AIM: To evaluate the accuracy of central conreal thickness ( CCT ) using EX500 Excimer Laser workstation (EX500) in laser in situ keratomileusis (LASIK) patients.METHODS:The CCT of 120 eyes (63 patients) who had LASIK between January 2013 and June 2013 were measured by A- scan and EX500. Three groups were classified: >550μm, 500 ~550μm, <500μm according the CCT value of A-scan. The CCT were measured again by corneal flap creating by moria SBK microkeratome. The thickness of the corneal bed stroma were measured by A-scan and EX500 after keratomileusis. All outcomes were analyzed with paired t test.
RESULTS: The average preoperative CCT value was 527. 9±34. 3μm measured by A-scan, 528. 5±34. 6μm measured by EX500. There was no significant difference between these two measurements (t=1. 736, P=0. 085). In group which CCT >550μm, the average preoperative CCT value was 571. 4±17. 3μm measured by A-scan, 572.7±15. 7μm measured by EX500. There was no significant difference between these two measurements (t=1. 857, P=0. 072). In group which CCT 500 ~ 550μm, the average preoperative CCT value was 523. 4±13. 1μm measured by A-scan, 524. 2±12. 4μm measured by EX500. There was no significant difference between these two measurements ( t=1. 934, P = 0. 058 ). In group which CCT <500μm, the average preoperative CCT value 484. 5±9.8μm measured by A-scan, 483. 7±8. 9μm measured by EX500. There was no significant difference between these two measurements (t=1. 395, P=0. 174). The average CCT value after corneal flap lifting was 401. 3 ± 34. 2μm measured by A-scan, 393. 4±38. 9μm measured by EX500. There was a significant difference between these two measurements ( t = 6. 669, P = 0. 000 ). The average thickness of the corneal bed stroma value after keratomileusis was 332. 6±38. 3μm measured by A-scan, 307. 3 ± 37. 1μm measured by EX500. There was a significant difference between these two measurements ( t=17. 165, P=0. 000).
CONCLUSION: There is no significant difference between preoperative CCT value measured by A-scan and EX500. After corneal flap lifting and keratomileusis, the CCT value measured by EX500 is smaller than measured by A-scan.

AIM: To evaluate the changes of corneal high - order aberration (including Coma, Spab, RMSh) after laser in situ keratomileusis (LASIK) with femtosecond laser, sub- Bowman keratomileusis ( SBK ) and laser epithelial keratomileusis (LASEK). METHODS: Of 82 myopic patients ( 164 eyes ), 31 patients (62 eyes) were treated by FS-LASIK, 31 patients (62 eyes) were treated by SBK, 20 patients (40 eyes) were treated by LASEK. Sirius system was used for measuring the coma aberration, spherical aberration, and high order aberration at 1, 15d,1, 3mo after surgery. RESULTS: 1) Vision: The uncorrected visual acuity of the three groups had no differences (P>0. 05). 2) Corneal aberrations: Three kinds of surgical procedure for patients with corneal aberration had significant impact. The C7, C8, C12 and RMSh of three groups were increased significantly (P<0. 05). The C7, C8, C12 and RMSh were not recovered to preoperative levels after 3mo. But the increase of patients after FS- LASIK was smaller than the other two groups, with statistical significance (P<0. 05). CONCLUSION: Compared with SBK and LASEK, FS - LASIK has better visual acuity in the early postoperative and corneal higher-order aberrations increase is relatively small.

OBJECTIVEThe determination of antigenicity and immunogenicity of Leptospira interrogans genus-specific outer envelope proteins (OEPs) will offer evidence for developing universal leptospiral genetic engineering vaccine and detection kit.

METHODSIn this study, Ni-NTA affinity chromatography is used to purify the recombinant products rLipL21, rOmpL1/1, rOmpL1/2, rLipL32/1, rLipL32/2, rLipL41/1 and rLipL41/2 expressed by the major genotypes of four leptospiral OEPs of 15 serogroups. SDS-PAGE is applied to examine the expression and purity of the recombinant proteins. Rabbits are intracutaneously immunized with the recombinant proteins to obtain antisera. Microscope agglutination test (MAT) is used to measure the cross inmmunoagglutination titers of antisera. The OMPs of the reference standard strains belonging to 15 serogroups of L. interrogans in China and L. biflexa strain Patoc I are prepared using salt-denature method. By each of the antisera as the first antibody, Western blot assay is established to detect the natural expressions and immunoreactivity of the four OEPs.

