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Resumen Se calcula que el cuerpo humano está conformado por billones de células, las cuales sufren cientos de miles de lesiones al día en su DNA. Aunque el DNA no es la única biomolécula que sufre daños, su importancia radica en que es la única que no puede ser sustituida por la célula, así que, cuando esta sufre daños, la célula debe repararlos, tolerarlos o, en el caso extremo, activar las vías que la llevarán a la muerte, ya que lo importante es mantener la integridad celular y la homeostasis del organismo. Hay miles de agentes que pueden dañar al DNA, algunos los produce la misma célula y se les denomina 'agentes endógenos', mientras que otros son agentes externos y se les conoce como 'agentes exógenos'. La célula no puede evitar el daño causado por los agentes endógenos, ya que son productos de la actividad metabólica, por ejemplo; así que, cuando suceden se activan de forma inmediata los mecanismos celulares para mitigarlos. Lo mismo pasa con los daños causados por agentes exógenos, ya que la célula hará todo lo posible por disminuir los efectos adversos que pueden causar. El problema se pone de manifiesto cuando la célula no puede reparar los daños o los repara mal o son tantos que los mecanismos de reparación se ven rebasados, es entonces cuando el daño permanece en el DNA y se genera un estado de inestabilidad cromosómica que puede conducir a la célula a la disfunción y a la malignización. Este estado de inestabilidad cromosómica se puede ver reflejado en el aumento de rompimientos de DNA o de micronúcleos en las células expuestas, lo que se puede cuantificar por medio de métodos especiales como el 'Ensayo Cometa' y el 'Ensayo de Micronúcleos', ya que identificar el daño en el DNA es una forma de evaluar el potencial tóxico que tienen los agentes a los que están expuestas las poblaciones, permite conocer los mecanismos de acción que tienen y, además, ayuda a comprender los factores que influyen en el detrimento de la salud poblacional.
Abstract It is estimated that the human body is made of trillions of cells, which suffer hundreds of thousands of DNA lesions every day. Although DNA is not the only biomolecule that suffers damage, its importance lies in the fact that it is the only biomolecule that cannot be replaced by the cell, so when it suffers damage, the cell must repair it, tolerate or, in a extreme case, activate pathways that will lead to death, since the objective is to maintain cell integrity and the homeostasis of the organism.There are thousands of agents that can damage DNA, some are produced by the cell and are called 'endogenous, while others are external agents and are known as 'exogenous. The cell cannot avoid the damage caused by endogenous agents, since they are products of its metabolic activity, for example, so when they occur, cellular mechanisms are immediately activated to mitigate them. The same happens with the damage caused by exogenous agents, since the cell will do everything possible to diminish the adverse effects they can cause. The problem becomes apparent when the cell is unable to repair the damage or poorly repairs it, or repairs so much that the mechanisms are overwhelmed, when the damage remains in the DNA and a state of chromosomal instability is generated that can lead the cell to dysfunction and malignization. This state of chromosomal instability can be reflected in increased DNA breaks or micronuclei in exposed cells, which can be quantified by special methods such as the 'Comet Assay' and the 'Micronucleus Assay'. Since identifying DNA damage is a way of evaluating the toxic potential of the agents to which populations are exposed, it allows us to know their mechanisms of action and helps to understand the factors that influence the detriment in population's health.
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@#Objective To develop a kinetic chromogenic quantitative method for the determination of endotoxin content in intermediate of pertussis antigen,and to verify the method so as to better control the quality of diphtheria,tetanus,and pertussis vaccine(DTP vaccine).Methods A kinetic chromogenic assay[Limulus Amebocyte Lysate(LAL)]was developed to detect the endotoxin content in the intermediate products of pertussis antigens after detoxification,and verified for the linearity,specificity,accuracy,reproducibility and intermediate precision. The quantitative detection results of kinetic chromogenic assay were compared with those of gel method.Results The absolute value of the linear correlation coefficient（|r|）of the kinetic chromogenic assay was more than 0. 99;under the maximum effective multiple dilution,the interference test recovery of the intermediate was within 50% — 200%,and pertussis toxin(PT)diluted to 10,100 and 1 000 times,filamentous hemagglutinin(FHA)diluted to 3 000,5 000 and 10 000 times,and pertussis adhesin(PRN)diluted to 50,75 and 100 times had no interference effect on the experiment after detoxification;the accuracy verification recovery rates of PT,FHA and PRN were 125%,110% and 99% respectively;and the CVs of reproducibility verification were 7. 21%,8. 31% and 5. 84%,and the CVs of intermediate precision verification were 6. 04%,16. 29% and 12. 23%,respectively.The bacterial endotoxin content of the three batches of pertussis antigen intermediates detected by kinetic chromogenic assay was consistent with that verified by gel method,both of which were less than the limit of bacterial endotoxin in the intermediates of pertussis antigen after detoxification.Conclusion The developed kinetic chromogenic assay has good linearity,specificity,accuracy and precision with accurate detection results,which can be used to detect the endotoxin content in intermediate products of component pertussis antigen after detoxification.
