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.

The doctor-patient relationship is a set of social relationships based on shared interests， mutual trust， and emotional bonds， to relieve illnesses and promote health. However， the doctor-patient relationship often falls into tensions and conflicts. How to build a trusting and harmonious doctor-patient destiny community has become one of the most important issues of concern to the whole society. Based on the biopsychosocial concept of disease， the emotion management technique （EMT） emphasizes that doctors take the patient’s emotion as a clue in clinical diagnosis and treatment， regard emotions as one of the important indicators for disease diagnosis， understand the emotional events behind the disease， and provide patients with appropriate empathy and emotional management， so as to provide clinical methods for managing diseases and building a trusting and harmonious doctor-patient relationship.

Objective:To evaluate the relationship between the mechanism of ω-3 polyunsaturated fatty acids (ω-3 PUFAs) in preventing brain neurotoxicity caused by multiple sevoflurane anesthesia and peroxisome proliferator-activated receptor gamma (PPARγ)/peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) signaling pathway in neonatal mice.Methods:This study was performed in 2 parts. Part Ⅰ Using a random number table method, 6 C57BL/6 pregnant mice were assigned to receive a standard diet, 3 pregnant mice were assigned to receive a diet supplemented with fish oil from day 2 of gestation to day 14 after parturition (ω-3 PUFAs 300 mg were added to every 20 g of conventional diet). Healthy C57BL/6 mice of both sexes, aged 6 days, weighing 3-5 g, were selected after parturition. Seventeen neonatal pups from 3 pregnant mice that were fed a conventional diet were assigned to control group (C group), 17 neonatal pups from 3 pregnant mice that were fed a conventional diet were assigned to sevoflurane group (S group), and 17 neonatal pups from pregnant mice that were fed a diet supplemented with fish oil were assigned to ω-3 PUFAs plus sevoflurane group (PS1 group) using the random number table method. Part Ⅱ Four C57BL/6 pregnant mice were assigned to receive a diet supplemented with fish oil from day 2 of gestation to day 14 after parturition. After parturition, 12 neonatal pups from 2 pregnant mice that were fed a diet supplemented with fish oil were assigned to ω-3 PUFAs plus sevoflurane group (PS2 group), and 12 neonatal pups from 2 pregnant mice that were fed a diet supplemented with fish oil were assigned to ω-3 PUFAs plus PPARγ inhibitor GW9662 plus sevoflurane group (PGS group) using a random number table method. GW9662 (2 mg/kg) was intraperitoneally injected at 30 min before exposure to sevoflurane in PGS group. All offspring mice were breastfed until 21 days of age, after which they were housed separately from the mother and allowed ad libitum access to a conventional diet. S, PS1, PS2 and PGS groups inhaled 3% sevoflurane in 40% oxygen for 2 h daily on postnatal days 6, 7 and 8. C group inhaled only 40% oxygen instead. Y maze test was performed on days 33 after birth. The rotarod test was performed on day 35 after birth. After the behavioral testing, the expression of PPARγ, PGC1α, mitofusin-1 (MFN1), MFN2, dynamin-related protein 1 (DRP1), interleukin-6 (IL-6), interleukin-1 β (IL-1 β), and tumor necrosis factor-α (TNF-α) was detected by Western blot, the ultrastructure of mitochondria in hippocampal neurons was observed with a transmission electron microscope, and the mitochondrial membrane potential (MMP), level of reactive oxygen species (ROS) and content of ATP were determined.Results:Part Ⅰ Compared with C group, the time of stay at the new-arm and time spent on the rotarod were significantly shortened, the percentage of movement distance in the new-arm was decreased, the expression of PPARγ, PGC1α, MFN1 and MFN2 in the hippocampus was down-regulated, the expression of DRP1, IL-6, IL-1β and TNF-α in the hippocampus was up-regulated, the MMP and content of ATP were decreased, and the level of ROS was increased in S group ( P<0.05). Compared with S group, the time of stay at the new-arm and time spent on the rotarod were significantly prolonged, the percentage of movement distance in the new-arm was increased, the expression of PPARγ, PGC1α, MFN1 and MFN2 in the hippocampus was up-regulated, the expression of DRP1, IL-6, IL-1β and TNF-α in the hippocampus was down-regulated, the MMP and content of ATP were increased, and the level of ROS was decreased in PS1 group ( P<0.05). Part Ⅱ Compared with PS2 group, the time of stay at the new-arm and time spent on the rotarod were significantly shortened, the percentage of movement distance in the new-arm was decreased, the MMP and content of ATP were decreased, the level of ROS was increased, and the expression of IL-6, IL-1β and TNF-α was up-regulated ( P<0.05). Conclusions:The mechanism by which ω-3 PUFAs prevent brain neurotoxicity caused by multiple sevoflurane anesthesia is related to the activation of the PPARγ/PGC1α signaling pathway and alleviation of mitochondrial dysfunction and neuroinflammation in neonatal mice.

