Objective:To compare the distribution differences of serum protein electrophoresis (SPE) among different gender and age individuals, and to explore the clinical application of SPE screening monoclonal gammopathy.Methods:A retrospective analysis was conducted based on the SPE results obtained from 533 989 cases enrolled from January 2018 to December 2019 at Zhongshan Hospital Affiliated to Fudan University. Among these patients, 435 479 inpatients were from departments of hematology, nephrology, spinal surgery, endocrinology, and rheumatology and immunology; and 98 510 were apparently healthy individuals. The distributions of albumin, α1 globulin, α2 globulin, β1 globulin, β2 globulin and γ globulin in different gender and age groups (≤20, 21-30, 31-40, 41-50, 51-60, 61-70, 71-80, 81-90, ≥91 years old) were compared. A total of 10 014 cases were selected by immunofixation electrophoresis (IFE). The positive detection rates of different SPE bands and IFE bands were analyzed. The sensitivity and specificity of SPE methods were determined according to IFE results as the gold standard.Results:No significant difference was examined in the proportion of SPE bands between different genders ( P>0.05). There were statistically significant differences in the proportion of albumin bands between apparently healthy individuals and hospitalized patients at different ages (apparently healthy individuals: F=5.12, P<0.05, inpatients: F=4.18, P<0.05), and all of them decreased with the increase of age. The proportion of γ globulin bands increased with age (apparently healthy individuals: F=1.34, P<0.05; inpatients: F=1.24, P<0.05). The sensitivity of SPE was 69% (2 098/3 051), and the specificity was 97% (6 721/6 963). Compared with IFE method, the positive detection rate of monoclonal gammopathy was significantly different (χ2=5 049.94, P<0.05). The positive rate of monoclonal gammopathy in γ globulin region (21.11%, 2 114/10 014) was higher than that in β globulin region (3.28%, 328/10 014) (χ2=90.74, P<0.05) and β-γ globulin region (1.63%, 163/10 014) (χ2=44.34, P<0.05). IgG and IgM bands are common in γ globulin region. Among them, IgG-κ type accounted for 94.1% (995/1 058), IgG-λ type accounted for 94.8% (690/728), IgM-κ type accounted for 89.2% (222/249), IgM-λ accounted for 83.8% (62/74). IgA bands are common in β region, of which IgA-κ accounted for 49.8% (103/207) and IgA-λ accounted for 51.6% (149/289). The positive rate of monoclonal gammopathy of IgG-κ type was the highest (10.57%, 1 058/10 014), and the positive rate of monoclonal gammopathy of IgM-λ type was the lowest (0.74%, 74/10 014). Conclusions:With increasing age, the proportion of albumin band in SPE decreased and the proportion of γ globulin band increased. IgG and IgM type monoclonal gammopathy is mostly found in the gamma region, with a higher detection rate in IgG type. IgA type monoclonal gammopathy is mostly found in the β region, with a lower detection rate.

