BACKGROUND:The phosphatidylinositol 3-kinase/protein kinase(PI3K/AKT)signaling pathway plays a pivotal role in regulating osteoclast activation,which is essential for maintaining bone homeostasis.Bone destruction in osteoarticular tuberculosis is caused by aberrant osteoclastogenesis induced by Mycobacterium tuberculosis infection.However,the role of the PI3K signaling pathway in Mycobacterium tuberculosis-induced aberrant osteoclastogenesis remains unclear. OBJECTIVE:To investigate the effects and mechanisms of the PI3K/AKT signaling pathway inhibitor BYL-719 on aberrant osteoclastogenesis induced by Mycobacterium tuberculosis. METHODS:RAW264.7 cells were infected with bovine Mycobacterium tuberculosis bacillus calmette-cuerin vaccine,and Ag85B was used for cellular immunofluorescence staining.The cell counting kit-8 assay was employed to determine the safe concentration of BYL-719.There were four groups in the experiment:blank control group,BYL-719 group,BCG group,and BCG+BYL-719 group.Under the induction of receptor activator of nuclear factor kappa-B ligand,the effects of BYL-719 on post-infection osteoclast differentiation and fusion were explored through tartrate-resistant acid phosphatase staining and phalloidin staining.RT-PCR and western blot were used to detect the expression of osteoclast-related genes and proteins,and further investigate the mechanism of action. RESULTS AND CONCLUSION:Immunofluorescence staining showed that RAW264.7 cells phagocytosed Mycobacterium tuberculosis.Cell counting kit-8 data indicated that 40 nmol/L BYL-719 was non-toxic to cells.Tartrate-resistant acid phosphatase staining and phalloidin staining showed that BYL-719 inhibited the generation and fusion ability of osteoclasts following infection.RT-PCR and western blot results also indicated that BYL-719 suppressed the upregulation of osteoclast-specific genes(including c-Fos,NFATc1,matrix metalloproteinase 9,and CtsK)induced by Mycobacterium tuberculosis infection(P<0.05).Western blot and immunofluorescence staining revealed that BYL-719 inhibited excessive osteoclast differentiation induced by Mycobacterium tuberculosis by downregulating the expression of IκBα-p65.To conclude,BYL-719 inhibits aberrant osteoclastogenesis induced by Mycobacterium tuberculosis through the downregulation of IκBα/p65.Therefore,the IκBα/p65 signaling pathway is a potential therapeutic target for osteoarticular tuberculosis,and BYL-719 holds potential value for the preventing and amelioration of bone destruction in osteoarticular tuberculosis.BYL-719 has the potential to prevent and ameliorate bone destruction in osteoarticular tuberculosis.

Diterpenoid ferruginol is a key intermediate in biosynthesis of active ingredients such as tanshinone and carnosic acid.However, the traditional process of obtaining ferruginol from plants is often cumbersome and inefficient. In recent years, the increasingly developing gene editing technology has been gradually applied to the heterologous production of natural products, but the production of ferruginol in microbe is still very low, which has become an obstacle to the efficient biosynthesis of downstream chemicals, such as tanshinone. In this study, miltiradiene was produced by integrating the shortened diterpene synthase fusion protein,and the key genes in the MVA pathway were overexpressed to improve the yield of miltiradiene. Under the shake flask fermentation condition, the yield of miltiradiene reached about(113. 12±17. 4)mg·L~(-1). Subsequently, this study integrated the ferruginol synthase Sm CYP76AH1 and Sm CPR1 to reconstruct the ferruginol pathway and thereby realized the heterologous synthesis of ferruginol in Saccharomyces cerevisiae. The study selected the best ferruginol synthase(Il CYP76AH46) from different plants and optimized the expression of pathway genes through redox partner engineering to increase the yield of ferruginol. By increasing the copy number of diterpene synthase, CYP450, and CPR, the yield of ferruginol reached(370. 39± 21. 65) mg·L~(-1) in the shake flask, which was increased by 21. 57-fold compared with that when the initial ferruginol strain JMLT05 was used. Finally, 1 083. 51 mg·L~(-1) ferruginol was obtained by fed-batch fermentation, which is the highest yield of ferruginol from biosynthesis so far. This study provides not only research ideas for other metabolic engineering but also a platform for the construction of cell factories for downstream products.
Saccharomyces cerevisiae/genetics* ; Diterpenes/metabolism* ; Metabolic Engineering ; Fermentation ; Abietanes