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@#Objective To develop and verify a method for detecting the activity of neutralizing antibodies in ELISA antibody positive serum of rats immunized with recombinant human interleukin-1 receptor antagonist(rhIL-1Ra). Methods The SD rats were subcutaneously immunized with 3,20 and 100 mg/kg rhIL-1Ra injection respectively,10 rats in each group,half male and half female,twice a day at an interval of at least 4 h between each dose for 13 consecutive weeks. The blood samples were collected from the jugular vein of rats during the administration period and the recovery period. The serum samples were isolated and detected for the antibody titers by ELISA,and the samples positive for rhIL-1Ra antibody were purified by Protein A chromatographic column. Based on,D10G4·1 cells biological activity assay,a method for the detection of neutralizing antibody activity was developed and verified for the specificity,sensitivity and precision. The neutralizing antibody activity of rhIL-1Ra antibody positive serum determined by ELISA was detected by using the developed method.Results With the increase of doses,the serum antibody titers of rats in various dose groups gradually increased,and there were still antibodies in the recovery period,and the titer was still high. Rabbit anti-rhIL-1Ra monoclonal antibody showed obvious neutralizing effect on rIL-1Ra,while rabbit anti-rIFN-2b monoclonal antibody had no dose-effect relationship with rIL-1Ra. The sensitivity of the method was 171. 93 μg/mL;The CVs of precision verification were not more than 20%. The positive antibody sera detected by ELISA all had neutralizing effect on rhIL-1Ra injection,which was consistent with the results detected by ELISA. Conclusion The method developed in this study has good specificity and high sensitivity in the detection of serum neutralizing antibody activity in rats immunized with rhIL-1Ra,which can be used to detect the serum neutralizing antibody activity of animals with rhIL-1Ra repeated administration.
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@#Objective To develop and verify a double-antibody sandwich ELISA method for the detection of process-specific E.coli residual protein in recombinant biological preparations.Methods Taking the production and purification process of glucagon-like peptide(GLP)expressed by E.coli as the specific process model,the same process was used to intercept the residual protein of empty E.coli(normal E.coli that does not express recombinant protein). One female New Zealand white rabbit and six female Kunming mice were immunized with the residual protein as the immunogen. Using the IgG antibody purified from rabbit immune serum as the coating antibody,mouse immune serum as the second sandwich antibody,and antimouse IgG-HRP as the enzyme-labeled secondary antibody,a double antibody sandwich ELISA method for process-specific residual protein of E.coli was established. The specificity,accuracy and precision of the method were verified,and the limit of detection(LOD)was determined. Simultaneously,the developed method and the commercial E.coli host protein residue detection kit were used to quantitatively determine the residual protein of purified GLP preparation.Results After a series of gradient dilution of process-specific residual protein with known concentration,the sensitivity of this ELISA method reached 338 pg/mL. No cross reaction occurred in the detection of CHO and yeast cell lysis protein by this method,the recoveries of samples with low,medium and high concentrations were all in the range of 80% — 120%,and the intra-assay and inter-assay CVs of the empty E.coli interception standard with low,medium and high concentrations were all less than15%. For the residual protein in GLP preparation,about 62% of the residual proteins were not detected by the commercial non-process-specific ELISA kit compared with the total amount of residual proteins detected by the developed method,and these residual proteins should be the process-specific residual proteins.Conclusion The double antibody sandwich ELISA method developed in this study has high sensitivity,strong specificity,good accuracy and precision for the detection of process-specific E.coli residual protein,which can meet the detection requirements that the residual protein is less than0. 01% — 0. 1% in biological preparations.
