Objective:Inflammation has been confirmed to play an important role in the occurrence and development of sudden sensorineural hearing loss（SSNHL）, and the neutrophil-to-lymphocyte ratio（NLR） is a biomarker positively correlated with the degree of inflammation. This study aims to identify the difference in serum NLR between patients with SSNHL and normal population, and to evaluate the predictive efficacy of NLR for the occurrence and prognosis of SSNHL, thereby guiding the clinical diagnosis and treatment of SSNHL. Methods:In this study, 96 patients diagnosed with SSNHL admitted to our department from January 2023 to March 2024 and 96 patients diagnosed with vocal cord polyps admitted to our department during the same period were recruited as a control group. Multivariate Logistic regression was used to evaluate independent related factors, and a nomogram was constructed to predict the probability of SSNHL. The receiver operating characteristic（ROC） curve and calibration curve were used to evaluate the accuracy of prediction. Results:Multivariate logistic regression analysis showed that a high level NLR（OR2.215; 95%CI1.597-3.073; P<0.001） were independently associated with the presence of SSNHL. High age（OR1.036; 95%CI1.009-1.067; P=0.012）, high FIB（OR2.35; 95%CI1.176-4.960; P=0.019） were the risk factor for SSNHL. Incorporating these 3 factors, a forest plot and a nomogram were generated. The ROC curve, nomogram and calibration curve showed that the model had good clinical practicability. A low NLR（OR0.598; 95%CI0.439-0.816; P<0.001） was significantly associated with a favorable prognosis of SSNHL. Conclusion:Elevated NLR can serve as an promising biomarker for assessing the risk of SSNHL. The nomograms calculation model may be utilized as a tool to estimate the probability of SSNHL. Low level NLR is significantly associated with a good prognosis of SSNHL.
Humans ; Neutrophils ; Female ; Male ; Lymphocytes ; Hearing Loss, Sensorineural/blood* ; Hearing Loss, Sudden/diagnosis* ; Middle Aged ; Prognosis ; Nomograms ; ROC Curve ; Adult ; Logistic Models ; Biomarkers/blood* ; Lymphocyte Count ; Inflammation/blood* ; Clinical Relevance

【Objective】 To investigate the level of serum bone metabolism and biochemical markers and bone density of plasmapheresis donors, and to provide scientific basis for ensuring the health and safety of plasmapheresis donors in China. 【Methods】 A total of 437 plasmapheresis donors from Linwu plasmapheresis station in Hunan Province from July 1 to September 30, 2022 were recruited to determine the levels of total serum calcium, albumin, serum 25-hydroxyvitamin D (25OHD), serum type I procollagen N-terminal propeptide (P1NP), and collagen type 1 crosslinked carboxyl-terminal peptide (β-CTX). Dual-energy X-ray method was used to measure the bone density of the anteroposterior lumbar spine (L1-L4) and bilateral femoral neck bone density of plasmapheresis donors. Plasmapheresis donors were grouped according to the type of plasma donation (first-time and repeat plasmapheresis donors) and the total number of plasma donations to assess the differences in bone density and serum bone metabolism biochemical markers between groups. The dose-response relationship between the total number of plasmapheresis donations and biochemical indexes was analyzed by limiting cubic spline, and the influencing factors of different indexes were explored by multiple linear regression. 【Results】 A total of 437 plasmapheresis donors were included in this study, including 187 first-time plasmapheresis donors and 250 repeat plasmapheresis donors. There were no significant differences in bone density and prevalence of osteoporosis between first-time donors and repeat donors (P>0.05). There was also no significant difference in bone density levels between groups of total number of plasmapheresis donations. The levels of albumin and 25OHD decreased with the increase of the total number of plasma donations, while the serum P1NP level was positively correlated with the total number of plasma donations. The results of the restriction cubic spline showed that the total number of plasmapheresis donations had a nonlinear dose-response relationship with 25OH and P1NP (P<0.05). The results of multiple linear regression showed that the frequency of plasmapheresis donation was the influencing factor of 25OHD, and the total number of plasmapheresis donation was the influencing factor of P1NP. 【Conclusion】 Plasmapheresis donation does not affect the bone health of donors and increase the risk of osteoporosis due to the use of long-term anticoagulants, but it will increase the osteogenic activity of plasmapheresis donors. It is recommended that middle-aged and elderly plasmapheresis donors supplement vitamin D appropriately.

