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Objective:To evaluate the capacity and level of melting point determination of chemical drugs in the laboratories par-ticipating in the proficiency testing. Methods:Two test samples were prepared, and the labs volunteered to participate in the proficien-cy testing program ( PTP) . The melting point determination was performed according to the general principle 0612 in part four of Chi-nese Pharmacopoeia (2015 edition), and the results were analyzed by robust statistics and the determination proficiency of the laborato-ries were evaluated by Z-score. Results:The analysis showed that two test samples were homogeneous and stable, which met the re-quirements of the PTP. Totally 31 laboratories had satisfactory results, which accounted for 83. 8%. Conclusion:The majority of the participant laboratories can accurately determine the melting points of test samples, and the information is very helpful to the next profi-ciency testing program.
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To find out whether the integration of PERV in HEK293 cells influence the expression profile of HERV-W,based on the gene sequences of H ERV-W in GenBank,the primers of H ERV-W gag,pol,env,env from locus 7q21.2 genes as well as humanβ-actin were synthesized respectively and were used to develop the means of SYBR Green I real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the mRNA expression of these genes in HEK293 cells and PERV-infected HEK293 cells.Experiments showed that these RT-qPCR methods were of good specificity,sensitivity and stability:the standard curves could ensure the correlation coefficients to be all above 0.99 and the amplification efficiency were between 95％ and 110％,which verified that these methods could be used to detect the mRNA of HERV-W and humanβ-actin in culture cells.Through the detection and analysis of relative gene expression data using the 2-△△ct method,we found that the mRNA level of HERV-W gag,pol,env,env from locus 7q21.2 genes as well as humanβ-actin in PERV-infected HEK293 cells increased,after integration,by 37.08,42.56,2.49 and 13.17 times than in non-infected HEK293 cells,respectively.Results provide a reference to further evaluate the safety of PERV pathogen in xenotransplantation.
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Objective To learn the levels of serum 25-(OH)D in 0 12 years old chindren in Kunming.Methods Serum 25-(OH)D levels of children (4 498) were measured by using Roche cobas 8 000 / E602 automatic electrochemical luminescence immunoassay.Results As the increase of age,the levels of serum 25-(OH)D were gradually decreased,different age groups had significant difference (P＜0.05).The levels of 25-(OH)D between boys and girls had no significant difference (P＞0.05);The average level of sernm 25-(OH)D was 83.48 nmol/L,the level of serum 25-(OH)D was distributed in 0 ～ 1 years old group,and most concentrated in the 6 ～ 9 years old group.Conclusion The levels of 25-(OH)D in 0 ～ 12 years old chindren are in a good status,but with the increase of age,the level of 25-(OH)D is decreasing gradually,especially the levels of 25-(OH)D decreased significantly after the age of three,in shortage or lack of status,this should cause everybody's attention.
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Objective To evaluate the values of combined detection of soluble mesothelin related proteins (SMRP)and carbohydrate antigen 1 25 (CA1 25)in the diagnosis of malignant peritoneal mesothelioma.Methods 20 patients with malignant peritoneal mesothelioma were selected as the malignant group,50 patients with benign ovar-ian tumor were selected as the benign group,60 healthy women were selected as the control group.Enzyme -linked immunosorbent assay(ELISA)was adopted to detect serum SMRP levels and electrochemiluminescence immunifaction (ECLI)was adopted to detect serum CA1 25 levels.The SMRP,CA1 25 levels were compared among all groups,the sensitivity,specificity of SMRP and CA1 25 in the diagnosis of peritoneal malignant mesothelioma were analyzed. Results Serum levels of SMRP,CA1 25 in malignant group were (1 4.8 ±1 .6 )μg/L,(1 89.1 ±1 5.3 )μg/Lrespectively,which were significantly higher than those in the benign group[(3.9 ±0.9)μg/L,(28.6 ±4.2)μg/L] (t =27.40,49.61 0,all P <0.01 )and the control group[(2.8 ±0.7)μg/L,(1 6.9 ±3.8)μg/L](t =29.877, 53.334,all P <0.01 ).The sensitivities of SMRP,CA1 25 detection alone for malignant peritoneal mesothelioma diag-nosis were 65.0%,50%,respectively,while their specificities were 83.6%,79.1 %.The sensitivity and specificity of combined detection of SMRP and CA1 25 for malignant peritoneal mesothelioma were 95.0% and 93.6%.Conclusion SMRP has some value in the diagnosis of peritoneal malignant mesothelioma.SMRP combined with CA1 25 detection can markedly improve the diagnostic sensitivity and specificity for malignant peritoneal mesothelioma.