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Objective To observe the effectiveness of fluorescence labeling-based assay bundle intervention in the prevention and control of multidrug-resistant organism(MDRO)infection.Methods Patients who were detected MDRO in a hospital from January to December 2022 were selected as the research subjects.MDRO monitoring data and implementation status of prevention and control measures were collected.Fluorescence labeling assay was adopted to monitor the cleaning and disinfection effectiveness of the surrounding object surface of the bed units.Based on the bundled prevention and control measures as well as management mode of the pre-intervention group,the post-intervention group implemented enhanced rectification measures for the problems found by the pre-interven-tion group.Changes in relevant indicators between January-June 2022(before intervention)and July-December 2022(after intervention)were compared.Results There were 136 MDRO-infected patients in the pre-intervention group,208 MDRO strains were detected and 10 healthcare-associated infection(HAI)occurred.There were 128 MDRO-infected patients in the post-intervention group,198 MDRO strains were detected and 9 HAI occurred.Af-ter intervention,the total detection rates of methicillin-resistant Staphylococcus aureus(MRSA),carbapenem-re-sistant Acinetobacter baumannii(CRAB),and total MDRO from patients decreased significantly compared to before intervention(all P＜0.05).After intervention,the detection rates of MRSA,carbapenem-resistant Pseudomonas aeruginosa(CRPA),CRAB,and total MDRO from the surrounding object surface were all lower than those before intervention(all P＜0.05).The detection rate of MDRO from surrounding object surface before intervention was 34.52％,which showed a decreased trend after intervention(P＜0.05).The clearance rate of fluorescent labeled markers before intervention was 41.84％,which showed an upward trend after implementing intervention measures(from July to December),and increased to 85.00％at the end of intervention(November-December).The comp-liance rates of issuing isolation medical orders,placing isolation labels,using medical supplies exclusively,and cor-rectly handling medical waste after intervention have all increased compared to before intervention(all P＜0.05).Conclusion Adopting fluorescence labeling-based assay bundle intervention can effectively improve the effectiveness of MDRO infection prevention and control.
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Objective To investigate doctors'knowledge and differences in islet function assessment methods in China.Methods This is a cross-sectional study that conducted by online questionnaire survey.Demographic data,examination items,blood collection point of OGTT,detection method,kit type and follow-up frequency were collected and compared among doctors in different regions,different levels of hospitals,different specialties and different titles.Results 79.2%and 85.1%of physicians believed that the levels of insulin and C-peptide should be measured at the same time to assess islet function in patients with newly diagnosed and follow-up diabetes mellitus patients.Endocrinologists preferred to access insulin and C-peptide at the same time(P<0.05).56.0%of physicians chose bread meal test for T1DM patients and 54.7%for T2DM patients.Compared with non-specialists,endocrinologists preferred to commit bread meal test to T1DM patients(61.4%vs 41.0%,P<0.05).In addition,for the islet function assessment of new-onset diabetes patients,7.6%of physicians chose the six-point method(0,30,60,90,120,180 min),27.3%selected the five-point method I(0,30,60,120,180 min),8.5%selected the five-point method II(0,30,60,90,120 min),9.8%selected the four-point method I(0,30,60,120 min),10.3%selected the four-point method II(0,60,120,180 min),13.8%chose the three-point method(0,60,120 min)and 13.4%chose the two-point method(0,120 min).At the time of follow-up assessment,the above selection rates were 5.3%,20.4%,6.4%,6.6%,9.4%,15.8%and 24.1%,respectively.In terms of the frequency of assessment,39.2%of doctors assessed islet function once a year and 24.7%once every six months.Specialists preferred to assess islet function once a year,and physicians with senior titles chose to assess islet function more variably.Conclusion At present,there are still great differences in assessment methods of islet function in China.It is of great significance for the clinical diagnosis and treatment of diabetes to understand the differences in the selection of islet function assessment methods among doctors in different regions,specialties and job titles.
