Objective To establish an indirect enzyme-linked immunosorbent assay(ELISA)method for testing the concentration of a monoclonal antibody target tumor necrosis factor-α(TNF-α)in animal serum.Methods The critical parameters of the method including coating concentration of human TNF-α,source,concentration and stability of HRP-labeled goat anti-human immunoglobulin G(IgG)were investigated.The specificity,accuracy,precision,linearity and Limited of Determination of the method were investigated.Results The critical parameters of the method were confirmed as below:TNF-α was coated at 400 ng·mL-1;HRP labeled goat anti-human IgG antibody was diluted at 1:3.0 ×105;the diluted horseradish peroxidase labeled goat anti-human IgG antibody is well stored at 4 ℃ for 3 days.Meanwhile the method was confirmed to have good specificity,the recovery rate ranged from 84.00％to 106.82％,the coefficient of variation of different antibody concentration levels were no more than 10％;the method had a good linearity and the standard curve was y=(-8.37×103-2.37 × 106)/[1+(x/29.80)106]+2.37 × 106(R2=0.999);the limit of quantification was 1 ng·mL-1,all of which met the requirements.Conclusion A accurate and robust ELISA method was developed to test the concentration of anti-TNF-α monoclonal antibody in serum.

Objective:To investigate the safety of early antiplatelet therapy for non-cardioembolic mild stroke patients with thrombocytopenia.Methods:Data of acute ischemic stroke patients with baseline National Institutes of Health Stroke Scale(NIHSS)score≤3 and a platelet count<100×109/L were obtained from a multicenter register.Those who required anticoagulation or had other contraindications to antiplatelet therapy were excluded.Short-term safety outcomes were in-hospital bleeding events,while the long-term safety outcome was a 1-year all-cause death.The short-term neurological outcomes were evaluated by modified Rankin scale(mRS)score at discharge.Results:A total of 1868 non-cardioembolic mild stroke patients with thrombocytopenia were enrolled.Multivariate regression analyses showed that mono-antiplatelet therapy significantly increased the proportion of mRS score of 0-1 at discharge(OR=1.657,95%CI:1.253-2.192,P<0.01)and did not increase the risk of intracranial hemorrhage(OR=2.359,95%CI:0.301-18.503,P>0.05),compared with those without antiplatelet therapy.However,dual-antiplatelet therapy did not bring more neurological benefits(OR=0.923,95%CI:0.690-1.234,P>0.05),but increased the risk of gastrointestinal bleeding(OR= 2.837,95%CI:1.311-6.136,P<0.01)compared with those with mono-antiplatelet therapy.For patients with platelet counts≤75×109/L and>90×109/L,antiplatelet therapy significantly improved neurological functional outcomes(both P<0.05).For those with platelet counts(>75-90)×109/L,antiplatelet therapy resulted in a significant improvement of 1-year survival(P<0.05).For patients even with concurrent coagulation abnormalities,mono-antiplatelet therapy did not increase the risk of various types of bleeding(all P>0.05)but improved neurological functional outcomes(all P<0.01).There was no significant difference in the occurrence of bleeding events,1-year all-cause mortality risk,and neurological functional outcomes between aspirin and clopidogrel(all P>0.05).Conclusions:For non-cardioembolic mild stroke patients with thrombocytopenia,antiplatelet therapy remains a reasonable choice.Mono-antiplatelet therapy has the same efficiency as dual-antiplatelet therapy in neurological outcome improvement with lower risk of gastrointestinal bleeding.

Objective This study aims to elucidate the protective role of Acacetin against apoptosis in HP-infected GES-1 cells and to delineate its potential underlying mechanisms.Materials and Methods GES-1 cells were subjected to in vitro treatment with HP and Acacetin.Cell viability was assessed utilizing the CCK-8 assay,alterations in cell migration and healing capacities through the wound healing assay,rates of apoptosis via flow cytometry,and expression levels of apoptosis-associated proteins through western blot analysis.Results HP infection led to a diminution in GES-1 cell viability,a suppression of cell migration,an augmentation in the rate of apoptosis,and an increase in the expression levels of Bax and cle-caspase3.Conversely,treatment with Acacetin was found to enhance cell viability,mitigate apoptosis induced by HP infection,and modulate the expression of apoptosis proteins by downregulating Bax and cle-caspase3.Discussion and Conclusion Acacetin significantly improves GES-1 cell vitality and inhibits apoptosis in HP-infected GES-1 cells,thereby offering a protective effect on gastric mucosal epithelial cells.

