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.

The mechanism of L-perilla alcohol(L-POH) in intervening hypoxic pulmonary hypertension(HPAH) was discussed based on network pharmacology, and experimental verification. The active components and potential targets of the volatile oil of Rhodiola tangutica(VORA) in the intervention of HPAH were screened by network pharmacology. The biological process of Gene Ontology(GO) and the signaling pathway enrichment of Kyoto Encyclopedia of Genes and Genomes(KEGG) were analyzed for the core targets, and a "component-common target-disease" network was constructed. Four active components were screened from VORA: L-POH, linalool, geraniol, and(-)-myrtenol. The core targets for treating HPAH were HSP90AA1, AKT1, ESR1, PIK3CA, EP300, EGFR, and JAK2. GO enrichment analysis mainly involved biological processes such as reaction to hypoxia, heme binding, and steroid binding. KEGG enrichment analysis mainly involved hypoxia-inducing factor 1(HIF-1) signaling pathway, phosphatidylinositol 3-kinase/protein kinase B(PI3K/AKT) signaling pathway, and Janus kinase/activator of signal transduction and transcription(JAK/STAT) signaling pathway. The vasodilation effects of the four active components were screened by perfusion experiment of extracorporeal vascular rings, and the mechanism of the main active component L-POH was studied by channel blockers. The inhibitory effects of the four active components on the proliferation of pulmonary artery smooth muscle cells(PASMCs) induced by hypoxia were screened by cell proliferation experiment, and the mechanism of the main active component L-POH was studied by flow cytometry, cell cycle experiment, and Western blot. The results showed that L-POH could directly act on vascular smooth muscle to relax pulmonary arterioles, induce ATP-sensitive potassium channels to open, and inhibit extracellular Ca~(2+) influx through voltage-gated calcium channels to relax blood vessels. In addition, L-POH could inhibit the abnormal proliferation of PASMCs induced by hypoxia and promote its apoptosis, and its mechanism may be related to the increase in Bax protein expression and the decrease in p-JAK2, p-STAT3, Bcl-2, and cyclinA2 protein expression. In summary, L-POH can interfere with HPAH by relaxing pulmonary arterioles and inhibiting the proliferation of smooth muscle cells.
Network Pharmacology ; Animals ; Hypertension, Pulmonary/physiopathology* ; Drugs, Chinese Herbal/administration & dosage* ; Rats ; Hypoxia/metabolism* ; Rhodiola/chemistry* ; Signal Transduction/drug effects* ; Humans ; Monoterpenes/chemistry* ; Male ; Cell Proliferation/drug effects* ; Rats, Sprague-Dawley

This study investigated the effects of total flavonoids of Dracocephalum moldavica(TFDM) on apoptosis in rat H9c2 cells induced by endoplasmic reticulum stress(ERS) established by oxygen-glucose deprivation and reoxygenation(OGD/R) injury and tunicamycin(TM), and explored the potential mechanisms. After successful modeling, the following groups were set in this experiment: control group, model(OGD/R or TM) group, and TFDM low-, medium-, and high-dose groups(12.5, 25, and 50 μg·mL~(-1)). The OGD/R injury model was constructed in vitro. Cell proliferation was assessed using the cell counting kit-8(CCK-8) method. The levels of lactate dehydrogenase(LDH) and creatine kinase MB isoenzyme(CKMB) in the cell supernatant were detected. Western blot was used to assess the expression of ERS-related proteins, including glucose regulatory protein 78(GRP78), C/EBP homologous protein(CHOP), activating transcription factor 6(ATF6), and apoptotic proteins B-cell lymphoma 2(Bcl-2) and Bcl-2-associated X protein(Bax). Apoptosis was detected using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling(TUNEL) method. In the TM-induced ERS model, Western blot was used to measure the expression of ERS pathway-related proteins GRP78, CHOP, inositol-requiring enzyme 1(IRE1), X-box binding protein 1(XBP1), protein kinase RNA-like endoplasmic reticulum kinase(PERK), eukaryotic initiation factor 2α(eIF2α), ATF6, p-ATF6, and apoptotic proteins Bcl-2, Bax, cysteinyl aspartate specific proteinase-12(caspase-12), and cleaved caspase-12. Gene expression of GRP78, CHOP, PERK, and ATF6 was detected by real-time fluorescence quantitative PCR(RT-qPCR). Apoptosis was again detected using the TUNEL method. The results showed that in the OGD/R model, compared with the control group, the levels of LDH and CKMB in the cell supernatant were significantly increased in the OGD/R group. Compared with the OGD/R group, the levels of LDH and CKMB in the TFDM group were significantly reduced. Western blot results revealed that compared with the control group, the expression of ERS-related proteins and Bax in the OGD/R group was significantly increased, while the expression of Bcl-2 was significantly decreased. Compared with the OGD/R group, the expression of ERS-related proteins and Bax in the TFDM groups was significantly reduced, and the expression of Bcl-2 was significantly increased. TUNEL assay showed that apoptosis was significantly decreased after TFDM treatment. In the TM-induced ERS experiment, compared with the control group, the expression of ERS-related genes, ERS-related proteins, and apoptotic proteins in the TM group was significantly increased, while the expression of Bcl-2 was significantly decreased. Compared with the TM group, the expression of ERS-related genes, ERS-related proteins, and apoptotic proteins in the TFDM group was significantly reduced, and the expression of Bcl-2 was significantly increased. These results suggest that ERS exists in the OGD/R-injured H9c2 cell model, and TFDM can effectively inhibit ERS-induced apoptosis. The mechanism may be related to the downregulation of ERS pathway-related proteins and apoptotic proteins.
Animals ; Endoplasmic Reticulum Stress/drug effects* ; Apoptosis/drug effects* ; Rats ; Flavonoids/pharmacology* ; Glucose/metabolism* ; Cell Line ; Lamiaceae/chemistry* ; Drugs, Chinese Herbal/pharmacology* ; Oxygen/metabolism* ; Reperfusion Injury/physiopathology* ; Myocytes, Cardiac/cytology*