Objective:To investigate the characteristics and distribution of monoclonal gammopathy in clinical patients.Methods:A total of 936 171 patients (508 449 males and 427 722 females) who received capillary zone electrophoresis in Zhongshan Hospital Affiliated to Fudan University from January 2012 to December 2021 were selected, from which 14 945 patients with abnormal bands were screened as the study subjects, including 10 173 males and 4 772 females and the age 21-102 (65±13) years old. According to the age, patients were divided into 8 groups: 21-30 years old (168 cases), 31-40 years old (405 cases), 41-50 years old (1 326 cases), 51-60 years old (3 068 cases), 61-70 years old (4 985 cases), 71-80 years old (3 288 cases), 81-90 years old (1 519 cases), and≥91 years old (186 cases). The diagnostic results of the 14 945 patients with abnormal bands were collected and were divided into tumor group (5 196 cases) and non-tumor disease group (9 749 cases) according to the presence of tumor. The distribution of abnormal bands in different gender, age, and disease groups were retrospectively analyzed. Among the 14 945 patients, 4 988 cases underwent immunofixation electrophoresis, excluding 336 negative cases and 412 cases of double clonal bands reaction, and 4 240 patients with monoclonal immunoglobulin (M protein) reaction were selected as the study subjects, including 2 794 males and 1 446 females aged 21-102 (67±12) years old. They were divided into 8 groups according to the age: 21-30 years old (18 cases), 31-40 years old (91 cases), 41-50 years old (364 cases), 51-60 years old (862 cases), 61-70 years old (1 455 cases), 71-80 years old (904 cases), 81-90 years old (486 cases), and≥91 years old (60 cases). The diagnostic results and immunoglobin subtypes (IgA-κ, IgA-λ, IgG-κ, IgG-λ, IgM-κ, IgM-λ, κ, λ) of patients were collected, and the distribution of monoclonal gammopathy in different gender, age and disease groups were retrospectively analyzed.Results:Among 936 171 patients, 14 945 cases showed abnormal bands in electropherograms with a detection rate of 1.60%; the detection rates of abnormal bands in males and females were 2.00% (10 173/508 449) and 1.12% (4 772/427 722), respectively, with a statistically significant difference ( P<0.01). There was a significant difference in the detection rate of abnormal bands among different age groups ( P<0.01); among them, the highest detection rate of abnormal band in group of ≥91 years old was 5.98%, and the ratio of male to female was 1.67∶1. Among the 14 945 cases of abnormal bands, patients aged 51-60, 61-70 and 71-80 accounted for 20.53% (3 068 cases), 33.36% (4 985 cases) and 22.00% (3 288 cases), respectively, and the differences among the age groups were statistically significant (χ 2=115.82, P<0.01). In the tumor group, the top 3 tumors with abnormal bands were plasmacytoma with 1 123 cases, lymphoma with 289 cases, and leukemia with 49 cases. The detection rate of abnormal bands in electropherograms of plasmacytoma was 89.92% (1 123/1 249), which was higher than that in lymphoma and leukemia [6.73% (289/4 296) and 6.40% (49/766), respectively, P<0.01]. Among 4 240 patients with positive M protein, the proportion of 51-60, 61-70 and 71-80 years old patients were 20.33% (862/4 240), 34.32% (1 455/4 240) and 21.32% (904/4 240), respectively, and the differences among age groups were statistically significant ( P<0.01). The results of M protein types showed that the proportion of IgG-κ type was the highest in both genders, with 32.28% (902/2 794) in males and 34.30% (496/1 446) in females. In the 21-30, 31-40, and 41-50 age groups, the proportion of IgG-λ was the highest, which were 38.89% (7/18), 36.26% (33/91) and 34.07% (124/364) in these groups respectively. However, the proportions of IgG-κ were the highest in either of the 51-60, 61-70, 71-80, 81-90 and ≥91 years old groups, which were 33.87% (292/862), 34.16% (497/1 455), 31.53% (285/904), 34.57% (168/486), 28.33% (17/60), respectively, and the differences among all age groups and gender groups had statistical significance ( P<0.01). Among patients with positive M protein in the tumor group, plasmacytoma accounted for 14.22% (603/4 240), followed by lymphoma 6.30% (267/4 240); among non-tumor diseases, M proteinemia accounted for the highest proportion (7.24%, 307/4 240), followed by pulmonary infection (5.47%, 232/4 240). Conclusions:The detection rate of abnormal bands in capillary zone electrophoresis may increase with age, and is higher in males than in females in the same age group; different malignant tumor diseases can also show abnormal bands in capillary zone electrophoresis, but they are still mainly hematological tumors. Among the positive results of M protein, 61-70 years old group accounts for the highest proportion; the most common type of monoclonal gammopathy is IgG type; in the age group of 21-50 years, the proportion of IgG-λ type is the highest; in the group of >50 years old, the proportion of IgG-κ type is the highest; in the diagnosis of positive monoclonal gammopathy, the top 3 diseases are all hematological diseases, including plasmacytoma, monoclonal gammopathies and lymphoma.

