OBJECTIVE: To assess the efficacy of Qingda Granule (QDG) in ameliorating hypertension-induced cardiac damage and investigate the underlying mechanisms involved. METHODS: Twenty spontaneously hypertensive rats (SHRs) were used to develope a hypertension-induced cardiac damage model. Another 10 Wistar Kyoto (WKY) rats were used as normotension group. Rats were administrated intragastrically QDG [0.9 g/(kg•d)] or an equivalent volume of pure water for 8 weeks. Blood pressure, histopathological changes, cardiac function, levels of oxidative stress and inflammatory response markers were measured. Furthermore, to gain insights into the potential mechanisms underlying the protective effects of QDG against hypertension-induced cardiac injury, a network pharmacology study was conducted. Predicted results were validated by Western blot, radioimmunoassay immunohistochemistry and quantitative polymerase chain reaction, respectively. RESULTS: The administration of QDG resulted in a significant decrease in blood pressure levels in SHRs (P<0.01). Histological examinations, including hematoxylin-eosin staining and Masson trichrome staining revealed that QDG effectively attenuated hypertension-induced cardiac damage. Furthermore, echocardiography demonstrated that QDG improved hypertension-associated cardiac dysfunction. Enzyme-linked immunosorbent assay and colorimetric method indicated that QDG significantly reduced oxidative stress and inflammatory response levels in both myocardial tissue and serum (P<0.01). CONCLUSIONS Both network pharmacology and experimental investigations confirmed that QDG exerted its beneficial effects in decreasing hypertension-induced cardiac damage by regulating the angiotensin converting enzyme (ACE)/angiotensin II (Ang II)/Ang II receptor type 1 axis and ACE/Ang II/Ang II receptor type 2 axis.
Animals ; Drugs, Chinese Herbal/therapeutic use* ; Hypertension/pathology* ; Renin-Angiotensin System/drug effects* ; Rats, Inbred SHR ; Oxidative Stress/drug effects* ; Male ; Rats, Inbred WKY ; Blood Pressure/drug effects* ; Myocardium/pathology* ; Rats ; Inflammation/pathology*

OBJECTIVE: To evaluate the efficacy and safety of Shexiang Tongxin Dropping Pill (STDP) in treating stable angina patients with phlegm-heat and blood-stasis syndrome by exercise duration and metabolic equivalents. METHODS: This multicenter, randomized, double-blind, placebo-controlled clinical trial enrolled stable angina patients with phlegm-heat and blood-stasis syndrome from 22 hospitals. They were randomized 1:1 to STDP (35 mg/pill, 6 pills per day) or placebo for 56 days. The primary outcome was the exercise duration and metabolic equivalents (METs) assessed by the standard Bruce exercise treadmill test after 56 days of treatment. The secondary outcomes included the total angina symptom score, Chinese medicine (CM) symptom scores, Seattle Angina Questionnaire (SAQ) scores, changes in ST-T on electrocardiogram and adverse events (AEs). RESULTS: This trial enrolled 309 patients, including 155 and 154 in the STDP and placebo groups, respectively. STDP significantly prolonged exercise duration with an increase of 51.0 s, compared to a decrease of 12.0 s with placebo (change rate: -11.1% vs. 3.2%, P<0.01). The increase in METs was significantly greater in the STDP group than in the placebo group (change: -0.4 vs. 0.0, change rate: -5.0% vs. 0.0%, P<0.01). The improvement of total angina symptom scores (25.0% vs. 0.0%), CM symptom scores (38.7% vs. 11.8%), reduction of nitroglycerin consumption (100.0% vs. 11.3%), and all domains of SAQ, were significantly greater with STDP than placebo (all P<0.01). The changes in Q-T intervals at 28 and 56 days from baseline were similar between the two groups (both P>0.05). Twenty-five participants (16.3%) with STDP and 16 (10.5%) with placebo experienced AEs (P=0.131), with no serious AEs observed. CONCLUSION STDP could improve exercise tolerance in patients with stable angina and phlegm-heat and blood stasis syndrome, with a favorable safety profile. (Registration No. ChiCTR-IPR-15006020).