【Objective】 To compare the desalination effects of five desalination methods and their effects on the components for human coagulation factor Ⅷ(FⅧ), and provide reference for selection of protein desalination methods. 【Methods】 Sephadex G-25 Medium gel, Fractogel EMD BioSEC gel, ultrafiltration, room temperature dialysis and 4℃ dialysis were used to desalt human FⅧ. The desalination effect was evaluated by the removal rate of Na +, citrate ion and glycine. FⅧ protein recovery, FⅧ activity (FⅧ∶C), VWF antigen (VWF∶Ag), VWF activity(VWF∶Ac), VWF polymers and SDS-PAGE analysis before and after desalination were compared to evaluate the effect of desalination on FⅧ components. 【Results】 In terms of desalination effect, the removal rate of Na⁺ was the lowest in ultrafiltration desalination, while that of Fractogel EMD BioSEC gel was the highest [(97.90±0.06) % vs (99.82±0.07) %]. Except that there was no statistical significance between Sephadex G-25 Medium gel desalination and Fractogel EMD BioSEC gel desalination (P=0.90), the removal rates of the other four methods were statistically significant. The removal rate of glycine was the lowest in ultrafiltration desalination, wihle that of Fractogel EMD BioSEC gel desalination was the highest [(95.78±0.42) % vs (99.81±0.08) %]. Significant difference in glycine removal was noticed in ultrafiltration desalination, but not among the other four desalination methods. There was no significant difference in the removal rate of citrate ions among the five methods (P=0.85). For the effect of FⅧ components, FⅧ∶C, VWF∶Ag, VWF∶Ac and protein recovery rates of ultrafiltration desalination were the highest, with (18.34±1.99) IU/mL, (11.81±0.33) IU/mL, (12.26±0.58) IU/mL and (97.13±1.37) %, respectively. There was no significant change in VWF∶Ac/VWF∶Ag before and after desalination by the five methods. SDS-PAGE and VWF polymer analysis showed that different desalination methods had no significant impact on protein composition. 【Conclusion】 Although different desalination methods had no significant effect on the composition of FⅧ protein, the desalination effect was different. Moreover, different desalination methods had significant effects on protein recovery, FⅧ∶C, VWF∶Ag and VWF∶Ac. The selection of desalination methods should be more considered during protein processing,