Peyronie's disease (PD) is a disorder characterized by fibrous plaque formation in the penile tissue that leads to curvature and complications in advanced stages. In this study, we aimed to compare four injectable induction agents for the establishment of a robust rat model of PD: transforming growth factor-β1 (TGF-β1), fibrin, sodium tetradecyl sulfate (STS) combined with TGF-β1, and polidocanol (POL) combined with TGF-β1. The results showed that injection of TGF-β1 or fibrin into the tunica albuginea induced pathological endpoints without causing penile curvature. The STS + TGF-β1 combination resulted in both histological and morphological alterations, but with a high incidence of localized necrosis that led to animal death. The POL + TGF-β1 combination produced pathological changes and curvature comparable to STS + TGF-β1 and led to fewer complications. In conclusion, fibrin, STS + TGF-β1, and POL + TGF-β1 all induced PD with a certain degree of penile curvature and histological fibrosis in rats. The POL + TGF-β1 combination offered comparatively greater safety and clinical relevance and may have the greatest potential for PD research using model rats.
Animals ; Male ; Penile Induration/drug therapy* ; Rats ; Transforming Growth Factor beta1/metabolism* ; Disease Models, Animal ; Fibrin ; Penis/drug effects* ; Polidocanol/administration & dosage* ; Rats, Sprague-Dawley ; Polyethylene Glycols/administration & dosage* ; Injections

OBJECTIVE: To evaluate the efficacy of shockwave therapy, acupuncture, hyperthermia, biofeedback therapy, electrical nerve stimulation, magnetotherapy and ultrasound therapy in the treatment of chronic prostatitis/chronic pelvic pain syndrome(CP/CPPS), and to provide evidence-based support for clinical decision-making. METHODS: Two researchers independently searched PubMed, Web of Science, Embase, Cochrane Library, CNKI, Wanfang, VIP and Chinese Biomedical Literature databases for randomized controlled trials(RCTs) on the effects of different interventions on CP/CPPS from the establishment of the databases to August 2024. We evaluated the quality of the included literature and extracted the relevant data according to the Cochrane Handbook for Systematic Reviews of Interventions, followed by network meta-analysis using Revman 5.3, R 4.33 and Stata17 software. RESULTS: A total of 25 RCTs involving 1 794 cases were included. The results of network meta-analysis showed that electrical nerve stimulation, shockwave therapy, biofeedback therapy, magnetotherapy, ultrasound therapy and acupuncture were significantly superior to conventional medication and placebo in the total NIH-CPSI scores(P< 0.05), and so were electrical nerve stimulation and shockwave therapy to acupuncture and hyperthermia(P< 0.05), magnetic therapy to hyperthermia, and ultrasound therapy to placebo(P< 0.05). Shockwave therapy, biofeedback therapy, electrical nerve stimulation, magnetotherapy and ultrasound therapy achieved remarkably better clinical efficacy than conventional medication and placebo in the treatment of CP/CPPS, and so did shockwave therapy than electrical nerve stimulation, hyperthermia, ultrasonic therapy, magnetotherapy and acupuncture. CONCLUSION For the treatment of CP/CPPS, electrical nerve stimulation is advantageous over the other interventions in improving total NIH-CPSI scores, and shockwave therapy is advantageous in relieving pain symptoms and clinical efficacy. This conclusion, however, needs to be further verified by more high-quality clinical studies.
Humans ; Acupuncture Therapy ; Biofeedback, Psychology ; Chronic Disease ; Electric Stimulation Therapy ; Extracorporeal Shockwave Therapy ; Magnetic Field Therapy ; Pelvic Pain/therapy* ; Prostatitis/therapy* ; Randomized Controlled Trials as Topic ; Ultrasonic Therapy

