Aim To investigate the intracellular up-take and target protein binding characteristics of two Bruton's tyrosine kinase inhibitors(BTKi)with differ-ent selectivities to provide further insights into the mechanisms of drug off-target-related bleeding risk.Methods Ibrutinib(non-selective BTKi)and za-nubrutinib(selective BTKi)were used as study drugs.After incubation of MEC-1 cells and human platelets with drugs,the cellular thermal shift assay(CETSA)was combined with Western blot to obtain the melting curve and isothermal curve to analyze the binding char-acteristics of the two drugs with the target protein BTK.After incubation of MEC-1 cells and human platelets with drugs,the concentrations of the two drugs were detected by liquid chromatography-tandem mass spectrometry(LC-MS/MS)to analyze the intracellular uptake of the two drugs.Results CETSA analysis confirmed that zanubrutinib was more selective for the target protein BTK compared to ibrutinib.LC-MS/MS analysis showed that both drugs were uptaken intracel-lularly by MEC-1 cells and platelets in a concentration-dependent manner.Conclusions While BTKi targe-ting BTK to B lymphocytes exerts therapeutic effects,off-target effects on platelets due to differences in their intracellular uptake,and target-binding characteristics may be one of the reasons for the differences in bleed-ing risk across selective BTKi.

Objective To explore the causes of postoperative revision for cubital tunnel syndrome(CTS),summarize the details of revision surgery for CTS and evaluate the efficacy,in order to reduce the revision probability of the disease.Methods The data of 14 patients undergoing revision surgery for CTS in our hospital from March 2019 to August 2022 were collected and analyzed.By comparing the first-stage surgical incision,examining the tension and shape of ulnar nerve,the compression site and the improvement of postoperative symptoms,the reasons for revision were summarized and the curative effect was evaluated.Result The reasons for revision included the poor outcome of the initial operation and no improvement of symptoms or symptom worsen within 3 months after surgery.The main cause for revision was irregular incision design(14/14),the secondary cause was incomplete release of nerve entrapment points(12/14),and the rare cause was insufficient hemostasis in the operative area(1/14)and insufficient protection of sensory nerve(1/14).According to Gu's functional evaluation criteria for CTS,the excellent rate of revision surgery was 9/14.Conclusion The main reason for revision after primary surgery for CTS is the inadequate management of surgical details in the first operation,and the revision rate of CTS can be reduced by standardized operation.Revision surgery is still effective after the failure of the initial operation.

Objective To investigate the effect of the long non-coding RNA FGD5 antisense RNA1(lncRNA FGD5-AS1)on myocar-dial cell injury induced by high glucose levels through regulation of the miR-195-5p/Pim-1 proto-oncogene(PIM1)axis.Methods The rat myocardial cell line H9c2 and human myocardial cells were randomly allocated to the control,model,lncRNA FGD5-AS1 overex-pression,miR-195-5p inhibitor,negative control,and lncRNA FGD5-AS1 overexpression+miR-195-5p mimic groups to determine the expression of lncRNA FGD5-AS1,miR-195-5p,and PIM1.In addition,cell proliferation,apoptosis,and pro-inflammatory cytokine levels of the cells in each group were analyzed.Results Compared with the control group,the protein and mRNA expressions of lncRNA FGD5-AS1 and PIM1 and the survival and proliferation rates in H9c2 and human myocardial cells in the model group decreased(P<0.05),and the expression of miR-195-5p,apoptosis rate,and production levels of tumor necrosis factorα(TNF-α)and interleukin-6(IL-6)increased(P<0.05).Overexpression of the lncRNA FGD5-AS1 reversed these pathological changes in model group cells,whereas upregulation of miR-195-5p could weaken the effects of overexpression of lncRNA FGD5-AS1.Conclusion Overexpression of lncRNA FGD5-AS1 enhances the expression of PIM1 by downregulating miR-195-5p,thereby inhibiting high glucose-induced myocyte inflamma-tion,promoting survival and growth,and alleviating apoptotic injury.

