Application of MALDI-TOF MS in clinical difficult blood group typing / 中国输血杂志

【Objective】 To explore the application of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) in the genotyping of difficult blood typing samples, and to provide evidence for clinical blood transfusion. 【Methods】 Three ambiguous blood group samples, submitted to Shanghai Blood Center by Shanghai regional hospitals, were studied, of which Sample1 included the proband and his parents. Serological methods were used to perform blood group typing, direct antibody test, unexpected antibody screening and identification test. Blood group genotyping was performed by using the MALDI-TOF MS detection systeme stablished in our laboratory. Sanger sequencing was used to confirm gene mutation sites, and serological or flow methods were used to verify specific samples′ phenotype. 【Results】 Serological results indicated the existence of antibodies against high frequency antigens in sample 1 (including proband and her mother), 2 and 3. The genotyping results of MALDI-TOF MS showed that the proband of sample 1 was Di(a+ b+ ), her father was Di(a-b+ ), her mother was Di(a+ b-), sample 2 was p, and sample 3 was Jr(a-). Sequencing results of three samples were consistent with mass spectrometry typing results. Serological results showed that sample 2 had a p phenotype. The flow cytometry results suggested that sample 3 had a Jr(a-) phenotype. 【Conclusion】 For the first time, we applied MALDI-TOF MS technology to blood type genotyping of ambiguous clinical samples in China. Compared with other genotyping methods such as PCR-SSP, MALDI-TOF MS has the advantages of rapid detection, high throughput and high specificity, which would contribute to identification of difficult blood typing samples in the future, as well as rare blood group screening.

Quantitative detection of red blood cell antibody-mediated complement activation / 中国输血杂志

【Objective】 To construct an in-vitro model of erythrocyte antibody-mediated complement activation, and establish quantitative detection methods based on flow cytometry and spectrophotometry, so as to explore the correlation of anti-body titers and complement activation speed, and provide a methodological basis for studying the adverse transfusion reactions of anti-body mediated complement hemolysis. 【Methods】 Mouse monoclonal antibody that recognized human C3b and fluorescent secondary antibody were used to label C3b fragments on erythrocytes, and the deposition of C3b fragments after complement activation was detected by flow cytometry. The absorbance at 540 nm of the supernatant in the complement activation reaction system was measured by spectrophotometry as the amount of hemoglobin released was related to the absorbance. 【Results】 The complement activation system was constructed according to the ratio of 3% red blood cell suspension (mixed for 6 people) 1∶anti-Tja 1∶complement 2. The repeatability was good (P value>0.05) as different red blood cell mixtures had been used to repeat the detection reaction system. When using 32×, 64× and 128× dilutions of anti-Tja mediated complement activation, the deposition of C3b fragments has been detected by flow cytometry at 30 s, 1 min and 2 min, respectively, and MFI peaked at 5 min, 10 min and 30 min, respectively. No obvious hemolysis has been observed within 1.5 h. 【Conclusion】 In vitro model of anti-Tja-mediated complement activation demonstrates the speed of complement activation is related to the concentration of antibody. At a certain antibody concentration, the speed of complement activation has been slowed down, and no obvious hemolysis observed.

Physiologically based pharmacokinetic modeling for children and its application in pediatric drug research / 药学学报

Physiologically based pharmacokinetic (PBPK) modeling is an important tool to predict pharmacokinetic or pharmacodynamic profiles in special populations, especially in children and infants where designing and conducting clinical studies is difficult. The application of PBPK modeling can effectively promote the development of pediatric drugs and their clinical use. At present, PBPK modeling of pediatric populations is mainly applied in clinical trial design, drug-drug interaction (DDI) risk assessment, and dose selection in children. This review discusses the advantages of PBPK modeling in pediatric drug research and summarizes how to extrapolate a PBPK model from adults to children. The theoretical basis for pediatric PBPK models, the modelling process and important physiological parameters during the modeling process are introduced. Some successful applications of PBPK modeling in pediatric drug research and development are also presented. This review also analyzes the current limitations and future directions of pediatric PBPK modeling.