Objective To investigate the predictive value of white blood cell count,LDL-C and fasting blood glucose on the occur-rence of AMI in elderly patients with coronary atherosclerotic heart disease(CAD).Methods A total of 210 elderly patients with CAD were selected and divided into the AMI group as the case group and the chronic CAD group as the control group based on clinical manifes-tations and laboratory and coronary angiography results.The general data,laboratory data,coronary angiography results,and other clinical information of the two groups were retrospectively analyzed.We used the Logistic regression analysis to explore the risk factors for acute myocardial infarction in elderly CAD patients and the receiver operating characteristic curve(ROC)to evaluate the predictive efficacy of white blood cell count,LDL-C and fasting blood glucose on the occurrence of AMI in elderly CAD patients.Results Compared with the chronic CAD group,the AMI group had a higher smoking rate,white blood cell count,CRP,ALT,AST,D-D dimer,fasting blood glu-cose,total cholesterol,LDL-C and Gensini score and lower LVEF(P＜0.05).The Logistic regression analysis showed that white blood cell count(β=0.859,P＜0.001,OR=2.361),LDL-C(β=1.003,P=0.009,OR=2.728)and fasting blood glucose(β=0.495,P=0.007,OR=1.641)were independent risk factors for AMI in elderly CAD patients.The ROC curve analysis showed that when the cut-off values of fasting blood glucose,white blood cell count,and LDL-C were 4.895mmol/L,7.19 x 109/L and 2.635 mmol/L respectively,the area under the curve for predicting AMI was the largest,with values of 0.734,0.859 and 0.735.The area un-der the ROC curve of the combination of the three is 0.902.Conclusion White blood cell count,LDL-C and fasting blood glucose are associated with AMI in elderly patients with CAD.Higher levels of white blood cell count,LDL-C and fasting blood glucose indicate a higher risk of AMI.The combination of the three can serve as a more valuable predictive index.

Objective To understand how warfarin inhibition is hindered by the vitamin K epoxide reductase(VKORC1)resistant mutations.Methods An electrophoretic mobility assay was conducted based on the principle that binding with warfarin resulted in changes in VKORC1 electrophoretic mobility.The activity of VKORC1 and the half maximal inhibitory concentration(ICs0)of warfarin inhibiting VKORC1 were detected by ELISA.These experiments evaluated the binding ability of VKORC1 resistant mutants to warfarin.Results With warfarin bound,VKORC1 was protected from N-ethylmaleimide(NEM)modification and showed increased electrophoretic mobility,which was dependent on an unmodified cysteine on the flexible transmembrane helix 1(TM1)of VKORC1.In-creasing the warfarin concentration could shift more VKORC1 towards the species with the fast mobility.Compared to the wildtype VKORC1,the fast mobility fraction became less or disappeared in warfarin-resistant mutants,indi-cating weakened binding of warfarin.In addition,VKORC1 mutants with the weaker electrophoretic mobility shift indicated the stronger warfarin resistant.Conclusion Weakened warfarin binding is the primary cause of warfarin resistance.