Diabetes mellitus(DM)is a disease syndrome characterized by chronic hyperglycaemia.A long-term high-glucose environment leads to reactive oxygen species(ROS)production and nuclear DNA damage.Human umbilical cord mesenchymal stem cell(HUcMSC)infusion induces significant antidiabetic effects in type 2 diabetes mellitus(T2DM)rats.Insulin-like growth factor 1(IGF1)receptor(IGF1R)is important in promoting glucose metabolism in diabetes;however,the mechanism by which HUcMSC can treat diabetes through IGF1R and DNA damage repair remains unclear.In this study,a DM rat model was induced with high-fat diet feeding and streptozotocin(STZ)administration and rats were infused four times with HUcMSC.Blood glucose,interleukin-6(IL-6),IL-10,glomerular basement membrane,and renal function were examined.Proteins that interacted with IGF1R were determined through coimmunoprecipitation assays.The expression of IGF1R,phosphorylated checkpoint kinase 2(p-CHK2),and phosphorylated protein 53(p-p53)was examined using immunohistochemistry(IHC)and western blot analysis.Enzyme-linked immunosorbent assay(ELISA)was used to determine the serum levels of 8-hydroxydeoxyguanosine(8-OHdG).Flow cytometry experiments were used to detect the surface markers of HUcMSC.The identification of the morphology and phenotype of HUcMSC was performed by way of oil red"O"staining and Alizarin red staining.DM rats exhibited abnormal blood glucose and IL-6/10 levels and renal function changes in the glomerular basement membrane,increased the expression of IGF1 and IGF1R.IGF1R interacted with CHK2,and the expression of p-CHK2 was significantly decreased in IGF1R-knockdown cells.When cisplatin was used to induce DNA damage,the expression of p-CHK2 was higher than that in the IGF1R-knockdown group without cisplatin treatment.HUcMSC infusion ameliorated abnormalities and preserved kidney structure and function in DM rats.The expression of IGF1,IGF1R,p-CHK2,and p-p53,and the level of 8-OHdG in the DM group increased significantly compared with those in the control group,and decreased after HUcMSC treatment.Our results suggested that IGF1R could interact with CHK2 and mediate DNA damage.HUcMSC infusion protected against kidney injury in DM rats.The underlying mechanisms may include HUcMSC-mediated enhancement of diabetes treatment via the IGF1R-CHK2-p53 signalling pathway.

Objective To explore the analgesic effect of different doses of dezocine combined with ropivacaine for thoracic paravertebral block(TPVB)on patients undergoing thoracoscopic radical resection of lung cancer and the influence on hemodynamics and immune function of patients.Methods Patients with lung cancer who underwent thoracoscopic radical resection were divided into low-dose group and high-dose group according to random number table method.Both groups of patients were given total intravenous anesthesia to complete the surgery.At 15 min before general anesthesia induction,the low-dose group was given TPBV with 0.1 mg·kg-1 dezocine+0.375％ropivacaine for a total of 20 mL,and the high-dose group was given TPBV with 0.15 mg·kg-1 dezocine+0.375％ropivacaine for a total of 20 mL.Comparisons were performed on both groups in terms of analgesic effect,hemodynamic parameters,immune function and occurrence of adverse drug reactions.Results There were 48 cases in low-dose group and 46 cases in high-dose group.In low-dose group,the heart rate values before TPVB,before skin incision,at 5 min after sectioning and at the end of surgery were(78.52±6.54),(70.79±7.07),(74.48±6.68)and(76.69±7.29)beat·min-1,the mean arterial pressure values were(93.16±5.72),(86.38±7.51),(92.15±6.36)and(91.14±6.13)mmHg.In high-dose group,the heart rate values at the above time points were(79.36±7.11),(71.68±6.49),(74.76±7.06)and(76.57±6.52)beat·min-1;the mean arterial pressure values were(93.89±7.18),(85.27±7.41),(90.34±6.52)and(92.43±6.34)mmHg,there were no statistical differences between the two groups(all P＞0.05).The resting state scores at 2,6 and 12 h after surgery were(1.38±0.19),(1.54±0.21)and(1.72±0.16)points,the pain scores at motion state were(1.88±0.15),(2.36±0.37)and(3.26±0.38)points in low-dose group;in high-dose group,the resting state scores were(1.32±0.17),(1.58±0.22)and(1.81±0.18)points,the pain scores at motion state were(1.81±0.13),(2.11±0.31)and(3.03±0.36)points,respectively,there were no statistical differences between the two groups(all P＞0.05).The number of analgesic pump compressions at 24 h after surgery and the number of cases with analgesic remedy were(5.12±1.26)times and 15 cases in low-dose group and were(4.74±1.03)times and 10 cases in high-dose group,with no statistical differences between the groups(all P＞0.05).The percentages of CD3+cells in low-dose group at the end of surgery and at 12 h and 24 h after surgery were(68.51±6.76)％,(54.22±5.43)％and(51.47±6.58)％,the percentages of CD4+cells were(40.29±5.02)％,(34.94±4.79)％and(30.48±5.11)％,CD4+/CD8+ratios were 1.54±0.34,1.36±0.28 and 1.16±0.23;the percentages of CD3+cells in high-dose group were(67.92±7.11)％,(56.58±6.36)％and(54.47±6.89)％,percentages of CD4+cells were(41.33±5.75)％,(35.86±5.21)％and(32.27±4.78)％,the CD4+/CD8+were 1.53±0.35,1.40±0.30 and 1.22±0.26,all with no significant difference(all P＞0.05).The incidence of postoperative adverse drug reactions in high-dose group and low-dose group were 32.61％and 14.58％,with significant difference(P＜0.05).Conclusion When TPVB regimen of dezocine combined with ropivacaine is used in thoracoscopic radical resection of lung cancer,the analgesic effect of low-dose dezocine is comparable to that of high-dose dezocine,with lower risk of adverse drug reactions.