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The immune response against infection is a multifaceted process encompassing the activation and migration of diverse immune cells, as well as the clearance of pathogens. The behaviors of immune cells and the identification of pathogens play pivotal roles as indicators for disease diagnosis and prediction. In recent years, the utilization of microfluidic chip technology has gained substantial attention within the areas of biology, pharmacology, and clinical research and diagnosis. This is primarily attributed to the numerous advantages it offers, including miniaturization, enhanced throughput, heightened sensitivity, expedited analysis, and reduced sample consumption. As a result, microfluidic technology has facilitated the development and utilization of immune cell behavioral assays, bacterial growth studies, and drug-screening assays. This paper is to review the application of microfluidic technology in the field of anti-infection immunity research, focusing on the analysis of migratory behavior of innate immune cells, deformation of their nuclei, and rapid identification of pathogenic bacteria and viruses. The primary objective of this review is to advance the application of microfluidic technology in research on both anti-infection immunity and clinical diagnosis.
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Objective:To investigate the factors influencing phytohemagglutinin (PHA) response in the detection of Mycobacterium tuberculosis infection by gamma interferon release assay (IGRA). Methods:A retrospective case-control study was conducted on 360 hospitalized patients who received IGRA in West China Hospital of Sichuan University from January 2019 to December 2021. According to PHA response (IFN-γ level), they were divided into three groups: negative mitogen response group (IFN-γ<2 pg/ml), weak positive mitogen response group (IFN-γ: 2-100 pg/ml), and normal mitogen response group (IFN-γ>400 pg/ml).Results:Immune diseases were independently associated with negative (OR=0.34, 95%CI: 0.17-0.72, P=0.004) and weak positive mitogen responses (OR=0.29, 95%CI: 0.16-0.55, P<0.001). Infections caused by pathogens other than Mycobacterium tuberculosis was independently associated with negative mitogen response (OR=0.266, 95%CI: 0.09-0.83, P=0.023), while immunodeficiency was independently associated with weak positive mitogen response (OR=0.280, 95%CI: 0.12-0.63, P=0.002). Mitogen response was significantly correlated with the levels of albumin and hemoglobin in serum and the counts of neutrophils and lymphocytes ( P<0.001). Conclusions:Immune diseases and immunodeficiency can affect mitogen response. Therefore, clinicians should give attention to mitogen response in the interpretation of IGRA test results to prevent misdiagnosis and underdiagnosis. Besides, to a certain extent, mitogen response can reflect the infection status of hospitalized patients.
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Objective:To analyze the genetic mutation characteristics of glucose-6-phosphate dehydrogenase (G6PD) deficiency among infants in Kunming.Methods:A total of 15 533 infants (7 994 males and 7 539 females) born in Kunming from January 1, 2018, to December 31, 2020, with an age range of 2 to 44 days, were selected. G6PD enzyme activity and gene mutation types were detected using fluorescence quantitative analysis, multicolor melting curve analysis (MMCA), and Sanger sequencing. Droplet digital PCR (ddPCR) was used for quantitative analysis of a newly identified variant family to determine the mutant allele proportion in family members. Meanwhile,the protein structure model and pathogenicity prediction of the novel variant were analyzed.Data analysis was conducted using SPSS 26.0. Specifically, chi-square tests were used for the detection rates of G6PD enzyme activity and gene mutations between different genders. One-way analysis of variance (ANOVA) was used for the comparison of enzyme activity among different mutation types.Results:Among 15 533 infants, 143 cases (129 males and 14 females) were tested positive for G6PD activity, with a detection rate of 0.92% (143/15 533). The difference in detection rates between males and females was statistically significant (χ 2=96.76, P<0.001). Out of 89 enzyme activity-positive cases (83 males and 6 females) underwent genetic testing, 77 (72 males and 5 females) were detected by MMCAand other 12 negative samples were underwent further Sanger sequencing, revealing mutations in 6 samples, all of which were males. Among the 83 individuals with gene mutations, 78 had heterozygous mutations, 1 had a homozygous mutation, and 4 had compound heterozygous mutations. A total of 12 mutation types were detected, with G6PD c.487G>A, c.1024C>T, c.1388G>A, and c.1376G>T being the most common, accounting for 74.70% (62/83) of all mutation types. The average G6PD enzyme activity of c.1376G>T was the lowest, and the differences were statistically significant compared to the average enzyme activity of the other three mutations ( P<0.05). One male infant with a newly identified G6PD c.242G>C mutation was detected, predicted to be pathogenic. ddPCR confirmed that the mother of the affected child was a c.242G>C mutant chimera, with a chimera proportion of 6.66%. Conclusions:In the Kunming region, the predominant G6PD deficiency gene mutation is c.487G>A, with the detection of a novel G6PD c.242G>C mutation. The application of ddPCR technology can assist in detecting the proportion of mutation chimeras.
