.0%) observed during follow-up. The VHD-ACI was derived after identifying 13 independent mortality predictors: cardiomyopathy, myocardial infarction, chronic obstructive pulmonary disease, pulmonary artery hypertension, low body weight, anaemia, hypoalbuminaemia, renal insufficiency, moderate/severe hepatic dysfunction, heart failure, cancer, NYHA functional class and age. The index exhibited good discrimination (AUC, 0.79) and calibration (Brier score, 0.062) in the total cohort, outperforming both EuroSCORE II and ACCI (P < 0.001 for comparison). Internal validation through 100 bootstrap iterations yielded a C statistic of 0.694 (95% CI: 0.665-0.723) for 2-year mortality prediction. VHD-ACI scores, as a continuous variable (VHD-ACI score: adjusted HR (95% CI): 1.263 (1.245-1.282), P < 0.001) or categorized using thresholds determined by the Yoden index (VHD-ACI ≥ 9 vs. < 9, adjusted HR (95% CI): 6.216 (5.378-7.184), P < 0.001), were independently associated with mortality. The prognostic performance remained consistent across all VHD subtypes (aortic stenosis, aortic regurgitation, mitral stenosis, mitral regurgitation, tricuspid valve disease, mixed aortic/mitral valve disease and multiple VHD), and clinical subgroups stratified by therapeutic strategy, LVEF status (preserved vs. reduced), disease severity and etiology. CONCLUSION The VHD-ACI is a simple 13-comorbidity algorithm for the prediction of mortality in VHD patients and providing a simple and rapid tool for risk stratification.

ObjectiveTo evaluate the quality of research and evidence related to antihypertensive Chinese patent medicines combined with western medicines for the treatment of hypertension， synthesize and update the evidence， form expert consensus， and provide evidence for clinical decision-making. MethodThe databases of China National Knowledge Infrastructure （CNKI）， WanFang Data Knowledge Service Platform （WanFang）， Vip Chinese Science and Technology Journal Database （VIP）， Chinese Biomedical Literature Service System （Sinomed）， National Library of Medicine （PubMed）， Cochrane Library， Web of Science， and US Clinical Trials Registry were searched for randomized controlled trials of antihypertensive Chinese medicine combined with western medicine for the treatment of hypertension from database construction to July 31， 2022. The quality of the literature was evaluated using the bias risk assessment tool in Cochrane Handbook 6.3. Evidence synthesis of main outcome indicators was performed using R software. The Grading of Recommendations Assessment， Development， and Evaluation profiler （GRADEprofiler） 3.6 was employed to evaluate the quality of evidence. Expert consensus was formed based on the Delphi method after two rounds of voting. Result64 pieces of literature were included， and the results of literature quality evaluation and risk of bias showed that 70.31% （45/64） of the studies indicated some risks， and 29.69% （19/64） indicated high risks. Compared with conventional western medicines， the combination of Chinese patent medicines with western medicines can significantly lower systolic pressure （SBP） and diastolic pressure （DBP）， increase the effective rate of antihypertensive， reduce the incidence of adverse reactions， endothelin-1， and traditional Chinese medicine syndrome scores. Egger's test showed that Songling Xuemaikang capsules reduced SBP and DBP. Tianma Gouteng granules reduced SBP and DBP and increased the effective rate of antihypertensive， and Xinmaitong capsules reduced SBP and increased the effective rate of antihypertensive， without significant publication bias. Songling Xuemaikang capsules increased the effective rate of antihypertensive， and Xinmaitong capsules decreased DBP， with significant publication bias. The results of the GRADE evidence quality evaluation showed that most evidence was at grades B and C. Finally， four strong recommendations and 14 weak recommendations were formed. ConclusionCompared with conventional western medicines for the treatment of hypertension， antihypertensive Chinese patent medicines combined with western medicines have advantages in reducing blood pressure and improving drug use safety， but they are mostly weak recommendations in terms of efficacy， and more high-quality evidence is needed.