Objective To develop an intelligent model for differential diagnosis of atrioventricular nodal re-entrant tachycardia(AVNRT)and atrioventricular re-entrant tachycardia(AVRT)using 12-lead wearable electrocardiogram devices.Methods A total of 356 samples of 12-lead supraventricular tachycardia(SVT)electrocardiograms recorded by wearable devices were randomly divided into training and validation sets using 5-fold cross validation to establish the intelligent classification model,and 101 patients with the diagnosis of SVT undergoing electrophysiological studies and radiofrequency ablation from October,2021 to March,2023 were selected as the testing set.The changes in electrocardiogram parameters before and during induced tachycardia were compared.Based on multiscale deep neural network,an intelligent diagnosis model for classifying SVT mechanisms was constructed and validated.The 3-lead electrocardiogram signals from Ⅱ,Ⅲ,and V1 were extracted to build new classification models,whose diagnostic efficacy was compared with that of the 12-lead model.Results Of the 101 patients with SVT in the testing set,68 were diagnosed with AVNRT and 33 were diagnosed with AVRT by electrophysiological study.The pre-trained model achieved a high area under the precision-recall curve(0.9492)and F1 score(0.8195)for identifying AVNRT in the validation set.The total F1 scores of the lead Ⅱ,Ⅲ,V1,3-lead and 12-lead intelligent diagnostic models in the testing set were 0.5597,0.6061,0.3419,0.6003 and 0.6136,respectively.Compared with the 12-lead classification model,the lead-Ⅲ model had a net reclassification index improvement of-0.029(P=0.878)and an integrated discrimination index improvement of-0.005(P=0.965).Conclusion The intelligent diagnostic model based on multiscale deep neural network using wearable electrocardiogram devices has an acceptable accuracy for classifying SVT mechanisms.

Objective To develop an intelligent model for differential diagnosis of atrioventricular nodal re-entrant tachycardia(AVNRT)and atrioventricular re-entrant tachycardia(AVRT)using 12-lead wearable electrocardiogram devices.Methods A total of 356 samples of 12-lead supraventricular tachycardia(SVT)electrocardiograms recorded by wearable devices were randomly divided into training and validation sets using 5-fold cross validation to establish the intelligent classification model,and 101 patients with the diagnosis of SVT undergoing electrophysiological studies and radiofrequency ablation from October,2021 to March,2023 were selected as the testing set.The changes in electrocardiogram parameters before and during induced tachycardia were compared.Based on multiscale deep neural network,an intelligent diagnosis model for classifying SVT mechanisms was constructed and validated.The 3-lead electrocardiogram signals from Ⅱ,Ⅲ,and V1 were extracted to build new classification models,whose diagnostic efficacy was compared with that of the 12-lead model.Results Of the 101 patients with SVT in the testing set,68 were diagnosed with AVNRT and 33 were diagnosed with AVRT by electrophysiological study.The pre-trained model achieved a high area under the precision-recall curve(0.9492)and F1 score(0.8195)for identifying AVNRT in the validation set.The total F1 scores of the lead Ⅱ,Ⅲ,V1,3-lead and 12-lead intelligent diagnostic models in the testing set were 0.5597,0.6061,0.3419,0.6003 and 0.6136,respectively.Compared with the 12-lead classification model,the lead-Ⅲ model had a net reclassification index improvement of-0.029(P=0.878)and an integrated discrimination index improvement of-0.005(P=0.965).Conclusion The intelligent diagnostic model based on multiscale deep neural network using wearable electrocardiogram devices has an acceptable accuracy for classifying SVT mechanisms.

