TEG evaluation and blood transfusion prediction model for patients with upper gastrointestinal bleeding / 中国输血杂志

【Objective】 To establish a blood transfusion outcome prediction model for comprehensivel evaluation of coagulation function of patients with upper gastrointestinal bleeding by thrombelastogram (TEG) and blood coagulation indicators. 【Methods】 The data of 101 patients with upper gastrointestinal hemorrhage, admitted to the Department of Gastroenterology of Zhejiang Provincial People′s Hospital and its Chun′an Branch from June 2018 to June 2021, were collected through Tongshuo blood transfusion management system and His system. Those patients were divided into blood transfusion group (n=56) and non-transfusion group (n=45), and into cirrhosis group (n=74) and non-cirrhosis group (n=27), and 40 patients, with non-upper gastrointestinal bleeding, were enrolled as the control. The results of TEG indicators (R, K, α, MA), coagulation function (PT, INR, APTT, TT, Fib), blood routine (Hb, Plt, WBC, NEUT%) and biochemical detection(Alb, SCr, ALT, AST, GGT) before transfusion were compared between groups and the correlation between TEG indicators and traditional coagulation parameters was analyzed. Single-factor and multi-factor analysis were used to screen blood transfusion-related factors to establish a predictive model. 【Results】 The comparisons of paremeters between transfusion and non-transfusion group were as follows, K (min), α (°), and MA (mm) was 3.86±3.12 vs 2.50±1.47, 54.00±14.08 vs 61.05±10.88, and 51.12±13.37 vs 58.26±11.08, respectively (P<0.01); PT (s) and Fib (g) was 16.36±7.45 vs 13.44±1.50 and 1.59±0.87 vs 2.35±1.09 (P<0.01); NEUT% and Hb (g/L) was 0.75 ±0.13 vs 0.66±0.15 and 68.04±14.49 vs 100.73±22.92 (P<0.01); Alb (g/L) and SCr (nmol/L) was 29.73±6.08 vs 33.73±7.19 and 99.50±53.55 vs 76.25±19.28 (P<0.01). Correlation analysis showed that APTT was positively correlated with R and K values, and negatively correlated with α and MA. Fib was negatively correlated with K values, and positively correlated with α and MA. Plt was negatively correlated with K values, and positively correlated with α and MA (P<0.01). Eight pre-transfusion indicators as K, MA, PT, Fib, NEUT%, Hb, Alb, and SCr were subjected to Logistic regression to establish a blood transfusion prediction model. The optimal ROC curve of blood transfusion threshold (blood transfusion predictive value of patients), sensitivity, specificity and AUC were 0.448, 92.9%, 88.9%, and 0.969, respectively. 【Conclusion】 The establishment of Logistic regression model by integrating detection indicators of TEG, coagulation function, blood routine and biochemistry in patients with upper gastrointestinal bleeding have showed significant correlation with blood transfusion prediction, and good clinical practicability.

Diagnosis of common blood stream infection pathogens based on central homo-sequence primer by multiplex PCR combined with MALDI-TOF MS / 中华检验医学杂志

Objective:Based on the high-throughput detection technique of multiplex PCR combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, constructing the characteristic SNP profiles of different strains, and establishing a rapid, accurate and highly sensitive method for the diagnosis of bloodstream infection pathogens.Methods:Seven kinds of pathogens such as common Escherichia coli were selected as target. The multiple PCR reaction conditions was optimized, and the characteristic peaks of each target bacteria were detected by MALDI-TOF MS to establish the joint detect system. Common primer pairs and central homo-sequence primer pairs were designed to analyse the formation of primer dimer. Using simulated bacterial infection blood samples with detection system to determine specificity and sensitivity. One hundred and fifty blood samples from suspected bacteremia patients were collected from June to September 2020 in a hospital in Beijing, and the identification results were compared to traditional identification method of clinical application that are using χ 2 test. Results:The cycle threshold (Ct) value of the central homo-sequence primers that were designed were more than 38, with a delay of 6-10 cycles. The joint mass spectrometry detection system could detect seven kinds of bacteria divided into two groups at the same time. The target bacteria can be detected specific product of the peak, and the clinical strains other than the target strains only had primer peaks. All maps had non-specific miscellaneous peaks. The sensitivity of Escherichia coli could reach 50 CFU/ml, and the detection limit of other bacteria was 100 CFU/ml. The detection results of 150 patients showed that 46 cases were positive by traditional method. The positive rate was 30.67% (46/150), including two cases of mixed infection. Forty-eight cases were positive by mass spectrometry, and the positive rate was 32.0% (48/150), including three cases of mixed infections. The negative coincidence rate was 100% (101/101). The comparison of the two methods showed that the P=0.625>0.01, the Kappa=0.938, the sensitivity and specificity was 97.82%(45/46) and 97.11%(101/104), respectively. There was no significant difference between the two methods, and the results of nucleic acid mass spectrometry could also be used in clinic. Conclusions:The established detection system can not only quickly and accurately detect seven common pathogens causing bloodstream infection, and effectively shorten the time needed for traditional culture and identification, but also can detect multiple bacterial mixed infections at the same time to make up for the possibility of missed detection. Besides, the method can also be used to identify other bacteria.

