A primary hallmark of pathological cardiac hypertrophy is excess protein synthesis due to enhanced translational activity. However, regulatory mechanisms at the translational level under cardiac stress remain poorly understood. Here we examined the translational regulations in a mouse cardiac hypertrophy model induced by transaortic constriction (TAC) and explored the conservative networks versus the translatome pattern in human dilated cardiomyopathy (DCM). The results showed that the heart weight to body weight ratio was significantly elevated, and the ejection fraction and fractional shortening significantly decreased 8 weeks after TAC. Puromycin incorporation assay showed that TAC significantly increased protein synthesis rate in the left ventricle. RNA-seq revealed 1,632 differentially expressed genes showing functional enrichment in pathways including extracellular matrix remodeling, metabolic processes, and signaling cascades associated with pathological cardiomyocyte growth. When combined with ribosome profiling analysis, we revealed that translation efficiency (TE) of 1,495 genes was enhanced, while the TE of 933 genes was inhibited following TAC. In DCM patients, 1,354 genes were upregulated versus 1,213 genes were downregulated at the translation level. Although the majority of the genes were not shared between mouse and human, we identified 93 genes, including Nos3, Kcnj8, Adcy4, Itpr1, Fasn, Scd1, etc., with highly conserved translational regulations. These genes were remarkably associated with myocardial function, signal transduction, and energy metabolism, particularly related to cGMP-PKG signaling and fatty acid metabolism. Motif analysis revealed enriched regulatory elements in the 5' untranslated regions (5'UTRs) of transcripts with differential TE, which exhibited strong cross-species sequence conservation. Our study revealed novel regulatory mechanisms at the translational level in cardiac hypertrophy and identified conserved translation-sensitive targets with potential applications to treat cardiac hypertrophy and heart failure in the clinic.
Animals ; Humans ; Cardiomegaly/physiopathology* ; Ribosomes/physiology* ; Protein Biosynthesis/physiology* ; Mice ; Cardiomyopathy, Dilated/genetics* ; Ribosome Profiling

This study aims to explore the effects and action mechanisms of the active ingredients in Buyang Huanwu Decoction(BYHWD), namely tetramethylpyrazine(TMP) and hydroxy-safflor yellow A(HSYA), on oxygen-glucose deprivation/reglucose-reoxygenation(OGD/R)-induced inflammation and oxidative stress of microglia(MG). Network pharmacology was used to screen the effective monomer ingredients of BYHWD and determine the safe concentration range for each component. Inflammation and oxidative stress models were established to further screen the best ingredient combination and optimal concentration ratio with the most effective anti-inflammatory and antioxidant effects. OGD/R BV2 cell models were constructed, and BV2 cells in the logarithmic growth phase were divided into a normal group, a model group, an HSYA group, a TMP group, and an HSYA + TMP group. Enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of inflammatory cytokines such as interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), and interleukin-6(IL-6). Oxidative stress markers, including superoxide dismutase(SOD), nitric oxide(NO), and malondialdehyde(MDA), were also measured. Western blot was used to analyze the protein expression of both inflammation-related pathway [Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB)] and oxidative stress-related pathway [nuclear factor erythroid 2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)]. Immunofluorescence was used to assess the expression of proteins such as inducible nitric oxide synthase(iNOS) and arginase-1(Arg-1). The most effective ingredients for anti-inflammatory and antioxidant effects in BYHWD were TMP and HSYA. Compared to the normal group, the model group showed significantly increased levels of IL-1β, TNF-α, IL-6, NO, and MDA, along with significantly higher protein expression of NF-κB, TLR4, Nrf2, and HO-1 and significantly lower SOD levels. The differences between the two groups were statistically significant. Compared to the model group, both the HSYA group and the TMP group showed significantly reduced levels of IL-1β, TNF-α, IL-6, NO, and MDA, lower expression of NF-κB and TLR4 proteins, higher levels of SOD, and significantly increased protein expression of Nrf2 and HO-1. Additionally, the expression of the M1-type MG marker iNOS was significantly reduced, while the expression of the M2-type MG marker Arg-1 was significantly increased. The results of the HSYA group and the TMP group had statistically significant differences from those of the model group. Compared to the HSYA group and the TMP group, the HSYA + TMP group showed further significant reductions in IL-1β, TNF-α, IL-6, NO, and MDA levels, along with significant reductions in NF-κB and TLR4 protein expression, an increase in SOD levels, and elevated Nrf2 and HO-1 protein expression. Additionally, the expression of the M1-type MG marker iNOS was reduced, while the M2-type MG marker Arg-1 expression increased significantly in the HSYA + TMP group compared to the TMP or HSYA group. The differences in the results were statistically significant between the HSYA + TMP group and the TMP or HSYA group. The findings indicated that the combined use of HSYA and TMP, the active ingredients of BYHWD, can effectively inhibit OGD/R-induced inflammation and oxidative stress of MG, showing superior effects compared to the individual use of either component.
Oxidative Stress/drug effects* ; Drugs, Chinese Herbal/pharmacology* ; Animals ; Mice ; Glucose/metabolism* ; Cell Line ; Inflammation/genetics* ; Oxygen/metabolism* ; Pyrazines/pharmacology* ; Microglia/metabolism* ; NF-E2-Related Factor 2/immunology* ; NF-kappa B/immunology* ; Toll-Like Receptor 4/immunology* ; Anti-Inflammatory Agents/pharmacology* ; Humans