Objective:To compare the anesthetic potency of dexmedetomidine combined with remifentanil for colonoscopy in the patients with different body mass indexes (BMIs) to assess the clinical significance of the influence of weight on the level of pain during the procedure.Methods:American Society of Anesthesiologists Physical Status classification Ⅰ or Ⅱ patients, aged 18-64 yr, undergoing elective colonoscopy, were divided into 3 groups based on the BMI: group Ⅰ (underweight group, BMI<18.5 kg/m 2), group Ⅱ (normal weight group, BMI 18.5-24.0 kg/m 2), and group III (overweight group, 24.0 kg/m 2 < BMI <30.0 kg/m 2). The prescribed dose of dexmedetomidine was infused within 2 min, then remifentanil was infused as a bolus of 1 μg/kg within 2 min followed by an infusion of 0.1 μg · kg -1 · min -1 throughout the surgery, and then colonoscopy was performed in patients of each group. The up-and-down sequential allocation was used to determine the dose of dexmedetomidine, the initial dose of dexmedetomidine in each group was 0.3 μg/kg, and the ratio between the two successive doses was 1.2. The positive response was defined as the Modified Observer′s Assessment of Alertness/Sedation Scale score > 1 and occurrence of body movement during the operation. Each time the dose of dexmedetomidine increased/decreased in the next patient depending on whether or not the response was positive. The median effective dose (ED 50) and 95% confidence interval ( CI) of dexmedetomidine were calculated using the Dixon-Massey formula. Results:Compared with group Ⅰ (0.42 [95% CI 0.38-0.47] μg/kg), the ED 50 of dexmedetomidine was significantly decreased in group II (0.23 [95% CI 0.19-0.32] μg/kg) and in group III (0.18 [95% CI 0.13-0.22] μg/kg) ( P<0.05). The ED 50 of dexmedetomidine was significantly decreased in group Ⅲ when compared with group Ⅱ ( P<0.05). Conclusions:With the increase of patients′ BMIs, the anesthetic potency of dexmedetomidine for colonoscopy is significantly enhanced when combined with remifentanil, indicating that clinicians should pay attention to the influence of weight on the level of pain during procedures.

This study aims to explore the chemical composition of Rehmanniae Radix braised with mild fire and compare the effect of processing method on the chemical composition of Rehmanniae Radix. To be specific, ultra-high performance liquid chromatography with linear ion trap-orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) was used to screen the chemical constituents of Rehmanniae Radix. The chemical constituents were identified based on the relative molecular weight and fragment ions, literature information, and Human Metabolome Database(HMDB). The ion peak area ratio of each component before and after processing was used as the index for the variation. SIMCA was employed to establish principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) models of different processed products. According to the PCA plot, OPLS-DA plot, and VIP value, the differential components before and after the processing were screened out. The changes of the content of differential components with the processing method were analyzed. A total of 66 chemical components were identified: 57 of raw Rehmanniae Radix, 55 of steamed Rehmanniae Radix, 55 of wine-stewed Rehmanniae Radix, 51 of repeatedly steamed and sundried Rehmanniae Radix Praeparata, 62 of traditional bran-braised Rehmanniae Radix, and 63 of electric pot-braised Rehmanniae Radix. Among them, the 9 flavonoids of braised Rehmanniae Radix were from Citri Reticulatae Pericarpium. PCA suggested significant differences in the chemical composition of Rehmanniae Radix Praeparata prepared with different processing methods. OPLS-DA screened out 32 chemical components with VIP value >1 as the main differential components. Among the differential components, 9 were unique to braised Rehmanniae Radix(traditional bran-braised, electric pot-braised) and the degradation rate of the rest in braised(traditional bran-braised, electric pot-braised) or repeatedly steamed and sundried Rehmanniae Radix was higher than that in the steamed or wine-stewed products. The results indicated the chemical species and component content of Rehmanniae Radix changed significantly after the processing. The 32 components, such as rehmapicrogenin, martynoside, jionoside D, aeginetic acid, hesperidin, and naringin, were the most important compounds to distinguish different processed products of Rehmanniae Radix. The flavonoids introduced by Citri Reticulatae Pericarpium as excipient may be the important material basis for the effectiveness of braised Rehmanniae Radix compared with other processed products.
Humans ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal/chemistry* ; Plant Extracts/chemistry* ; Rehmannia/chemistry* ; Flavonoids/analysis*

Management and treatment of terminal metastatic castration-resistant prostate cancer (mCRPC) remains heavily debated. We sought to investigate the efficacy of programmed cell death 1 (PD-1) inhibitor plus anlotinib as a potential solution for terminal mCRPC and further evaluate the association of genomic characteristics with efficacy outcomes. We conducted a retrospective real-world study of 25 mCRPC patients who received PD-1 inhibitor plus anlotinib after the progression to standard treatments. The clinical information was extracted from the electronic medical records and 22 patients had targeted circulating tumor DNA (ctDNA) next-generation sequencing. Statistical analysis showed that 6 (24.0%) patients experienced prostate-specific antigen (PSA) response and 11 (44.0%) patients experienced PSA reduction. The relationship between ctDNA findings and outcomes was also analyzed. DNA-damage repair (DDR) pathways and homologous recombination repair (HRR) pathway defects indicated a comparatively longer PSA-progression-free survival (PSA-PFS; 2.5 months vs 1.2 months, P = 0.027; 3.3 months vs 1.2 months, P = 0.017; respectively). This study introduces the PD-1 inhibitor plus anlotinib as a late-line therapeutic strategy for terminal mCRPC. PD-1 inhibitor plus anlotinib may be a new treatment choice for terminal mCRPC patients with DDR or HRR pathway defects and requires further investigation.
Male ; Humans ; Prostate-Specific Antigen ; Treatment Outcome ; Prostatic Neoplasms, Castration-Resistant/drug therapy* ; Immune Checkpoint Inhibitors/therapeutic use* ; Retrospective Studies