Analysis of TWNK variant in a family affected with Perrault syndrome / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic etiology of two patients with Perrault syndrome (PRLTS) in a family.@*METHODS@#Whole exome sequencing (WES) was carried out to screen potential variants within genomic DNA extracted from the proband. Suspected variants were validated by clinical data and results of Sanger sequencing.@*RESULTS@#WES has identified two heterozygous variants of TWNK gene, namely c.1172G>A (p.Arg391His) and c.1844G>C (p.Gly615Ala). Sanger sequencing confirmed that the c.1172G>A (p.Arg391His), a known pathogenic variant, was derived from her father, while the c.1844G>C (p.Gly615Ala), a novel variant, was derived from her mother. Her brother, who was similarly affected, has carried the same compound heterozygous variants.@*CONCLUSION@#The compound heterozygous variants c.1172G>A (p.Arg391His) and c.1844G>C (p.Gly615Ala) of the TWNK gene probably underlie PRLTS in the sib pair. The above results have facilitated genetic counseling and prenatal diagnosis for the affected family.

Variant analysis for a pedigree affected with limb-girdle muscular dystrophy type 2D / 中华医学遗传学杂志

OBJECTIVE@#To analyze variant of SGCA gene in a Chinese pedigree affected with limb-girdle muscular dystrophy type 2D with whole exome sequencing (WGS).@*METHODS@#Multiplex ligation-dependent probe amplification (MLPA) was employed to detect large fragment deletion or duplication of the DMD gene. FastTarget next generation sequencing was used to detect variants of the DMD gene, and the result was verified by Sanger sequencing. After excluding the diagnosis of DMD for the proband, WGS was applied to test the proband and his parents. Suspected pathogenic variants were validated by Sanger sequencing.@*RESULTS@#No variant, deletion or duplication of the DMD gene was detected. Whole exome sequencing showed that the proband has carried compound heterozygous missense variants c.409G>A (p.Glu137Lys) and c.409G>C (p.Glu137Gln) in exon 5 of the SGCA gene, which were respectively inherited from his mother and father. Neither variant was found in DNA derived from the cord blood sample.@*CONCLUSION@#The c.409G>A (p.Glu137Lys) and c.409G>C (p.Glu137Gln) compound heterozygous missense variants probably underlie the disease in the proband. Above finding has facilitated genetic counseling and prenatal diagnosis for the family.

Whole exome sequencing analysis for a Chinese pedigree affected with X-Linked intellectual disability / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the clinical features and genetic mutation in a family affected with non-syndrome X-linked intellectual disability (NS-XLID) using whole exome sequencing (WES).</p><p><b>METHODS</b>Multiplex ligation-dependent probe amplification (MLPA) was applied to screen potential mutations of Fragile X syndrome (FXS). Whole exome sequencing (WES) and Sanger sequencing were screen for pathological mutations.</p><p><b>RESULTS</b>FXS was excluded by MLPA analysis. WES has discovered in the proband an ARX gene mutation c.88G>T, which was confirmed by Sanger sequencing. Combining his clinical phenotype with information from the OMIM database, it was inferred that the ARX mutation probably underlies the NS-XLID in the proband. The same mutation was found in his mother and two uncles but not in his father and sister.</p><p><b>CONCLUSION</b>WES is capable of revealing the mutation underlying NS-XLID and can facilitate genetic counseling for the affected families.</p>

Analysis of genome-wide copy number variations among fetuses with abnormalities detected by prenatal ultrasouography / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the genetic etiology of fetal abnormalities detected by prenatal ultrasound through single nucleotide polymorphism (SNP array) analysis.</p><p><b>METHODS</b>Two hundred and eight fetuses were tested with SNP array and conventional karyotyping. Complex copy number variations (CNVs) were verified with fluorescence in situ hybridization (FISH), multiplex ligation-dependent probe amplification (MLPA) and quantitative fluorescence polymerase chain reaction (QF-PCR).</p><p><b>RESULTS</b>For the 208 cases, the diagnostic yields of conventional karyotping and SNP assay were 8.2%(17/208) and 13.9%(29/208), respectively. For fetuses with malformations of the cardiovascular system, central nervous system or multiple systems, pathogenic CNVs was detected in 4.6% (8/174), 2.3%(4/174), and 1.1% (2/174) of all fetuses, respectively. No pathogenic CNVs was detected among those with abnormalities of the renal system, digestive system, skeletal system, facial dysmorphism or respiratory system.</p><p><b>CONCLUSION</b>CNVs are significantly related with birth defects. Compared with conventional karyotyping, SNP array is a better platform for CNVs detection and can provide more clues for genetic counseling, recurrence risk assessment and prenatal diagnosis.</p>