Jinqi Jiangtang Capsule (JQJTC) is clinically used for the prevention and treatment of type 2 diabetes, but the contents of its main chemical components are not yet clear. In this study, an ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was established for the determination of 15 components in JQJTC, including new chlorogenic acid, chlorogenic acid, cryptochlorogenic acid, formononetin, ononin, calycosin, calycosin-7-glucoside, astragaloside IV, berberine, epiberberine, berberrubine, coptisine, jatrorrhizine, palmatine and magnoflorine. The method was used to determine the contents of 15 components in the capsule and then to investigate the influence of excipients on the contents of the components in JQJTC. The separation was performed on a ACQUITY UPLC BEH C₁₈ column (100 mm × 2.1 mm, 1.7 μm) with a mobile phase consisting of 0.1% acetic acid and 5 mmol·L^-1 ammonium acetate (A) and acetonitrile (B) with gradient elution at a flow rate of 0.3 mL·min^-1 and a column temperature at 40 ℃. Electron spray ionization was used for mass spectrometry in positive ion mode. The established method meets the requirements of methodology of content determination in Chinese pharmacopoeia. The contents of 15 components in JQJTC varied from high to low. The top 5 contents were berberine, chlorogenic acid, magnoflorine, coptisine, and cryptochlorogenic acid, accounting for 87.31% of the total content. The contents of 10 components, including the alkaloids of coptidis rhizoma (berberine, epiberberine, berberrubine, coptisine, jatrorrhizine, palmatine and magnoflorine) and the organic acids of honeysuckle (new chlorogenic acid, chlorogenic acid, and cryptochlorogenic acid) in the whole formula extract without excipients was significantly lower than that in the capsule. These components accounted for 99.20% of the determined component contents. In this experiment, an accurate, sensitive and efficient UHPLC-MS/MS method for the determination of multi-components in JQJTC was established, which stably and reliably detected the contents of 15 components in the capsule and could provide the basis for more comprehensive quality analysis. It was also found that excipients had an increasing effect on the contents of detected alkaloid and organic acid components, which may be beneficial to the effectiveness of the capsules.

Objective:We applied a hypoxia-induced model of human fetal retinal microvascular endothelial cell (RMEC) to study the effect of carbonic anhydrase 9 (CA9) on cell proliferation.Methods:The eyeballs of spontaneously aborted fetuses in Guangdong Women and Children's Hospital were obtained, and the retinas were isolated. RMEC was obtained by trypsin and collagenase two-step enzyme digestion, and endothelial cells were identified by CD34. The fetal RMEC and the purchased adult RMEC were cultured in normoxic and hypoxic incubators (1%O 2+5%CO 2+94%N 2), and the expression of CA9 was detected by qPCR and Western blot. After knocking down the CA9 by small interference RNA technique, the cell proliferation was detected by CCK-8 method, and the cell viability was detected by CCK-8 after adding CA9 inhibitor U-104. Results:The primary RMEC was extracted successfully. Immunofluorescence staining showed the percentage of CD34 positive cells in the third-generation cells was nearly 100%. The expression of CA9 mRNA in immature fetus and adult RMEC under hypoxia culture was higher than that under normoxic culture (fetal 1% O 2 group vs. fetal 21% O 2 group: 67.80±10.31 vs. 1.00±0.04, P<0.001; adult 1% O 2 group vs. adult 21% O 2 group: 1.72±0.22 vs. 1.00±0.02, P=0.014). Western blot analysis showed significantly increased expression of CA9 in the fetal RMEC exposed to hypoxia, which aligned with the expression of CA9 mRNA. When fetal RMEC was transfected with siCA9 20 nM, the knockdown rate of CA9 was 95% ( P<0.001). CCK-8 assay showed significantly lower proliferation of fetal RMEC cells in siCA9 group compared to siNC group (0.57±0.05 vs. 0.90±0.03, P<0.001), which was reflected by the OD value. With the addition of 100 μM CA9 inhibitor U-104, the viability of fetal RMEC in the treated groupwas significantly lower than that in the untreated group (99.16%±3.82% vs. 119.10% ±1.72%, P=0.002). Conclusions:The expression of CA9 differed between adult and preterm fetus in our hypoxia-induced RMEC model. Inhibiting CA9 can inhibit the proliferation of retinal microvascular endothelial cells of preterm fetus.