Humans ; Double-Blind Method ; Drugs, Chinese Herbal/adverse effects* ; Male ; Female ; Middle Aged ; Angina, Stable/physiopathology* ; Aged ; Syndrome ; Treatment Outcome ; Placebos ; Tablets

OBJECTIVES: To investigate the effects of periodontitis induced by Prevotella nigrescens (Pn) combined with ligation on cognitive functions in mice. METHODS: Twenty-four C57BL/6J mice were randomly divided into control group, ligation group, and ligation + Pn treatment (P+Pn) group. Experimental periodontitis was induced by silk ligation of the first molars followed by topical application of Pn for 6 weeks. After modeling, alveolar bone resorption was assessed using micro-CT and histological analysis. Learning and memory abilities of the mice were evaluated using open field test (OFT), novel object recognition test (NORT), and Morris water maze test (MWM). Seven weeks after the start of modeling, the mice were sacrificed for examining histopathological changes in the hippocampus using HE and Nissl staining. RESULTS: After 6 weeks of molar ligation, micro-CT revealed horizontal alveolar bone resorption and furcation exposure in the mice, and histological analysis showed apical migration of the junctional epithelium, epithelial ridge hyperplasia, and lymphocyte infiltration, and these changes were obviously worsened in P+Pn group. Alveolar bone height decreased significantly in both ligation groups compared to the control group. Cognitive tests showed that the mice in both of the ligation groups traveled shorter distances in OFT, showed reduced novel object preference in NORT, and exhibited longer escape latencies in MWM, and the mice in P+Pn group had significantly poorer performances in the tests. Histologically, obvious neuronal cytoplasmic degeneration, necrosis, nuclear pyknosis, vacuolation, and reduced Nissl bodies and viable neurons were observed in the hippocampal regions of the mice in the two ligation groups. CONCLUSIONS Pn infection aggravates alveolar bone destruction, accelerates necrosis and causes morphological abnormalities of neuronal cells in the hippocampus to reduce cognitive functions of mice with periodontitis.
Animals ; Periodontitis/microbiology* ; Mice ; Mice, Inbred C57BL ; Cognition ; Alveolar Bone Loss ; Hippocampus/pathology* ; Male ; Inflammation ; Maze Learning

Objective:To investigate the value of gadoxetate disodium-enhanced MRI combined with T 1 mapping in preoperative prediction of cytokeratin 19 (CK19) and glypican-3 (GPC3) positive dual-phenotype hepatocellular carcinoma (DPHCC). Methods:This case-control study included retrospectively enrolled patients with pathologically confirmed HCC from Central People′s Hospital of Zhanjiang (training set, n=85; December 2020 to July 2022) and the Second Affiliated Hospital, School of Medicine, South China University of Technology (test set, n=35; April 2023 to April 2024). Patients were categorized into CK19 and GPC3 positive DPHCC group (training set=19, test set=11) and non-DPHCC group (training set=66, test set=24) based on postoperative immunohistochemical staining. All patients received preoperative MRI scans, including gadoxetate disodium-enhanced imaging and T 1 mapping. Clinical data were collected, qualitative MRI features were evaluated, and quantitative parameters were measured, including signal intensity, T 1 values, apparent diffusion coefficient (ADC), tumor-to-liver ADC ratio (rADC), tumor-to-liver signal intensity ratio, and T 1 relaxation time reduction rate (ΔT 1%). Statistical comparisons between groups were performed using independent t-tests, Mann-Whitney U tests, or χ2 tests. Multivariate logistic regression identified independent predictors of CK19 and GPC3 positive DPHCC, and a combined model was constructed. Predictive performance was evaluated using area under the receiver operating characteristic curve (AUC), with DeLong test comparing model performance. Results:Significant intergroup differences were observed in alpha-fetoprotein (AFP), total bilirubin, direct bilirubin, DWI target sign, rADC, hepatobiliary-phase T 1 (T 1HBP), and ΔT 1% ( P<0.05). Multivariate analysis identified AFP>20 