Objective:To investigate the impact of Tumor-Infiltrating Lymphocytes (TILs) density in the tumor stroma on the long-term prognosis of hepatocellular carcinoma (HCC) liver transplant patients meeting the Hangzhou criteria.Methods:This study is a retrospective cohort study. The clinical data of 83 patients with hepatocellular carcinoma who met the Hangzhou criteria and underwent allogeneic liver transplantation from January 2018 to December 2023 in the Department of Hepatobiliary Surgery, Beijing Chaoyang Hospital affiliated to Capital Medical University were collected and analyzed. Hematoxylin and Eosin (HE) staining was used to study the density of TILs in the resected liver grafts. Patients were divided into TILs-negative group (TILs<10%, n=31) and TILs-positive group (TILs≥10%, n=52) based on whether the TILs density exceeded 10%. Clinical and pathological characteristics were analyzed, and the significance of alpha-fetoprotein (AFP), TILs density, and microvascular invasion on the prognosis of HCC patients who met the Hangzhou criteria was studied. Measurement data with normal distribution were expressed as mean±standard deviation ( ± s) and compared between groups using t-test. Measurement data with skewed distribution were expressed as M ( Q1, Q3) and compared using rank-sum tests. Categorical data were compared using chi-square test. Kaplan-Meier method was used to study the relationship between various observation indicators and overall survival, and survival curves were plotted. Log-rank test was used to compare the survival rates between groups, and multivariate Cox regression model was used to adjust for the distribution of risk factors between groups. Results:The preoperative AFP level in the TILs-negative group was (15.69±1.21) U/mL, and in the TILs-positive group was (12.17±0.13) U/mL, with a statistically significant difference between the two groups ( P<0.05). In the TILs-negative group, 8 cases had microvascular invasion, and the number of low, moderate, and high differentiation tumors was 8, 23, and 0, respectively. In the TILs-positive group, 3 cases had microvascular invasion, and the number of low, moderate, and high differentiation tumors was 2, 31, and 19, respectively. The results indicated that patients in the TILs-negative group were more likely to have microvascular invasion and poorer tumor differentiation ( P<0.05). All patients were regularly followed up, and the 1-, 2-, and 3-year survival rates in the TILs-negative group and TILs-positive group were 84.0%, 77.6%, 69.8%, and 94.7%, 91.7%, 86.6%, respectively ( P<0.01). Cox proportional hazards model indicated that microvascular invasion ( RR=4.474, 95% CI: 1.172-17.072, P=0.028) and TILs-negative status ( RR=5.081, 95% CI: 1.420-18.184, P=0.012) were independent risk factors for the long-term prognosis of HCC patients who met the Hangzhou criteria. Conclusions:Among HCC patients meeting the Hangzhou criteria, the density of TILs in the tumor stroma is related to AFP levels, tumor differentiation, and the presence of microvascular invasion. TILs-negative status indicates a poorer prognosis for these patients.

Objective To compare the clinical outcomes of using elastic intramedullary nail and plate to fix fibular frac-ture.Methods The 60 patients with tibiofibular fractures admitted from January 2015 to December 2022 were divided into two groups:intramedullary nail group and plate group,30 cases each,intramedullary nail group was treated with elastic in-tramedullary nail fixation group,plate group was treated with steel plate and screw fixation group.Intramedullary nail group,there were 18 males and 12 females,aged from 22 to 75 years old with an average of(39.4±9.8)years old,including 24 cases of traffic accidents injury,6 cases of falling injury,23 cases of closed fractures,7 cases of open fractures.Steel plate group,there were 15 males and 15 females,aged from 24 to 78 years old with an average of(38.6±10.2)years old.The 22 cases were injured by traffic accident,8 cases were injured by falling.The 24 cases were closed fractures and 6 cases were open fractures.The operation time,intraoperative bleeding,American Orthopedic Foot and Ankle Society(AOFAS)ankle and hind foot scores,clinical healing time of fibula and the incidence of wound complications were compared between the two groups.Re-sults The patients in both groups were followed up for 6 to 21 months,with an average of(14.0±2.8)months.Compared with plate group,intramedullary nail group had shorter operative time,less bleeding,shorter clinical healing time of fibula,and low-er infection rate of incision,and the difference was statistically significant(P＜0.05).There were 2 cases of delayed healing in intramedullary nail group,1 case of nonunion in plate group,and 2 cases of delayed healing in plate group,and there was no statistically significant difference between the two groups(P＞0.05).In the last follow-up,according to the AOFAS scoring standard,the ankle function in intramedullary nail group was excellent in 17 cases,good in 12 cases,fair in 1 case,with an av-erage of(88.33±4.57)points,while in plate group,excellent in 16 cases,good in 10 cases,fair in 4 cases,with an average of(87.00±4.14)points;There was no statistical difference between the two groups(P＞0.05).Conclusion Elastic intramedullary nail has the advantages of short operation time,less intraoperative bleeding,short fracture healing time and less incision com-plications in the treatment of fibular fracture,which is worthy of clinical application.