Objective To investigate the relieving effects of knockdown of long non-coding RNA(lncRNA)taurine up-regulated gene 1 (TUG1) on inhibiting nucleotide binding oligomerization domain like receptor protein 1 (NLRP1) inflammasome and the progression of Alzheimer’ s disease. Methods Wild-type (WT group, 10 mice) or amyloid precursor protein (APP) / presenilin-1 (PS1) transgenic mice (30 mice) with a genetic background of C57 / BL6 aged 9-10 weeks were used in this study. APP / PS1 transgenic mice were randomly divided into model group, model+lncRNA TUG1 short hairpin RNA (shRNA) group and model + shRNA non target (NT) group (n = 10) . Blood samples, cerebral cortex tissues, primary microglial cells and primary astrocytes were collected from mice 12 weeks of age on day 1 (3-month-old) and 32 weeks of age on day 1 (8-month-old), with 5 mice per group at each time point. Real-time PCR analysis was used to detect the expression levels of lncRNA TUG1 and macrophage migration inhibitory factor (MIF) mRNA in cerebral cortex tissues and primary microglial cells, and C1r and C1s mRNA levels in primary astrocytes of 3-month-old and 8-month-old mice in the above 4 groups, respectively. ELISA was used to determine the MIF in plasma samples of the above 4 groups of mice. Primary microglia and astrocytes from the cerebral cortex of 3-month-old and 8-month-old mice were co-cultured. CCK-8 method was used to determine the proliferation ability of the above cells. Western blotting was used to determine the expression levels of MIF, pro interleukin-1β (pro-IL-1β), apoptosis associated speck-like protein containing a caspase recrult domain(ASC), Caspase-1 (p20), Caspase-1 (full), NLRP1 and NLRP3 in cerebral cortex tissues of 3-month-old and 8-month-old mice. Immunofluorescent staining was used to determine amyloid beta(Aβ) in cerebral cortex of 8-month-old mice. Results At the age of 3-month-old and 8-month-old, compared with the WT group, the relative expression level of lncRNA TUG1 and MIF in cerebral cortex tissues and primary microglia of model group mice was significantly up-regulated, with primary microglial cells and astrocytes proliferation ability enhanced (P<0. 05) . Compared with the model group, the relative expression level of lncRNA TUG1 and MIF cerebral cortex tissues and primary microglia of model + lncRNA TUG1 shRNA group were significantly down-regulated, with primary microglial cells and astrocytes proliferation ability decreased (P<0. 05) . Compared with the WT group, MIF factor in the peripheral plasma of model group increased significantly, with pro-IL-1β,ASC,Caspase-1 (p20),Caspase-1 (full), NLRP1 and NLRP3 expression level up-regulated in the model group mice cerebral cortex tissues, with increased Aβ immunofluorescent indensity (P<0. 05) . Compared with the model group, MIF factor in the peripheral plasma, and pro-IL-1β, ASC, Caspase-1 (p20), Caspase-1 (full) and NLRP1 expression in the model + lncRNA TUG1 shRNA group mice cerebral cortex tissues were down-regulated, and Aβ immunofluorescent indensity decreased (P<0. 05), while NLRP3 expression level were not changed (P>0. 05) . There was no significant difference between the model group and the model+shRNA NT group mice of all the above factors (P>0. 05) . Conclusion In APP / PS1 transgenic mice, up-regulation of lncRNA TUG1 and MIF are positively associated with the activation of NLRP1 inflammasome in mice cerebral cortex tissues and primary microglia. Knock-down of lncRNA TUG1 can ameliorate the progression of Alzheimer’ s disease.

