Background In recent years, a growing number of studies have indicated that perfluorooctanoic acid (PFOA) exposure can impact heart development, though the specific mechanisms remain elusive. The aryl hydrocarbon receptor (AHR) is a critical environmental sensor capable of inducing oxidative stress and cell apoptosis. Objective To explore the role of AHR in the cardiac developmental toxicity of PFOA by using zebrafish embryo as an in vivo model. Methods Zebrafish embryos at 2 h post-fertilization (2 hpf) were exposed to dimethyl sulfoxide (DMSO) control, 1, 10, 100, and 1000 μg·L⁻¹ of PFOA. The AHR inhibitor CH223191 (CH) was added to the high-concentration group (1000 μg·L⁻¹) to form an intervention group. Under a dissecting microscope, the survival rate, mortality rate, heart rate, and heart malformation rate of 72 hpf zebrafish larvae were assessed. The activity of AHR was measured using 7-ethoxyresorufin-O-dealkylation (EROD) staining. The levels of intracellular and mitochondrial ROS in the heart of zebrafish larvae were assessed using dichloro-dihydro-fluorescein diacetate and MitoSOX™ Red, respectively. Apoptosis was examined using acridine orange (AO) staining and Immunofluorescence method. Total RNA was extracted from dissected hearts, and mRNA expression levels of oxidative stress-related genes (sod2, cat) and apoptosis-related gene (p53) were analyzed using quantitative PCR. Results Compared to the DMSO control group, PFOA at even the lowest concentration (1 μg·L⁻¹) increased heart malformation rate (P＜0.05) and reduced heart rate (P＜0.01) in the zebrafish larvae at 72 hpf, and the results showed a clear concentration-response relationship. Adding the AHR inhibitor CH significantly decreased heart malformation rate and restored heart rate to the control group level. The EROD results showed that PFOA at 1000 μg·L⁻¹ increased AHR activity (P＜0.001). Further studies revealed that PFOA caused a dose-dependent increase in intracellular ROS (P<0.001) and an increase in mitochondrial ROS (P<0.001) in the heart region of zebrafish larvae. A high dose of PFOA (1000 μg·L⁻¹) also induced elevated mRNA expression levels of oxidative stress-related genes sod2 and cat (P<0.05). Addition of the AHR inhibitor CH effectively antagonized PFOA-induced (1000 μg·L⁻¹) oxidative stress in the heart of zebrafish larvae (P<0.001). In addition, PFOA exposure induced a concentration-dependent increase in apoptotic bodies in the heart of zebrafish larvae (P<0.05). Moreover, PFOA at 1000 μg·L⁻¹ caused an increase in the cleaved-caspase 3 immunofluorescence signal (P<0.05) as well as overexpression of the apoptosis-associated gene p53 (P<0.001).which were attenuated by the CH supplementation. Conclusion PFOA triggers oxidative stress and apoptosis via AHR activation in the heart of zebrafish larvae, resulting in cardiac defects.

Objective To investigate the predictive value of preoperative frailty for pulmonary com-plications(PPCs)after cardiac surgery in elderly patients.Methods A total of 162 elderly patients,109 males and 53 females,aged 65-83 years,BMI 18-36 kg/m2,ASA physical status Ⅱ-Ⅳ,underwent elec-tive open heart surgery from July 2022 to January 2023 were collected.The patients were divided into two groups according to the occurrence of PPCs:the PPCs group(n=57)and the non-PPCs group(n=105).General information,smoking history,alcohol consumption history,EuroSCORE Ⅱ,frailty,chronic comorbidities(hypertension,diabetes mellitus,myocardial infarction,pulmonary hypertension,chronic ob-structive pulmonary disease,sleep apnea syndrome,etc.),Hb,creatinine,albumin,pulmonary function indices,left ventricular ejection fraction,type of surgery,duration of surgery,aortic clamping time,and cardiopulmonary bypass time were collected.Factors with P＜0.2 and clinically significant in the univariate regression analysis were included in the multivariate logistic regression analysis,and the predictive efficacy of the Fried frailty scale and EuroSCORE Ⅱ for PPCs were compared by the area under the ROC curve(AUC).Results PPCs occurred in 57 patients(35.2％).Multifactorial Logistic regression analysis showed that frailty(OR=3.14,95％CI 1.05-9.37,P＜0.05)and EuroSCORE Ⅱ(OR=2.16,95％CI 1.01-4.60,P＜0.05)were risk factors for the development of PPCs.The predictive power of Fried frailty scale(AUC=0.76,95％CI 0.68-0.82)was significantly higher than that of EuroSCORE Ⅱ(AUC=0.65,95％CI 0.57-0.72)(P＜0.05).Conclusion Preoperative frailty is the independent risk factors for pulmonary complications after cardiac surgery in elderly patients,and the Fried frailty scale has a better predictive efficacy compared to EuroSCORE Ⅱ,a traditional risk predictor.