Objective To investigate the binding and carrying effects of human serum albumin(HSA)from various sources on sphingosine-1-phosphate(S1P).Methods Utilizing human plasma-derived HSA(pHSA)and recombinant HSA(rHSA)samples as the focal points of our investigation,LC-MS/MS technology was employed to meticulously compare and an-alyze the disparities in S1P content among the aforementioned samples.Subsequently,under physiological concentration condi-tions,S1P was directly introduced to HSA samples for loading processing,facilitating a comprehensive comparison of the bind-ing efficacy of HSA from different sources to S1P.Within a serum-free culture setting,HSA samples from various sources were co-cultured with HUVEC cells.The alterations in S1P content within the cell culture supernatant across different treatment groups were meticulously analyzed,allowing for a nuanced comparison of the S1P carry effects exerted by HSA from different sources on cells.The interaction between HSA and S1P molecules from different sources was analyzed and their affinity was cal-culated using surface plasmon resonance(SPR)technology.Furthermore,leveraging AutoDock Vina software and the Mol-prophet platform,the molecular docking analysis of HSA and S1P was conducted,aiming to predict the key binding pocket do-main of S1P within HSA.Results All pHSA samples exhibited detectable levels of S1P(ranging from 3.31±0.03 to 30.35±0.07 μg/L),with significant variations observed among pHSA samples from different manufacturers(P＜0.001).Conversely,S1P was undetectable in all rHSA samples.Upon load treatment,the binding affinity of HSA from diverse sources to S1P dem-onstrated significant discrepancies(P＜0.001),with rHSA exhibiting approximately double the average S1P loading compared to pHSA(ΔCrHSA=801.75±142.45 μg/L vs ΔCpHSA=461.94±85.73 μg/L;P＜0.001,t=5.006).Co-culture treatment out-comes revealed a significant elevation in S1P concentration within the supernatant after 6 hours of co-culture across all HSA sample processing groups with HUVEC cells,while no changes were observed in the supernatant of the blank control group.Notably,significant differences in supernatant S1P concentration were observed among treatment groups at 6 h,12 h,and 24 h(P＜0.001).SPR analysis unveiled a stronger affinity of pHSA for S1P compared to rHSA(KDpHSA-S1P:2.38E-06,KDrHSA-S1P:3.72E-06).Molecular docking analysis and binding pocket prediction suggested that the key binding pocket of HSA and S1P may reside in the IB subdomain of the HSA molecule.Conclusion HSA from various sources exhibits distinct binding and carrying effects on S1P,which appear to be closely associated with the IB subdomain of the HSA molecule.

The aim of this study was to prepare tandem multimeric proteins of BmSPI38, a silkworm protease inhibitor, with better structural homogeneity, higher activity and stronger antifungal ability by protein engineering. The tandem multimeric proteins of BmSPI38 were prepared by prokaryotic expression technology. The effects of tandem multimerization on the structural homogeneity, inhibitory activity and antifungal ability of BmSPI38 were explored by in-gel activity staining of protease inhibitor, protease inhibition assays and fungal growth inhibition experiments. Activity staining showed that the tandem expression based on the peptide flexible linker greatly improved the structural homogeneity of BmSPI38 protein. Protease inhibition experiments showed that the tandem trimerization and tetramerization based on the linker improved the inhibitory ability of BmSPI38 to microbial proteases. Conidial germination assays showed that His₆-SPI38L-tetramer had stronger inhibition on conidial germination of Beauveria bassiana than that of His₆-SPI38-monomer. Fungal growth inhibition assay showed that the inhibitory ability of BmSPI38 against Saccharomyces cerevisiae and Candida albicans could be enhanced by tandem multimerization. The present study successfully achieved the heterologous active expression of the silkworm protease inhibitor BmSPI38 in Escherichia coli, and confirmed that the structural homogeneity and antifungal ability of BmSPI38 could be enhanced by tandem multimerization. This study provides important theoretical basis and new strategies for cultivating antifungal transgenic silkworm. Moreover, it may promote the exogenous production of BmSPI38 and its application in the medical field.
Animals ; Antifungal Agents/pharmacology* ; Escherichia coli/metabolism* ; Proteins/metabolism* ; Protease Inhibitors/chemistry* ; Bombyx/chemistry* ; Saccharomyces cerevisiae/metabolism* ; Peptide Hydrolases

【Objective】 To develop methods to display the IgG autoantibody repertoire of intravenous immunoglobulin (IVIG) products, analyze the different types of antibodies and study the diversity of IgG autoantibody in 4 IVIG preparations from different Chinese manufacturers. 【Methods】 Two-dimensional gel electrophoresis and immunoblotting with human umbilical vein endothelial cell (HUVEC) proteins were used to demonstrate the IgG autoantibody repertoire and the human protein microarray with bioinformatics analysis was employed to profile the immune reactive autoantigens of the 4 IVIG preparations. 【Results】 The methods to showcase the autoantibody repertoire and study the antibody diversity of IVIG were successfully established. High-quality repertoires of IVIG autoantibodies and biological information about self-proteins that can be recognized were obtained. There was a significant difference in the recognition of the quantity and variety of the self-antigens by different IVIG products. The number of antibodies against HUVEC proteins in four products ranged from 241-386. The number of proteins recognized on the human protein chip ranged from 292-435, with 172 human self-proteins recognized by all four products. 【Conclusion】 Demonstration of antibody repertoire and protein chip technology can be used to analyze IVIG products′ IgG autoantibody repertoire. All four preparations tested in this study exhibited a broad spectrum of antibodies against HUVEC proteins and human proteome microarray, each product had its unique antibody repertoire characteristics.