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.

Abstract: Objective　To evaluate the value of bronchoalveolar lavage fluid Aspergillus-specific fluorescent PCR assay combined with galactomannan (GM) assay in the diagnosis of patients with non-neutropenic invasive pulmonary aspergillosis (IPA). Methods From March 2022 to December 2023, 113 hospitalized patients with clinically suspected IPA were selected from the Zibo First Hospital of Zibo City, Shandong Province. Bronchoalveolar lavage fluid samples from each patient were simultaneously subjected to potassium hydroxide microscopy, fungal culture, GM assay, and Aspergillus-specific fluorescence PCR assay. According to the diagnostic criteria of IPA, patients were divided into clinically diagnosed IPA and non-IPA groups. The values of these four methods for the diagnosis of IPA were compared. Results According to the diagnostic criteria for IPA, 37 out of the 113 suspected patients were clinically diagnosed as IPA. The proportion of diabetic patients was significantly higher in the IPA group compared to the non-IPA group (χ2=7.494, P=0.006); similarly, the proportion of patients using glucocorticoids was significantly higher in the IPA group (χ2=6.981, P=0.008). Patients in the IPA group more frequently showed cavitation within consolidation areas on imaging, which was statistically significant (χ2=15.603, P<0.001). There were significant differences in the sensitivity of the four fungal detection methods in the diagnosis of IPA (χ2=45.803, P<0.001), with Aspergillus-specific PCR assay showing the highest sensitivity at 94.59%. Specificity also varied significantly across the four methods (χ2=31.511, P<0.001), with the highest specificity being seen in potassium hydroxide microscopy and fungal culture at 100.00%. There were significant differences in the clinical coincidence rate of the four methods in the diagnosis of IPA (χ2=11.768, P=0.008), with Aspergillus-specific fluorescence PCR assay having the highest coincidence rate at 90.27%. The AUC of the ROC curve of Aspergillus-specific fluorescent PCR assay combined with the GM assay was 0.976 7, higher than 0.913 8 by Aspergillus-specific fluorescent PCR assay merely. Conclusions The combination of Aspergillus-specific fluorescent PCR assay and GM assay using bronchoalveolar lavage fluid could significantly improve the accuracy of IPA diagnosis in patients without neutropenia.

Nucleic acid-based molecular diagnostic methods are considered the gold standard for detecting infectious pathogens.However,when applied to portable or on-site rapid diagnostics,they still face various limitations and challenges,such as poor specificity,cumbersome operation,and portability difficulties.The CRISPR(Clustered regularly interspaced short palindromic repeats)/CRISPR-associated protein(Cas)-fluorescence detection method holds the potential to significantly enhance the specificity and signal-to-noise ratio of nucleic acid detection.In this study,we developed a portable grayscale reader detection system based on loop-mediated isothermal amplification(LAMP)-CRISPR/Cas.On one hand,in the presence of CRISPR RNA(crRNA),the CRISPR/Cas12a system was employed to achieve precise fluorescent detection of self-designed LAMP amplification reactions for influenza A and influenza B viruses.This further validated the high selectivity and versatility of the CRISPR/Cas system.On the other hand,the accompanying independently developed portable grayscale reader allowed for low-cost collection of fluorescence signals and high-reliability visual interpretation.At the end of the detection process,it directly provided positive or negative results.Practical sample analyses using this detection system have verified its reliability and utility,demonstrating that this system can achieve highly sensitive and highly specific portable analysis of influenza viruses.