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Objective:To establish an HPLC method for the content determination of gliquidone tablets to improve the specificity of the content determination and the rationality of the preparation of test solution. Methods: A UPLC-MS system was used to analyze the degradation products with positive and negative ion scanning and sub-ion scanning. An ACQUITY UPLC BEH C18 column(2. 1 mm × 50 mm,1. 7 μm) was employed with the mobile phase consisting of water (adjustting pH to 3. 5 with formic acid)-acetonitrile with gradient elution. The HPLC method was performed on an Agilent Zorbax SB-C18 column(150 mm × 4. 6 mm,5 μm). Water (adjusing pH to 3. 5 with formic acid)-acetonitrile(37. 5∶62. 5) was used as the mobile phase. The detection wavelength was set at 230 nm. The column temperature was set at 30℃. The flow rate was 1. 0 ml·min-1 with the injection volume of 20μl. Results:A good linear re-lationship was obtained within the range of 60. 200-140. 400μg·ml-1(r=0. 999 5), and the average recovery was 98. 60% with RSD of 0. 6% (n=9). Conclusion:The method is accurate, reliable, specific and reproducible, which can be used in the determination of content and content uniformity of gliquidone tablets.
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Objective To investigate the condition of nutrition in child with cerebral palsy (CP) and the effect of nutritional intervention. Methods 49 CP children and other 60 health children (controls) were measured their bodies, hemoglobin, serum trace elements, and bone mineral density (with ultrasonic), and the feeding behavior was also investigated. Results The incidence of malnutrition was 48.97%, in which 26.53%for low weight. The levels of serum iron and zinc were poor in the CP children, and the incidence of iron deficiency anemia was 34.67%in the CP children, different from the controls (P0.05). Feeding problems were found in 44.9%of CP children. About 50%of malnutrition was cor-rected, especially the body weight after 4 months of Intervention, with anemia corrected in 88.2%, and bone mineral density recovered in 50%. Conclusion It is a problem for many CP children with malnutrition and nutritional disorders, and need nutrition intervention as the content of the rehabilitation.
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This paper is to report the polymorphism of raw materials of clopidogrel bisulfate at home and abroad. By the analysis of Fourier transform infrared spectroscopy (FTIR) and powder X-ray diffraction (p-XRD), samples are roughly classified into two groups, except one patent material. And the differential scanning calorimeter (DSC) examination showed more detailed information for these materials. The results of the study could provide comprehensive basis for the quality evaluation of clopidogrel bisulfate.