The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease, including the devastating COVID-19. Novel effective antivirals with broad-spectrum coverage are urgently needed. Herein, we reported a novel broad-spectrum antiviral compound PAC5. Oral administration of PAC5 eliminated HBV cccDNA and reduced the large antigen load in distinct mouse models of HBV infection. Strikingly, oral administration of PAC5 in a hamster model of SARS-CoV-2 omicron (BA.1) infection significantly decreases viral loads and attenuates lung inflammation. Mechanistically, PAC5 binds to a pocket near Asp49 in the RNA recognition motif of hnRNPA2B1. PAC5-bound hnRNPA2B1 is extensively activated and translocated to the cytoplasm where it initiates the TBK1-IRF3 pathway, leading to the production of type I IFNs with antiviral activity. Our results indicate that PAC5 is a novel small-molecule agonist of hnRNPA2B1, which may have a role in dealing with emerging infectious diseases now and in the future.
Animals ; Mice ; Antiviral Agents/pharmacology* ; COVID-19 ; Hepatitis B virus ; Interferon Type I/metabolism* ; SARS-CoV-2/drug effects* ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B/antagonists & inhibitors*

OBJECTIVE: To evaluate the feasibility and tolerability of metoprolol standard dosing pathway (MSDP) in Chinese patients with acute coronary syndrome (ACS). METHODS: In this multicenter, prospective, open label, single-arm and interventional study that was conducted from February 2018 to April 2019 in fifteen Chinese hospitals. A total of 998 hospitalized patients aged ≥ 18 years and diagnosed with ACS were included. The MSDP was applied to all eligible ACS patients based on the standard treatment recommended by international guidelines. The primary endpoint was the percentage of patients achieving the target dose at discharge (V2). The secondary endpoints included the heart rate and blood pressure at V2 and four weeks after discharge (V4), and percentage of patients experiencing bradycardia (heart rate < 50 beats/min), hypotension (blood pressure < 90/60 mmHg) and transient cardiac dysfunction at V2 and V4. RESULTS: Of the 998 patients, 29.46% of patients achieved the target dose (≥ 95 mg/d) at V2. The total population was divided into two groups: target group (patients achieving the target dose at V2) and non-target group (patients not achieving the target dose at V2). There was significant difference in the reduction of heart rate from baseline to discharge in the two groups (-4.97 ± 11.90 beats/min vs. -2.70 ± 9.47 beats/min, P = 0.034). There was no significant difference in the proportion of bradycardia that occurred in the two groups at V2 (0 vs. 0, P = 1.000) and V4 (0.81% vs. 0.33%, P = 0.715). There was no significant difference in the proportion of hypotension between the two groups at V2 (0.004% vs. 0.004%, P = 1.000) and V4 (0 vs. 0.005%, P = 0.560). No transient cardiac dysfunction occurred in two groups during the study. A total of five adverse events (1.70%) and one serious adverse event (0.34%) were related to the pathway in target group. CONCLUSIONS In Chinese ACS patients, the feasibility and tolerability of the MSDP have been proved to be acceptable.

Mammals exhibit limited heart regeneration ability, which can lead to heart failure after myocardial infarction. In contrast, zebrafish exhibit remarkable cardiac regeneration capacity. Several cell types and signaling pathways have been reported to participate in this process. However, a comprehensive analysis of how different cells and signals interact and coordinate to regulate cardiac regeneration is unavailable. We collected major cardiac cell types from zebrafish and performed high-precision single-cell transcriptome analyses during both development and post-injury regeneration. We revealed the cellular heterogeneity as well as the molecular progress of cardiomyocytes during these processes, and identified a subtype of atrial cardiomyocyte exhibiting a stem-like state which may transdifferentiate into ventricular cardiomyocytes during regeneration. Furthermore, we identified a regeneration-induced cell (RIC) population in the epicardium-derived cells (EPDC), and demonstrated Angiopoietin 4 (Angpt4) as a specific regulator of heart regeneration. angpt4 expression is specifically and transiently activated in RIC, which initiates a signaling cascade from EPDC to endocardium through the Tie2-MAPK pathway, and further induces activation of cathepsin K in cardiomyocytes through RA signaling. Loss of angpt4 leads to defects in scar tissue resolution and cardiomyocyte proliferation, while overexpression of angpt4 accelerates regeneration. Furthermore, we found that ANGPT4 could enhance proliferation of neonatal rat cardiomyocytes, and promote cardiac repair in mice after myocardial infarction, indicating that the function of Angpt4 is conserved in mammals. Our study provides a mechanistic understanding of heart regeneration at single-cell precision, identifies Angpt4 as a key regulator of cardiomyocyte proliferation and regeneration, and offers a novel therapeutic target for improved recovery after human heart injuries.
Humans ; Mice ; Rats ; Cell Proliferation ; Heart/physiology* ; Mammals ; Myocardial Infarction/metabolism* ; Myocytes, Cardiac/metabolism* ; Pericardium/metabolism* ; Single-Cell Analysis ; Zebrafish/metabolism*