Objective:To evaluate the role of docosahexaenoic acid (DHA) in long-term cognitive impairment after multiple sevoflurane anesthesia in newborn mice.Methods:Clean-grade healthy male C57BL/6 mice, aged 6 days, were used in this study. Ten mice were divided into 2 groups ( n=5 each) by the random number table method: control group (group C) and sevoflurane group (group S). The animals inhaled 3% sevoflurane for 2 h at 6, 7 and 8 days after birth. The DHA content was detected by ultra-high performance liquid chromatography-mass spectrometry at 9 days of age. Fifty-two mice were selected and divided into 4 groups ( n=13 each) by a random number table method: control+ normal saline group (group C+ S), sevoflurane anesthesia + normal saline group (group S+ S), control+ DHA group (group C+ D), and sevoflurane anesthesia+ DHA group (group S+ D). The sevoflurane anesthesia method was the same as the one mentioned above. DHA 50 mg/kg was administered by intragastric gavage from postnatal days 6-19 (at 6, 7 and 8 days after birth, 2 h before anesthesia) in C+ D and S+ D groups. The equal volume of normal saline was given instead in C+ S group and S+ S group. The novel object recognition test was conducted at 37 days of age, and the Morris water maze test was performed at 42 days of age. The corpus callosum and hippocampal tissues were isolated at 47 days of age for examination of the ultrastructure of myelin (with a transmission electron microscope) and for determination of the expression of myelin basic protein (MBP) in hippocampal tissues (by Western blot). The G-ratio was calculated. Results:Compared with group C, the content of DHA in hippocampal tissues was significantly decreased in group S ( P<0.05). Compared with group C+ S, the discrimination index was significantly decreased, the percentage of duration of staying at the target platform quadrant and the number of crossing the original platform were decreased, the expression of MBP was down-regulated, and the G-ratio in the original platform and hippocampus was increased in S+ S group ( P<0.05). Compared with group S+ S, the discrimination index was significantly increased, the percentage of duration of staying at the target platform quadrant and the number of crossing the original platform were increased, the expression of MBP was up-regulated, and the G-ratio in the original platform and hippocampus was decreased in S+ D group ( P<0.05). Conclusions:The mechanism of long-term cognitive impairment following multiple sevoflurane anesthesia may be related to a decrease in the content of DHA, which subsequently leads to myelin structural damage in neonatal mice.

Objective:To evaluate the effect of multiple sevoflurane anesthesia on the metabolism of long-chain fatty acids in the hippocampus of newborn mice and the role of peroxisome proliferator-activated receptor-beta (PPARβ).Methods:Clean-grade healthy male C57BL/6 mice, aged 6 days, weighing 3-5 g, were divided into 2 groups ( n=8 each) by a random number table method: control group (group C) and multiple sevoflurane anesthesia group (group S). This study was performed in 2 parts. PartⅠ Sixteen newborn mice were divided into 2 groups ( n=8 each) using a random number table: control group (C group) and multiple sevoflurane anesthesia group (S group). Anesthesia was performed with sevoflurane on postnatal days 6, 7 and 8. The hippocampus was obtained at postnatal day 9 for determination of the content of long-chain fatty acids (by ultra-high performance liquid mass spectrometry), expression of PPARβ (by Western blot), and expression of stearoyl-CoA desaturase-2 (Scd2) and fatty acid desaturase 2 (Fads2) mRNA (using quantitative real-time polymerase chain reaction). Part Ⅱ Twenty-one newborn mice were divided into 3 groups ( n=7 each) using a random number table: control+ normal saline group (group C+ S), sevoflurane + normal saline group (group S+ S), and sevoflurane+ PPARβ specific agonist KD3010 group (group S+ K). Anesthesia was carried out with sevoflurane on postnatal days 6, 7 and 8. KD3010 25 mg/kg was intraperitoneally injected once a day from postnatal day 6 to 13 in S+ K group. The novel object recognition test was performed on postnatal day 37, and the Morris water maze test was performed on postnatal day 42. The hippocampal tissues were obtained on postnatal day 47 for detection of the expression of Scd2 mRNA and Fads2 mRNA by fluorescent quantitative real-time polymerase chain reaction. Anesthesia was carried out with sevoflurane as follows: Mice were exposed to 3% sevoflurane in 40% oxygen-60% nitrogen in an induction chamber for 2 h at a flow rate of 1 L/min. Results:PartⅠ Compared with group C, the total content of long-chain fatty acids, contents of saturated fatty acids, monounsaturated fatty acids and polyunsaturated fatty acids were significantly decreased, the percentage of saturated fatty acids was increased, the percentage of monounsaturated fatty acids and polyunsaturated fatty acids was decreased, the expression of Scd2 mRNA and Fads2 mRNA was down-regulated, and the expression of PPARβ was down-regulated in group S ( P<0.05). Part Ⅱ Compared with group C+ S, the discrimination index in the novel object recognition test and percentage of time spent in the target quadrant were significantly decreased, the number of crossing the original platform was reduced, and the expression of Scd2 and Fads2 mRNA was down-regulated in group S+ S ( P<0.05). Compared with group S+ S, the discrimination index in the novel object recognition test and percentage of time spent in the target quadrant were significantly increased, the number of crossing the original platform was increased, and the expression of Scd2 and Fads2 mRNA was up-regulated in group S+ K ( P<0.05). Conclusions:Multiple anesthesia with sevoflurane can lead to the disorder of long-chain fatty acid metabolism in the hippocampus of neonatal mice, resulting in long-term cognitive dysfunction. The mechanism may be related to inhibiting the activity of hippocampal PPARβ signaling pathway.