Objective:To explore the effects of the fat mass and obesity-associated gene(FTO)on apoptosis and the inflammatory response of chondrocytes in osteoarthritis(OA).Methods:Differences in FTO expression between normal human cartilage tissue samples and OA cartilage tissue samples were examined.Primary OA chondrocytes were isolated and cultured, and a rat OA model was constructed.The expression of FTO was detected in clinical, animal and cellular samples.Cells were treated with an FTO knockdown lentivirus vector(sh-FTO)and an m 6A methylation inhibitor(cycloleucine). The amount of m 6A and the expression levels of inflammatory cytokines, interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α), were detected.Flow cytometry was used to detect apoptosis in OA chondrocytes, and Western blot was used to detect the expression levels of B-cell lymphoma 2(Bcl-2)and Bcl-2-associated X protein(Bax). Results:Compared with the normal control group, FTO mRNA and protein expression in human OA cartilage tissue, rat OA cartilage tissue and OA chondrocytes was significantly increased(all P<0.05). After FTO knockdown, the level of m 6A increased, the levels of IL-6 and TNF-α decreased considerably, the apoptosis rate decreased, the expression of the proapoptotic protein Bax decreased considerably, and the expression of Bcl-2 increased considerably in primary OA chondrocytes.However, cycloleucine intervention clearly reduced the level of m6A, increased the levels of IL-6 and TNF-α, promoted cell apoptosis and the expression of apoptosis-related proteins, and reversed the effect induced by the FTO knockdown lentivirus in OA chondrocytes(all P<0.05). Conclusions:FTO may be involved in mechanisms related to the action of m 6A to promote OA chondrocyte apoptosis and the inflammatory response, thus accelerating the progression of OA.

The pathogenesis of urolithiasis is not yet clear， and there are obvious limitations in the prevention and treatment of urolithiasis by either Chinese or western medicine. The microscopic pathological changes of the kidney from anatomical perspective have a certain internal connection with viewpoint of “kidney storing insufficiency， and kidney deficiency as the root” in the traditional Chinese medicine （TCM） zang-fu （脏腑） theory. Accordingly， the prevention and treatment of urolithiasis by integrated traditional Chinese and western medicine based on “shen-kidney” theory has been proposed. It is believed that the prevention and treatment of urolithiasis can be divided into two stages， that is expelling stones and preventing stones. In terms of preventing stones from kideny， it is recommended to focus on the early pathological changes of the kidney； for preventing stones from shen， it is advised to prevent and treat urolithiasis from kidney deficiency. The treatment should be time-based and stage by stage. Adhering to the principle of “prevention before disease occurs， prevention is more important than treatment” aims to advance the intervention targets for the prevention and treatment of urolithiasis. Emphasizing on the simultaneous treatment of kidney disease and urolithiasis， it is critical to put focus on the development of calcium-containing crystalline nephropathy in the early stage of stone formation， as well as the fundamental pathogenesis of kidney deficiency in TCM. Shen-kidney theory aims to further promote the integration of traditional Chinese and western medicine in the prevention and treatment of urolithiasis， which may provide certain reference for solving the current dilemma of urolithiasis prevention and treatment.

Serum cholinesterase(ChE)level is important for the diagnosis and prognostic evaluation of various diseases such as liver diseases and toxic diseases,and butyrylcholinesterase(BuChE)is an important component of ChE.Mutations in the BCHE gene can cause a significant reduction in the level of BuChE,with extensive reports in European and American populations and relatively few reports in Eastern countries,particularly China.This study describes a male patient,aged 35 years,who was misdiagnosed with organophosphorus pesticide poisoning due to an extreme reduction in ChE level and was given detoxification therapy,but such diagnosis was excluded by various biochemical examinations.Finally whole-exome sequencing and Sanger sequencing revealed the complex heterozygous mutations of c.1027dup and c.401dup at exon 2 of the BCHE gene,and hereditary BuChE deficiency due to these mutations is the cause of the extreme reduction in ChE level.