Objective:To establish a set of artificial intelligence (AI) verification rules for blood routine analysis.Methods:Blood routine analysis data of 18 474 hospitalized patients from the First Hospital of Jilin University during August 1st to 31st, 2019, were collected as training group for establishment of the AI verification rules,and the corresponding patient age, microscopic examination results, and clinical diagnosis information were collected. 92 laboratory parameters, including blood analysis report parameters, research parameters and alarm information, were used as candidate conditions for AI audit rules; manual verification combining microscopy was considered as standard, marked whether it was passed or blocked. Using decision tree algorithm, AI audit rules are initially established through high-intensity, multi-round and five-fold cross-validation and AI verification rules were optimized by setting important mandatory cases. The performance of AI verification rules was evaluated by comparing the false negative rate, precision rate, recall rate, F1 score, and pass rate with that of the current autoverification rules using Chi-square test. Another cohort of blood routine analysis data of 12 475 hospitalized patients in the First Hospital of Jilin University during November 1sr to 31st, 2023, were collected as validation group for validation of AI verification rules, which underwent simulated verification via the preliminary AI rules, thus performance of AI rules were analyzed by the above indicators. Results:AI verification rules consist of 15 rules and 17 parameters and do distinguish numeric and morphological abnormalities. Compared with auto-verification rules, the true positive rate, the false positive rate, the true negative rate, the false negative rate, the pass rate, the accuracy, the precision rate, the recall rate and F1 score of AI rules in training group were 22.7%, 1.6%, 74.5%, 1.3%, 75.7%, 97.2%, 93.5%, 94.7%, 94.1, respectively.All of them were better than auto-verification rules, and the difference was statistically significant ( P<0.001), and with no important case missed. In validation group, the true positive rate, the false positive rate, the true negative rate, the false negative rate, the pass rate, the accuracy, the precision rate, the recall rate and F1 score were 19.2%, 8.2%, 70.1%, 2.5%, 72.6%, 89.2%, 70.0%, 88.3%, 78.1, respectively, Compared with the auto-verification rules, The false negative rate was lower, the false positive rate and the recall rate were slightly higher, and the difference was statistically significant ( P<0.001). Conclusion:A set of the AI verification rules are established and verified by using decision tree algorithm of machine learning, which can identify, intercept and prompt abnormal results stably, and is moresimple, highly efficient and more accurate in the report of blood analysis test results compared with auto-vefication.

AIM:To explore the roles and associations of hepatocyte nuclear factor 4 alpha(HNF4A)and mu-protocadherin(MUCDHL)in the kidney of diabetic mice.METHODS:(1)A cohort of six 12-week-old db/m mice and six db/db mice were selected and maintained on a standard diet until 16 weeks.The protein levels of fibronectin(FN),collagen type III(Col-III),E-cadherin,α-smooth muscle actin(α-SMA),HNF4A,Snail and MUCDHL in renal tissues were scrutinized using Western blot.Immunohistochemical staining was conducted to observe the distribution and expres-sion of FN,HNF4A and MUCDHL.(2)Mouse renal tubular epithelial cells(mRTEC)were cultured in vitro and catego-rized into groups:normal glucose(NG)group,high glucose(HG)group,overexpression control groups(NG+vector and HG+vector),overexpression groups(NG+OE-MUCDHL,HG+OE-MUCDHL,NG+OE-HNF4A and HG+OE-HNF4A),knockdown control groups(NG+control and HG+control),and knockdown groups(NG+si-MUCDHL,HG+si-MUCDHL,NG+si-HNF4A and HG+si-HNF4A).The relevant protein levels were also detected by Western blot.RESULTS:(1)In db/db group,elevated body weight,blood glucose and urine albumin-to-creatinine ratio(UACR)indicated significant re-nal injury.Compared with db/m group,the mice in db/db group exhibited increased expression of FN,Col-III,α-SMA and Snail,and decreased expression of E-cadherin,HNF4A and MUCDHL.MUCDHL was predominantly expressed in the apical membrane of renal tubular epithelial cells,FN in the tubular mesenchyme,and HNF4A in the plasma and nu-cleus of renal tubular cells.(2)In HG group,there was an up-regulation in the expression of fibrosis-related proteins and a down-regulation in the expression of E-cadherin,HNF4A and MUCDHL compared with NG group.Overexpression of MUCDHL led to a decrease in the expression of FN,Col-III,α-SMA and Snail proteins,an increase in the expression of E-cadherin and MUCDHL proteins,and unaltered expression of HNF4A.Knockdown of MUCDHL resulted in a reversal of the aforementioned effects,with HNF4A expression remaining unaltered.Overexpression of HNF4A led to an increased ex-pression of MUCDHL,and the expression changes of the remaining indicators were consistent with the overexpression of MUCDHL.Knockdown of HNF4A reversed the aforementioned effects.MUCDHL may represent a downstream target gene of HNF4A.CONCLUSION:The diminished expression of HNF4A and MUCDHL in the renal tubules of diabetic mice implies their involvement in the progression of renal fibrosis in diabetic kidney disease(DKD).HNF4A may potentially impede the progression of renal fibrosis in DKD by up-regulating the expression of MUCDHL.