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Objective:To investigate the effect of estrogen-related receptor α (ESRRA)-mediated lipophagy on the proliferation and migration abilities of nasopharyngeal carcinoma cells.Methods:A total of 16 clinical samples diagnosed by pathology in the Affiliated Hospital of Nantong University from 2021 to 2023 were selected, including 8 normal nasopharyngeal mucosa tissues and 8 nasopharyngeal carcinoma tissues. Immortalized normal nasopharyngeal epithelial cell line NP69 and nasopharyngeal carcinoma cell lines C666-1, CNE2, TW03-EBV and TW03 were selected. The cell lines C666-1 and CNE2 were divided into the siR-NC group (transfected with small interfering RNA negative control sequence) and siR-ESRRA group (transfected with small interfering RNA against ESRRA gene). The relative expression levels of ESRRA were detected by Western blotting and immunohistochemical assay. EdU assay was used to detect the proliferation ability of C666-1 and CNE2 cells, and Transwell assay was used to detect the migration ability. Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expression levels of ESRRA and perilipin 3 (PLIN3) mRNA. The formation of lipophagy in C666-1 and CNE2 cells was observed by transmission electron microscopy. The co-localization of LC3, PLIN3 and LAMP2 with lipid droplets labeling with Bodipy was detected by immunofluorescence assay. Dual-luciferase reporter gene assay was used to verify the targeting relationship between ESRRA and PLIN3.Results:The relative expression level of ESRRA in nasopharyngeal carcinoma tissues was higher than that in normal nasopharyngeal mucosa tissues(1.15±0.75 vs. 0.32±0.21, t = 3.02, P = 0.009). The relative expression level of ESRRA in nasopharyngeal carcinoma cell lines C666-1 (1.539±0.044), CNE2 (1.420±0.030), TW03-EBV (2.867±0.044), and TW03 (1.323±0.022) were higher than that in normal nasopharyngeal epithelial cell line NP69 (0.094±0.002), and the difference was statistically significant ( F = 34.08, P < 0.001).The results of EdU assay showed that the proportions of EdU labeled positive cells in CNE2 cells of siR-NC group and siR-ESRRA group were (70.44±4.06)% and (51.51±0.92)% ( t = 7.88, P = 0.001), and the proportions in C666-1 cells were (62.25±3.89)% and (54.91±0.27)% ( t = 3.26, P = 0.031). The results of Transwell assay showed that the number of migrating cells in CNE2 and C666-1 cells was less than that in siR-NC group [CNE2 cells: (181±7) cells vs. (261±21) cells; C666-1 cells: (201±16) cells vs. (256±7) cells], and the differences were statistically significant ( t = 6.30, P = 0.003; t = 5.43, P = 0.006). According to qRT-PCR results, the relative expression level of PLIN3 mRNA in the siR-ESRRA group was higher than that in the siR-NC group (CNE2 cells: 1.58±0.16 vs. 0.83±0.17, t = 5.59, P = 0.005; C666-1 cells: 1.37±0.12 vs. 1.06±0.06, t = 3.86, P = 0.018). The dual-luciferase reporter gene assay results indicated a targeted binding interaction between PLIN3 and ESRRA. Transmission electron microscopy observation showed that the lipid droplets in nasopharyngeal carcinoma cells increased and the binding to autophagosomes decreased after knockdown of ESRRA. The results of immunofluorescence assay demonstrated that, in contrast to the siR-NC group, there was a decrease in the co-localization of LC3 and LAMP2 and an increase in the co-localization of lipid droplets with PLIN3. Conclusions:ESRRA is highly expressed in nasopharyngeal carcinoma tissues and cells. As a transcription repressor, ESRRA may work to prevent PLIN3 from being transcribed, decrease lipid droplet stability, mediate lipophagy, and promote proliferation and migration of nasopharyngeal carcinoma cells.