ng/ml ( OR=5.835, 95% CI 1.019-33.397, P=0.048), DWI target sign ( OR=13.408, 95% CI 2.216-81.131, P=0.005), and ΔT1%≤31% ( OR=14.429, 95% CI 2.166-96.125, P=0.006) as independent predictors of DPHCC. The AUC values of the aforementioned three independent predictors and the combined model for predicting DPHCC were 0.641 (95% CI 0.530-0.742), 0.679 (95% CI 0.569-0.777), 0.740 (95% CI 0.634-0.829), and 0.886 (95% CI 0.799-0.945) in the training set, and 0.568 (95% CI 0.390-0.743), 0.669 (95% CI 0.490-0.818), 0.689 (95% CI 0.511-0.843), and 0.824 (95% CI 0.658-0.931) in the test set, respectively. The DeLong test results showed that in the training set, the diagnostic performance of the combined model was superior to those of the three individual features ( Z=3.68, P<0.001; Z=3.15, P=0.002; Z=3.15, P=0.002). In the test cohort, the combined model demonstrated better diagnostic performance than AFP>20 ng/ml and ΔT 1%≤31% ( Z=2.15, P=0.032; Z=2.12, P=0.034), while no statistically significant difference was observed compared with the DWI target sign ( Z=1.77, P=0.076). Conclusion:The integrated model incorporating clinical data, gadoxetate disodium-enhanced MRI, and T 1 mapping parameters effectively predicts CK19 and GPC3 positive DPHCC.

AIM:To construct and breed infertility model mice with ATP/GTP binding carboxypeptidase 1(Agtpbp1)gene knockout homozygote(Agtpbp1-/-)by CRISPR/Cas9 technology,so as to provide an animal model for the subsequent exploration of the pathogenesis of Agtpbp1 gene in male sterility.METHODS:The CRISPR/Cas9 technology was used to obtain Agtpbp1 gene knockout heterozygote(Agtpbp1+/-)mice according to the data of the main protein functional region of Agtpbp1 gene and combined with Cas9 nuclease.The obtained Agtpbp1+/-mice were mated,and their offspring were genotyped by PCR and agarose gel electrophoresis.The expression of Agtpbp1 at different levels was detected by RT-PCR,Western blot and immunohistochemical staining to support the identification results.The HE staining was used to observe the mouse cerebellum and eyeball structure to analyze the effect of Agtpbp1 gene knockout on Purkinje cells and photoreceptor cells.The symptoms of ataxia in mice were observed in combination with behavioral tests.The growth of mice was observed,and the changes of testicular tissue volume and weight of male mice were analyzed.The HE staining was used to observe the changes of testicular structure,and PAS staining was used to observe the changes of testicular germ cell cycle.Finally,sperm analyzer was used to analyze the sperm motility,so as to analyze the growth and develop-ment of the mice.RESULTS:The male and female Agtpbp1+/-mice could continue to mate,and three genotypes,Agtpbp1 wild-type(Agtpbp1+/+),Agtpbp1+/-and Agtpbp1-/-,were obtained.The genotypes of the offspring mice were successfully identified by PCR.The results of RT-PCR,Western blot and immunohistochemical staining verified the successful con-struction of Agtpbp1-/-mouse model at different levels(P<0.05).The results of HE staining showed that Purkinje cells were lost in the cerebellum of Agtpbp1-/-mice and the number of photoreceptor cells in the eyeball was reduced.Behavioral tests confirmed that Agtpbp1-/-mice had ataxia symptoms such as motor dysfunction and uncoordinated movements.Com-pared with control group,the testicular volume and weight of Agtpbp1-/-mice were significantly reduced.The results of HE staining showed a very small amount of sperm in the testis of Agtpbp1-/-mice.Combined with the sperm analyzer,it was ob-served that the sperm motility,vitality and movement rate of Agtpbp1-/-mice were significantly lower than those of the con-trol mice.Testicular sections with PAS staining showed cell cycle arrest of the sperm from Agtpbp1-/-mice.CONCLU-SION:In this study,Agtpbp1 knockout mice were successfully bred.The deletion of Agtpbp1 caused the arrest of sper-matogenic cell differentiation and the decrease in sperm motility in adult male mice,resulting in infertility.At the same time,it provides a new experimental tool for further exploring the molecular mechanism of Agtpbp1-induced male sterility.