ObjectiveTo investigate the application of endoscopy in obtaining the great saphenous vein （GSV） during coronary artery bypass grafting （CABG） and explore the learning curve， with a particular focus on common challenges encountered during the learning process and their impact on early clinical outcomes. MethodsA retrospective analysis was conducted on clinical data from 83 patients who underwent off-pump CABG with endoscopic GSV harvesting at the First Affiliated Hospital of Zhengzhou University from July 2013 to April 2014. Patients were categorized into four groups based on the chronological order of their hospitalization： Group A （novice group， n=20）， Group B （proficient group， n=20）， Group C （progressive group， n=20）， and Group D （mature group， n=23）. Differences in perioperative and midterm follow-up outcomes among the groups were analyzed to determine the learning curve period. ResultsThe study population had a mean age of （60.22±8.06） years and a mean body weight of （69.77±11.66） kg. Comorbidities included hypertension （24 cases）， diabetes （26 cases）， and subacute cerebral infarction （14 cases）. The novice group exhibited significantly shorter GSV length-to-harvest time ratio relative to the other three groups （P<0.001） and a significantly higher incidence of main vein damage （P=0.006）. However， there was no statistically significant difference in graft patency at the 1-year follow-up. ConclusionThorough and reliable technical training in endoscopic GSV harvesting is essential to minimize vascular injury caused by novice operators. Approximately 20 cases of hands-on experience and a careful self-analysis of procedural challenges are likely required to achieve proficiency in GSV harvesting.

Objective To investigate the relationship between serum levels of E-cadherin and β-catenin and calcium phosphorus metabolism and carotid artery calcification in patients with diabetic nephropathy.Methods A total of 112 patients with diabetic nephropathy admitted to the hospital from May 2019 to No-vember 2022 were selected as the study group,and were divided into a carotid artery calcification group(n=44)and a non-carotid artery calcification group(n=68)according to the results of bilateral carotid artery col-or Doppler ultrasound.In addition,90 healthy people who underwent physical examination in the hospital dur-ing the same period were selected as the control group.Serum E-cadherin,β-catenin,calcium phosphorus me-tabolism levels were detected and compared.Pearson correlation analysis was used to explore the relationship between serum E-cadherin,β-catenin and calcium phosphorus metabolism in patients with diabetic nephropa-thy.Receiver operating characteristic(ROC)curve was used to evaluate the predictive value of serum E-cad-herin and β-catenin for carotid artery calcification in patients with diabetic nephropathy.Multivariate Logistic regression analysis was used to explore the influencing factors of carotid artery calcification in patients with di-abetic nephropathy.Results The levels of glycosylated hemoglobin,serum phosphorus,calcium-phosphorus product,parathyroid hormone(iPTH),creatinine,alkaline phosphatase and β-catenin in the study group were higher than those in the control group,and the level of E-cadherin was lower than that in the control group(P＜0.05).The levels of serum phosphorus,serum calcium,calcium phosphorus product,iPTH,creatinine,al-kaline phosphatase and β-catenin in the carotid artery calcification group were higher than those in the non-ca-rotid artery calcification group,and the level of E-cadherin was lower than that in the non-carotid artery calci-fication group(P＜0.05).Pearson correlation analysis showed that serum E-cadherin level in patients with di-abetic nephropathy was negatively correlated with serum phosphorus,serum calcium,calcium phosphorus product,iPTH,creatinine and alkaline phosphatase(r=-0.453,-0.654,-0.365,-0.490,-0.411,-0.377,all P＜0.001).The level of serum β-catenin was positively correlated with serum phosphorus,serum calcium,calcium phosphorus product,iPTH,creatinine,and alkaline phosphatase(r=0.444,0.345,0.421,0.398,0.651,0.622,all P＜0.001).ROC curve analysis showed that the area under the curve of serum E-cad-herin,β-catenin and their combination for predicting carotid artery calcification in diabetic nephropathy were 0.844(95％CI 0.795-0.894),0.853(95％CI 0.801-0.901)and 0.901(95％CI 0.801-0.901),respec-tively.Multivariate Logistic regression analysis showed that serum E-cadherin(OR=3.789,95％CI 2.055-6.983),β-catenin(OR=4.104,95％CI 1.795-9.385),calcium phosphorus product(OR=2.998,95％CI 1.895-4.743)and iPTH(OR=2.713,95％CI 1.787-4.118)were the influencing factors of carotid artery calcification in patients with diabetic nephropathy(P＜0.05).Conclusion The level of β-catenin is increased and the level of E-cadherin is decreased in patients with diabetic nephropathy.β-catenin and E-cadherin are closely related to calcium phosphorus metabolism and carotid artery calcification,which could be used as effec-tive indicators to evaluate carotid artery calcification in patients with diabetic nephropathy.The combination ofβ-catenin and E-cadherin has a higher predictive value for carotid artery calcification.

