BACKGROUND:Overactive osteoclasts disrupt bone homeostasis and play a bad role in the pathological mechanisms of related skeletal diseases,such as osteoporosis,fragility fractures,and osteoarthritis.Studies have confirmed that ellagic acid and ellagtannin have the potential to inhibit osteoclast differentiation.As their natural metabolites,urolithin A has antioxidant,anti-inflammatory,anti-proliferative and anti-cancer effects,but its effect on osteoclast differentiation and its underlying molecular mechanisms remain unclear. OBJECTIVE:To explore the effect of urolithin A on osteoclast differentiation induced by receptor activator for nuclear factor-κB ligand and its mechanism. METHODS:Mouse mononuclear macrophage leukemia cells(RAW264.7)that grew stably were cultured in vitro.Toxicity of urolithin A(0,0.1,0.5,1.5,2.5 μmol/L)to RAW264.7 cells were detected by cytotoxic MTS assay to screen out the safe concentration.Different concentrations of urolithin A were used again to intervene with receptor activator for nuclear factor-κB ligand-induced differentiation of RAW264.7 cells in vitro.Then,tartrate-resistant acid phosphatase staining and F-actin ring and nucleus staining were performed to observe its effect on the formation and function of osteoclasts.Finally,the expressions of urolithin A on upstream and downstream genes and proteins in the MAPK signaling pathway were observed by western blot and RT-qPCR assays. RESULTS AND CONCLUSION:Urolithin A inhibited osteoclast differentiation and F-actin ring formation in a concentration-dependent manner and 2.5 μmol/L had the strongest inhibitory effect.Urolithin A inhibited the mRNA expression of Nfatc1,Ctsk,Mmp9 and Atp6v0d2 and the protein synthesis of Nfatc1 and Ctsk,related to osteoclast formation and bone resorption.Urolithin A inhibited the activity of osteoclasts by downregulating the phosphorylation of p38 protein to inhibit the mitogen-activated protein kinase signaling pathway.

Objective To investigate the clinical effects and pregnancy outcomes of using luteal phase long protocol and GnRH antagonist protocol in patients with polycystic ovary syndrome(PCOS)who have failed their first GnRH antagonist protocol therapy.Methods The clinical data of 163 PCOS patients who underwent IVF/ICSI-ET were retrieved.After the failure of their first GnRH antagonist protocol treatment,they were divided into two groups in the second controlled ovarian hyperstimulation(COH)cycle:Luteal phase long protocol group(n=95)and Gn-RH antagonist protocol group(n=68).A retrospective analysis and comparison of basic clinical data,clinical and laboratory indicators,and pregnancy outcomes between two groups were conducted.Results ① There was no sta-tistically significant difference in basic clinical indicators between two group except LH.② Compared the first and second cycle treatments of patients in the luteal phase long protocol group,the initiation dose of gonadotropin(Gn),total number of Gn days,total Gn usage,estradiol(E2)on the day of hCG injection,number of retrieved eggs,oocyte maturation rate,2PN fertilization rate,2PN cleavage rate,blastocyst formation rate,high-quality blas-tocyst formation rate,and moderate to severe OHSS rate were significantly higher than those in the first GnRH an-tagonist cycle(P＜0.05).The GnRH antagonist protocol group also showed similar improvements.③ The com-parison of the second COH cycle between two groups showed that the total number of Gn days,total Gn usage,and total Gn cost in the luteal phase long protocol group were significantly higher(P＜0.05),while the E2 and LH on the day of hCG injection,and the maturation rate of eggs were significantly lower than those in the GnRH antagonist protocol group(P＜0.05).However,there was no statistically significant difference in the number of retrieved eggs,2PN fertilization,2PN cleavage,blastocyst formation rate,high-quality blastocyst formation rate,and OHSS rate between the two groups;④ The comparison of fresh transplantation cycles for the second COH cycle between the two groups showed that the luteal phase long protocol fresh transplantation rate,implantation rate,clinical preg-nancy rate,and live birth rate were slightly higher than those of the GnRH antagonist protocol group,but the differ-ence was not statistically significant.Comparing the outcomes of pregnancy following the initial frozen-thawed em-bryo transfer(FET)between two groups,the biochemical pregnancy rate and clinical pregnancy rate of the GnRH antagonist protocol group were higher than those of the luteal phase long protocol group(P＜0.05).However,no significant statistical variations were found in implantation rate,live birth rate,neonatal gestational age,and birth weight.Conclusion For PCOS patients who fail the first GnRH antagonist protocol,an appropriate increase in the initiating dose and usage of Gn can achieve satisfactory pregnancy outcomes with both protocols.Compared with change to a luteal phase long protocol,reusing the GnRH antagonist protocol still maintains its long-standing advan-tages,such as shorter total Gn days,lower costs,and better patient compliance.