Objective To explore the differential diagnostic indicators of adult-onset Still's disease(AOSD)from other fever of unknown origin(FUO).Methods The clinical data and laboratory indicators of 177 AOSD patients and 163 FUO patients who were hospitalized in the First Affiliated Hospital of Army Medical University from January 2010 to May 2021 were collected,and the patients were randomly divided into training group and verification group.Statistically significant variables were extracted from univariate analysis for receiver operating characteristic(ROC)curve analysis and the best cut-off value of the variables was obtained.The differential diagnostic indicators were extracted by multivariate logistic regression analysis and nomogram model was constructed.ROC curve,calibration curve,and decision curve analysis were used to evaluate the accuracy and stability of nomogram.Results Univariate analysis revealed that there were significant differences in 4 clinical features(arthralgia,rash,pharyngeal pain,myalgia)and 14 laboratory parameters[white blood cell count(WBC),monocyte percentage,neutrophil percentage,lymphocyte percentage,platelet count,C-reactive protein,interleukin-6(IL-6),ferritin,globulin,immunoglobulin A,and immunoglobulin G(IgG),creatine kinase,creatinine and complement C3].Multivariate analysis suggested that arthralgia,WBC≥9.995×109/L,IL-6≥98.13 ng/L,ferritin≥507.37 ng/ml,globulin≤36.58g/L,IgG≤13.59g/L,complement C3≥1.27 g/L were related with AOSD.The area under curve(AUC)values of training group and verification group were 0.917(95%CI 0.883-0.951)and 0.869(95%CI 0.802-0.936),respectively.The calibration curves showed good consistency.The decision curve analysis showed that training group and verification group had a large positive rate of return in the wide risk range of 5%-85%and 10%-85%,respectively.Conclusions This study has established a relatively accurate AOSD differential diagnosis model.The combination of arthralgia,WBC,IL-6,ferritin,globulin,IgG and complement C3 may help to distinguish AOSD from other causes of FUO.

Surgical operation is the main treatment of oral and maxillofacial tumors.Dysphagia is a common postoperative complication.Swal-lowing disorder can not only lead to mis-aspiration,malnutrition,aspiration pneumonia and other serious consequences,but also may cause psychological problems and social communication barriers,affecting the quality of life of the patients.At present,there is no systematic evalua-tion and rehabilitation management plan for the problem of swallowing disorder after oral and maxillofacial tumor surgery in China.Combining the characteristics of postoperative swallowing disorder in patients with oral and maxillofacial tumors,summarizing the clinical experience of ex-perts in the field of tumor and rehabilitation,reviewing and summarizing relevant literature at home and abroad,and through joint discussion and modification,a group of national experts reached this consensus including the core contents of the screening of swallowing disorders,the phased assessment of prognosis and complications,and the implementation plan of comprehensive management such as nutrition management,respiratory management,swallowing function recovery,psychology and nursing during rehabilitation treatment,in order to improve the evalua-tion and rehabilitation of swallowing disorder after oral and maxillofacial tumor surgery in clinic.

Cryoablation therapy with explicit anti-tumor mechanisms and histopathological manifestations has a long history.A large number of clinical practice has shown that cryoablation therapy is safe and effective,making it an ideal tumor treatment method in theory.Previously,its efficacy and clinical application were constrained by the limitations of refrigerants and refrigeration equipment.With the development of the new generation of cryoablation equipment represented by argon helium knives,significant progress has been made in refrigeration efficien-cy,ablation range,and precise temperature measurement,greatly promoting the progression of tumor cryoablation technology.This consensus systematically summarizes the mechanism of cryoablation technology,indications for oral mucosal melanoma(OMM)cryotherapy,clinical treatment process,adverse reactions and management,cryotherapy combination therapy,etc.,aiming to provide reference for carrying out the standardized cryoablation therapy of OMM.