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Objective To establish high performance liquid chromatography(HPLC)characteristic chromatograms of Xinqingduyin Granules(composed of Taraxaci Herba,Lonicerae Japonicae Flos,Chrysanthemi Indici Flos,etc.)and content determination of chicory acid and glycyrrhizic acid,and to optimize the preparation process of Xinqingduyin Granules.Methods Using the characteristic chromatograms of Xinqingduyin and the retention rate of chicory acid and glycyrrhizic acid as indexes,we carried out orthogonal experiment to optimize the extraction process of Xinqingduyin,and studied the concentration process.The molding process of Xinqingduyin Granules was conducted by screening the types and dosage of auxiliary materials,then three batches of pilot experiments were carried out.Results HPLC characteristic chromatograms of Xinqingshuyin Granules and the determination methods of chicory acid and glycyrrhizic acid were established.The optimal preparation technology was as follows:8 times amount of water was added,the drug was decocted for 3 times,with 1 hour per time.After the extract was concentrated under reduced pressure at 80℃,the appropriate amount of steviol glycoside and lactose was added into the extract and mixed.One-step granulation and packaging were adopted.The retention rates of chicoric acid and glycyrrhizic acid in the 3 batches of Xinqingduyin Granules,which were prepared on the pilot scale,were(54.56±1.63)%and(54.96±1.08)%,and the rate of finished product was(87.47±0.49)%,respectively.The quality is uniform,and the characteristic map of Xinqingduyin Granules showed high similarity with that of decoction prepared from the same batch of slices.Conclusion The optimized preparation technology is reasonable,feasible and reproducible.This preparation can be used to obtain the granule with similar materials of Xinqingduyin decoction.
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Objective To provide a toxicological basis for HMS-01 in future clinical trials, through genotoxicity testing of safety evaluation. Methods The genotoxicity of HMS-01 was evaluated by Bacterial Reverse Mutation Assay （Ames test） with Salmonella typhimurium. Results HMS-01 was non-mutagenic against Salmonella typhimurium at six doses of 20.6, 61.7, 185.2, 555.6, 1666.7, and 5000 μg/dish with （or without） a metabolic activation system. Conclusion HMS-01 was not found to be mutagenic in the dose range of this experiment.
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@#Bullous pemphigoid (BP) is a rare autoimmune blistering disorder primarily affecting older adults, with limited occurrences in children. BP in children typically manifests as large, tense blisters on the skin, often on flexural areas. It also more often affects the oromucosal areas and the face in children than in adults. Diagnosis involves histopathological examination revealing eosinophilic spongiosis or subepidermal split, immunofluorescence tests highlighting immunoglobulin G (IgG) and C3 depositions, and immunological assays detecting BP180 and BP230 IgG autoantibodies. This report presents two cases of childhood BP (CBP) with atypical immunopathological findings. Clinically, the two cases had generalized plaques and bullae, including the face. The first case exhibited the characteristic linear deposits of IgG and C3 on the basement membrane through direct immunofluorescence (DIF) and revealed negative anti‑BP180 antibodies on enzyme‑linked immunosorbent assay (ELISA). In contrast, the second case showed negative DIF results, despite clinical suspicion, but had positive anti‑BP180 IgG antibodies on ELISA. It is, therefore, crucial to consider the complete clinical presentation of the patient, in conjunction with the histological findings and immunopathologic assessments to diagnose CBP.
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Pemphigoïde bulleuseRÉSUMÉ
@#Objective To compare the immunogenicity of varicella-zoster virus(VZV)Oka-7S strain and VZV Oka strain.Methods Female BALB/c mice were subcutaneously immunized with VZV Oka-7S and Oka strains at high,medium and low doses(1 000,250 and 62.5 PFU/dose)for total two doses at an interval of 14 d,separately,with 12 mice in each group.The VZV-specific humoral immunity was evaluated based on fluorescent antibody to membrane antigen assay(FAMA),and the VZV-specific cellular immunity was evaluated by enzyme-linked immunospot assay(ELISPOT).Results Both VZV Oka-7S and VZV Oka strains induced specific humoral and cellular immune responses,and showed considerable immune effects;The level of anti-VZV specific IgG in serum of mice in Oka-7S-2.2 group increased significantly with the increase of immunization times and immunization doses.Conclusion The VZV Oka-7S strain had comparative immunogenicity with VZV Oka strain.