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Objective To investigate the relationship between Atherogenic Index of Plasma(AIP) and the dorsalis pedis artery atherosclerosis in type 2 diabetes. Methods One hundred and twenty-seven patients with type 2 diabetes and 33 healthy individuals were included in the study. The diabetic patients were devided into 2 groups, with( n = 66) or without ( n = 61 ) atherosclerosis. The intima media thickness (IMT) of dorsalis pedis artery were measured by B-mode Doppler ultrasonography. The triglyceride(TG) ,high density lipoproteincholesterol(HDL-C) and other glucolipid metabolic indices were measured in all patients. The AIP was defined as the log value of TG/HDL-C ratio. The one-way ANOVA test was performed to determine the differences among three groups. The relationships between IMT and the other indicators were assessed using Pearson bivariate correlations analysis. The optimal operating point of AIP to diagnose the dorsalis pedis artery atherosclerosis in type 2 diabetes was obtained by drawing ROC curve. Results The AIP value in atherosclerosis diabetic patients,no-atherosclerosis diabetic patients and healthy subjects were 0. 229 ± 0. 132,0. 112 ± 0. 074 and -0. 045 ± 0. 033 respectively. The IMT value were (0. 71 ± 0. 24 ) mm, (0. 49 ± 0. 09 ) mm and (0. 34 ± 0. 15 ) mm respectively. Both indices shown statistical differences among the three groups (Ps = 0. 001 respectively). After adjusting for potential influence factors, AIP showed positive correlation with IMT( r = 0. 315, P = 0. 001 ). The OOP of AIP was 0. 1671 ,the area under the curve was 0. 783. Conclusion AIP was well related to the dorsalis pedis artery atherosclerosis in type 2 diabetes,and the OOP of AIP had good diagnostic reference value in these patients.
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Objective To analyze the differential expression of Stathmin in human cells infected with human-tropic porcine endogenous retrovirus(PERV)and to explore the potential molecular effect of human-tropic PERV on human cells.Methods HEK293 cells were infected with the human-tropic PERV infectious molecular clone.PCR,real-time RT-PCR and immunofluorescence analysis were applied to confirm that HEK293 cells were infected.Then real-time RT-PCR and Western blot were carried out to analyze the differential expression of Stathmin at the mRNA level and protein level,respectively.Results HEK293 cells were infected by human-tropic PERV.Real-time RT-PCR and Western blot analysis showed that Stathmin was up-regulated in HEK293 cells infected with PERV compared with the control cells.Conclusion Stathmin was up-regulated in HEK293 cells infected with human-tropic PERV.These studies will be helpful for revealing the interaction of PERV and human cells,and for understanding the molecular effect of humantropic PERV on human cells.In addition,it suggested that PERV infection may infect cell growth and physiological functions,even be pathogenic.These will help to clarify the biologic characteristics of PERV and evaluate the safety of PERV in pig to human xenotransplantation.
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An HPLC-UV method has been developed for the determination of valibose, miglitol, voglibose and acarbose, the four anti-diabetic drugs. The separation was accomplished successfully by using reversed phase chromatography (Prevail carbohydrate column, 250 mm x 4.6 mm, 5 microm) with a gradient acetonitrile-phosphate buffer solution (pH 8.0) at a wavelength of 210 nm. Furthermore, the method of a high-performance liquid chromatography coupled with ESI-MS in positive ionization mode has been established. These two methods were successfully applied to the assay and qualitative detection of four alpha-glucosidase inhibitors in the potential counterfeit anti-diabetic drugs.
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The aim of this study is to construct a bifunctional fusion protein, which can conjugate both human red blood cells and antibodies against classical swine fever virus (CSFV). We respectively amplified 2E8ScFv and mE2 genes from different recombinant vectors, in which 2E8ScFv gene is the single chain Fv gene against H antigen of human red blood cells, whereas mE2 gene is the main antigen coding region gene of CSFV E2 protein. We used overlap extension PCR to obtain an artificial fusion gene segment 2E8mE2 containing genes of Both 2E8ScFv and mE2, then ligated into the expression vector pET-DsbA and expressed in Escherichia coli BL21(DE3) PlysS host cells, after induced with IPTG the target fusion protein was successfully expressed and identified in inclusion bodies by SDS-PAGE and Western blotting. We purified the fusion protein and renatured it from inclusion bodies to obtain a native state of well biological activity. The Erythrocyte agglutination test results indicated that the fusion protein can conjugate both human red blood cells and antibodies of CSFV.