Tropane alkaloids (TAs) are a class of anticholinergic drugs widely used in clinical practice and mainly extracted from plant, among which Atopa belladonna is the main commercial drug source. It is of great industrial value to obtain TAs in large quantities by plant metabolic engineering. In TAs pathway, cytochrome oxidase CYP82M3 catalyze the synthesis of tropinone and then tropinone reductase I (TRI) compete with TRII for tropinone to form tropine leading to the TAs synthesis (drainage). In this study, based on the "increasing flow and drainage" metabolic engineering strategy, two genes, namely HnCYP82M3 and DsTRI from Hyoscyamus niger and Datura stramonium, respectively, were overexpressed in the hair roots of A. belladonna, with a view to promote the TAs accumulation. The HnCYP82M3 gene was cloned from the root of H. niger, and it encoded amino acid with 91.7% sequence identity with AbCYP82M3 from A. belladonna. Overexpression of HnCYP82M3 alone did not affect the content of TAs in hair roots of A. belladonna, indicating that CYP82M3 was not a key enzyme in TAs biosynthesis. Simultaneous overexpression of HnCYP82M3 and DsTRI greatly promoted the accumulation of the three TAs, and the contents of hyoscyamine, anisodamine and scopolamine were 4.97 times, 2.83 times and 2.19 times that of the control, respectively, and the increase amplitude was greater than that of single overexpression of DsTRI. This study showed that the "increasing flow and drainage" strategy of enzyme genes co-expression at branch points was a promising metabolic engineering method to effectively improve the biosynthesis of TAs in A. belladonna, and laid a theoretical and technical foundation for the large-scale industrial acquisition of TAs.