Soy is a vital source of plant carbohydrates.However,it poses significant allergenic risks,particularly to young children and animals.Among the various proteins in soy,β-conglycinin,which con-stitutes approximately 30％of total soy carbohydrates,is a primary allergen.Undigested β-conglycinin can lead to intestinal damage by inhibiting cell growth,disrupting the cytoskeleton,and inducing apopto-sis.It can also enter the lymphatic and circulatory systems,triggering allergic reactions.Conventional ELISA methods for detecting β-conglycinin rely on polyclonal or monoclonal antibodies,which are limited by their large molecular weight,difficulty in accessing the protein core,and sensitivity to acidic and bas-ic conditions.To address these limitations,this study aimed to develop nanobodies(Nbs)against β-con-glycinin.Nbs,derived from the variable regions of heavy-chain antibodies found in camelids,have a mo-lecular weight approximately one-tenth that of conventional antibodies.They offer advantages such as small size,stable structure,high specificity,and strong affinity.A female alpacas was immunized five times using β-conglycinin,which showed a heavy chain antibody potency of 1∶16 000 by ELISA.Pe-ripheral blood lymphocytes were subsequently isolated and total RNA was extracted.The variable region of the heavy-chain antibody was amplified via PCR,and recombinant plasmids were constructed and transformed into the E.coli competency strain ER2738.The resulting library contained about 3.5×108 CFU/mL,which increased to 1.15×1012 PFU/mL after phage rescue,with a 100％Nbs gene insertion rate,indicating high diversity.Its Nbs phage output was significantly enriched by four rounds of solid-phase elution with an enrichment rate of 155.9.Four rounds of solid-phase panning yielded 35 positive clones,all of which shared the same amino acid sequence upon sequencing.The selected Nb was ex-pressed in a prokaryotic system,and its binding ability to β-conglycinin was confirmed using Western blotting and ELISA.The results demonstrated excellent specificity and affinity.This research lays the groundwork for developing a rapid and efficient detection method for β-conglycinin using Nbs,potentially enhancing food safety and allergen management.