Molecular cloning and expression of OspC protein of a Chinese Borrelia afzelli FP1 strain and pre-liminary study on the immune protectivity of the rOspC protein / 中华微生物学和免疫学杂志

Objective To clone and express the outer surface protein C ( OspC) from a Chinese Borrelia afzelli FP1 strain and to evaluate the immune protectivity of the recombinant OspC protein ( rOspC) . Methods The gene encoding OspC protein of Borrelia afzelli FP1 strain was amplified by polymerase chain reaction (PCR) and then inserted into pET-30a plasmid to construct the recombinant expression plasmid pET-30a-OspC. The transformed E. coli BL21 strains carrying pET-30a-OspC plasmid were induced by IPTG to express OspC protein. The expressed proteins were purified by Ni-IDA resin chromatography and analyzed by SDS-PAGE and Western blot assay. Indirect immunofluorescence assay ( IFA) was performed to detect anti-rOspC protein antibodies in serum samples from rabbits immunized with rOspC protein. In vitro neutral-ization test was performed for evaluation the immune protectivity of rOspC protein. Results The recombi-nant expression plasmid pET-30a-OspC was successfully constructed and highly expressed in E. coli BL21. A strong antigen-antibody reaction between the rOspC protein and polyclonal antibody against Borrelia afzelli FP1 strain was detected by Western blot assay. The titers of IgG in serum samples from rabbits immunized with rOspC protein were significantly elevated. The in vitro neutralization test indicated that 106/ml of Borre-lia afzelli FP1 strains were neutralized by every anti-OspC protein serum sample from the experiment group. Conclusion The rOspC protein showed a strong immune protectivity against Borrelia afzelli, which could be used in the development of polyvalent subunit vaccine against lyme disease.

Effect of mitochondrial DNA 5178 C/A polymorphism on risks for type 2 diabetes mellitus and its complications / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the role of mitochondrial DNA 5178 C/A (Mt5178) polymorphism of NADH-dehydrogenase subunit 2 (ND2) gene in type-2 diabetes mellitus (T2DM) among ethnic Han Chinese through a case-control study.</p><p><b>METHODS</b>The Mt5178C/A polymorphism was determined by sequencing 1103 T2DM patients and 791 healthy controls. Logistic regression analysis was conducted to estimate odds ratios (OR) and 95% confidence intervals (CI). To confirm the results, a meta-analysis was conducted based on published literature on the association of Mt5178 variant with T2DM.</p><p><b>RESULTS</b>No significant association was found between the Mt5178C/A variant and T2DM either by our study or the meta-analysis which included eight published studies. Nevertheless, it was found that the T2DM patients with 5178C genotype were at a higher risk for nephropathy complication (OR=1.49, 95%CI: 1.005-2.197, P<0.05) and at significantly lower risk for hypertension complication (OR=0.744, 95%CI: 0.556-0.996, P<0.05) compared with those carrying a 5178A genotype.</p><p><b>CONCLUSION</b>No association was found between the Mt5178C/A polymorphism of mitochondrial ND2 gene with the increased risk of T2DM. However, the polymorphism may affect the development of nephropathy and hypertension complications among T2DM patients.</p>

Progress of new biological markers based on proteomic approaches in colon cancer / 国际肿瘤学杂志

Colon cancer is the most common malignant tumor in the worldwide scale and it is one of the leading causes of cancer-related deaths.Finding accurate and informative cancer markers will provide significant insights in diagnosis and treatment strategy of colon cancer.Proteomic technology is an important tool in cancer research.In recent years,many studies have found a great of potential biomarkers of colon cancer.