【Objective】 To explore the risk of Alzheimer′s disease (AD) transmitted by blood transfusion. 【Methods】 There were 10 APP/PS1 mice of 3, 6 and 9 months old, half female and half male, and the cognitive and behavioral abilities of C57 mice of the same age were measured, and the blood of the oldest APP/PS1 mice with no behavioral changes were collected to detect the contents of Aβ40 and Aβ42. The polymers Aβ40 and Aβ42 were prepared and Western blotting analysis was conducted. Kunming mice aged from 6 to 7 months were randomly divided into 6 groups (10 mice/ group, half male and half female). The blood of APP/PS1 mice was injected intravenously in experimental group 1-2(100 μL/mouse) with high frequency injection (3 times/week) and low frequency injection (1 time/week), respectively. In experimental group 3-4, Aβ40 and Aβ42 polymerized mixture (100 μL/mouse) were injected in high frequency and low frequency, respectively. The control group 1-2 was injected with the same amount of normal saline, with high frequency and low frequency, respectively. The above groups were injected for 4 weeks, and the cognitive and behavioral abilities were tested and analyzed one week after injection. Finally, the contents of Aβ40 and Aβ42 in blood of Kunming mice were detected. 【Results】 Change in cognitive and behavioral ability showed in 9 months old APP/PS1 mice, but not in 3 and 6 months old APP/PS1 mice. The contents of Aβ40 and Aβ42 (pg/mL) in blood of 6-7 months old APP/PS1 mice were 418.40±2.18 and 15.68±0.20, respectively. Except for monomers, most of the polymerized mixtures of Aβ40 and Aβ42 were dimers and trimers. In both high frequency and low frequency, Kunming mice transfused with blood of APP/PS1 mice (experimental group 1-2) showed a certain degree of anxiety-like behavior and short-term memory shortening in open-field test and conditioned fear test, but without significant difference. There was no significant difference in open field test, new object recognition, Barnes maze and cognitive behavior analysis of conditioned fear between experimental group 3-4 and the control group. The levels of blood Aβ40 and Aβ42(pg/mL) of Kunming mice detected by ELISA were 10.30±0.08 and 3.360±0.005, respectively, and there was no significant difference between the two groups. 【Conclusion】 Blood transfusion of APP/PS1 mice and the mixture of Aβ40 and Aβ42 have no significant effect on the cognitive function of healthy Kunming mice in a short time, and the risk of AD transmission is relatively low.

【Objective】 To investigate the effect of immunoglobulin G (IgG) dimer concentration of intravenous immunoglobulin (IVIG) on the binding ability of IgG Fc fragment to THP-1 cell surface receptors. 【Methods】 Firstly, protein purification and high performance liquid chromatography (HPLC) were used to prepare different concentrations of IgG dimers. After that, IgG dimer was added to IVIG to prepare IVIG containing different concentrations of IgG dimer. Finally, based on the method established in our laboratory, we analyzed the effect of IgG dimer concentration in IVIG on the binding ability of IgG Fc fragment to THP-1 cell surface receptors. 【Results】 When the concentration of IgG dimer in IVIG was 1.11%-10.30%, its binding ability to Fc receptors on the surface of THP-1 cell was 97.67%-135.33%, and this binding ability was positively correlated with the concentration of IgG dimer. When the IgG dimer concentration exceeded 13.22%, the binding ability had no correlation with the IgG dimer concentration. 【Conclusion】 A certain concentration of IgG dimer can promote the binding ability of the IgG Fc fragment in IVIG to receptors on the surface of THP-1 cells, which needs further verification from animal experiments and clinical data.