Objective:To explore the impact of sleep duration, physical exercise, and their interactions on the risk of dyslipidemia in older adults aged ≥80 (the oldest old) in China.Methods:The study subjects were the oldest old from four rounds of Healthy Aging and Biomarkers Cohort Study (2008-2009, 2011-2012, 2014 and 2017-2018). The information about their demographic characteristics, lifestyles, physical examination results and others were collected, and fasting venous blood samples were collected from them for blood lipid testing. Competing risk model was used to analyze the causal associations of sleep duration and physical exercise with the risk for dyslipidemia. Restricted cubic spline (RCS) function was used to explore the dose-response relationship between sleep duration and the risk for dyslipidemia. Additive and multiplicative interaction model were used to explore the interaction of sleep duration and physical exercise on the risk for dyslipidemia.Results:The average age of 1 809 subjects was (93.1±7.7) years, 65.1% of them were women. The average sleep duration of the subjects was (8.0±2.5) hours/day, 28.1% of them had sleep duration for less than 7 hours/day, and 27.2% had sleep for duration more than 9 hours/day at baseline survey. During the 9-year cumulative follow-up of 6 150.6 person years (follow-up of average 3.4 years for one person), there were 304 new cases of dyslipidemia, with an incidence density of 4 942.6/100 000 person years. The results of competitive risk model analysis showed that compared with those who slept for 7-9 hours/day, the risk for dyslipidemia in oldest old with sleep duration >9 hours/day increased by 22% ( HR=1.22, 95% CI: 1.07-1.39). Compared with the oldest old having no physical exercise, the risk for dyslipidemia in the oldest old having physical exercise decreased by 33% ( HR=0.67, 95% CI: 0.57-0.78). The RCS function showed a linear positive dose-response relationship between sleep duration and the risk for hyperlipidemia. The interaction analysis showed that physical exercise and sleep duration had an antagonistic effect on the risk for hyperlipidemia. Conclusion:Physical exercise could reduce the adverse effects of prolonged sleep on blood lipids in the oldest old.

Objective:To investigate the association of urinary cadmium levels with peripheral leukocyte classification counts among middle-aged and older adults aged 40 to 89 years in selected areas of China.Methods:The research was based on the survey of the impact of soil quality of agricultural land on human health in typical areas conducted in 2019-2020. A total of 5 600 middle-aged and older adults aged 40 to 89 years were included by using a multi-stage stratified random sampling method. Baseline characteristics of the subjects were collected and physical examinations were performed. Random midstream urine was collected to measure urinary cadmium and urinary creatinine and fasting venous blood was collected to measure the leukocyte count, neutrophil count, lymphocyte count, monocyte count and eosinophil count. The linear mixed effect model was used to analyse the association of urinary cadmium levels with leukocyte classification counts, and the dose-response relationship between them was analyzed by using the restricted cubic spline (RCS) function.Results:The age of the subjects was (63.17±12.02) years; 2 851 (50.91%) were males; and the M ( Q 1, Q 3) of urinary creatinine-corrected urinary cadmium levels was 2.69 (1.52, 4.69) μg/g·creatinine. After adjusting for confounding factors, the results of linear mixed effects model analysis showed that for each 1-unit increase in urinary creatinine-corrected urinary cadmium level, the percentage change [% (95% CI)] of leukocyte count and lymphocyte count was -1.70% (-2.61%, -0.79%) and -1.57% (-2.86%, -0.26%), respectively. RCS function showed a negative linear relationship between urinary creatinine-corrected urinary cadmium levels and leukocyte counts and lymphocyte counts, respectively (all Pnon-linear>0.05). Conclusion:Urinary cadmium levels are negatively associated with leukocyte count and lymphocyte count among middle-aged and older adults aged 40 to 89 years in selected areas of China.