RESULTSThe outputs of rLipL21, rLipL32/1, rLipL32/2, rLipL41/1l, rLipL41/2, rOmpL1/1 and rOmpL1/2 are 10%, 40%, 35%, 15%, 10%, 30% and 15%, respectively. Each the purified recombinant proteins shows a single fragment after SDS-PAGE. Each the rabbit antisera displays extensive cross immunoreactivity between the products expressed by different genotypes of the same gene and the MAT titers ranging from 1:2-1:128. All the four OEPs can be detectable in the OEPs preparations. However, LipL21 is found to exist only in L. interrrogans.

CONCLUSIONThe results of this study indicate that all the four OEPs are superficial genus-specific antigens of Leptospira which can be used as the candidate antigens of leptospiral universal vaccine and detection kit.

Animals ; Antibody Formation ; Antigens, Bacterial ; immunology ; Electrophoresis, Polyacrylamide Gel ; Genetic Engineering ; Immunization ; Leptospira interrogans ; classification ; immunology ; Membrane Proteins ; Rabbits ; Recombinant Proteins ; immunology ; Serotyping

OBJECTIVETo construct lipL32/1-lipL41/1 fusion gene and its prokaryotic expression system and to determine frequencies of carrying and expression of lipL32 and lipL41 genes in L.interrogans wild strains and specific antibody levels in sera from leptospirosis patients.

METHODSlipL32/1-lipL41/1 fusion gene was constructed using linking primer PCR method and the prokaryotic expression system of the fusion gene done with routine techniques. SDS-PAGE was used to examine expression of the target recombinant protein rLipL32/1-rLipL41/1. Immunogenicity of rLipL32/1-rLipL41/1 was identified by Western blot. PCR and MAT were performed to detect carrying and expression of lipL32 and lipL41 genes in 97 wild L.interrogans strains. Antibodies against products of lipL32 and lipL41 genes in serum samples from 228 leptospirosis patients were detected by ELISA method.

RESULTSThe homogeneity of nucleotide and putative amino acid sequence of lipL32/1-lipL41/1 fusion gene were 99.9 % and 99.8 % in comparison with the reported sequences. Expression output of the target recombinant protein rLipL32/1-rLipL41/1, mainly present in inclusion body, accounted for 10 % of the total bacterial proteins. Both the rabbit antisera against rLipL32/1 and rLipL41/1 could combine to rLipL32/1-rLipL41/1. 97.9 % and 87.6 % of the L.interrogans wild strains had lipL32 and lipL41 genes, respectively. 95.9 % and 84.5 % of the wild strains were positive for MAT with titers of 1:4 - 1:128 using rabbit anti-rLipL32s or anti-rLipL41s sera, respectively. 94.7 % - 97.4 % of the patients'serum samples were positive for rLipL32s antibodies, while 78.5 % - 84.6 % of them were rLipL41s antibodies detectable.

CONCLUSIONlipL32/1-jlipL41/1 fusion gene and its prokaryotic expression system were successfully constructed. The expressed fusion protein had qualified immunogenicity. Both the lipL32 and lipL41 genes are extensively carried and frequently expressed by different serogroups of L.interrogans, and their expression products exhibit cross-antigenicity.

Antibodies, Bacterial ; blood ; Antigens, Bacterial ; genetics ; immunology ; Bacterial Outer Membrane Proteins ; genetics ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Gene Expression Regulation, Bacterial ; Genes, Bacterial ; genetics ; Humans ; Leptospira interrogans ; genetics ; Leptospirosis ; immunology ; microbiology ; Lipoproteins ; genetics ; Prokaryotic Cells ; metabolism ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology

OBJECTIVETo reconstruct the nucleotide sequence of Leptospira interrogans lipL21 gene for increasing the output of prokaryotic expression and to understand the changes on immunogenicity of the expression products before and after reconstruction, and to determine the position of envelope lipoprotein LipL21 on the surface of leptospiral body.