Objective:To retrospectively compare the clinical effects of the 2021 Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation and the European Kidney Function Consortium (EKFC) equation based on creatinine to estimate glomerular filtration rate (eGFRcr) in a Chinese population.Methods:Patients and healthy individuals who visited the Zhongshan Hospital, Fudan University, between January 1, 2023, and December 31, 2023, were enrolled. The exclusion criteria were age<18 years, amputees, pregnant women, patients with muscle-related diseases, and patients who had undergone ultrafiltration or dialysis. The final study population included 660 078 individuals with a median age of 58 years (44, 68), 56.51% (372 987/660 078) of them were men. eGFRcr was calculated using the 2021 CKD-EPI, EKFC equations and initial 2009 CKD-EPI equation. Results were compared by differences (P 10), charges in population distribution and impact on staging of chronic kidney disease (CKD) by gender, age and creatinine results. Results:Compared with the results of 2009 and 2021 CKD-EPI eGFRcr, the P 10was 99.99% ( R 2=0.997). When applying the 2021 CKD-EPI eGFRcr equation, there was a slight overestimation of 3.77 (2.92, 4.47) ml/(min×1.73 m 2). 86.42% (570 455/660 078)participants had higher eGFRcr owing to the utilization of the 2021 CKD-EPI equation, which did not cause CKD stage change. A total of 12.35% (81 514/660 078) of subjects had different CKD stage with the higher 2021 CKD-EPI eGFRcr. 1.23% (8 109/660 078) had lower eGFRcr but no change in the CKD stage with the 2021 equation. Compared to the 2009 CKD-EPI eGFRcr results, the EKFC eGFRcr demonstrates P 10 values of 89.79%, with a correlation coefficient of R 2=0.976. The overall eGFRcr values using the EKFC equation were decreased by 4.63(-2.22, -7.04) ml/(min×1.73 m 2). There were 75.14% (495 992/660 078) individuals whose eGFRcr results were lower in EKFC equation, which did not cause CKD stage change. There were 15.63% (103 161/660 078) leading to a different CKD stage. Additionally, 8.80% (58 077/660 078) individuals with a higer eGFRcr result did not change CKD stage, while 0.43% (2 848/660 078) experienced a change in CKD staging due to the overestimation. Conclusions:Compared with the 2009 CKD-EPI eGFRcr results, the 2021 CKD-EPI eGFRcr results were slightly higher and had good consistency in different genders and ages. However, the EKFC eGFRcr was generally lower than 2009 CKD-EPI eGFRcr, showing poor consistency. The differences between eGFR equations should be considered when judging the CKD stage of patients.

Objective:To develop a prototype artificial intelligence immunofluorescence image recognition system for classification of antinuclear antibodies in order to meet the growing clinical requirements for an automatic readout and classification of immunof luorescence patterns for antinuclear antibody (ANA) images.Methods:Immunofluorescence images with positive results of ANA in Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine from April 2020 to December 2021 were collected. Three senior technicians independently and in parallel interpreted the Immunofluorescence images to determine the ANA results. Then the images were labeled according to the ANA International Consensus on Fluorescence Patterns (ICAP) classification criteria. There were 7 labeled groups: Fine speckled, Coarse speckled, Homogeneous, nucleolar, Centromere, Nuclear dots and Nuclear envelope. Each group was randomly divided into training dataset and validation dataset at a ratio of 9∶1 by using random number table. On the deep learning framework PyTORCH 1.7, the convolutional neural network (CNN) training platform was constructed based on ResNet-34 image classification network, and the automatic ANA recognition system was established. After the model was established, the test set was set up separately, the judgment results of the model were output by ranking the prediction probability, with the results of the 2 senior technicians was taken as "golden standard". Parameters such as accuracy, precision, recall and F1-score were used as indicators to evaluate the performance of the model.Results:A total of 23138 immunofluorescence images were obtained after segmentation and annotation. A total of 7 models were trained, and the effects of different algorithms, image processing and enhancement methods on the model were compared. The ResNet-34 model with the highest accuracy andswas selected as the final model, with the classification accuracy of 93.31%, precision rate of 91%, and recall rate of 90% and F1-score of 91% in the test set. The overall coincidence rate between the model and manual interpretation was 90.05%, and the accuracy of recognition of nucleolus was the highest, with the coincidence rate reaching 100% in the test set.Conclusion:The current AI system developed based on deep learning of the ANA immunofluorescence images in the present study showed the ability to recognize ANA pattern, especially in the common, typical, simple pattern.

