1.Effects of E23K polymorphism in KCNJ11 gene on membrane current.
Xiao-Hui XIA ; Ai-Hong YANG ; Yang HU
Chinese Journal of Applied Physiology 2014;30(1):23-26
OBJECTIVEE23K polymorphism in KCNJ11 gene is associated with cardiovascular disease and diabetes. In order to explore the mechanism of E23K correlation to related diseases, the effect of E23K polymorphism in KCNJ11 gene on membrane current was investigated.
METHODSThe exon of KCNJ11 was obtained by PCR amplification and the G-->A mutation was completed by overlap extension PCR. The sequences of KCNJ11 exon contained 23E or 23K was inserted into pcDNA3.1/CT-GFP vector respectively. The recombinant plasmid, pcDNA3.1-KCNJ11(E) and pcDNA3.1-KCNJ11(K), were transfected into HEK293T cells by lipofectamine and the membrane current density was determined by whole-cell patch clamp technique.
RESULTS1,173 bp sequences of KCNJ11 gene's exon were amplified by PCR and the recombinant expression plasmid, pcDNA3.1-KCNJ11(E) and pcDNA3.1-KCNJ11(K), were constructed successful. Positive and negative currents were detected in HEK293T cells transfected with difference plasmid by whole-cell patch clamp technique. Results showed that the reversed voltage was 50mV. The current in HEK293T cells with pcDNA3.1-KCNJ11(E) was significantly greater than that with pcDNA3.1-KCNJ11(K) (P < 0.05, n = 10).
CONCLUSIONThe polymorphism of E23K in exon of KCNJ11 gene changed the membrane currents in HEK293T cells. It could be an experiment support for the possible mechanism between the locus and related diseases.
Exons ; HEK293 Cells ; Humans ; Membrane Potentials ; Patch-Clamp Techniques ; Polymorphism, Genetic ; Potassium Channels, Inwardly Rectifying ; genetics
2.Studies on Screening and Conditions of Strains Highly Producing Neutral Phytase
Zhao-Xia LI ; Ai-Min WANG ; Xiao-Min LI ;
Microbiology 1992;0(04):-
A highly phytase-producing strain B.licheniformis LL8 was obtained by several mutagenesis of UV with B.licheniformis as starting strain.The new strain produced about two folds of phytase activity as compared with the starting strain. The production performance of the strain was stable. The cultivation conditions were optimized by single factor and orthogonal experiment. When the mutant B.licheniformis LL8 was cultivated at 55℃, initial pH 7.5 with the inoculation size of 10% for 30h in WBE medUm, the phytase activity was up to 2268.4U/mL.
3.Investigation and analysis on current situation of clinical teaching
ming-min, GU ; ming-de, ZHU ; ai-li, NUER ; xiao-bo, LI ; hui, XIA ; gang, HUANG
Journal of Shanghai Jiaotong University(Medical Science) 2008;0(S1):-
Objective To investigate the clinical teaching situation by using developmental inspection of School of Medicine,Shanghai Jiaotong University(SJTU-SM),and to put forward some suggestions. Methods By checking questionnaires and informal discussions,the relevant information was collected and analyzed by using SPSS statistics sofware. Results The clinical teaching quality of SJTU-SM was basically satisfied.The satisfaction from internship of grade 2004 was better than that of grade 2003.However,some problems in clinical teaching must be improved.Conclusion The investigation showed that the clinical teaching quality of SJTU-SM is being improving.However,in order to achieve the international accreditation standards,the quality guarantee system of clinical teaching need to be further perfected.