Subject(s)
Humans , Erythrocytes , Allergy and Immunology , Escherichia coli , Genetics , Metabolism , Immunoglobulin Fragments , Genetics , Allergy and Immunology , Immunoglobulin Variable Region , Genetics , Allergy and Immunology , Recombinant Fusion Proteins , Genetics , Single-Chain Antibodies , Genetics , Viral Envelope Proteins , GeneticsABSTRACT
Objective To compare the clinical effects of hemiarthroplasty and dynamic hip screw (DHS) in treatment of unstable femoral intertrechanterie fractures in the elderly. Methods A total of 62 elderly patients with unstable femoral intertrochanteric fractures treated from January 2004 to April 2007 were randomly divided into DHS group and hemiarthroplasty group, with 31 patients in each group. The patients were foLlowed up to compare the clinical effects between two groups. Results Of all, 58 patients ( including 28 patients in the hemiarthroplasty group and 30 in DHS group) were followed up for 13-49 months. The mean operation time was ( 76.07 ± 6.29 ) minutes in hemiarthroplasty group and ( 117.50 ± 8.88 ) minutes in DHS group ( P < 0.05 ). The mean blood loss during operation was (305.36±34.26) ml in hemiarthroplasty group and (431.67±53.31) ml in DSH group (P<0.05). The postoperative complications occurred in three patients in hemiarthroplasty group and 10 in DHS group, with statistical difference ( P < 0. 05 ). The Harris score for hip function was ( 85.46 ± 3.05 ) points in hemiarthroplnsty group and ( 79.73 ± 2.57 ) points in DHS group, with no statistical difference between two groups ( P > 0.05 ). Conclusions DSH is still one of effective methods for patients with unstable femoral intertrochanterie fractures. Hemiarthroplasty can diminish bleeding during operation, help walk earlier and reduce complications and is a better selection for the elderly patients with osteoporosis and unstable femoral intertrochanteric fractures. However, hemiarthroplasty can not replace DHS.
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Objective The aim of this study was to establish a new method for genotyping ATP7B Arg778Leu gene mutation that does not require RFLP PCR or sequencing.Method 4-primer amplification refractory mutation system (ARMS)-PCR was performed to screen the Arg778Leu mutation in 47 unrelated Wilson's disease (WD) patients and 30 unrelated healthy controls.Direct sequencing was used to confirm the specific amplification products.Results PCR products were visualized on agarose gel electrophoresis.Among the 47 WD patients,4 were homozygous and 14 were heterozygous for this mutation.The total mutation rate was 38.3% (18/47).The results of direct sequencing completely consisted with the results of 4-primer ARMS-PCR.Conclusions The ATP7B Arg778Leu gene mutation is a hot spot for the research of Chinese WD patients.4-primer ARMS-PCR is a fast convenient and accurate method for typing mutation in high throughput population screening.This approach can be used to detect other point mutations.
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The expression of synaptotagmin Ⅱ (Syt2) in RBL-2H3 (RBL) and its role during exocytosis of RBL was investigated. The expression of Syt2 in RBL was detected by western blot and Syt2 gene was amplified by PCR. The anti-sense full length Syt2 cDNA expression vector was constructed with pEGFP-N1 and transfected into RBL by electroporation, and stable transfectants were selected by using G418. To analyze the role of Syt2 during exocytosis of RBL, the release of cathepsin D was assayed by immunoblotting. The results showed that Syt2 was expressed in RBL.The anti-sense expression vector pEGFP-N1-Syt2-AS was constructed and the sequence of insertion was completely consistent with rat Syt2 (accession number in GeneBank: NM012665). The stable transfectants (RBL-Syt2-AS) were obtained. Western blot showed that RBL-Syt2-AS expressed a lower level of Syt2 (8 % and 10 % of control cells), indicating that the expression of Syt2 in RBLSyt2-AS was markedly down-regulated by anti-RNA. Compared with control, the release of cathepsin D by RBL-Syt2-AS was increased. It was concluded that Syt2 expressed in RBL and could inhibit exocytosis of lysosomes in RBL.