Objective To investigate the relationship between single nucleotide polymorphisms(SNPs)in the eNOS gene and genetic susceptibility to systemic lupus erythematosus(SLE)in Hainan.Methods Blood samples were collected from SLE patients(SLE group,n=214)and healthy controls(control group,n=214)from January 2020 to December 2022 at the First Affiliated Hospital of Hainan Medical College and Hainan Provincial People's Hospital.The bases of eNOS gene rs3918188,rs1799983 and rs1007311 loci in each group were detected by SNaPshot sequencing technology.Logistic regression was used to analyze the correlation between genotypes,alleles and gene models(dominant model,recessive model,and overdominant model)of the above 3 target loci of the eNOS gene and genetic susceptibility to SLE.Haplotype analysis was conducted using HaploView 4.2 software to investigate the relationship between haploid and genetic susceptibility to SLE at each site.Results The results of logistic regression analysis revealed that the CC genotype and the C allele at rs3918188 locus were risk factors for genetic susceptibility to SLE(CC vs.AA:OR=2.449,P<0.05;C vs.A:OR=2.133,P<0.001).In recessive model at rs3918188 locus,CC genotype carriers had an increased risk of SLE development compared with AA+AC genotype carriers(OR=2.774,P<0.001).In contrast,in overdominant model at this locus,AC genotype carriers had a decreased risk of SLE occurrence compared with AA+CC genotype carriers(OR=0.385,P<0.001).In addition,polymorphisms of rs1799983 and rs1007311 were not associated with susceptibility to SLE in genotype,allele type and the 3 genetic models(P>0.05).Haplotype analysis revealed a strong linkage disequilibrium between the rs1007311 and rs1799983 loci of the eNOS gene,but no significant correlation was found between haplotype and genetic susceptibility to SLE(P>0.05).Conclusion The CC genotype and C allele at rs3918188 locus of eNOS gene may be risk factors for SLE in Hainan,while the risk of SLE occurrence is reduced in carriers of AC genotype under the overdominant model.

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.

OBJECTIVE: Melittin, a cell-penetrating peptide, improves the efficiency of many non-viral gene delivery vectors, yet its application in viral vectors has not been well studied. The non-pathogenic recombinant adeno-associated virus (rAAV) vector is an ideal in vivo gene delivery vector. However, its full potential will only be achieved after improvement of its transduction efficiency. To improve the transduction efficiency of rAAV2 vectors, we attempted to develop a melittin-based rAAV2 vector delivery strategy. METHODS: The melittin peptide was inserted into the rAAV2 capsid either in the loop VIII of all viral proteins (VPs) or at the N terminus of VP2. Various rAAV2-gfp or -fluc vectors were subjected to quantitative real-time polymerase chain reaction and Western blot assays to determine their titers and integrity of capsid proteins, respectively. Alternatively, the vectors based on wild-type capsid were pre-incubated with melittin, followed by transduction of cultured cells or tail vein administration of the mixture to C57BL/6 and BALB/c nude mice. In vivo bioluminescence imaging was performed to evaluate the transgene expression. RESULTS: rAAV2 vectors with melittin peptide inserted in the loop VIII of VPs had low transduction efficiency, probably due to dramatically reduced ability to bind to the target cells. Fusing the melittin peptide at the N-terminus of VP2 produced vectors without the VP2 subunit. Interestingly, among the commonly used rAAV vectors, pre-incubation of rAAV2 and rAAV6 vectors with melittin significantly enhanced their transduction efficiency in HEK293 and Huh7 cells in vitro. Melittin also had the ability to increase the rAAV2-mediated transgene expression in mouse liver in vivo. Mechanistically, melittin did not change the vector-receptor interaction. Moreover, cell counting kit-8 assays of cultured cells and serum transaminase levels indicated melittin had little cytotoxicity. CONCLUSION Pre-incubation with melittin, but not insertion of melittin into the rAAV2 capsid, significantly enhanced rAAV2-mediated transgene expression. Although further in vivo evaluations are required, this research not only expands the pharmacological potential of melittin, but also provides a new strategy to improve gene therapy mediated by rAAV vectors.
Mice ; Animals ; Humans ; Melitten/genetics* ; Dependovirus/genetics* ; Serogroup ; HEK293 Cells ; Mice, Nude ; Mice, Inbred C57BL ; Transgenes ; Genetic Vectors/genetics*