Objective To study the traditional Chinese medicine(TCM)constitution characteristics of overweight/obese patients in Shanghai region and to investigate the correlation of TCM constitution with body composition.Methods Relevant data were collected from the patients with complete information of TCM constitution and human body composition analysis who visited the specialized outpatient clinic of acup-moxibustion catgut embedding therapy in the Department of Endocrinology,Shanghai Tenth People's Hospital from August 2020 to December 2022.The patients were divided into a normal body mass group(BMI＜24 kg/m2),an overweight group(24 kg/m2≤BMI＜28 kg/m2)and obesity group(BMI≥28 kg/m2),and then the distribution of TCM constitution types in the three groups of patients were analyzed.After that,the correlation between TCM constitution and body composition were explored with multiple regression analysis.Results(1)A total of 315 patients were included,of which 43 patients had normal body mass,85 patients were overweight and 187 patients were obese.(2)The TCM constitution types in descending order of the composition ratio in the normal body mass group and in the overweight group were spleen deficiency constitution,liver stagnation constitution,damp-heat constitution,yang deficiency constitution,and yin deficiency constitution,in the obese group were spleen deficiency constitution,yang deficiency constitution,damp-heat constitution,liver stagnation constitution,and yin deficiency constitution,and in the overweight/obese group were spleen deficiency constitution,damp-heat constitution,yang deficiency constitution,liver stagnation constitution,and yin deficiency constitution.No statistically significant differences of the distribution of TCM constitution types were shown between normal body bass population and overweight/obese population(P＞0.05).In both normal body mass population and overweight/obese population,the single body constitution type was common,and biased constitution was rare,and there was no statistically significant difference when comparing between the two groups(P＞0.05).(3)The results of multiple regression analysis showed that the basal metabolism of all patients was positively correlated with yang deficiency constitution and was negatively correlated with damp-heat constitution,and the differences were statistically significant(P＜0.01).It is indicated that if the score of yang deficiency constitution rose by one point,the basal metabolism would increase by 0.54 kcal,and if the score of damp-heat constitution decreased by one point,the basal metabolism would decrease by 1.005 kcal.Conclusion In Shanghai region,obesity may be the main indication of the variation of the body constitution.In addition to spleen deficiency constitution,the proportions of yang deficiency constitution,damp-heat constitution and liver stagnation constitution are also higher in obese patients.In terms of the correlation between TCM constitution and body composition,basal metabolism is positively correlated with yang deficiency constitution and is negatively correlated with damp-heat constitution.Therefore,for the patients with yang deficiency constitution and damp-heat constitution,the influence of the basal metabolism level on the development of the disease should be taken into account.

Objective:To develop and validate a postoperative infection nomogram of hepatitis B-associated hepatocellular carcinoma (HCC) after hepatectomy.Methods:Clinical data of 229 patients with HCC undergoing hepatectomy at the Department of Hepatobiliary Surgery of Tianjin Third Central Hospital from January 2014 to December 2022 were retrospectively analyzed, including 174 males and 55 females, aged (58.2±11.4) years. LASSO regression analysis screened the factors associated with hepatitis B-associated HCC infection after hepatectomy, which were further incorporated into multivariate logistic regression analysis. A nomographic prediction model was established based on the results of multivariate logistic regression analysis. Concordance index (C-index), calibration curve and receiver operating characteristic (ROC) curve were used to evaluate the model, and decision curve analysis (DCA) was used to analyze the clinical applicability of the model. Internal validation of the model was performed using bootstrap method.Results:A total of nine variables were screened as factors associated with the postoperative infections using LASSO regression, including gender, smoking history, body mass index (BMI), serum level of alpha fetoprotein, resection fashion (anatomical or non-anatomical), intraoperative blood loss, surgical method (laparoscopy or open), serum level of creatinine, and postoperative biliary fistula. Multivariate logistic regression analysis showed that BMI, resection fashion, intraoperative blood loss >500 ml, and postoperative biliary fistula were risk factors for postoperative infection (all P<0.05). Based on the above risk factors, a postoperative infection nomogram of hepatitis B-associated HCC after hepatectomy was established. The C-index was 0.839 (95% CI: 0.768-0.910), and the area under ROC curve was 0.853 (95% CI: 0.795-0.912), indicating that the model had a good predictive ability. The calibration curve was basically consistent with the ideal curve. The DCA showed that the model had a good clinical applicability. Internal validation C-index was 0.829 (95% CI: 0.766-0.892). Conclusion:The nomogram based on BMI, surgical resection fashion, intraoperative blood loss >500 ml, and postoperative biliary fistula has a high predictive accuracy and can be used to predict postoperative infections after hepatectomy for HCC.