Objective:To explore the ability of sequential immunization regimen inducing neutralizing antibodies against severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) Wuhan-Hu1 and Omicron variants in mice.Methods:Groups of 6-8-week-old BALB/c mice were primed with two doses of Wuhan-Hu-1 inactivated vaccine, and then boosted with Omicron or Wuhan-Hu-1 inactivated vaccine, respectively. Binding antibodies were tested by enzyme linked immunosorbent assay; and neutralizing antibodies against Wuhan-Hu1 and Omicron variants were analyzed by vesicular stomatitis virus pseudovirus assay system; SARS-CoV-2 specific cellular immune responses were quantified by enzyme-linked immunosorbent spot assay.Results:IgG antibodies against Wuhan-Hu1, Delta and Omicron RBD were enhanced after the second dose of Wuhan-Hu1 inactivated vaccine. Compared with Wuhan-Hu1 inactivated vaccine, the group boosted with Omicron inactivated vaccine improved Wu-RBD and Omic-RBD specific IgG antibodies 1.41 and 1.26 times, and serum neutralizing antibodies against BA.1, BA.2, BA.4/5 and BF.7 were elevated 4.5, 3.4, 12.1 and 6.5 folds, respectively, by sequential immunization. After booster immunization with inactivated Wuhan-Hu1 or Omicron vaccines, Wu-RBD IgA titer was significantly higher than that of one dose inactivated Wuhan-Hu1 vaccine ( P=0.005 7, P=0.006 1). Conclusions:Neutralizing antibodies against Omicron variants were enhanced by sequential immunization with Omicron inactivated vaccine. Specific IgA was significantly enhanced after the third dose of inactivated vaccine.

Objective:To evaluate the effects of the alpha-lipoic acid on hepatic ischemia/reperfusion (I/R) injury and the role of (ALDH2).Methods:This experiment was performed in two parts in vivo and in vitro experiments. In vivo experiment Twenty-four male Sprague-Dawley rats, aged 8-10 weeks, weighing 250-300 g, were divided into 4 groups ( n=6 each) by the random number table method: sham operation group (Sham group), hepatic I/R group (IR group), and hepatic I/R plus α-lipoic acid group (IR+ ALA group) and hepatic I/R+ α-lipoic acid+ daidzin group (IR+ ALA+ D group). Hepatic I/R was induced by occlusion of the left and middle hepatic lobes for 60 min, followed by 6 h of reperfusion in anesthetized rats.In IR+ ALA+ D group, ALDH2 inhibitor daidzin 50 mg/kg was intraperitoneally injected at 45 min before ischemia.Alpha-lipoic acid 100 mg/kg was intraperitoneally injected at 30 min before ischemia in IR+ ALA group and IR+ ALA+ D group.Blood samples from the inferior vena cava were collected at the end of reperfusion to determine serum AST and ALT activities.Then the rats were sacrificed, and livers were removed for microscopic examination of pathological changes of the lung tissues which were scored and for determination of ALDH2 activity, level of reactive oxygen species (ROS) and expression of 4-hydroxy-trans-2-nonenal (4-HNE) and malondialdehyde (MDA) (by immuno-histochemistry). In vitro experiment Rat BRL-3A hepatocytes cultured in vitro were divided into 4 groups ( n=15 each) by the random number table method: control group (C group), hypoxia-reoxygenation group (HR group), hypoxia-reoxygenation+ α-lipoic acid group (HR+ ALA group) and hypoxia-reoxygenation+ α-lipoic acid+ daidzin group (HR+ ALA+ D group). BRL-3A hepatocytes were exposed to 95% N 2-5% CO 2 in an incubator at 37 ℃ for 6 h followed by reoxygenation with 95% O 2-5% CO 2 for 24 h. At 60 min before hypoxia, alpha-lipoic acid 100 μmol/L was addded in HR+ ALA group, and alpha-lipoic acid 100 μmol/L and daidzin 60 μmol/L were added in HR+ ALA+ D group.At 24 h of reoxygenation, cell viability was measured by CCK-8 method, ALDH2 activity was determined by spectrophotometry, ROS level was detected by DCFH-DA fluorescent probe method, and mitochondrial membrane potential (MMP) was measured by JC-1 method. Results:In vivo experiment Compared with Sham group, the serum AST and ALT activities, liver injury score, level of ROS in liver tissues and expression of 4-HNE and MDA were significantly increased ( P<0.05), and no significant change was found in ALDH2 activity in IR group ( P>0.05). Compared with IR group, the serum AST and ALT activities, liver injury score, level of ROS in liver tissues and expression of 4-HNE and MDA were significantly decreased, and the ALDH2 activity was increased in IR+ ALA group ( P<0.05). Compared with IR+ ALA group, the serum AST and ALT activities, liver injury score, level of ROS in liver tissues and expression of 4-HNE and MDA were significantly increased, and the ALDH2 activity was decreased in HR+ ALA+ D group ( P<0.05). In vitro experiment Compared with C group, the cell viability and MMP were significantly decreased, and the level of ROS was increased ( P<0.05), and no significant change was found in the activity of ALDH2 in HR group ( P>0.05). Compared with HR group, the cell viability, ALDH2 activity and MMP were significantly increased, and the level of ROS was decreased in HR+ ALA group ( P<0.05). Compared with HR+ ALA group, the cell viability, ALDH2 activity and MMP were significantly decreased, and the level of ROS was increased in HR+ ALA+ D group ( P<0.05). Conclusions:Alpha-lipoic acid can reduce hepatic I/R injury in rats, and the mechanism is related to activation of ALDH2, reduction of accumulation of toxic aldehyde and restoration of MMP.

