Objective:To explore the effect of hairy and enhancer of split related protein 2 (Hey2) on osteoclast differentiation through the activation of nuclear factor of activated T cells cytoplasmic 1 (NFATc1).Methods:RAW264.7 cells were induced with receptor activator of NF-κB ligand (RANKL) to differentiate into osteoclasts. Experimental groups were divided by different concentrations of RANKL (0, 10, 20, 50 μg/L) and different processing time (0, 3, 5, 7 days). Hey2 overexpression experiment was grouped as follows: blank control group, RANKL group, empty plasmid vector control group (Hey2-NC+RANKL), Hey2 overexpression group (Hey2-OE+RANKL); similarly, groups in Hey2 knockdown experiment were as follows: blank control group, RANKL group, negative control group (si-NC+RANKL), Hey2 knockdown group (si-Hey2+RANKL). Chromatin immunoprecipitation experiment groups were divided as non-specific IgG control group (IgG control group), non-specific IgG group (IgG RANKL group), Hey2-specific antibody control group (anti-Hey2 control group), Hey2-specific antibody group (anti-Hey2-RANKL group). For the different RANKL concentration groups and different induction time groups, real-time fluorescent quantitative PCR (RT-qPCR) was used to detect the mRNA expressions of nuclear factor of NFATc1, cathepsin K (CTSK), and cellular feline osteosarcoma oncogene (c-Fos) and tartrate-resistant acid phosphatase (TRAP) staining was used to assess the formation of multinucleated osteoclasts. After Hey2 overexpression or knockdown, RT-qPCR and Western blotting were used to detect the gene and protein expressions of NFATc1, c-Fos, and CTSK. TRAP staining was used to evaluate the formation of multinucleated osteoclasts. Bioinformatics prediction (NCBI, JASPAR) and chromatin immunoprecipitation (ChIP) assay were used to validate the binding of Hey2 to the NFATc1 promoter region.Results:During the osteoclastic differentiation of RAW 264.7 cells induced by RANKL, the expression of Hey2 could be detected, and the expression level of Hey2 decreased with the increase of RANKL concentration and induction time. In the 50 μg/L RANKL group, the expression levels of Hey2 gene (0.18±0.00) and protein (0.22±0.02) were significantly lower than those in the control group (1.00±0.00, 0.52±0.01) ( t=41.67, 12.88; both P<0.001). In the 50 μg/L RANKL group inducted for 5 days, the expression levels of Hey2 gene (0.27±0.02) and protein (0.79±0.01) were significantly lower than those in the control group (1.00±0.00, 1.15±0.02) ( t=11.47, 108.60; both P<0.001). Hey2 overexpression significantly reduced the gene and protein expressions of NFATc1, c-Fos, and CTSK, as well as the production of TRAP-positive cells (all P<0.05). Hey2 knockdown significantly increased the gene and protein expressions of NFATc1, c-Fos, and CTSK, as well as the production of TRAP-positive cells (all P<0.05). After inducing RAW264.7 cells with 50 μg/L RANKL for 1 day, ChIP results showed that among the two sample groups treated with Hey2 antibody, the detection level of the NFATc1 promoter region (-400 to -200 bp) in the anti-Hey2-RANKL group (18.06±0.06) was significantly higher than that in the anti-Hey2 control group (13.37±0.36) ( t=12.56, P<0.001). Conclusions:Hey2 can bind to the downstream target gene NFATc1 at -400 to -200 bp region of the promoter. As a transcriptional repressor, Hey2 inhibits osteoclast differentiation.