NK4 growth inhibition of human Raji lymphoma xenografts by competitive interrupting HGF/Met signal pathway / 中华病理学杂志

<p><b>OBJECTIVE</b>To observe the inhibition of NK4 protein in the proliferation of human Raji lymphoma xenografts in nude mice, and to explore its molecular mechanism.</p><p><b>METHODS</b>Models of human Raji lymphoma xenograft transfected with HGF gene were established by subcutaneous inoculation in nude mice. After establishment of the models, the mice received continuous NK4 protein via tail vein for 4 weeks, and the weight and tumor growth were monitored every week. After 8 weeks, the expression of HGF mRNA and c-Met mRNA of tumor tissues was measured by real-time fluorescent quantitation PCR. The apoptotic index (AI) and microvessel density (MVD) were evaluated by terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) and immunohistochemistry, respectively.</p><p><b>RESULTS</b>The models of human Raji lymphoma xenograft were successfully established. Although the animal weights of all groups declined, especially in the groups with NK4 protein injection, there was no statistical significance (P > 0.05). The tumor volume in HGF gene transfected group was larger than those of the control groups (P < 0.01), and there was no statistical significance among the control groups (P > 0.05). However, the tumor volume of the NK4 protein injection group decreased significantly (P < 0.01). Expression of HGF mRNA and c-Met mRNA in HGF gene transfected group increased significantly after injection of NK4 protein (P < 0.01). AI in HGF gene transfected group (33.5% ± 12.3%) was significantly lower than that of control groups (89.1% ± 22.3% vs. 81.9% ± 27.0%, P < 0.05), but became significantly higher (119.1% ± 18.9%) after NK4 protein injection (P < 0.01). MVD in HGF gene transfected group (28.5 ± 2.0) was higher than that of control groups (12.2 ± 1.4, 13.8 ± 1.3, P < 0.01), although declined (15.5 ± 2.5) after NK4 protein injection (P < 0.01).</p><p><b>CONCLUSIONS</b>NK4 protein suppresses significantly the growth of human Raji lymphoma xenografts transfected with HGF gene. The pathogenesis may be involved in promoting tumor cell apoptosis and restraining tumor angiogenesis through competitive interrupting HGF/Met signal pathway.</p>

Establishment and application of quantitative detection for heteroplasmic mtDNA mutation 3243A→G / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To develop a rapid, simple, cost-effective, accurate and sensitive method for quantitative detection of mitochondrial DNA (mtDNA) 3243A→G mutation in order to provide reference for selecting the best detection method under different conditions.</p><p><b>METHODS</b>Genomic DNA was extracted from peripheral leucocytes of 17 individuals from a Wenzhou family featuring maternally inherited diabetes and deafness (MIDD). Heteroplasmic level of mtDNA 3243A→G mutation was determined respectively with polymerase chain reaction-restriction fragment length polymorphism (PCR-RLFP), real time-amplification refractory mutation system-quantitative PCR (RT-ARMS-qPCR) and pyrosquencing. Eleven plasmids with various heteroplasmic levels of the 3243A→G mutation (ranging from 0 to 100%)were constructed as the standards. The reliability of above methods was compared by correlation coefficient based on observed and expected values.</p><p><b>RESULTS</b>For all three methods, measurement of the standards showed a linear correlation between the expected and detected values, i.e., PCR-RFLP (R(2)=0.828), RT-ARMS-qPCR (R(2)=0.998) and pyrosquencing (R(2)=0.997). For the MIDD family, it was consistent that there are 13 members carrying the A3243G mutation with different heteroplasmic levels. And there was no significant difference between the results by RT-ARMS-qPCR and pyrosquencing.</p><p><b>CONCLUSION</b>PCR-RFLP is not appropriate for the quantitative detection but could be used for early clinical screening. Both RT-ARMS-qPCR and pyrosquencing are suitable for the detection of low heteroplasmic level of A3243G mutation. Compared with pyrosquencing, RT-ARMS-qPCR is rapid, reliable and cost-effective, and is the best choice for detecting low mutation loads.</p>

Investigation of a family with Kennedy disease by genetic analysis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To report on a Chinese family from Wenzhou with genetically confirmed Kennedy disease and describe its clinical and genetic features.</p><p><b>METHODS</b>The clinical phenotype and the level of relevant biochemical markers were assessed. To determine the number of CAG repeats in the exon 1 of androgen receptor (AR) gene, genomic DNA was extracted from peripheral blood samples of the family members, amplified by PCR and identified by DNA sequencing.</p><p><b>RESULTS</b>The proband showed predominantly proximal limb weakness, fasciculation, muscle atrophy, gynecomastia, sexual dysfunction and increased serum creatine kinase. Myopathy and neuropathy were identified by electromyography. Two other affected males and 2 affected female carriers were identified to carry an expanded CAG repeat in the AR gene. The numbers of CAG repeats were found to be 43 in the proband, 43 and 42 in the other two affected males, one of which had similar clinical symptoms to the proband.</p><p><b>CONCLUSION</b>The family was diagnosed with Kennedy disease by analysis of the AR gene.</p>