Objective To investigate the expression levels of serum hemoglobin β(HBB)in hepatocellular carcinoma(HCC)patients and its correlation with topoisomeraseⅡα(TOP2A)gene.Methods A total of 48 HCC patients visited the Second Affiliated Hospital of Nantong University from August 2023 to September 2024 were selected as the HCC group,and 32 healthy individuals who underwent physical examination during the same period were selected as the healthy control group.Their blood samples were collected,and the ex-pression levels of serum HBB and TOP2A genes were detected by real-time fluorescence quantitative polymerase chain reaction(qRT-PCR).Then,the relationship between the expression of HBB gene and clinicopathological parameters of the patients was ana-lyzed.The Kaplan-Meier Plotter database was used to evaluate the relationship between the expression of HBB gene and the prognosis of HCC.The diagnostic value of the expressions of serum HBB and TOP2A genes for HCC was assessed by the receiver operating charac-teristic(ROC)curve.Spearman rank correlation analysis was used to evaluate the correlation between the expressions of serum HBB and TOP2A genes in HCC patients.The regulatory effect of HBB gene on TOP2A gene was verified by the cell experiment.Results The expression levels of serum HBB gene in HCC patients(0.097[0.055,0.155])were significantly lower than that in healthy controls(1.029[0.625,1.434],U=19,P＜0.001).The expression levels of serum TOP2A gene in HCC patients(1.810[0.825,3.623])were significantly higher than that in healthy controls(1.047[0.604,1.364],U=495,P=0.007).The expression level of serum HBB gene in HCC patients was significantly negatively correlated with that of TOP2A gene(ρ=-0.384,P=0.007).The analysis results of clinicopathological parameters showed that the expression level of HBB gene was only related to tumor size(χ2=4.090,P＜0.05).The Kaplan-Meier survival analysis showed that the 5-year overall survival rate of the patients with low expression of HBB gene was signifi-cantly lower than that with high expression(HR=0.680,95%CI:0.470-0.970,P＜0.05).The analysis of the ROC curve showed that the area under the ROC curve(AUCROC)of HBB gene in diagnosing HCC was 0.987(95%CI:0.963-1.000).When the cut-off value was 0.228,its sensitivity was 100%and specificity was 97%.The AUCROC of TOP2A gene for the diagnosis of HCC was 0.677(95%CI:0.559-0.797).When the cut-off value was 1.285,its sensitivity was 65%and specificity was 75%.The combined detection of HBB and TOP2A genes for the diagnosis of HCC had an AUCROC of 0.988(95%CI:0.965-1.000).When the cut-off value was 0.657,its sensitivity was 100%and specificity was 97%.The cell experiment results showed that the overexpression of HBB gene could inhibit the expression of TOP2A gene,while the knockout of HBB gene had the opposite effect.Conclusion HBB gene is lowly expressed in the serum of HCC patients and is significantly negatively correlated with the expression of TOP2A gene.