METHODSAccording to the preferred codons of E.coli, the nucleotide sequence of lipL21 gene was designed and synthesized, and then its prokaryotic expression system was constructed. By using SDS-PAGE plus BioRad agarose image analysor, the expression level changes of lipL21 genes before and after reconstruction were measured. A Western blot assay using rabbit anti-TR/Patoc I serum as the first antibody was performed to identify the immunoreactivity of the two target recombinant proteins rLipL21s before and after reconstruction. The changes of cross agglutination titers of antisera against two rLipL21s before and after reconstruction to the different leptospiral serogroups were demonstrated using microscope agglutination test (MAT). Immuno-electronmicroscopy was applied to confirm the location of LipL21s.

RESULTThe expression outputs of original and reconstructed lipL21 genes were 8.5 % and 46.5 % of the total bacterial proteins, respectively. Both the two rLipL21s could take place immune conjugation reaction with TR/Patoc I antiserum. After immunization with each of the two rLipL21s in rabbits, the animals could produce specific antibody. Similar MAT titers with 1:80 - 1:320 of the two antisera against rLipL21s were present. LipL21 was confirmed to locate on the surface of leptospiral envelope.

CONCLUSIONLipL21 is a superficial antigen of Leptospira interrogans. The expression output of the reconstructed lipL21 gene is remarkably increased. The expression rLipL21 maintains fine antigenicity and immunoreactivity and its antibody still shows an extensive cross immunoagglutination activity. The high expression of the reconstructed lipL21 gene will offer a favorable condition to use its product for further developing a novel universal vaccine as well as detection kit of leptospirosis.

Amino Acid Sequence ; Animals ; Antigens, Bacterial ; genetics ; immunology ; metabolism ; Bacterial Outer Membrane Proteins ; genetics ; immunology ; metabolism ; Bacterial Vaccines ; immunology ; Base Sequence ; Blotting, Western ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Immune Sera ; immunology ; Leptospira interrogans ; genetics ; immunology ; ultrastructure ; Lipoproteins ; genetics ; immunology ; metabolism ; Microscopy, Immunoelectron ; Molecular Sequence Data ; Rabbits ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Sequence Analysis, DNA ; Vaccines, DNA ; immunology

OBJECTIVEThe aim of this study was to investigate how to model an accuracy 3-dimension Finite Element Analysis (3D-FEA) model.

METHODSBased on computed tomography (CT) scan data of a woman, a 3D finite element model of the first molar on the left was rebuilt by computer imagines process and computer aided design (CAD). Analysis of the stress distribution on a cylinder dental implant and in the bone around it was conducted.

RESULTSThe stress distribution showed extremely asymmetry in bucco-lingual section: stress concentrated on the lingual side of the mandible; stress mainly was tensile in buccal side, and on the contrary compressive in lingual side.

CONCLUSIONThe results were more reliable because this model more really displayed the mandible.

Computer Simulation ; Computer-Aided Design ; Dental Implants ; Dental Stress Analysis ; Female ; Finite Element Analysis ; Humans ; Mandible ; Molar ; Stress, Mechanical

OBJECTIVETo probe the function of interleukin-17 (IL-17) in rheumatoid arthritis (RA) patients of accumulated dampness-heat obstruction in joints syndrome (ADOJS) by detecting levels of IL-17 in serum and the synovial fluid and analyzing its correlation with erythrocyte sedimentation rate (ESR) and C reactive protein (CRP).

METHODSFrom January 2011 to January 2013, recruited were 90 RA inpatients of ADOJS at Department of Integrative Medical Rheumatism, General Hospital of Chengdu Military Region, of which 28 patients had knee joint effusion. Besides, 30 healthy volunteers who received physical examination at our hospital were recruited as the normal control group, and 30 patients with osteoarthritis (OA) who had knee joint effusion were recruited as the synovial fluid control group. The expression levels of IL-17 in serum and the synovial fluid were detected by enzyme linked immunosorbent assay (ELISA), and contents of ESR and CRP were detected in RA patients. Then correlation analyses were performed between levels of IL-17 and contents of ESR and CRP.