RNA-based gene therapy has been widely used for various diseases, and extensive studies have proved that suitable delivery routes greatly help the development of RNA therapeutics. Identifying a safe and effective delivery system is key to realizing RNA therapeutics' clinical translation. Inhalation is a non-invasive pulmonary delivery modality that can enhance the retention of therapeutic agents in the lungs with negligible toxicity, thereby improving patient compliance. Inhaled RNA therapeutics are increasingly becoming an area of focus for researchers; however, only several clinical trials have explored inhaled delivery of RNA for pulmonary diseases. This review presents an overview of recent advances in inhaled delivery systems for RNA therapeutics, including viral and nonviral systems, highlighting state of the art regarding inhalation in the messenger RNA (mRNA) field. We also summarize the applications of mRNA inhalants in infectious and other lung diseases. Simultaneously, the research progresses on small interfering RNAs (siRNAs), antisense oligonucleotides (ASOs), and different types of RNA are also discussed to provide new strategies for developing RNA inhalation therapy. Finally, we clarify the challenges inhaled RNA-based therapeutics face before their widespread adoption and provide insights to help advance this exciting field to the bedside.

OBJECTIVE: To investigate the association between long-term glycemic control and cerebral infarction risk in patients with diabetes through a large-scale cohort study. METHODS: This prospective, community-based cohort study included 12,054 patients with diabetes. From 2006 to 2012, 38,272 fasting blood glucose (FBG) measurements were obtained from these participants. FBG trajectory patterns were generated using latent mixture modelling. Cox proportional hazards models were applied to assess the subsequent risk of cerebral infarction associated with different FBG trajectory patterns. RESULTS: At baseline, the mean age of the participants was 55.2 years. Four distinct FBG trajectories were identified based on FBG concentrations and their changes over the 6-year follow-up period. After a median follow-up of 6.9 years, 786 cerebral infarction events were recorded. Different trajectory patterns were associated with significantly varied outcome risks (Log-Rank P < 0.001). Compared with the low-stability group, Hazard Ratio ( HR) adjusted for potential confounders were 1.37 for the moderate-increasing group, 1.23 for the elevated-decreasing group, and 2.08 for the elevated-stable group. CONCLUSION Sustained high FBG levels were found to play a critical role in the development of ischemic stroke among patients with diabetes. Controlling FBG levels may reduce the risk of cerebral infarction.
Humans ; Cerebral Infarction/blood* ; Middle Aged ; Male ; Female ; Blood Glucose/analysis* ; Fasting/blood* ; Aged ; Prospective Studies ; Risk Factors ; Diabetes Mellitus/blood* ; Adult ; Proportional Hazards Models

Ambient air pollution is increasingly being recognized as a risk factor for heart failure; however, its effects on cardiac biomarkers remain unclear. This scoping review assessed the existing evidence on the association between air pollution and cardiac biomarkers in heart failure, described the key concepts, synthesized data, and identified research gaps. Following the PRISMA-ScR guidelines, PubMed, Embase, Web of Science, and CNKI databases were searched for studies on air pollution, heart failure, and biomarkers. A total of 765 records were screened, and 81 full texts were assessed for eligibility, resulting in 15 studies. The results showed that the exposure to particulate matter was associated with elevated N-terminal pro-B-type natriuretic peptide and troponin levels. Several studies have linked particulate matter exposure to a higher cardiovascular risk and heart failure biomarkers. Inflammatory and oxidative stress markers were consistently elevated across studies, supporting the biological relevance of these associations. However, few studies have focused specifically on populations with heart failure or clinically relevant biomarkers, and the evidence for gaseous pollutants remains inconclusive. These findings highlight the need to integrate environmental risk assessment into heart failure care and inform policy efforts to reduce the pollution-related cardiovascular burden. Further research should address these gaps through improved exposure assessments and the integration of mechanistic evidence.
Heart Failure/epidemiology* ; Biomarkers/metabolism* ; Humans ; Air Pollution/adverse effects* ; Air Pollutants/adverse effects* ; Particulate Matter/adverse effects* ; Environmental Exposure ; Natriuretic Peptide, Brain/blood* ; Oxidative Stress ; Troponin/blood*

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.