Objective:To explore the effect of hairy and enhancer of split related protein 2 (Hey2) on osteoclast differentiation through the activation of nuclear factor of activated T cells cytoplasmic 1 (NFATc1).Methods:RAW264.7 cells were induced with receptor activator of NF-κB ligand (RANKL) to differentiate into osteoclasts. Experimental groups were divided by different concentrations of RANKL (0, 10, 20, 50 μg/L) and different processing time (0, 3, 5, 7 days). Hey2 overexpression experiment was grouped as follows: blank control group, RANKL group, empty plasmid vector control group (Hey2-NC+RANKL), Hey2 overexpression group (Hey2-OE+RANKL); similarly, groups in Hey2 knockdown experiment were as follows: blank control group, RANKL group, negative control group (si-NC+RANKL), Hey2 knockdown group (si-Hey2+RANKL). Chromatin immunoprecipitation experiment groups were divided as non-specific IgG control group (IgG control group), non-specific IgG group (IgG RANKL group), Hey2-specific antibody control group (anti-Hey2 control group), Hey2-specific antibody group (anti-Hey2-RANKL group). For the different RANKL concentration groups and different induction time groups, real-time fluorescent quantitative PCR (RT-qPCR) was used to detect the mRNA expressions of nuclear factor of NFATc1, cathepsin K (CTSK), and cellular feline osteosarcoma oncogene (c-Fos) and tartrate-resistant acid phosphatase (TRAP) staining was used to assess the formation of multinucleated osteoclasts. After Hey2 overexpression or knockdown, RT-qPCR and Western blotting were used to detect the gene and protein expressions of NFATc1, c-Fos, and CTSK. TRAP staining was used to evaluate the formation of multinucleated osteoclasts. Bioinformatics prediction (NCBI, JASPAR) and chromatin immunoprecipitation (ChIP) assay were used to validate the binding of Hey2 to the NFATc1 promoter region.Results:During the osteoclastic differentiation of RAW 264.7 cells induced by RANKL, the expression of Hey2 could be detected, and the expression level of Hey2 decreased with the increase of RANKL concentration and induction time. In the 50 μg/L RANKL group, the expression levels of Hey2 gene (0.18±0.00) and protein (0.22±0.02) were significantly lower than those in the control group (1.00±0.00, 0.52±0.01) ( t=41.67, 12.88; both P<0.001). In the 50 μg/L RANKL group inducted for 5 days, the expression levels of Hey2 gene (0.27±0.02) and protein (0.79±0.01) were significantly lower than those in the control group (1.00±0.00, 1.15±0.02) ( t=11.47, 108.60; both P<0.001). Hey2 overexpression significantly reduced the gene and protein expressions of NFATc1, c-Fos, and CTSK, as well as the production of TRAP-positive cells (all P<0.05). Hey2 knockdown significantly increased the gene and protein expressions of NFATc1, c-Fos, and CTSK, as well as the production of TRAP-positive cells (all P<0.05). After inducing RAW264.7 cells with 50 μg/L RANKL for 1 day, ChIP results showed that among the two sample groups treated with Hey2 antibody, the detection level of the NFATc1 promoter region (-400 to -200 bp) in the anti-Hey2-RANKL group (18.06±0.06) was significantly higher than that in the anti-Hey2 control group (13.37±0.36) ( t=12.56, P<0.001). Conclusions:Hey2 can bind to the downstream target gene NFATc1 at -400 to -200 bp region of the promoter. As a transcriptional repressor, Hey2 inhibits osteoclast differentiation.

Objective To explore the clinical value of bedside gastrointestinal ultrasound exacerbations in evaluating gastro-intestinal function in patients with chronic obstructive pulmonary disease(AECOPD).Methods A total of 128 patients with AECOPD hospitalized in the Department of Intensive Care and Respiratory Medicine of Lanzhou First People's Hospital from June 2019 to December 2021 were selected and divided into mild to moderate group(63 cases without invasive respiratory sup-port treatment)and severe group(65 cases with invasive respiratory support treatment)according to the severity of their condi-tions.Fifty-four healthy subjects were selected during the same period.The general data,gastric emptying time(GET),anstral contraction frequency(ACF),anstral contraction amplitude(ACA),anstral motility index(MI),small intestine diameter,colon diameter,intestinal peristalsis,intestinal mucosal thickness,and colon mucosal thickness were compared among these pa-tients.Logistic regression analysis was performed to determine whether these indicators were associated with gastrointestinal function in patients with AECOPD.ROC curve and Jorden index were used to define the diagnostic threshold of these indica-tors,so as to evaluate the value of gastrointestinal ultrasound in the diagnosis of gastrointestinal dysfunction in AECOPD pa-tients.Results There was no significant difference in the general data among the three groups(P＞0.05).Compared with the healthy group,the GET of AECOPD patient was significantly prolonged,ACF was significantly reduced,ACA was significantly reduced,MI was significantly decreased,small intestine diameter and colon diameter were significantly increased,intestinal peri-stalsis was significantly slowed down,and intestinal mucosal thickness and colon mucosal thickness were significantly reduced(P＜0.01).Compared with normal group and light-medium group,GET was significantly longer,ACF was significantly lower,ACA was significantly reduced,MI was significantly decreased,small intestine diameter and colon diameter were significantly in-creased,intestinal peristalsis was significantly slowed down,and intestinal mucosal thickness and colon mucosal thickness were significantly reduced(P＜0.01)in severe group.These indexes were also associated with the degree of AECOPD(P＜0.01).Lo-gistic regression analysis showed that GET,ACF,ACA,MI,small intestine diameter,colon diameter and intestinal peristalsis were significantly correlated with gastrointestinal function in AECOPD patients.ROC curve analysis was performed in the indi-cators related to gastrointestinal function in AECOPD patients,and the area under ROC curve of all of them was greater than 0.5(P＜0.05).The comprehensive evaluation of gastrointestinal function by these indexes had a good diagnostic value,with good sensitivity and specificity.Conclusion Gastrointestinal ultrasound can be used to evaluate gastrointestinal motility dys-function in patients with AECOPD.Gastrointestinal ultrasound measurements of GET,ACF,ACA,MI,small intestine diame-ter,colon diameter and intestinal peristalsis can be used for early detection of gastrointestinal dysfunction in patients with AE-COPD.