RESULTSCompared with the normal serum control group, the expression levels of IL-17 in serum of RA patients significantly increased (P < 0.05). Compared with the serum of RA patients and the synovial fluid of OA patients, the expression levels of IL-17 in the synovial fluid of RA patients significantly increased (P < 0.05). The expression levels of IL-17 in serum of RA patients were not correlated with ESR or CRP (r = 0.092, -0.082; P > 0.05), and the expressional levels of IL-17 in the synovial fluid of RA patients were not correlated with ESR or CRP (r = 0.113, -0.034; P > 0.05).

CONCLUSIONSIL-17 was the main effector cytokine of Th17 cells. The expressional levels of IL-17 significantly increased in serum and the synovial fluid of RA patients of ADOJS, but with no correlation to ESR or CRP. It indicated that IL-17 participated in the occurrence and development of RA. Concrete mechanisms needed to be further proved in larger samples.

Aged ; Arthritis, Rheumatoid ; blood ; diagnosis ; metabolism ; Blood Sedimentation ; C-Reactive Protein ; metabolism ; Case-Control Studies ; Female ; Humans ; Interleukin-17 ; blood ; metabolism ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Synovial Fluid ; metabolism

OBJECTIVETo study the effect of bitter-cold herbs easing dampness method (BCHEDM) plus Sanhuang Yilong Decoction (SYD) combined with methotrexate (MTX) on expression levels of interleukin-1 (IL-1), IL-6, and IL-17 in rheumatoid arthritis (RA) patients of accumulated dampness-heat syndrome (ADHS).

METHODSFrom January 2011 to January 2013 recruited were 90 RA inpatients of ADHS at Department of Integrative Medicine on Rheumatoid Disease, General Hospital of Chengdu Military Region. They were assigned to the treatment group (45 cases) and the control group (45 cases) according to the random digit table produced by SPSS 11.5 Software. Patients in the treatment group were treated by heavy bitter-cold herbs plus SYD combined with MTX, while those in the control group were treated by MTX alone. Expressional levels of IL-1, IL-6, and IL-17 in serum were detected by enzyme linked immunosorbent assay (ELISA) before treatment, at week 2 and 4 after treatment. Erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and disease activity score in 28 joints (DAS28) were detected as well.

RESULTSAfter two or four weeks of treatment, ESR, CRP, and DAS28 decreased more in the treatment group than in the control group with statistical difference (P < 0.05, P < 0.01). After four weeks of treatment, IL-1, IL-6, IL-17, ESR, CRP, and DAS28 in the treatment group were all lower than before treatment and those of the control group at corresponding time points with statistical difference (P < 0.01).

CONCLUSIONSYD combined MTX could play roles of improving inflammatory indices within 2 weeks, and inhibiting the expression of IL-1, IL-6, and IL-17 within 4 weeks.

Arthritis, Rheumatoid ; blood ; drug therapy ; immunology ; Blood Sedimentation ; C-Reactive Protein ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; therapeutic use ; Hot Temperature ; Humans ; Interleukin-1 ; blood ; Interleukin-17 ; blood ; Interleukin-6 ; blood ; Methotrexate ; therapeutic use ; Syndrome ; Treatment Outcome

OBJECTIVETo observe the effects of Yijingfang on CatSper1 in the mouse model of cyclophosphamide-induced oligoasthenospermia.

METHODSForty Kunming male mice were randomly divided into a control group (CG), a model group (MG), a small-dose Yijingfang group (SG), and a large-dose Yijingfang group (LG). The mice of CG were intraperitoneally injected with normal saline at 60 mg/kg once a day, while those of MG, SG and LG with cyclophosphamide, all for 5 days. During the next 34 days, the mice of SG and LG received intragastric administration of Yijingfang once a day, the former at a dose 2 times and the latter 5 times that of human routine usage, those of MG given the same volume of normal saline, and CG normally fed. At 35 days, we measured the sperm count, percentages of grades a + b and a + b + c sperm, and the expression of CatSper1 in the epididymal sperm of the mice.