COVID-19 is caused by a novel coronavirus-severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), which has being spreading around the world, posing a serious threat to human health and lives. Neutralizing antibodies and small molecule inhibitors for virus replication cycle are the main antiviral treatment for novel coronavirus recommended in China. To further promote the rational use of antiviral therapy in clinical practice, the National Center for Infectious Diseases (Beijing Ditan Hospital Capital Medical University and the First Affiliated Hospital, Zhejiang University School of Medicine) invited experts in fields of infectious diseases, respiratory and intensive care to develop an Expert Consensus on Antiviral Therapy of COVID-19 based on the Diagnosis and Treatment Guideline for COVID-19 ( trial version 10) and experiences in the diagnosis and treatment of COVID-19 in China. The consensus is concise, practical and highly operable, hopefully it would improve the understanding of antiviral therapy for clinicians and provide suggestions for standardized medication in treatment of COVID-19.

Objective:To investigate the effect and molecular mechanism of tumor necrosis factor-α-inducible protein 8-like 2(TIPE2)on lipopolysaccharide(LPS)or interleukin-4(IL-4)-induced phenotypic switch of adipose tissue macrophages(ATM).Methods:The expression levels of TIPE2, inducible nitric oxide synthase(iNOS), monocyte chemoattractant protein 1(MCP-1), CD206, and arginase 1(Arg-1)in the visceral adipose tissue of obese mice, TIPE2-knockout(KO)mice, and control mice were detected by immunohistochemistry, Western blotting, and real-time PCR(RT-qPCR). Peritoneal macrophages isolated from KO and wild-type mice and RAW 264.7 mouse macrophage cell line were cultured, and then stimulated with LPS(100 ng/mL)or IL-4(20 ng/mL)for 6 hours. The expression levels of TIPE2, iNOS, MCP-1, CD206, and Arg-1 were detected by Western blotting and RT-qPCR.Results:Obese mice showed down-regulated TIPE2 expression, up-regulated pro-inflammatory markers iNOS and MCP-1 expressions, and down-regulated anti-inflammatory markers CD206 and Arg-1 expressions. LPS decreased the expression of TIPE2 in RAW 264.7 cells and peritoneal macrophages from mice, increased the expression of the classically activated macrophages(M1 phenotype)markers iNOS and MCP-1, and decreased the expression of the substituting activated macrophages(M2 phenotype)markers CD206 and Arg-1. IL-4 increased the expression of TIPE2 in RAW 264.7 cells and peritoneal macrophages, decreased the expression of iNOS and MCP-1, and increased the expression of CD206 and Arg-1. During the M1 polarization of macrophages, LPS increased toll-like receptor(TLR4)expression as well as nuclear transcription factor κBα suppressor protein(IκBα) and NF-κB phosphorylations in macrophages. Knockout of TIPE2 further increased the expression of the TLR4/IκBα/NF-κB signaling pathway and M1 macrophage markers, and further reduced the expression of the M2 macrophage markers.Conclusion:TIPE2 regulates ATM phenotypic transformation through inhibition of the TLR4/IκBα/NF-κB signaling pathway, which ameliorates adipose tissue inflammation in obese states.