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Objective @#To investigate the effect of cysteine rich acidic secretory protein like protein 1 (SPARCL1) on atherosclerosis (AS) plaque formation .@*Methods @#A case control study design was used , 394 patients with con firmed AS were selected as the case group , and 394 healthy medical examiners matched for age and gender were se lected as the control group . The expression level of serum SPARCL1 was determined by enzyme linked immunosor bent assay; immunohistochemistry was used to assess the expression level and localization of SPARCL1 protein in the AS plaque region , and the expression of SPARCL1 protein was also detected in the neutrophils and monocytes of peripheral blood of AS patients and normal controls; SPARCL1 overexpressing and the recombinant adenoviral vec tors were constructed to inhibit SPARCL1 overexpression and expression , and the effects of SPARCL1 on cell mi gration were ob served in the cell scratch assay using mouse macrophage cells (J774A.1) as target cells .@*Results@#Serum SPARCL1 levels in the AS patient group were lower than those in the healthy group ( P < 0.05 ) ; high SPARCL1 expression was detected in AS plaques and was mainly expressed in the cytoplasm of foamy cells; SPARCL1 expression levels in peripheral blood neutrophils and monocytes were lower than those in normal controls in AS patients (P < 0.05) ; recombinant SPARCL1 overexpression and inhibition of expression of adenovirus was successfully constructed; the cell migration rate was decreased in J774A.1 cells that inhibited SPARCL1 expression and increased in J774A.1 cells that overexpressed SPARCL1 ( P < 0.05) .@*Conclusion @#SPARCL1 is highly expressed in foam cells at the site of AS lesions , which may result from compensatory recruitment of peripheral blood monocytes and neutrophils , and SPARCL1 may be involved as a protective factors for blood vessels in inhibiting the development of AS plaques .
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Background The benchmark dose (BMD) method calculates the dose associated with a specific change in response based on a specific dose-response relationship. Compared with the traditional no observed adverse effect level (NOAEL) method, the BMD method has many advantages, and the 95% lower confidence limit of benchmark dose lower limit (BMDL) is recommended to replace NOAEL in deriving biological exposure limits. No authority has yet published any health-based guideline for rare earth elements. Objective To evaluate genotoxicity threshold induced by acute exposure to neodymium nitrate in mice using BMD modeling through micronucleus test and comet assay. Methods SPF grade mice (n=90) were randomly divided into nine groups, including seven neodymium nitrate exposure groups, one control group (distilled water), and one positive control group (200 mg·kg−1 ethyl methanesulfonate), 10 mice in each group, half male and half female. The seven dose groups were fed by gavage with different concentrations of neodymium nitrate solution (male: 14, 27, 39, 55, 77, 109, and 219 mg·kg−1; female: 24, 49, 69, 97, 138, 195, and 389 mg·kg−1) twice at an interval of 21 h. Three hours after the last exposure, the animals were neutralized by cervical dislocation. The bone marrow of mice femur was taken to calculate the micronucleus rate of bone marrow cells, and the liver and stomach were taken for comet test. Results The best fitting models for the increase of polychromatophil micronucleus rate in bone marrow of female and male mice induced by neodymium nitrate were the exponential 4 model and the hill model, respectively. The BMD and the BMDL of female mice were calculated to be 31.37 mg·kg−1 and 21.90 mg·kg−1, and those of male mice were calculated to be 58.62 mg·kg−1 and 54.31 mg·kg−1, respectively. The best fitting models for DNA damage induced by neodymium nitrate in female and male mouse hepatocytes were the exponential 5 model and the exponential 4 model, respectively, and the calculated BMD and BMDL were 27.15 mg·kg−1 and 11.99 mg·kg−1 for female mice, and 16.28 mg·kg−1 and 10.47 mg·kg−1 for male mice, respectively. The hill model was the best fitting model for DNA damage of gastric adenocytes in both female and male mice, and the calculated BMD and BMDL were 36.73 mg·kg−1 and 19.92 mg·kg−1 for female mice, and 24.74 mg·kg−1 and 14.08 mg·kg−1 for male mice, respectively. Conclusion Taken the micronucleus rate of bone marrow cells, DNA damage of liver cells and gastric gland cells as the end points of genotoxicity, the BMDL of neodymium nitrate is 10.47 mg·kg−1, which can be used as the threshold of genotoxic effects induced by acute exposure to neodymium nitrate in mice.