4.Influence of coal-arsenic exposure on human T cells proliferation and its mechanism
Jun-ying, GU ; Ai-hua, ZHANG ; Bi-xia, ZHANG ; Xiao-xin, HUANG
Chinese Journal of Endemiology 2009;28(1):20-23
Objective To explore the influence of coal-arsenic exposure on human T cells proliferation and its mechanism.Methods Blood samples colleoted from individuals which lived in arsenism area of coal-burning type and non-arsenism area in Guizhou Province were divided into exposed group(17),mild(35),moderate(38) and severe arsenism group(19)and control group(35)according to Diagnosis Smndard for Endemic Arsenism (WS/T 211-2001).T cell stimulation index wag determined by methyl thiazolyl tetrazolium(MTT)colorimetric method.The intracellular Ca2+ exponential(IECa2+)in peripheral blood mononuclear cell(PBMC)was analyzed by Fho-3/AM dye and flow cytometry.DNA binding activity of actively T cells nuclear factor(NF-AT)in PBMC was evaluated by electrophoretie mobility shift assay(EMSA).Results Concanavalin A(ConA)stimulation decreased the T cells stimulation indexes in exposed group,mild,moderate and severe arsenism groups(1.315±0.962, 1.611±1.224,1.114±0.545,1.289±0.875)compared with control group(2.322±1.241),all the differences being statistically significant(P<0.01).After stimulated by anti-CD3 monoclonal antibody(McAb),the T cells stimulation index in exposed group,mild,moderate and severe arsenism group(0.997±0.177,1.103±0.291,1.007±0.221, 0.957±0.205) were lower than that of control group(1.842±0.429,P < 0.01 ). IECa2+ of PBMC after treated by anti-CD3 McAb in mild,moderate and severe arsenism group( 110.130±49.637,92.429±31.191,77.640± 35.372) were lower compared with control group(145.986±59.450,P <0.01 ). Moreover,IECa2+ in moderat and severe arsenism group were lower than exposed group(121.337±46.410,P < 0.05). DNA binding activity of PBMC NF-AT in mild,moderate and severe arsenism group(1.354±0.446,1.290±0.291,1.159±0.411 ) were lowered than that of control group(1.722±0.291,P < 0.01) and exposed group(1.611±0.294,P < 0.05). Conclusions The coal-arsenic exposure can reduce the human T cells stimulation indexes,IECa2+ in PBMC and the DNA binding activity of NF-AT. It suggest that arsenic may suppress the proliferation ability of human T cells,which may be partly related to the influence of arsenic on T cell receptor(TCR)/CD3 signal transduetion pathway.
5.Role of protein kinase C/NF-kappa B signal pathway In activation of T lymphocytes In human exposed to coal-arsenic
Jun-ying, GU ; Ai-hua, ZHANG ; Bi-xia, ZHANG ; Xiao-xin, HUANG
Chinese Journal of Endemiology 2010;29(1):33-36
Objective To investigate the activation of T lymphocytes in human peripheral blood and the signaling molecules in protein kinase C/nuclear factor KB(PKC/NF-κB) pathway expressivity or activity changes in human peripheral blood mononuclear cells(PBMCs) exposed to coal-arsenic,to explore the role of PKC/NF-κB signal pathway in activation of T cells in human exposed to coal-arsenic. Methods Blood samples were collected from individuals who lived in arsenism area of coal-burning in Guizhou province, and were divided into asymptomatically exposed group (12),mild arsenism group (33),moderate arsenism group (34) and severe arscnism group (15) according to Diagnosis Standard for Endemic Arsenism (WS/T 211-2001). The individuals who lived in non-arsenism area were control group(27). The ratio of activated T ceils was analyzed by flow cytometry. DNA binding activity of NF-κB in PBMCs was evaluated by electrophoretic mobility shift assay(EMSA). The expression of PKCθ and phospho-PKCθ(pPKCθ) in PBMCs were detected with western blotting analysis. Results The ratio of activating T cells in asymptomatically exposed group[(21.76±15.31)%],mild arsenism group[(18.41±11.36)%],moderate arsenism group[(17.78±11.93)%]and severe arsenism group[(18.79±13.38)%]were all higher than that of control group[(3.19±2.12)%],the difference among all groups being statistically significant(F = 7.893,P < 0.05). DNA binding activity of PBMCs NF-κB in asymptomatically exposed group,mild arsenism group,moderate arsenism group and severe arsenism group(1.49±0.24,1.58±0.30,1.57±0.34,1.51±0.16) were higher than that of the control group(1.30±0.17),the difference being statistically sign/ficant(P < 0.05 or < 0.01). The expression of PBMCs pPKCθ in mild arsenism group,moderate arsenism group and severe arsenism group(0.64± 0.14,0.64±0.27,0.62±0.12) were all lower than that of the control group(0.93±0.20),the difference being statistically significant(P < 0.05). There were significant negative correlations between the expression of pPKCθ and the activity of NF-κB(r =-0.565,P < 0.01). There were significant positive correlations between the activity of NF-κB and the ratio of activating T cells(r = 0.546,P < 0.01). Conclusion Coal-arsenic enhances the DNA binding activity of NF-κB,reduces the expression of PBMCs pPKCθ in human PBMCs and up-regulates the activity of T cells. It suggests that the PKC/NF-κB signal might be one of transduction pathway via activating of T cells by coal-arsenic.