Cerebral microbleeds (CMBs) are a subclinical terminal microvascular disease in which the blood exudates or leaks out from the tiny blood vessels and the small lesions were formed by the deposition of hemosiderin in the brain tissue. The pathogenesis of cerebral microbleeds is different depending on the location, with lobar CMBs attributed to cerebral amyloid angiopathy (CAA), while cerebrovascular diseases caused by hypertension are an important cause of deep and subtentorium CMBs. The prevalence of CMBs in stroke patients is high, especially in patients with ischemic stroke treated with oral antiplatelet drugs, and long-term (>5 years) treatment may be related to CMBs and intracerebral hemorrhage (ICH) events. At the same time, a certain burden of microbleeds may cause risk of ICH in the future, but whether the bleeding risk of antiplatelet treatment overweighs the clinical benefit of antithrombotic therapy remains unclear. How to better instruct antiplatelet therapy in patients with ischemic stroke warrants further clinical investigations.

Objective:To investigate the clinical value of continuous increase of blood lactic acid (Lac) in prognosis evaluation of patients with sepsis.Methods:From January 2016 to December 2018, 84 patients with sepsis in the Department of Critical Medicine of Lanzhou First People's Hospital were retrospectively analyzed. According to the final outcome, the patients were divided into death group and survival group. The general condition, initial Lac, Lac at 6, 12, 18, 24 h, and the duration of Lac>2 mmol/L (T Lac>2) were compared between the two groups. Receiver operating characteristic curve (ROC) was used to analyze the sensitivity and specificity of gender, age, acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ), initial lac, 6, 12, 18, 24 h lac, T Lac>2 in evaluating the prognosis of patients with sepsis. At the same time, the relationship between these parameters and the prognosis of patients was analyzed by Cox regression analysis to evaluate the clinical value of the time of continuous increase of blood lactate in the prognosis of patients with sepsis. Results:There was no significant difference in age, gender, APACHE Ⅱ score and initial lac between the two groups ( P>0.05). The Lac of death group was higher than that of survival group at 6, 12, 18 and 24 h after treatment ( P<0.05), and T Lac>2 in death group was significantly longer than that in survival group ( P<0.05). Cox regression analysis showed that age, gender, APACHE Ⅱ score, initial Lac, 6, 12, 18, 24 h lac had no significant effect on the prognosis of sepsis patients, while T Lac>2 was significantly correlated with survival rate and death risk. The longer T Lac>2 was, the lower the survival rate and the higher the risk of death. At the same time, ROC curve analysis showed that gender, age, APACHE Ⅱ score and area under the curve (AUC) of initial lac showed that these indicators could not effectively evaluate the prognosis of patients ( P>0.05). The area under the curve of T Lac>2 was the largest, and the evaluation of prognosis was the best, followed by 24 h Lac and 18 h, 12 h and 6 h Lac. In addition, the sensitivity of T Lac>2, 24, 18, 12, 6 h for sepsis mortality risk assessment were 90.9%, 81.8%, 81.8%, 81.8%, 88.6%, and the specificity were 71.4%, 52.5%, 52.5%, 47.7% and 25.2%, respectively. Conclusions:Transient increase of lactic acid can not evaluate the prognosis of patients with sepsis, but the duration of lactic acid increase has a significant impact on the prognosis of patients with sepsis. The longer the increase of Lactic acid (T Lac>2) is, the lower the survival rate and the higher the risk of death. The sensitivity and specificity of lactic acid duration in evaluating the risk of death were higher than those of other parameters, and the prognostic efficacy was the best.