Objective To develop plantar force model of patellofemoral pain (PFP), so as to provide theoretical references for the assessment of PFP rehabilitation. Methods The case-control study was conducted, and a total of 126 patients with PFP and 126 healthy controls matched by gender and age were enrolled in the study. The participants were tested for plantar force and pressure during level walking, and twelve plantar regions were divided and recorded. Whether the participants suffered PFP was analyzed as dependent variable, meanwhile the peak force and peak pressure in 12 plantar regions of participants at selected speed during level walking were analyzed as independent variables. Conditional logistic regression (CLR) equations of peak force and peak pressure with PFP were established, respectively. The receiver-operating characteristic (ROC) curve of the corresponding equations was derived, and the area under ROC curve was calculated to analyzed the validity of different equations on PFP assessment. Results The CLC equation of peak force in 12 plantar regions of the participants with FFP was constructed, and only peak force of lateral heel was in the equation. The CLC equation of peak pressure in each plantar region included medial heel, midfoot, 1st and 2nd metatarsals. Meanwhile, the area under ROC curve of the pressure equation was larger than that of the force equation. Conclusions Peak force and pressure at different plantar regions can be used to assess PFP during level walking, and peak pressure is more effective for assessment.

OBJECTIVE: To study the effect of rapamycin on scar formation in rabbit eyes following filtering operation and explore the possible mechanism. METHODS: Ninety-six healthy adult rabbits were subjected to trabeculectomy of the left eye and subsequently randomly divided into 4 groups (=24) for treatment with castor oil (control) or rapamycin (1%, 3%, or 5%) eye drops of the operated eyes 4 times a day. The morphology and function of the filtering blebs of the rabbits were compared at 7, 14, 21 and 28 days after the operation; at each of the time points, 6 rabbits from each group were euthanized for detection of expressions of proliferating cell nuclear antigen (PCNA) and α-smooth muscle actin (α-SMA) in the tissues in the surgical area using immunohistochemistry. Cultured rabbit subconjunctival fibroblasts (RTFSs) were treated with different concentrations of rapamycin (0.06, 0.25, 1, and 4 mg/L) and the cell apoptosis was detected using flow cytometry. RESULTS: In the first, second and third weeks after the operation, the rate of functional follicle formation was significantly higher in the 3 rapamycin groups than in the control group ( < 0.05), and the number of α- SMA-positive fibroblasts decreased over time in the 3 rapamycin groups. In cultured RTFSs, treatment with rapamycin at different concentrations resulted in increased apoptosis of the cells, and rapamycin above 0.25 mg/L significantly increased the cell apoptosis in a dose-dependent manner. CONCLUSIONS Rapamycin can inhibit hyperplasia of the filtering passage tissue, helps to preserve the functional filtering blebs and prolong their life span, and induces apoptosis of RTFS.