6.Expression of P14ARF, MDM2 and mutant type P53 in skin tissue of coal-burning-type of endemic arseniasis patients
Yu-jie, XIA ; Ai-hua, ZHANG ; Xue, HAN ; Xiao-xin, HUANG
Chinese Journal of Endemiology 2012;31(1):24-27
Objective To determine the protein expression of P14ARF,MDM2 and mutant type P53 (P53mt) in skin specimens of coal-burning-type of endemic arseniasis patients and to reveal the molecular mechanism of the disease.Methods Sixty skin specimens from 60 endemic arseniasis patients including 35 of skin lesions patients,19 of precancerous lesion and 6 of skin cancer and 9 normal skin specimens from non-cancer patients were studied.Expression of P14~,MDM2 and P53mt was evaluated by immunohistochemistry using corresponding monoclonal antibodies.Results There was significant difference in the positive rates of P14ARF,MDM2 and P53mt among the 4 groups(x2 =9.39,6.21,20.64,all P < 0.05).The positive rates of P14ARF in precancerous lesion and skin cancer specimens were 46.1% (6/19) and 33.3% (2/6),respectively,which were significantly lower than that of the normal skin specimens [88.9%(8/9),all P < 0.05].Decreased expression of P14ARF was correlated with the development of dermopathy (P < 0.05).The positive rates of MDM2 and P53mt in skin lesions,precancerous lesion and skin cancer specimens were 54.2% ( 19/35 ),63.2% (10/19),66.7% (4/6) and 25.7%(9/35),73.7%(14/19),83.3%(5/6),respectively,which were significantly higher than those of the control (0,0,all P< 0.05).The expression of MDM2 and P53mt increased with the development of dermopathy(all P < 0.05).Conclusions P53mt protein in skin tissue of coal-burning-type of endemic arseniasis patients is over expressed.Abnormal expression of P14ARF and MDM2 may be one of the reasons lead to abnormal cell cycle control disorders and may play a role in the development of endemic arseniasis.
7.Relationship between myeloperoxidase and catalase genetic polymorphism and their activities with arsenic poisoning caused by coal-burning
Bing, LIANG ; Ai-hua, ZHANG ; Xu-guang, XI ; Bi-xia, ZHANG ; Xiao-xin, HUANG
Chinese Journal of Endemiology 2009;28(3):272-275
Objective To detect genetic polymorphism of myeloperoxidase (MPO) gene and catalase (CAT) gene and their activities, and to analyze their relationship with arsenic poisoning caused by coal-burning. Methods One hundred and thirty arsenic poisoning patients were chosen as case group in Jiaole Village, Xingren County, Guizhou Province(an endemic area). One hundred and forty healthy residents living in 13 km away were chosen as control group. Their blood was collected. Polymerase chain reaction-restriction fragment length polymorphism technique(PCR-RFLP) was used to detect polymorphism of MPO-463G/A and CAT-262C/T. Ultraviolet spectmphotometer method was used to detect myeloperoxidase activity. Chromatometry method was used to detect catalase activity. Results The genotype frequency of MPO-463G/A at GG, GA, AA site was 47.24%(60/127), 44.09%(56/127),8.67% (11/127) in case group and 42.34% (58/137),48.17% (66/137)1,9.49% (13/137) in control group, respectively. The difference between the two groups was not significant(χ2 = 0.642, P > 0.05). The genotype frequency of CAT-262C/T, at CC, CT, TT site was 65.60%(82/125),28.80%(36/125),5.60%(7/125) in case group and 76.51%(101/132), 18.94% (25/132) ,4.55% (6/132) in control group, respectively, without significant difference (χ2 =3.845, P>0.05). The relationship between polymorphism of MPO-463G/A and CAT-262C/T and the risk of arsenic poisoning was not found in this study(ORadj= 1.36, 95%CI: 0.74-2.50 for MPO; ORadj=1.35, 95%CI: 0.69-2.63 for CAT). The activities of MPO and CAT were (25.30±8.70)U/L and (2.80± 1.09)×103 U/L in case group, while (22.76±7.59)U/L and (3.90±1.01)×103U/L in control group with a significant difference(F=0.760 for MPO, F=0.855 for CAT, all P < 0.05). The genotype of MPO-463G/A and CAT-262C/T was not found to have relationship with the activities of MPO, CAT(F=1.312,2.822 for MPO; F= 0.151,0.036 for CAT, P>0.05). Conclusions Genetic polymorphism of MPO-463G/A and CAT-262C/T is not found to have relationship with arsenic poisoning. Arsenic can lead to the change of MPO and CAT activity, which, however, may not be affected by MPO-463G/A and CAT-262C/T polymorphism.