Objective To investigate the relationship between serum levels of growth differentiation factor-11(GDF-11)and S100 calcium binding protein A4(S100A4)and the severity and disease outcome in diabetic nephropathy(DN)patients.Methods A total of 95 DN patients admitted to the hospital from May 2021 to January 2023 were selected as the study group,and 110 healthy people were selected as the healthy group.The DN patients were divided into mild group(n=66)and severe group(n=29)according to the severity of the disease.The patients were followed up for half a year after discharge,and were divided into good prognosis group(n=64)and poor prognosis group(n=31)according to the prognosis.Serum GDF-11 and S100A4 lev-els were detected by enzyme-linked immunosorbent assay.Spearman rank correlation analysis was used to ex-plore the relationship between serum GDF-11,S100A4 levels and the severity of the disease.Receiver operat-ing characteristic(ROC)curve was used to evaluate the predictive value of serum GDF-11 and S100A4 for dis-ease outcome in DN patients,and multivariate Logistic regression was used to analyze the influencing factors of disease outcome in DN patients.Results The levels of serum GDF-11 and S100A4 in mild group and severe group were higher than those in healthy group,and those in severe group were higher than those in mild group(P＜0.05).Serum GDF-11 and S100A4 levels were positively correlated with the severity of DN patients(P＜0.05).The good prognosis group had significantly lower serum levels of GDF-11 and S100A4 than the poor prognosis group(P＜0.05).The area under the curve(AUC)of serum GDF-11 and S100A4 in predicting the outcome of DN patients was 0.785 and 0.839,respectively,and the AUC of combined prediction was 0.902.The proportion of type 2 diabetes(T2DM)duration,glomerular grade Ⅲ-Ⅳ,interstitial inflammation score 2,interstitial fibrosis and tubular atrophy(IFTA)score 2-3 points,estimate glomerular filtration rate＜60 mL/(min·1.73 m2)and the levels of total cholesterol,triglyceride,low-density lipoprotein choles-terol,24 h urinary protein,glycosylated hemoglobin,C-peptide,hematocrit,and erythrocyte sedimentation rate in the good prognosis group were higher than those in the good prognosis group(P＜0.05).Multivariate Lo-gistic regression analysis showed that the duration of T2DM ≥12.0 years,IFTA score ≥2 points,GDF-11≥700.82 ng/mL,S100A4 ≥ 211.53 ng/L were risk factors for poor prognosis in DN patients(P＜0.05).Conclusion The levels of serum S100A4 and GDF-11 are highly expressed in patients with diabetes mellitus,and are related to the severity and outcome of the disease,which are expected to be potential markers for eval-uating the condition and prognosis of diabetes mellitus.

Objective:To explore the clinical and genetic features of two Chinese pedigrees affected with Autosomal dominant intellectual developmental disorder 49 (MRD49).Methods:Two MRD49 pedigrees which were admitted to the Children′s Hospital Affiliated to Shandong University respectively on January 28, 2021 and November 10, 2022 were selected as the study subjects. Clinical data of the two pedigrees were collected and analyzed. Genomic DNA was extracted from peripheral blood samples of the probands and their family members. The probands were subjected to mutational analysis by high-throughput sequencing. Candidate variants were validated using real-time fluorescence quantitative PCR (q-PCR) or Sanger sequencing and bioinformatic analysis. This study was approved by the Medical Ethics Committee of the Children′s Hospital Affiliated to Shandong University (No. SDFE-IRB/T-2022002).Results:Proband 1 had presented with language delay, motor retardation and intellectual disability, and his maternal grandmother, mother, aunt and cousin all had various degrees of intellectual disability. Sequencing results showed that proband 1 had deletion of exons 3 ~ 7 of the TRIP12 gene. q-PCR verification showed that his mother, aunt, maternal grandmother and cousin had all harbored the same deletion. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as pathogenic (PVS1+ PM2_Supporting+ PP1). Proband 2, who had mainly presented with language delay, motor retardation and intellectual disability, and was found to harbor a heterozygous c.3010C>T (p.Arg1004*) variant of the TRIP12 gene, which was verified to be de novo in origin. Based on the guidelines from the ACMG, the variant was classified as pathogenic (PVS1+ PS2+ PM2_Supporting). Conclusion:This study had diagnosed two MRD49 families through high-throughput sequencing. Above findings have enriched the phenotypic and mutational spectrum of MRD49 in China, which has also facilitated genetic counseling for the two pedigrees.

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.