Inhibitory Effect of Feiji Recipe on IDO Induced Immune Escape on the Murine Model of Lewis Lung Carcinoma / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study the effect of Feiji Recipe (FR) intervening indoleamine 2,3-dioxygenase (IDO) induced immune escape on the murine model of Lewis lung carcinoma. Methods Totally 48 C57BL/6 mice inoculated with Lewis lung cancer cells transfected with human (enhanced green fluorescent protein,EGFP)-IDO gene were divided into four groups according to radom digit table, i.e., the model group (administered with normal saline by gastrogavage) , the Chinese medicine group (treated with FR Decoction at the daily dose of 100 mg/g by gastrogavage), the 1-methyl-D-trytaphan (1-MT) group (administered with 1-MT mixed liquor at the daily dose of 100 mg/kg by gastrogavage), and the Paclitaxel group (treated with Paclitaxel at the daily dose of 15 mg/kg by peritoneal injection), 12 in each group. The intervention was started from the 2nd day of modeling. The survival time was observed in 24 of them. Ratios of CD4+ CD25+ FoxP3+ regulatory T cells (Treg) in the spleen were detected in the rest 24 mice by flow cytometry respectively.</p><p><b>RESULTS</b>Compared with the model group, the survival time was significantly prolonged in the Chinese medicine group and the 1-MT group (P < 0.01); ratios of Treg cells remarkably decreased in the Chinese medicine group, the 1-MT group, and the Paclitaxel group (P < 0. 01). Compared with the Paclitaxel group, the survival time was significantly prolonged in the Chinese medicine group and the 1-MT group (P < 0.01); ratios of Treg cells decreased significantly in the 1-MT group (P < 0.05).</p><p><b>CONCLUSION</b>FR could inhibit the proliferation of lung cancer cells and immune eseape, improve the immune function, and prolong the survival of tumor-bearing mice.</p>

Establishment and application of cell models with stable expression of hOCTN1/2 / 药学学报

Human carnitine/organic cation transporter 1 and 2(hOCTN1 and hOCTN2) mediate transport of endogenous and exogenous compounds. The present study aimed to establish cell models with stable expression of hOCTN1 or hOCTN2 to study interactions with compounds and transporters. MDCK cells were transfected with pcDNA3.1(+) plasmid vector containing hOCTN1 or hOCTN2(pcDNA3.1(+)-hOCTN1/2), several stable transfected clones were obtained after G418 screening. hOCTN1 and hOCTN2 clones were screened with ergothioneine and mildronate respectively as substrates to identify the best candidates. We explored interactions of endogenous substances, alkaloids, flavonoids and ACEIs with hOCTN1/2. As a result, the cellular accumulation of ergothioneine in MDCK-hOCTN1 or mildronate in MDCK-hOCTN2 was 122 and 108 folds of the control cells, respectively. The kinetic parameters, Km and Vmax of ergothioneine, mediated by MDCK-hOCTN1, were 8.19±0.61 μmol·L-1 and 1427±49 pmol·mg-1(protein)·min-1; while Km and Vmax of mildronate by MDCKhOCTN2 were 52.3±4.3 μmol·L-1 and 2454±64 pmol·mg-1(protein)·min-1. Dopamine, glutamine, piperine, berberine, nuciferine, lisinopril and fosinopril could inhibit ergothioneine or mildronate uptake by MDCKhOCTN1/2. In conclusion, cell models with good stable hOCTN1 and hOCTN2 functions have been established successfully, which can be applied to the study of interactions between compounds and transporters of hOCTN1 and hOCTN2.