Abstract: Viral shedding of SARS-CoV-2 is a continuous dynamic process, which can be divided into latent stage, initial stage, peak stage and decreasing stage according to the characteristics of viral shedding. After being infected with SARS-CoV-2, the infected person generally stays in the latent period for 1-3 days, which is characterized by continuous negative nucleic acid test results and no infectiousness, and the risk of infection for close contacts is very low. At the initial stage of viral shedding is characterized by a rapid decline in the Ct value of nucleic acid tests in a short time, and clinical symptoms gradually appear. The infectiousness of the infected person gradually increases during this period, and the risk of infection for close contacts also gradually increases, but it is still in the early stage of infection, the possibility of viral shedding is low, and the risk of infection of secondary close contacts is low. The peak of viral shedding is characterized by low Ct value in nucleic acid test and obvious clinical symptoms; during this period, the infected person is the most infectious, and the risk of infection of the contact is the highest, so the scope of close contacts should be expanded appropriately. The decreasing period is characterized by the gradual increase of Ct value of nucleic acid test and the gradual disappearance of clinical symptoms; during this period, the infectiousness of the infected person gradually decreases to disappear. In an outbreak, an infected person in the decreasing phase is more likely to be an early infected person in the transmission chain. If infected individuals in the decreasing phase are found in an area without a SARS-CoV-2 epidemic, it suggests that the local outbreak epidemic has been spreading for some time and may be larger in scale. According to the characteristics of viral shedding, risk personnel can be determined more scientifically and accurately, so as to minimize the risk and reduce the waste of epidemic prevention resources.