Objective To investigate the relationship between insomnia,gastrointestinal symptoms,and glycosylated hemoglobin(HbA1c)levels in individuals diagnosed with type 2 diabetes mellitus(T2DM),as well as the related influencing factors.Methods A total of 910 T2DM patients treated in our multicenter from January 2022 to December 2022 were enrolled in this study.General information(gender,age,smoking and drinking history,exercise,course of disease,treatment and complications),HbA1c,Athens Insomnia Scale(AIS)scores and Gastrointestinal Symptoms Rating Scale(GSRS)scores of patients were collected.The differences of sleep and gastrointestinal symptoms between groups were analyzed,and the correlation between the differences and HbA1c was analyzed.Furthermore,the risk factors for non-standard HbA1c were analyzed.Results The AIS score and GSRS score in the HbA1c control group were less than those in the non-standard group(P＜0.01).Insomnia was reported by 37.0％of T2DM patients,and the HbA1c level in the insomnia group was significantly higher than that in the non-insomnia group(10.00％±2.38％vs.8.26％±1.73％,P＜0.01).Gastrointestinal symptoms were present in 57.5％of T2DM patients,and the HbA1c levels in the group with gastrointestinal symptoms were significantly higher than those in the group without gastrointestinal symptoms(9.26％±2.23％vs.8.43％±1.98％,P＜0.01).Furthermore,26.3％of T2DM patients experienced both insomnia and gastrointestinal symptoms.Remarkably,the HbA1c levels in the group with both insomnia and gastrointestinal symptoms were significantly higher than those in the group without either condition(10.18％±2.44％vs.8.45％±1.86％,P＜0.01).Correlation analysis demonstrated a significant association between sleep quality,gastrointestinal function,and HbA1c levels(P＜0.01).The logistic regression analysis result revealed that age,GSRS score,AIS score,and the presence of insomnia combined with gastrointestinal symptoms were independent risk factors for predicting HbA1c≥6.5％(P＜0.01).Having both insomnia and gastrointestinal symptoms concurrently was the strongest risk factor for substandard HbA1c control,and the risk of blood sugar control may increase about 5 times when both appear together.Conclusion Insomnia and gastrointestinal symptoms are common comorbidities in T2DM patients,showing a cross-interfering relationship,and they appear together with poor blood sugar control,interact causally,and amplify each other.

BACKGROUND:Bone relies on the close connection between blood vessels and bone cells to maintain its integrity.Bones are in a physiologically hypoxic environment.Therefore,the study of angiogenesis and osteogenesis in hypoxic environment is closer to the microenvironment in vivo. OBJECTIVE:To explore the influence of Wenshen Tongluo Zhitong(WSTLZT)formula on H-type bone microvascular endothelial cell/bone marrow mesenchymal stem cell co-culture system in hypoxia environment and its related mechanism. METHODS:Enzyme digestion method and flow sorting technique were used to isolate and identify H-type bone microvascular endothelial cells.Mouse bone marrow mesenchymal stem cells were isolated and obtained by bone marrow adhesion method.H-type bone microvascular endothelial cell/bone marrow mesenchymal stem cell hypoxic co-culture system was established using Transwell chamber and anoxic culture workstation.WSTLZT formula powder was used to intervene in each group at a mass concentration of 50 and 100 μg/mL.The angiogenic function of H-type bone microvascular endothelial cells in the co-culture system was evaluated by scratch migration test and tube formation test.The osteogenic differentiation ability of bone marrow mesenchymal stem cells in the co-cultured system was evaluated by alkaline phosphatase staining and alizarin red staining.The protein and mRNA expression changes of PDGF/PI3K/AKT signal axis related molecules in H-type bone microvascular endothelial cells in the co-cultured system were detected by Western Blotting and q-PCR,respectively. RESULTS AND CONCLUSION:(1)Compared with the normal oxygen group,the scratch mobility and new blood vessel length of H-type bone microvascular endothelial cells were significantly higher(P<0.05);the osteogenic differentiation capacity of bone marrow mesenchymal stem cells was higher(P<0.05);the expression of PDGF/PI3K/AKT axis-related molecular protein and mRNA increased(P<0.05)in the hypoxia group.(2)Compared with the hypoxia group,scratch mobility and new blood vessel length were significantly increased in the H-type bone microvascular endothelial cells(P<0.05);bone marrow mesenchymal stem cells had stronger osteogenic function(P<0.05);the expression of PDGF/PI3K/AKT axis-related molecular proteins and mRNA further increased(P<0.05)after treatment with different dose concentrations of WSTLZT formula.These findings conclude that H-type angiogenesis and osteogenesis under hypoxia may be related to the PDGF/PI3K/AKT signaling axis,and WSTLZT formula may promote H-type vasculo-dependent bone formation by activating the PDGF/PI3K/AKT signaling axis,thereby preventing and treating osteoporosis.