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In order to investigate whether Arg110Gln polymorphism in the coding region of the IL-13 gene is associated with asthma and total plasma IgE level in Han nationality in Hubei Chinese population, the allele frequency of 4257(g/a) site and Arg110Gln genotype of IL-13 was detected by using restriction fragment length polymorphism in Han nationality in Hubei Chinese population including 43 asthmatic children, 45 asthmatic adults, 31 control children and 46 control adults. Total plasma IgE was measured by Chemiluminescence assay. The results showed that the frequency of allele A at 4257 bp of IL-13 in children and adults was 0.39 and 0.32, respectively. The GlnGln form of Arg110Gln polymorphism of IL-13 gene was associated with susceptibility of asthma and elevated total plasma IgE in children (P=0.030 and 0.0009, respectively), but not with them in adults (P=0.219 and 0.174, respectively). Our results suggest that the Arg110Gln polymorphism of IL-13 gene is associated with susceptibility of asthma and elevated total plasma IgE in Chinese children of Han nationality in Hubei, but not with them in adults.
Subject(s)
Adult , Female , Humans , Male , Asthma , Genetics , China , Ethnology , Chromosomes, Human, Pair 5 , Genetics , Gene Frequency , Genetic Predisposition to Disease , Genetics , Immunoglobulin E , Blood , Interleukin-13 , Genetics , Polymorphism, Genetic , GeneticsABSTRACT
In order to investigate whether Arg110Gln polymorphism in the coding region of the IL-13 gene is associated with asthma and total plasma IgE level in Han nationality in Hubei Chinese population, the allele frequency of 4257(g/a) site and Arg110Gln genotype of IL-13 was detected by using restriction fragment length polymorphism in Han nationality in Hubei Chinese population including 43 asthmatic children, 45 asthmatic adults, 31 control children and 46 control adults. Total plasma IgE was measured by Chemiluminescence assay. The results showed that the frequency of allele A at 4257 bp of IL-13 in children and adults was 0.39 and 0.32, respectively. The GlnGln form of Arg110Gln polymorphism of IL-13 gene was associated with susceptibility of asthma and elevated total plasma IgE in children (P=0.030 and 0.0009, respectively), but not with them in adults (P=0.219 and 0.174, respectively). Our results suggest that the Arg110Gln polymorphism of IL-13 gene is associated with susceptibility of asthma and elevated total plasma IgE in Chinese children of Han nationality in Hubei, but not with them in adults.
Subject(s)
Asthma/genetics , China/ethnology , Chromosomes, Human, Pair 5/genetics , Gene Frequency , Genetic Predisposition to Disease/genetics , Immunoglobulin E/blood , Interleukin-13/genetics , Polymorphism, Genetic/geneticsABSTRACT
<p><b>OBJECTIVE</b>To investigate the link between the polymorphism of -109 and Glu237 in the high-affinity IgE receptor beta (FcepsilonRIbeta) gene and susceptibility to allergic asthma in a Chinese population.</p><p><b>METHOD</b>Blood samples from 216 allergic asthma patients and 198 age- and sex-matched controls were studied. A -109C/T and a coding variant Glu237Gly in FcepsilonRIbeta were detected with polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).</p><p><b>RESULTS</b>The genotype frequencies were 0.403 for -109T/T, 0.491 for -109T/C and 0.106 for -109C/C in allergic asthma in a Chinese population. No significant difference in the distribution of -109C/T polymorphism was found between allergic asthma subjects and healthy controls, however, homozygosity for the -109T allele was associated with increased total plasma IgE levels in subjects with allergic asthma (F = 4.020, P < 0.05). The allele frequency of Gly237 in the patients and control was 0.236 and 0.136 respectively. There was a significant association between the Gly/Gly genotype and allergic asthma. Among allergic asthma patients Gly237 was significantly associated with high IgE levels.</p><p><b>CONCLUSIONS</b>These results suggest that the Gly237 variant of the FcepsilonRIbeta gene is involved in the development of allergic asthma. The -109C/T and Glu237Gly polymorphisms are two of the genetic factor identified thus far, which affect total plasma IgE levels of allergic asthma patients in a Chinese population.</p>