Objective: To analyze the clinical features, efficacy and prognosis factors of core binding factor (CBF) acute myeloid leukemia (AML) children in South China. Methods: This was a retrospective cohort study. Clinical data of 584 AML patients from 9 hospitals between January 2015 to December 2020 was collected. According to fusion gene results, all patients were divided into two groups: CBF-AML group (189 cases) and non-CBF-AML group (395 cases). CBF-AML group were divided into AML1-ETO subgroup (154 cases) and CBFβ-MYH11 subgroup (35 cases). Patients in CBF-AML group chosen different induction scheme were divided into group A (fludarabine, cytarabine, granulocyte colony stimulating factor and idarubicin (FLAG-IDA) scheme, 134 cases) and group B (daunorubicin, cytarabine and etoposide (DAE) scheme, 55 cases). Age, gender, response rate, recurrence rate, mortality, molecular genetic characteristics and other clinical data were compared between groups. Kaplan-Meier method was used for survival analysis and survival curve was drawn. Cox regression model was used to analyze prognostic factors. Results: A total of 584 AML children were diagnosed, including 346 males and 238 females. And a total of 189 children with CBF-AML were included, including 117 males and 72 females. The age of diagnosis was 7.3 (4.5,10.0)years, and the white blood cell count at initial diagnosis was 21.4 (9.7, 47.7)×10⁹/L.The complete remission rate of the first course (CR1) of induction therapy, relapse rate, and mortality of children with CBF-AML were significantly different from those in the non-CBF-AML group (91.0% (172/189) vs. 78.0% (308/395); 10.1% (19/189) vs. 18.7% (74/395); 13.2% (25/189) vs. 25.6% (101/395), all P<0.05). In children with CBF-AML, the CBFβ-MYH11 subgroup had higher initial white blood cells and lower proportion of extramedullary invasion than the AML1-ETO subgroup, with statistical significance (65.7% (23/35) vs. 14.9% (23/154), 2.9% (1/35) vs. 16.9% (26/154), both P<0.05). AML1-ETO subgroup had more additional chromosome abnormalities (75/154), especially sex chromosome loss (53/154). Compared with group B, group A had more additional chromosome abnormalities and a higher proportion of tumor reduction regimen, with statistical significance (50.0% (67/134) vs. 29.1% (16/55), 34.3% (46/134) vs. 18.2% (10/55), both P<0.05). Significant differences were found in 5-years event free survival (EFS) rate and 5-year overall survival (OS) rate between CBF-AML group and non-CBF-AML group ((77.0±6.4)%vs. (61.9±6.7)%,(83.7±9.0)%vs. (67.3±7.2)%, both P<0.05).EFS and OS rates of AML1-ETO subgroup and CBFβ-MYH11 subgroup in children with CBF-AML were not significantly different (both P>0.05). Multivariate analysis showed in the AML1-ETO subgroup, CR1 rate and high white blood cell count (≥50×10⁹/L) were independent risk factors for EFS (HR=0.24, 95%CI 0.07-0.85,HR=1.01, 95%CI 1.00-1.02, both P<0.05) and OS (HR=0.24, 95%CI 0.06-0.87; HR=1.01, 95%CI 1.00-1.02; both P<0.05). Conclusions: In CBF-AML, AML1-ETO is more common which has a higher extramedullary involvement and additional chromosome abnormalities, especially sex chromosome loss. The prognosis of AML1-ETO was similar to that of CBFβ-MYH11. The selection of induction regimen group FLAG-IDA for high white blood cell count and additional chromosome abnormality can improve the prognosis.
Male ; Female ; Humans ; Child ; Retrospective Studies ; RUNX1 Translocation Partner 1 Protein/genetics* ; Core Binding Factor Alpha 2 Subunit/therapeutic use* ; Prognosis ; Leukemia, Myeloid, Acute/genetics* ; Cytarabine/therapeutic use* ; Oncogene Proteins, Fusion/genetics* ; Chromosome Aberrations

This study aims to establish a method for the simultaneous determination of 7 active components in Dracocephalum tanguticum and to evaluate the quality of medicinal materials from different habitats. The method was established with high performance liquid chromatography(HPLC) and the gradient elution was performed with the mobile phase of acetonitrile-methanol-0.2% phosphoric acid solution at a column temperature of 35 ℃, an injection volume of 15 μL, and a flow rate of 0.6 mL·min~(-1). The detection wavelength was set as 215 nm. With rosmarinic acid as the internal reference, the relative correction factors and the content of other 6 components were calculated. The results were compared with those obtained with the external standard method. The results showed that the samples from Huangzhong county, Qinghai province had the best quality, with the highest content of p-hydroxybenzoic acid, cosmosiin, rosmarinic acid, oleanolic acid, and ursolic acid(9.29, 12.14, 6.02, 3.11, 17.67 mg·g~(-1) respectively). The samples from Chaya county, Tibet autonomous region ranked the second, with the highest content of betulin and betulinic acid(15.53, 7.17 mg·g~(-1), respectively). The method is accurate, reliable, and repeatable and suitable for the simultaneous determination of multiple components in D. tanguticum. The content of functional components varied in the samples from different producing areas and can be used as the indicator for the quality evaluation of medicinal materials.
Cinnamates ; Drugs, Chinese Herbal/analysis* ; Lamiaceae ; Chromatography, High Pressure Liquid/methods* ; Rosmarinic Acid

Humans ; Mesothelioma/pathology* ; Mesothelioma, Malignant ; Pleural Neoplasms/diagnosis* ; Pleural Effusion, Malignant ; Pleural Effusion/pathology*