OBJECTIVE To study the intervention effects of Wenshen Tongluo Zhitong Recipe(WTZR)on macrophage senes-cence and senile osteoporosis.METHODS The senescent macrophage model was established using hydrogen peroxide(H2 O2)and subsequently divided into four groups:control,model,low-dose drug-treated serum,and high-dose drug-treated serum.β-galactosidase staining,Western blot and qPCR were employed to evaluate the mRNA expression of senescence markers p21 and p53.ROS staining and JC-1 staining were applied to assess mitochondrial function in macrophages.The mRNA levels of Interleukin(IL)-6,IL-10,CD206,and inducible nitric oxide synthase(iNOS)were determined by qPCR analysis.Immunofluorescence was used to evaluate argi-nase(ARG1)and iNOS protein expressions for assessing the impact of drug-containing serum on macrophage polarization.qPCR analy-sis was conducted to measure osteocalcin(OCN),collagen type Ⅰ alpha 1(Col1a1),runt-related transcription factor 2(Runx2)mRNA levels as osteoblast-related markers;ALP staining along with alizarin red staining were performed to evaluate the effect of macrophage conditioned medium treated with drug-containing serum on bone marrow mesenchymal stromal cell(BMSC)osteogenic differentiation.C57BL/6J mice were randomly allocated into four groups:control group,model group,WTZR low-dose group,and WTZR high-dose group.The senile osteoporosis(SOP)mouse model was established by D-galactose.Micro-CT scanning analyzed fe-mur microstructure while HE staining detected pathological changes in femur bone tissue samples collected from each experimental con-dition.Furthermore,Western blot was used to detect the senescence-related molecules p21 and p53 and the osteogenesis-related mark-ers OCN and Runx2,qPCR analysis measured tibial expression levels of senescence-related molecules(p21,p53)as well as macro-phage polarisation-related molecules(IL-6,iNOS,CD206,and IL-10)to assess the effect of this compound on a mouse model simula-ting SOP.RESULTS Following intervention with serum containing WTZR,there was a significant decrease in the number of senes-cent positive cells compared to the model group.Additionally,there was a notable decrease in p21 and p53 mRNA and proteins expres-sion(P＜0.05,P＜0.01,P＜0.001).Furthermore,drug-containing WTZR effectively inhibited ROS production induced by H2 O2 and mitigated mitochondrial membrane potential reduction in macrophages(P＜0.05,P＜0.001).Treatment with drug-containing WTZR re-sulted in down-regulated mRNA expression of M1-related gene iNOS(P＜0.05)while up-regulating mRNA expression of M2-related genes CD163 and CD206(P＜0.05).The drug-containing WTZR significantly reduced fluorescence intensity for iNOS(P＜0.01)while increasing ARG1(P＜0.05)fluorescence intensity.Moreover,conditioned medium from macrophages treated with drug-containing ser-um increased ALP positive cell count(P＜0.01,P＜0.001),alizarin red positive area(P＜0.05),as well as Col1a1,Runx2 and OCN mRNA expression levels(P＜0.05,P＜0.01).The Tb.N,BMD,and BV/TV were significantly higher in the WTZR group compared to the model group(P＜0.05,P＜0.01,P＜0.001);meanwhile,Tb.Sp was notably lower than that observed in the model group(P＜0.05,P＜0.01);bone trabeculae were significantly improved,increased in number and widened.Additionally,the compound could significantly inhibit the D-galactose induced up-regulation of tibial senescence-related genes and proteins p21 and p53(P＜0.05,P＜0.001,P＜0.0001),promote the expression of osteogenesis-related markers OCN and Runx2 protein(P＜0.01,P＜0.0001),promote the down-regulation of M1 related genes IL-6 and iNOS(P＜0.05),and promote the expression of M2 related genes IL-10 and CD206(P＜0.05,P＜0.01).CONCLUSION Wenshen Tongluo Zhitong Recipe may play an anti-osteoporosis effect by inhibiting macrophage senescence and promoting the osteogenic differentiation of BMSC.