【Objective】 To investigate the effect of intravenous immunoglobulin(IVIG) with different Aβ antibody content on the cognitive function of Alzheimer′s disease model mice. 【Methods】 IVIG from 8 domestic blood products companies were selected. Enzyme-linked immunosorbent assay(ELISA) was used to detect the content of Aβ40/42 antibody. Three kinds of IVIG with high, middle and low Aβ42/40 antibody levels were selected to treat 3xTg-AD mice. Forty 3-month-old 3xTg-AD mice were randomly divided into 4 groups with 10 mice in each group(half male and half female). Three treat groups were intraperitoneally injected with three kinds of IVIG with 1g·kg^-1 for 12 weeks(twice a week). The controls were injected with the same volume of saline. Behavioral tests were performed immediately by using the mouse behavior analysis system after a total of 24 injections. 【Results】 The concentrations of antibodies(μg/mL) against Aβ40 monomer in IVIG ranged 0.7±0.05 to 3.1±0.05, concentrations of antibodies against Aβ40 oligomer ranged 11.7±0.7 to 32.0±2.2, concentrations of antibodies against Aβ42 monomer ranged 1.8±0.1 to 27.9±0.3, and concentrations of antibodies against Aβ42 oligomeric ranged 2.3±0.1 to 49.4±2. High(IVIG-1), medium(IVIG-8) and low(IVIG-6) IVIG were selected for mice study. In the open field test, the time of four groups of mice entering the central area(s) was 0.5±0.9, 23.4±6.1(P<0.0001), 4.6±2.8 and 2.6±2.3, respectively; the number of feces(grains) was 1.6±0.7(P<0.0001), 1.2±0.4(P<0.0001), 2.4±0.5(P<0.001) and 3.8±0.8, respectively. In the novel object recognition test, the scores of exploring new objects were 71.3±29.5(P<0.05), 71.8±20.5(P<0.05), 75.9±26.9(P<0.01) and 25.6±23.7, respectively. In the Barnes maze test, the time of exploring the target hole in the IVIG-8 group was significantly longer than that in the control on the 6^th day(50.3±19.3 vs 21±14.6, P<0.05) and the 13^th day(58.2±20.9 vs 19.2±15.9, P<0.005), but there was no significant difference between the IVIG-1, 6 groups and the control. 【Conclusion】 There is a significant difference in the level of Aβ40/42 antibody among 8 kinds of domestic IVIG. Domestic IVIG could improve the cognitive function of 3-month-old 3xTg-AD mice after continuous intervention for 3 months. The improvement effect, however, was related to the Aβ antibody in IVIG, but not to the antibody concentration.

【Objective】 To obtain the quality information of von Willebrand factor (vWF) in coagulation factor Ⅷ (FⅧ) concentrates in China. 【Methods】 FⅧ concentrates produced by 7 domestic blood product manufactures and 1 foreign manufacture were collected, then FⅧ and vWF contained in FⅧ concentrates were evaluated. 【Results】 The activity loss of vWF was more than 25% in 2 of the 7 domestic FⅧ concentrates. The ratio of vWF activity to FⅧ activity in FⅧ concentrates from different domestic manufactures was significantly different (P<0.05). The ratio in FⅧ concentrates prepared by C, D, F manufacturer was greater than 1, which was similar to that in willate^@ approved abroad for the treatment of vWD. The ratio in FⅧ concentrates prepared by E manufacturer was greater than 0.7 and less than 1, and by A, B, G manufacturers was less than 0.5. In addition, the specific activities of FⅧ and vWF were significantly different among different FⅧ concentrates in China (P<0.05), and the specific activities of FⅧ and vWF were much lower than that of willate^@. 【Conclusion】 The variation of vWF quality between domestic FⅧ concentrates and willate^@ is mainly due to the different in vWF content. After the comprehensive consideration of various indicators, the FⅧ concentrates made by C and D manufacturers may be used in the treatment of vWD.