ObjectiveThis study leverages structural data from antigen-antibody complexes of the influenza A virus neuraminidase (NA) protein to investigate the spatial recognition relationship between the antigenic epitopes and antibody paratopes. MethodsStructural data on NA protein antigen-antibody complexes were comprehensively collected from the SAbDab database, and processed to obtain the amino acid sequences and spatial distribution information on antigenic epitopes and corresponding antibody paratopes. Statistical analysis was conducted on the antibody sequences, frequency of use of genes, amino acid preferences, and the lengths of complementarity determining regions (CDR). Epitope hotspots for antibody binding were analyzed, and the spatial structural similarity of antibody paratopes was calculated and subjected to clustering, which allowed for a comprehensively exploration of the spatial recognition relationship between antigenic epitopes and antibodies. The specificity of antibodies targeting different antigenic epitope clusters was further validated through bio-layer interferometry (BLI) experiments. ResultsThe collected data revealed that the antigen-antibody complex structure data of influenza A virus NA protein in SAbDab database were mainly from H3N2, H7N9 and H1N1 subtypes. The hotspot regions of antigen epitopes were primarily located around the catalytic active site. The antibodies used for structural analysis were primarily derived from human and murine sources. Among murine antibodies, the most frequently used V-J gene combination was IGHV1-12*01/IGHJ2*01, while for human antibodies, the most common combination was IGHV1-69*01/IGHJ6*01. There were significant differences in the lengths and usage preferences of heavy chain CDR amino acids between antibodies that bind within the catalytic active site and those that bind to regions outside the catalytic active site. The results revealed that structurally similar antibodies could recognize the same epitopes, indicating a specific spatial recognition between antibody and antigen epitopes. Structural overlap in the binding regions was observed for antibodies with similar paratope structures, and the competitive binding of these antibodies to the epitope was confirmed through BLI experiments. ConclusionThe antigen epitopes of NA protein mainly ditributed around the catalytic active site and its surrounding loops. Spatial complementarity and electrostatic interactions play crucial roles in the recognition and binding of antibodies to antigenic epitopes in the catalytic region. There existed a spatial recognition relationship between antigens and antibodies that was independent of the uniqueness of antibody sequences, which means that antibodies with different sequences could potentially form similar local spatial structures and recognize the same epitopes.

Objective To investigate the effects of periodontitis induced by Prevotella nigrescens(Pn)combined with ligation on cognitive functions in mice.Methods Twenty-four C57BL/6J mice were randomly divided into control group,ligation group,and ligation+Pn treatment(P+Pn)group.Experimental periodontitis was induced by silk ligation of the first molars followed by topical application of Pn for 6 weeks.After modeling,alveolar bone resorption was assessed using micro-CT and histological analysis.Learning and memory abilities of the mice were evaluated using open field test(OFT),novel object recognition test(NORT),and Morris water maze test(MWM).Seven weeks after the start of modeling,the mice were sacrificed for examining histopathological changes in the hippocampus using HE and Nissl staining.Results After 6 weeks of molar ligation,micro-CT revealed horizontal alveolar bone resorption and furcation exposure in the mice,and histological analysis showed apical migration of the junctional epithelium,epithelial ridge hyperplasia,and lymphocyte infiltration,and these changes were obviously worsened in P+Pn group.Alveolar bone height decreased significantly in both ligation groups compared to the control group.Cognitive tests showed that the mice in both of the ligation groups traveled shorter distances in OFT,showed reduced novel object preference in NORT,and exhibited longer escape latencies in MWM,and the mice in P+Pn group had significantly poorer performances in the tests.Histologically,obvious neuronal cytoplasmic degeneration,necrosis,nuclear pyknosis,vacuolation,and reduced Nissl bodies and viable neurons were observed in the hippocampal regions of the mice in the two ligation groups.Conclusion Pn infection aggravates alveolar bone destruction,accelerates necrosis and causes morphological abnormalities of neuronal cells in the hippocampus to reduce cognitive functions of mice with periodontitis.