Effect of Interaction between PLIN Gene Polymorphisms and Open Lifestyle Intervention on Weight-loss in Chinese Han Adults / 中国医学科学院学报

<p><b>OBJECTIVE</b>To explore the effect of the interaction between PLIN gene polymorphisms and open lifestyle intervention on weight-loss in Chinese Han adults.</p><p><b>METHODS</b>Totally 109 overweight or obese subjects were assigned by random number table to the intervention group (n=56) or control group (n=53),and subjects in the intervention group received 22-week open lifestyle intervention. Anthropometric and metabolic indicators were measured for all subjects before and after intervention,and the PLIN1,PLIN4,and PLIN6 genotypes were amplified by polymerase chain reaction and sequenced through the first-generation sequencing technologies.</p><p><b>RESULTS</b>Among all these subjects,the rare allele C was dominant at PLIN1 (0.619),the common allele G was dominant at PLIN4 (0.606),and the common allele A was dominant at PLIN6 (0.564),in which PLIN1 and PLIN4 as well as PLIN4 and PLIN6 were in strong linkage disequilibrium (D'>0.9). After intervention,the body mass index,waist circumference,and body fat percent of female subjects were significantly decreased in intervention group and were lower than in control group;in male subjects,however,only the waist circumference showed significant difference with the control group (P<0.05). Subjects carrying rare allele homozygote of PLIN6 got less weight/fat loss than those carrying common alleles in intervention group,while subjects carrying rare allele of PLIN1 had more weight/fat increase than those with common allele homozygote in control group (P<0.05). Females in intervention group carrying any one of rare allele homozygotes of PLIN1,PLIN4 and PLIN6 got less weight/fat loss than those with common alleles,and female subjects carrying the rare allele homozygote haplotype of PLIN1/PLIN4 or PLIN4/PLIN6 got less weight/fat loss than those with other haplotypes (P<0.05).</p><p><b>CONCLUSION</b>The interaction between open lifestyle intervention and PLIN gene polymorphisms can directly influence weight-loss in Chinese Han overweight and obese adults.</p>

Establishment of platelet antigen panel and its application in the identification of platelet specific antibodies / 中华检验医学杂志

Objective To establish the platelet antigen panel for identifying the specificity of platelet antibodies which cause platelet transfusion refractoriness and neonatal alloimmune thrombocytopenia and provide evidence for clinical therapy and platelet genotyping research.Methods Based on the frequency distribution of human platelet alloantigen (HPA)-1 to HPA-16 gene in China, the frequencies of HPA-1 to HPA-6,HPA-15 alleles in blood group O donors were genotyped by the polymerase chain reaction with sequence-specific primers (PCR-SSP) method, and suitable donors were chosen to establish platelet-specific antigen panel.Using the established platelet-specific antigen panel, the specificity of platelet antibodies caused by alloimmune reaction was identified by using simplified sensitized erythrocyte platelet serology assay (SEPSA).Results Eleven ptatelet donors with blood group O were chosen to establish platelet-specific antigen panel which can identify specificity of HPA-1 to HPA-6, HPA-15 antibodies.One case of HPA-4b (Penb) and two cases of HPA-15a (Govb) platelet specific antibodies were detected in 1 120 samples.Conclusion Identifying the specific platelet antibodies using platelet specific antigen panel has profound significance on increasing the safety and effectiveness of clinical platelet transfusion and prevention of neonatal alloimmune thrombocytopenia.

Effects of Stroke Unit on the Compliance of Secondary Prevention in Patients with Stroke / 国际脑血管病杂志

Objective: To investigate the impact of stroke unit (SU) on the compliance of secondary prevention in patients with stroke at 12 months after stroke. Methods: Research subjects were stroke patients who were treated in SU (n = 500) and in general ward (GW) (n =445) using a design of retrospective study. The patients in the SU group were followed up by hospital, telephone and home interviews for 12 months, and the patients in the GW group were followed up by telephone interview for 12 months. The main outcome measures were the rate of using antithrombotics, the rate of smoking cessation, and the rates of awareness of early stroke symptom and stroke risk factors of patients. Results: he rate of using antithrombotics was 92.76% in the SU group, and it was significantly higher than 51.49% in the GW group (P ＜0.01); the rate of smoking cessation, and the rates of awareness of early stroke symptom and stroke risk factors of patients were 82.33%, 91.04%, and 94.03% respectively in the SU group, and they were significantly higher than 54.75%, 6.53%, and 70.37% in the GW group(P all ＜ 0.01 ). Conclusions: SU attaches importance to the secondary stroke prevention and emphasizes standardized treatment, and the compliance of the secondary stroke prevention in patient with stroke is improved significantly.