Objective:To evaluate the relationship between preoperative gut microbiota and post-operative ventilator-associated pneumonia (VAP) in patients undergoing coronary artery bypass grafting.Methods:This was a secondary analysis of a previous research project study. Patients who received invasive mechanical ventilation treatment after elective off-pump coronary artery bypass grafting at the Second Hospital of Hebei Medical University from April to September 2023 were selected and divided into VAP group and non-VAP group based on whether VAP occurred after surgery. Fecal samples were collected from patients before surgery, and 16S rRNA gene sequencing was used to analyze the characteristics of preoperative gut microbiota in the two groups. The differences in the diversity of gut microbiota between the two groups were compared. The linear discriminant analysis was used to identify the gut microbiota with significant differences between groups (differential bacteria), and logistic regression analysis was conducted to assess the association between differential bacteria and VAP. Receiver operating characteristic curves were drawn to analyze the predictive value of the differential bacteria for VAP.Results:A total of 79 patients were finally included, with 25 in VAP group and 54 in non-VAP group. The Beta diversity analysis showed statistically significant differences between VAP group and non-VAP group (pseudo- F=2.00, P=0.002). The linear discriminant analysis indicated that Bifidobacterium, Blautia and Megamonas were enriched in non-VAP group, while Klebsiella was enriched in VAP group. Multivariate logistic regression analysis showed that the relative abundance of Bifidobacterium was a protective factor for postoperative VAP ( OR=0.32, 95% confidence interval [ CI] 0.15-0.71, P=0.005), and the relative abundance of Klebsiella was a risk factor for postoperative VAP ( OR=2.49, 95% CI 1.143-5.43, P=0.022). The area under the receiver operating characteristic curve of the relative abundance of Bifidobacterium for predicting VAP was 0.80 (95% CI 0.69-0.90, P<0.001) and of the relative abundance of Klebsiella was 0.70 (95% CI 0.57-0.83, P=0.005). Conclusions:Bifidobacterium is a protective factor, while Klebsiella is a risk factor for postoperative VAP in patients undergoing coronary artery bypass grafting, and the relative abundance of both bacteria has a certain predictive value for VAP.

Spleen-stomach damp-heat syndrome is currently the most prevalent TCM pattern in patients with chronic atrophic gastritis (CAG), with internal damp-heat accumulation regarded as a key factor contributing to its prolonged and refractory course. This syndrome represents a critical stage in the progressive pathogenesis of CAG, characterized by a deepening pathological evolution. Modern TCM practitioners generally agree that its core pathogenesis lies in "deficiency in root and excess in superficiality, with internal damp-heat retention", and emphasize a treatment strategy that combines eliminating pathogenic factors and reinforcing the body's healthy qi through dynamic syndrome differentiation. Chinese materia medica used in treating CAG with spleen-stomach damp-heat syndrome can effectively relieve clinical symptoms, improve the internal damp-heat environment, mitigate gastric mucosal atrophy and intestinal metaplasia, and delay the inflammation-to-cancer transformation. Its mechanisms may involve eradication of Helicobacter pylori, repair of gastric mucosal injury, regulation of immune inflammatory response and other aspects, which has the advantages of multi-channel and multi-target.

Senile osteoporosis is a growing public health challenge with significant impacts on the daily life of the elderly population as a result of its hidden nature,high prevalence,and high risk of disability.Suitable animal models that simulate senile osteoporosis are crucial for understanding its pathological mechanism and to facilitate the development of anti-osteoporosis drugs and identify new therapeutic targets.This review considers the most commonly used method for creating animal models of senile osteoporosis,analyzes their advantages and limitations,and discusses research progress in animal models in terms of evaluation indicators,to provide references for research using animal models of senile osteoporosis.

Nonalcoholic steatohepatitis （NASH） is a chronic liver disease with the main pathological features of hepatic steatosis， inflammatory cell infiltration， and interstitial fibroplasia， and it is an important risk factor for liver fibrosis， liver cirrhosis， and hepatocellular carcinoma. NOD-like receptor protein 3 （NLRP3） inflammasome is the core of innate immunity， and the abnormal activation of NLRP3 inflammasome is closely associated with the development and progression of NASH， which involves multiple links such as inflammatory response and oxidative stress. A large number of studies have shown that the active ingredients of traditional Chinese medicine （TCM） and TCM compound prescriptions can improve oxidative stress， regulate lipid metabolism， and alleviate liver inflammation by regulating NLRP3 inflammasome. TCM treatment applied in clinical practice has achieved a good therapeutic effect， while inflammasome is one of the key pathways or targets for TCM in improving NASH. This article reviews the mechanism of action of NLRP3 inflammasome in NASH and the research advances in TCM intervention of NLRP3 inflammasome， in order to provide ideas for the clinical TCM treatment of NASH， as well as reference targets and research directions for the research and development of new TCM drugs.