8.Study on Measuring Cord Blood Leptin Level and the Ralatioship between Leptin and Insulin in Premature Infants
xue-ai, FAN ; yan-min, LI ; xiao-mei, HAN ; zhao-xia, TIAN
Journal of Applied Clinical Pediatrics 1986;0(02):-
0.05).2.Neonates umbilical serum leptin concentration was positively correlated with body mass index(r=0.520 P
9.Correlation between Serum Interleukin -4,12 and Cellular Immunity in Children with Asthma
Ai-hua, CUI ; Yi-zhen, FENG ; Xiao-jing, SUN ; Jun, LIANG ; Ming-xia, ZHANG
Journal of Applied Clinical Pediatrics 2006;0(16):-
Objective To study the changes of serum interleukin(IL)-4,IL-12 and correlation with cellular immunity in children with asthma of different stages.Methods Fifty asthmatic children were randomly selected, including 30 cases in attack stage (group A) and 20 cases in remission stage (group R). At the same time, 22 healthy children were studied as normal controls (group N).The levels of IL-12 and IL-4 ,T cells subgroups and erythrocyte immunity were detected.Results 1.Serum IL-12 levels were (24.44? 13.26 ),(42.30?12.65),(44.68?28.28) ng/L in group A, R and N,respectively. There was significant difference in three groups (F=8.92 P
10.Intervention of Shenkangling Decoction on the renal injury of primary nephrotic syndrome children patients of Shen deficiency blood stasis syndrome: a clinical observation.
Jian ZHENG ; Si AI ; Fan YANG ; Cai-Xia QIU ; Xiao-lu LU
Chinese Journal of Integrated Traditional and Western Medicine 2014;34(5):541-544
OBJECTIVETo observe the intervention of Shenkangling Decoction (SD) on the renal injury of primary nephrotic syndrome (PNS) children patients of Shen deficiency blood stasis syndrome (SDBSS) and to explore its mechanism.
METHODSTotally 65 PNS children patients were randomly assigned to the combined group (33 cases, treated by SD +Western medicine) and the Western medicine group (32 cases, treated by Western medicine). Meanwhile, 30 healthy children were recruited as the healthy control group from the medical examination center. Those in the Western medicine group were treated with prednisone (5 mg per tablet) at the daily dose of 1.5 -2.0 mg/kg till two weeks after their urine protein turned to negative. Then the dosage was reduced once daily per every other day. The therapeutic course lasted for more than 1 year. For those with no effect of prednisone or partial effect, cyclophosphamide intravenous pulse therapy was additionally applied for 2 successive days per week, a total of 6 times, or they took cyclosporine A. Patients in the combined group additionally took SD while starting treatment of prednisone. SD was decocted in water for oral dose, once daily, taken in two portions until 2 months after prednisone was discontinued. Efficacy was evaluated based on serum levels of chemotactic factor CXCL16, disintegrin metalloproteinase 10 ( ADAM10 ), disintegrin metalloproteinase 17 (ADAM17), albumin (ALB), total cholesterol (TC), and 24-h urine protein excretion (UPE) detected by ELISA before and after treatment.
RESULTSCompared with before treatment in the same group, levels of CXCL16, ADAM10, ADAM17, TC, and 24-h UPE were significantly lower in the two treatment groups (P <0. 01). Compared with the control group, levels of CXCL16, ADAM10, ADAM17, TC, and 24-h UPE significantly increased, and the serum ALB level decreased in the two treatment groups (P <0.01). Compared with the Western medicine group at the same time point, levels of CXCL16, ADAM10, ADAM17, TC, and 24-h UPE significantly decreased in the combined group. The 1 -year recurrence rate and the recurrence times decreased in the combined group (P <0.01). The complete remission rate increased in the combined group (P <0.01).
CONCLUSIONSD could effectively improve the clinical prognosis of PNS children patients possibly by reducing the release of inflammatory mediators such as CXCL16, ADAM10, and ADAM17, decreasing UPE and the TC level, and elevating the serum ALB level.
Child ; Drugs, Chinese Herbal ; therapeutic use ; Humans ; Medicine, Chinese Traditional ; Nephrotic Syndrome ; drug therapy ; Prednisone ; Syndrome