Objective 　To evaluate the role of surgical resection on synchronous multiple pulmonary nodules identified difficultly in clinics. Methods 　The clinical data of 97 patients with synchronous multiple pulmonary nodules who received surgical resection between 2012 and 2019 in Hunan Cancer Hospital were retrospectively analyzed. There were 72 males and 25 females, aged 58.1卤9.0 years. Among these patients, there were 78 patients with ipsilateral and 19 patients with bilateral pulmonary nodules. Clinicopathological parameters between main nodules and secondary nodules were evaluated. Perioperative morbidity was also assessed. Results 　The operation was successfully completed on all patients for the ipsilateral and bilateral lesions. Totally, 71.1% of mian lesions was mostly removed by lobectomy, and the completion rate of video-assisted thoracoscopic surgery (VATS) was 69.1% (67/97); 80.4% of secondary lesions were mostly removed by wedge resection, and the completion rate of VATS was 71.1% (69/97). The incidence of grade 3 or higher complications after unilateral or bilateral surgery was 12.8% and 5.3%, respectively. Postoperative pathology confirmed that the main lesions were malignant in 65 patients (67.0%), mainly adenocarcinoma (63.1%), of which 43.1%were in the stage Ⅰ; 32 patients were benign, mainly tuberculoma (56.3%). There were 29 patients of malignant secondary lesions, 67 benign, and 1 both benign and malignant; the pathological agreement rate of primary and secondary lesions was 54.6% (lung cancer metastases in the lung and all the benign). When the primary lesion was malignant with its diameters of <3 cm, 3-<5 cm, 5-7 cm, >7 cm, the metastatic rate of secondary lesions was 42.5%, 15.8%, 20.0%, 0, respectively. When the primary lesion was malignant with lymph node metastasis, the probability of the secondary lesion being a metastatic tumor was higher than that without lymph node metastasis (46.7% vs. 30.0%, P>0.05). When the primary lesion was malignant and the primary and secondary lesions were located in the same lobe, the secondary lesions were more likely to metastasize (54.5%), while when they were located on different lobes on the same side or different sides, the secondary lesions were more likely to be benign (58.1%, 72.7%), and the possibility of metastasis was small ( 32.6%, 9.1%). When the primary lesion was benign and clinical differential diagnosis was difficult, the secondary lesion was benign. Conclusion    For synchronous multiple pulmonary nodules, the diameter of the primary lesion is large, the metastatic rate of secondary lesions tends to decrease. In ipsilateral synchronous multiple pulmonary nodules, especially with node metastasis, the risk of metastatic nodule increases. Bilateral surgical resection does not significantly increase the perioperative morbidity.

Objective:To investigate the stiffness characteristic of triple-negative human breast cancer at different size in a implantation nude female mice model using shear wave elastography(SWE) and to compare the clinical pathologic features of tumors with elasticity variables.Methods:Human breast cancer MDA-MB-231 cells were injected into 30 nude female mice and 27 transplanted tumors were successfully found in nude female mice. Ultrasound and SWE were longitudinally performed on maximum diameter plane of 21 tumours in 21 nude mice. The elastic parameters of maximal elasticity(Emax), mean elasticity (Emean) and standard deviation of elasticity(Esd) were recorded. The mice were divided into 3 groups according to the tumor size. They were group A with tumor size less than or equal to 5 mm, group B with tumor size greater than 5 mm and less than or equal to 10 mm, group C with tumor size larger than 10 mm and smaller than or equal to 15 mm. Compared with pathology, the relationships between Ki67 of transplanted tumor and elastic parameters were analyzed.Results:As the transplanted tumors increased, the values of Emax, Esd, Ki67 all increased. The lesions maximal size, Emax, Esd, Ki67 were significant higher in group B ( P<0.001, P=0.006, P=0.002, P=0.026) and group C ( P<0.001, P<0.001, P<0.001, P=0.028) than group A. The other parameters were not significantly different among the groups(all P>0.05). The size of transplanted tumors was significantly and positively correlated with Emax ( rs=0.673, P=0.001), Esd ( rs=0.661, P=0.001), and Ki67 ( rs=0.509, P=0.018). Conclusions:SWE Emax and Esd can reflect the tumor tissue stiffness change and biological activity during the tumor growth.

Objective: The aim of this study is to evaluate the commutability of 16 processed materials for 17-hydroxyprogesterone by using 2 commutability assessment approaches. Methods: 52 serum specimens were collected in Clinical Laboratory Department of Beijing Hospital from February 2018 to June 2019. According to the report of the Clinical and Laboratory Standards Institute (EP14-A3) document and the recommendations of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) working group on commutabilityassessment, serum 17-hydroxyprogesterone isotope diluent chromatogram tandem mass spectrometry (ID-LC/MS/MS) was used for comparison. Three clinical routine analysis systems (1 radioimmunoassay, 2 LC/MS analysis methods) were used to determine the concentration of 17-hydroxyprogesterone in 52 human serum samples and 16 processed materialsfor commutabilityassessment. Results: Combined with the results of the two commutability assessment, all accuracy verification materials and national steroid hormone standards showed good commutability in the LC/MS analysis system, and 6/9 EQA materials showed commutability in the three routine analysis systems.All materials showed good commutability in the LC/MS analysis system of bias difference method. Conclusions The two kinds of commutability assessment results are different. Bias difference method has more clinical value, but it has certain application limitations. The use of fresh frozen human serum as a quality assessment materialfor serum 17-hydroxyprogesterone is meets the commutability requirement.