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This guideline aims to provide comprehensive and practical guidance for clinical management of tuberculosis in kidney transplant recipients. First, it summarizes the particularity of tuberculosis in kidney transplant recipients, and highlights the high incidence and diverse clinical manifestations. To better understand the patients' conditions, relevant assessment of tuberculosis is recommended before kidney transplantation. Extensive attention should be paid to the monitoring of tuberculosis after kidney transplantation. Regarding the diagnosis, the guideline explicitly introduces common diagnostic approaches for tuberculosis, and evaluates the applicability in kidney transplant recipients. After the diagnosis is confirmed, it discusses how to balance the treatment and rejection of tuberculosis under the background of immunosuppressants, and focuses upon the potential drug interaction. In terms of prevention, it emphasizes the screening of tuberculosis prior to kidney transplantation. This guideline is designed to deepen the understanding of medical staff for tuberculosis management in kidney transplant recipients, promote more effective clinical practice and improve the quality of life of the recipients.
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@#Objective To establish and verify a universal and stable potency test method in vitro for SARS-CoV-2 mRNA vaccine,so as to use it for the quality control of SARS-CoV-2 mRNA vaccine.Methods ELISA kits that could bind well to S protein of SARS-CoV-2 variants,as well as transfected cells,cell plating concentrations and doses for transfection were screened,and then a potency test method for SARS-CoV-2 mRNA vaccine in vitro was established and verified.Results An ELISA kit was found with good binding ability to S protein of each variant,and HEK293T cells were determined as the transfection cells,with the plating concentration of 2.5 × 10~5 cells/mL and the transfection dose of 4 μg/well in the 6-well plate.An universal and stable potency test method for SARS-CoV-2 mRNA vaccine in vitro was established.The verification results showed that the method met the quality control needs.Conclusion The established potency test method in vitro for SARS-CoV-2 mRNA vaccine has good relative accuracy,linearity,intermediate precision and range,and can be applied to the quality control of SARS-CoV-2 mRNA vaccines
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Objective To detect cell DNA damage more effectively by optimizing the influencing factors of Comet assay and establishing a positive control.Methods Based on the traditional alkaline comet assay,the gel prepara-tion method,gel concentration,agarose solvent,lysis time,cell density and electrophoresis voltage were opti-mized.HL-60 cells were induced by hydrogen peroxide(H2O2)as a DNA damage inducer as a positive control.Results The optimized experimental results showed that the gel preparation method improved from three-step meth-od to two-step method.PBS was used as agarose solvent,0.8％normal melting point agarose was used to prepare the bottom layer,and 0.7％low melting point agarose was mixed with cell suspension as the second layer.The gel preparation effect was better,the operation was simpler,the time was saved,and the degumming problem was bet-ter solved.After lysis for 1 h and electrophoresis at 1 V/cm voltage,representative comet images were obtained.The results were easy to interpret and could effectively avoid false positive results.The positive control of comet as-say was successfully established by comparing the DNA damage of HL-60 cells induced by different concentrations of H2 O2.Conclusion Compared with the traditional method,the optimized comet assay method is simpler,faster,more accurate,lower cost and has better repeatability,which can quickly detect DNA damage in cells.
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Objective To investigate the changes and clinical significance of serum tumor necrosis fac-tor-α(TNF-α),adiponectin(ADP)and their ratio in patients with depression.Methods A total of 40 patients with depression admitted to Shandong Daizhuang Hospital from January to December 2020 were selected as the observation group,and 26 healthy volunteers were selected as the control group.The observation group was treated with oral antidepressants for four weeks,and the severity of depressive symptoms was assessed by Hamilton depression scale(HAMD).The levels of serum TNF-α and ADP in the observation group before and after treatment and the control group were detected by ELISA,and the change of TNF-α/ADP was ana-lyzed.Results HAMD scores of the observation group were 25.5(21.0,30.0)before treatment and 2.0(1.0,4.0)after treatment,the difference was statistically significant(P<0.05).The levels of TNF-α and TNF-α/ADP in the observation group before treatment were higher than those in the control group,which af-ter treatment were also higher than those before treatment(P<0.05).The level of ADP in the observation group after treatment was lower than that before treatment and in the control group,the differences were sta-tistically significant(P<0.05).Conclusion The level of TNF-α/ADP in patients with depression is more ob-vious than TNF-α.