Objective:To investigate the risk factors for detecting clinically significant prostate cancer (CSPCa) in patients with Prostate Imaging Reporting and Data System (PI-RADS) score≤3 on multi-parameter magnetic resonance imaging (mpMRI).Methods:Retrospective analysis was performed on the case data of 335 patients with suspected prostate cancer and PI-RADS score ≤3 who were admitted to Beijing Friendship Hospital, Capital Medical University from January 2013 to October 2022. All patients underwent 24-needle prostate biopsy. Clinical data such as age, body mass index, past medical history, serological laboratory indicators, and mpMRI imaging data were collected. The patients were grouped according to whether the puncture pathology was CSPCa or not, and the differences in clinical data between the two groups were analyzed by t-test, rank sum test and Chi-test. Multivariate Logistic regression analysis was further used to determine independent risk factors for MRI invisible prostate cancer, and receiver operating characteristics (ROC) curves were drawn. At the same time, further subgroup analysis was conducted based on whether prostate-specific antigen (PSA) was positive before puncture and PI-RADS score, respectively, and the same statistical method was used to further determine the influence of different serological indicators and PI-RADS score on the analysis results of risk factors. Results:Among all patients, 81 were CSPCa patients and 254 were non-CSPCa patients. Multivariate Logistic regression analysis showed that prostate-specific antigen density (PSAD) and PI-RADS score of 3 were independent risk factors for MRI invisible prostate cancer. At the same time, compared with suspected lesions located only in the transitional zone, the incidence of CSPCa in patients with suspected lesions located in the peripheral zone would increase, and the incidence of CSPCa would further increase when suspected lesions were found in both the transitional zone and the peripheral zone. In PSA-negative patients, only suspected lesion location was an independent risk factor for MRI invisible prostate cancer, while in PSA-positive patients, prostate volume, PSAD, and PI-RADS scores were independent risk factors. In subgroup analysis with different PI-RADS scores, suspicious lesions in both the transitional zone and peripheral zone indicate a higher likelihood of CSPCa. For patients with PI-RADS scores of 1 to 2, suspicious lesions in the peripheral zone alone may also indicated CSPCa, while for patients with PI-RADS scores of 3, the lower free prostate-specific antigen/total prostate-specific anti-principle was more accurate in predicting CSPCa.Conclusions:For patients who are clinically suspected of prostate cancer but whose PI-RADS score is less than or equal to 3 points indicated by mpMRI, it is necessary to further focus on the results of different serological indicators according to whether their PSA is positive and PI-RADS score respectively to judge whether patients should receive systemic prostate puncture, instead of using PSA level as a single indication for puncture. At the same time, clinicians should also pay full attention to the location of suspected lesions, when they are located in the peripheral zone, or there are suspected lesions in both the peripheral zone and the transitional zone, the possibility of CSPCa should be fully considered.

OBJECTIVE: Circadian rhythm disruption (CRD) is a risk factor that correlates with poor prognosis across multiple tumor types, including hepatocellular carcinoma (HCC). However, its mechanism remains unclear. This study aimed to define HCC subtypes based on CRD and explore their individual heterogeneity. METHODS: To quantify CRD, the HCC CRD score (HCCcrds) was developed. Using machine learning algorithms, we identified CRD module genes and defined CRD-related HCC subtypes in The Cancer Genome Atlas liver HCC cohort (n = 369), and the robustness of this method was validated. Furthermore, we used bioinformatics tools to investigate the cellular heterogeneity across these CRD subtypes. RESULTS: We defined three distinct HCC subtypes that exhibit significant heterogeneity in prognosis. The CRD-related subtype with high HCCcrds was significantly correlated with worse prognosis, higher pathological grade, and advanced clinical stages, while the CRD-related subtype with low HCCcrds had better clinical outcomes. We also identified novel biomarkers for each subtype, such as nicotinamide n-methyltransferase and myristoylated alanine-rich protein kinase C substrate-like 1. CONCLUSION We classify the HCC patients into three distinct groups based on circadian rhythm and identify their specific biomarkers. Within these groups greater HCCcrds was associated with worse prognosis. This approach has the potential to improve prediction of an individual's prognosis, guide precision treatments, and assist clinical decision making for HCC patients. Please cite this article as: Li XJ, Chang L, Mi Y, Zhang G, Zhu SS, Zhang YX, et al. Integrated-omics analysis defines subtypes of hepatocellular carcinoma based on circadian rhythm. J Integr Med. 2025; 23(4): 445-456.
Humans ; Carcinoma, Hepatocellular/pathology* ; Liver Neoplasms/pathology* ; Circadian Rhythm/genetics* ; Prognosis ; Male ; Female ; Biomarkers, Tumor/genetics* ; Middle Aged ; Machine Learning ; Computational Biology