Subject(s)
Adolescent , Adult , Aged , Child , Child, Preschool , Female , Humans , Male , Middle Aged , Asian People , Genetics , Asthma , Genetics , China , Polymorphism, Genetic , Receptors, IgE , GeneticsABSTRACT
The relationship between 3 polymorphisms sites [interleulin-4 (IL-4), IL-4 receptor (IL-4R) alpha chain and activation-induced cytidine deaminase (AICDA)] and adult allergic asthma in China was studied. By using case-control method, DNA and clinical data were obtained from allergic asthmatic patients and compared with those in the control subjects. The subjects were genotyped for the IL-4 C-589T promoter polymorphism, the IL-4R alpha chain Q576R and the AICDA C8408T by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The results showed that the IL-4 C-589T was not associated with adult allergic asthma in China. However, the IL-4R alpha chain 576R/R and AICDA 8408T/T frequency was significantly increased in allergic asthma group as compared with that in the control group [odd ratio (OR) = 3.797 and 9.127, respectively; P < 0.01)] and was correlated with the increased plasma total IgE. These data suggested that the IL-4R alpha chain 576R/R and AICDA 8408T/T genotypes confer genetic susceptibility to adult allergic asthma in China.
Subject(s)
Adult , Humans , Alleles , Asthma , Genetics , Cytidine Deaminase , Genetics , Immunoglobulin E , Blood , Interleukin-4 , Genetics , Phenotype , Polymorphism, Restriction Fragment Length , RNA Processing, Post-Transcriptional , Receptors, Interleukin-4 , GeneticsABSTRACT
The relationship between 3 polymorphisms sites [interleulin-4 (IL-4), IL-4 receptor (IL-4R) alpha chain and activation-induced cytidine deaminase (AICDA)] and adult allergic asthma in China was studied. By using case-control method, DNA and clinical data were obtained from allergic asthmatic patients and compared with those in the control subjects. The subjects were genotyped for the IL-4 C-589T promoter polymorphism, the IL-4R alpha chain Q576R and the AICDA C8408T by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The results showed that the IL-4 C-589T was not associated with adult allergic asthma in China. However, the IL-4R alpha chain 576R/R and AICDA 8408T/T frequency was significantly increased in allergic asthma group as compared with that in the control group [odd ratio (OR) = 3.797 and 9.127, respectively; P < 0.01)] and was correlated with the increased plasma total IgE. These data suggested that the IL-4R alpha chain 576R/R and AICDA 8408T/T genotypes confer genetic susceptibility to adult allergic asthma in China.
Subject(s)
Alleles , Asthma/etiology , Asthma/genetics , Cytidine Deaminase/genetics , Immunoglobulin E/blood , Interleukin-4/genetics , Phenotype , Polymorphism, Restriction Fragment Length , RNA Processing, Post-Transcriptional , Receptors, Interleukin-4/geneticsABSTRACT
Aim To study the effects of dexamethasone(DXM)and peroxisome-proliferation activated receptor?(PPAR?)agonist pioglitazone on intracellular cytokines expression of T_H1/T_H2 and the mechanism of that in asthma model,and compare the difference between two drugs.Methods 24 Balb/c mice were divided into 4 groups randomly: control group,asthma group,DXM group,and pioglitazone group,with 6 mice in each group.The expression of T-bet and GATA3 of lung were detected using western blot,and the intracellular cytokines interluekin 4 and interferon ? expressions of CD~(4+)Tcell were measured using flowcytometry.Results Compared with control group,the level of the expression of T-bet of lung in asthma group increased significantly(P0.05),while both of GATA3 and T-bet decreased after DXM treatment,and the decreasing degree of GATA3 was more than that of T-bet.The results of flowcytometry indicated that the ratio of intracellular cytokines IL-4/IFN? of CD~(4+) T cell in asthma group was much higher than that of control group(P