Aldosterone and renin are important clinical indicators in the diagnosis and treatment of hypertension. There are two kinds of methods for aldosterone detection, including immunoassay and mass spectrometry. The coefficients of variation within the methods, as well as the deviation between methods are large.And the results of immunoassay are much higher than that of mass spectrometry. Clinically, renin tests mainly include renin concentration and renin activity, both of which have advantages and disadvantages in the diagnosis and treatment of hypertension. The disordered test results of aldosterone and renin brings great trouble to the clinical diagnosis and treatment. Standardization/consistency is urgently needed to make the test results of each laboratory more accurate and comparable. The standardization project for renin and aldosterone in China will greatly promote the standardization process in order to provide accurate and comparable clinical tests.

Objective To evaluate the learning curve of video-assisted thoracoscopic sleeve lobectomy in patients with central lung cancer.Methods A total of 86 consecutive patients with resected central lung cancer in the second department of thoracic surgery of Hunan Cancer Hospital between Apirl 2016 and July 2018 were retrospectively enrolled.Video-assisted tho-racoscopic tracheoplasty with sleeve resection and lobectomy were performed in 56 patients, video-assisted thoracoscopic tra-cheoplasty with wedge resection and lobectomy were performed in 20 patients, and 10 patients transit to thoracotomy.Surgical parameter of patients who underwent video-assisted thoracoscopic sleeve lobectomy were investigated to assess the learning curve, including operation duration, bleeding volume, amount of lymph nodes examined(medianstinal and intrapulmonary). Lowess smoothing method was performed to fit curve to evaluate the variation tendency of surgical parameters .Cut-off point, as well as the confidence interval, were identified using piecewise regression analysis.Results Surgical duration tend to be stable (almost 200 min) when the cumulative case amount of video-assisted thoracoscopic sleeve lobectomy reach 40.Surgical bleed-ing tend to be stable( almost 200 ml) when the cumulative case amount of video-assisted thoracoscopic sleeve lobectomy reach 20.There is no significant correlation between the amount of lymph node harvest and surgical volume .Conclusion The cut-off point for video-assisted thoracoscopic sleeve lobectomy is approximately 40 cases.

Objective To evaluate the comparability and consistency of two kinds of triglycerides reference methods, one of which is the isotope dilution liquid chromatography-mass spectrometry (LC/MS) in the Cholesterol Reference Method Laboratory Network (CRMLN), the other isthehigh-performance liquid chromatography (HPLC) method for triglyceride detection in China. Methods 52 fresh frozen sera with triglycerides levels among 0.45-4.52 mmol/L were determined by LC/MS and HPLC. After evaluation the precision and accuracy of the two methods,a series of analyses were conducted including plotting to scatter plots and deviation graphs, testing outliers, selecting the best fitting regression models and calculating the regression equations and parameters, and so on. The expected deviation at the level of medical decision of triglycerides and the 95％confidence range were statistically analyzed.Results For HPLC method, the CV of instrument measurement was 0.29％(0％-1.16％), the total CV of samples measurement was 0.54％(0.04％-1.28％), and the average bias of the reference materials was 0.22％(-0.43％-0.68％). ForLC/MSmethod,the CV of instrument measurement was 0.55％(0％-1.68％),the total CV of samples measurement was 0.79％(0％-1.93％), and the average bias of the NIST reference materials was 0.09％(-0.73％-1.29％). No outlier was found from the scatter plots and the statistical analysis and the linear regression was fitted to analyze the results of the two methods. The linear regression parameters of two methods for 52 fresh frozen human sera were as follows:the slope was 0.9988,the standard error of slope was 0.0035, the intercept was 0.0037mmol/L, the standard error of intercept was 0.0030 mmol/L, the standard error of Y-estimate was 0.0236 mmol/L,and the correlation coefficient was 0.9997. Compared with the LC/MS method,the absolute deviation of fresh sera by HPLC method was-0.001 mmol/L (-0.070-0.056 mmol/L), with a relative deviation of 0.13％ (-2.01-2.83％). T-test showed no statistically significant difference between the two methods. The expected deviations at the triglycerides medicine decision level were within the 95％confidence range,and the expected deviations were far less than the allowable error. Conclusions The HPLC method of triglyceridesdetetion has good consistency and comparability with LC/MS method as one of the reference methods of CRMLN. Because of the advantages of HPLC method such as low cost, simplicity,less technical need,and better precision,HPLC method is expected to play an important role in the process of standardization and traceability of serum triglycerides.