RESULTSThe sperm counts of CG, MG, SG and LG were (5.20 +/- 1.34), (1.73 +/- 0.03), (2.08 +/- 0.01) and (3.31 +/- 0.56) x 10(6)/ml, respectively, significantly lower in MG than in CG (P < 0.05), but higher in LG than in MG (P < 0.05). The grade a + b sperm constituted (14.49 +/- 0.30), (6.64 +/- 1.88), (11.99 +/- 1.01) and (19.40 +/- 3.13)% in CG, MG, SG and LG, respectively, remarkably lower in MG than in CG (P < 0.05) but higher in LG than in MG (P < 0.05); the grade a + b + c sperm accounted for (68.39 +/- 15.13), (39.96 +/- 4.89), (62.28 +/- 4.43) and (73.61 +/- 5.05)%, respectively, obviously lower in MG than in CG (P < 0.05) but higher in LG than in MG (P < 0.05); the CatSper1 expressions were 0.76 +/- 0.05, 0.73 +/- 0.03, 0.75 +/- 0.12 and 0.85 +/- 0.04, respectively, markedly higher in LG than in MG (P < 0.05).

CONCLUSIONIntraperitoneal injection of cyclophosphamide decreases the sperm count, percentages of grades a + b and a + b + c sperm, and the expression of CatSper1 in mice, while large-dose Yijingfang can increase the above parameters, and hence contributes to the treatment of oligoasthenospermia.

Animals ; Calcium Channels ; drug effects ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Male ; Mice ; Mice, Inbred Strains ; Sperm Motility ; Sperm Tail ; drug effects ; metabolism

OBJECTIVETo construct lipL32/1-lipL21-OmpL1/2 fusion gene of Leptospira interrogans and its prokaryotic expression system, and to identify the immunogenicity of its products.

METHODSPCR using linking primers was applied to construct lipL32/1-lipL21-OmpL1/2 fusion gene and a prokaryotic expression system of the fusion gene was then established using routine genetic engineering technique. SDS-PAGE was used to examine output of the target recombinant protein rLipL32/1-LipL21-OmpL1/2. Double immunodiffusion and Western Blot assay were applied to identify immunogenicity of rLipL32/1-LipL21-OmpL1/2.

RESULTlipL32/1-lipL21-OmpL1/2 fusion gene with correct sequence and its prokaryotic expression system E.coli BL21DE3pET42a-lipL32/1-lipL21-ompL1/2 was obtained in this study. The output of rLipL32/1-LipL21- OmpL1/2 after optimisation was 37.78 mg/L. The immunodiffusion titer of rabbit antiserum against rLipL32/1-LipL21-OmpL1/2 was 1:4. The rLipL32/1-LipL21-OmpL1/2 antiserum was able to recognize rLipL32/1-LipL21-OmpL1/2, rLipL32/1, rLipL21 and rOmpL1/2. Positive Western hybridization signals were found among rLipL32/1-LipL21-OmpL1/2 and rabbit antiserum against whole cell of strain 56601 and serum from patients infected with L.interrogans serogroups Icterohaemorrhagiae, Grippotyphosa, Autumnalis and Pomona.

CONCLUSIONThe fusion gene lipL32/1-lipL21-OmpL1/2 and its prokaryotic expression system were successfully constructed in this study. The expressed fusion protein can be used as the antigen for developing universal genetic engineering vaccine and universal serological tests of leptospirosis.

Animals ; Antigens, Bacterial ; biosynthesis ; genetics ; Bacterial Outer Membrane Proteins ; biosynthesis ; genetics ; Bacterial Vaccines ; immunology ; Escherichia coli ; genetics ; metabolism ; Humans ; Leptospira interrogans ; genetics ; immunology ; Lipoproteins ; biosynthesis ; genetics ; Rabbits ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Vaccines, Synthetic ; immunology