AIM To study the chemical constituents from the leaves of Cyanocarya paliurus(Batalin)Iljinskaja and their α-glucosidase inhibitory activities.METHODS The 95%ethanol extract from the leaves of C.paliurus was isolated and purified by macroporous resin,silica gel,Sephadex LH-20,polyamide,C18 reversed-phase silica gel and semi-preparative HPLC,then the structures of obtained compounds were identified by physicochemical properties and spectral data.Their α-glucosidase inhibitory activities were evaluated by PNPG.RESULTS Fifteen compounds were isolated and identified as cyclopaloside C(1),cyclopaloside A(2),juglanosides E(3),vaccinin A(4),ent-murin A(5),kaempferol 3-O-α-L-rhamnopyranoside(6),kaempferol-3-O-β-D-glucopyranoside(7),kaempferol-3-O-β-D-glucuronide methyl ester(8),kaempferol-3-O-β-D-glucuronide ethyl ester(9),kaempferol-3-O-β-D-glucuronide butyl ester(10),quercetin-3-O-α-L-rhamnopyranoside(11)quercetin-3-O-β-D-glucopyranoside(12),quercetin-3-O-β-D-galactopyranoside(13),quercetin-3-O-β-D-glucuronide butyl ester(14),dihydrokaempferol(15).The IC50 value of total extracts ihibited α-glucosidase was(1.83±0.04)μg/mL,and the IC50 values of compounds 1,4-5 were(29.48±1.86),(0.50±0.07),(0.71±0.07)μmol/L,respectively.CONCLUSION Compound 1 is a new tetrahydronaphthalene glycoside.Compounds 4-5,8-10 and 14 are isolated from the leaves of C.paliurus for the first time.Compounds 4-5 are relatively rare flavonoid lignans with potential inhibitory activities against α-glucosidase.

Objective To explore the effect of quercetin combined with iron death inhibitor Ferrostain-1 on oxalate-induced HK-2 cell injury.Methods HK-2 cells were randomly divided into control group(normal cultured cells),model group(0.5 mmol·L-1 calcium oxalate crystals),quercetin group(0.5 mmol·L-1 calcium oxalate crystals+100 μmol·L-1 quercetin),inhibitor group(0.5 mmol·L-1 calcium oxalate crystals+8 μmol·L-1 Ferrostain-1)and combination group(0.5 mmol·L-1calcium oxalate crystals+100 μmol·L-1quercetin+8 μmol·L-1 Ferrostain-1).Cell counting kit-8(CCK-8)assay was used to detect cell survival rate;Western blot was used to detect iron death related protein expression such as glutathione peroxidase 4(GPX4);flow cytometry and Tunel assay were used to detect cell apoptosis,and assay kit was used to detect cellular iron ions and antioxidant levels.Results The cell survival rates of control group,model group,quercetin group,inhibitor group and combination group were(100.00±2.55)％,(54.49±4.11)％,(64.26±6.30)％,(58.03±3.04)％and(79.37±4.29)％,respectively;GPX4 protein expression levels were 0.98±0.11,0.33±0.05,0.56±0.05,0.78±0.07 and 1.11±0.11,respectively;cell apoptosis rates were(4.15±0.28)％,(23.12±2.49)％,(17.28±1.07)％,(15.08±1.41)％and(8.95±0.75)％,respectively;Fe2+levels were(100.00±0.87)％,(162.55±14.70)％,(149.09±9.50)％,(144.95±11.12)％and(131.76±12.18)％,respectively;superoxide dismutase(SOD)levels were(58.67±3.46),(21.56±1.32),(33.60±2.03),(35.15±3.02)and(44.27±3.89)U·mL-1,respectively.The above indicators of the model group were compared with the control group,and the above indicators of the quercetin group,inhibitor group,and combination group were compared with the model group,the above indicators of the combination group were compared with the quercetin group,and inhibitor group,all they all showed statistical significance(all P＜0.05).Conclusion Iron death inhibitors can enhance the inhibitory effect of quercetin in vitro on oxalate-induced renal tubular epithelial cell injury.