Objective To construct four types of glucagon-like peptide-1 (GLP-1) and human serum albumin (HSA) fusion proteins that can be realeased at different rate in vivo by introducing protease cleavage sites between these two moieties.The therapeutic effect and release rate are studied to achieve balanced pharmacokinetics ( PK) and pharmacody-namics ( PD) of GLP-1 and HSA fusion proteins.Methods The gene with different polypeptide joint of GLP-1 and HSA fusion proteins were synthesized by overlap extension PCR amplification, cloned into expression vector pPIC9 and transformed into Pichia pastoris GS115.Then, fusion proteins were obtained by protein purification after being induced by methanol.The preliminary PK and PD of the fusion proteins were studied after purification.Results The fusion protein Gly2-GLP-1-GGGGG-HSA showed no release while Gly2-GLP-1-VTR-HSA, Gly2-GLP-1-SARSVRA-HSA, and Gly2-GLP-1-GRSRVTRSV-HSA showed a slow, medium and fast release rate, respectively, after incubation with furin.In vitro biological activity test results dispalyed that each type of fusion protein promoted insulin secretion of MIN6 cells.In vivo PK test indicated the half-life size of fusion proteins was the largest in Gly2-GLP-1-GGGGG-HSA, followed by Gly2-GLP-1-VTR-HSA, Gly2-GLP-1-SARSVRA-HSA, and Gly2-GLP-1-GRSRVTRSV-HSA.In vivo PD test exhibited hypoglycemic activity that was the highest in Gly2-GLP-1-VTR-HSA, followed by Gly2-GLP-1-SARSVRA-HSA, Gly2-GLP-1-GRSRVTRSV-HSA, and Gly2-GLP-1-GGGGG-HSA.Conclusion GLP-1 can be released from fusion proteins with full activity after the introduction of protease cleavage sites.Releasable fusion proteins at an appropriate release rate have the most balanced PK and PD.

OBJECTIVETo evaluate the effectiveness of locking plate external fixator in treating middle and distal tibial fractures.

METHODSFrom January 2010 to January 2013,18 patients suffered from middle and distal tibial fractures were treated by locking plate external fixator,including 11 males and 7 females, with an average age of 53.5 (ranged from 13 to 80) years old,the course of disease ranged from 2 h to 3 d. According to AO classification, 4 cases were type A,11 cases were type B and 3 were type C. Among them,6 patients were open fracture, including 2 cases with type I, 3 cases with type II and 1 case with type III, according to Gustilo classification), 12 patients were close fracture. Operation time, postoperative complications were observed, and Johner-Wruhs scoring were used to evaluate clinical outcomes.

RESULTSAll patients were followed up from 6 to 15 (meaned 11) months. Two cases occurred skin necrosis (1 case occurred bone exposure), 2 cases occurred delayed union (all were open fracture), and 1 case occurred nail infection. No screw loosening or broken occurred. According to Johner-Wruhs scoring, 10 cases obtained excellent result,6 cases good,and 2 cases fine.

CONCLUSIONLocking plate external fixator for the treatment of middle and distal tibial fractures, which has advantages of lessen damage, shorter operative time, less complications and rapid functional recovery, is one of good choice.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Bone Plates ; utilization ; External Fixators ; utilization ; Female ; Fracture Fixation ; instrumentation ; methods ; Humans ; Male ; Middle Aged ; Tibial Fractures ; surgery ; Treatment Outcome ; Young Adult

OBJECTIVETo assess the prognostic value of lymph node ratio (LNR) in patients with stage III rectal cancer after curative resection.

METHODSA retrospective review of the clinicopathological data was performed in 161 patients with stage III rectal cancer who received curative surgical excision in our hospital from June 2005 to June 2010. The variables including LNR, age, gender, T stage, N stage, total number of dissected lymph nodes, number of metastatic lymph nodes, and positive rate of lymph node metastasis were studied through univariate and multivariate analyses, and the survival analysis was performed using Kaplan-Meier method and Log rank test.

RESULTSMultivariate analysis revealed that LNR, but not number of positive nodes or number of harvested lymph nodes, had independent prognostic value for overall survival and disease-free survival for patients with stage III rectal cancer. The overall survival in the LNR < 0.43 and LNR ≥ 0.43 groups was 75.8% and 41.3%, respectively (P < 0.01), while the disease-free survival was 68.8% and 40.3%, respectively (P = 0.001).

CONCLUSIONSThe LNR is an independent prognostic factor for survival of patients with stage III rectal cancer, and is more efficient than the number of positive nodes and total number of dissected lymph nodes in the survival prediction.

Adenocarcinoma ; pathology ; surgery ; Adult ; Aged ; Disease-Free Survival ; Female ; Follow-Up Studies ; Humans ; Lymph Node Excision ; Lymph Nodes ; pathology ; surgery ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Rectal Neoplasms ; pathology ; surgery ; Retrospective Studies ; Survival Rate

Objective To investigate the effects of standardized tertiary rehabilitation (STR) on limb motor function (LMF) after stroke.Methods Eighty-two patients were divided into a primary cerebral infarction group (PCI group) and a primary cerebral hemorrhage group (PCH group),and then randomly further divided into experi- mental and control sub-groups.All patients received routine internal medicine treatment,supplemented with stand- ardized tertiary" rehabilitation in the experimental groups.All patients were assessed with the simplified Fugl-Meyer motor function assessment (S-FMMFA) at enrollment,and 1,3 and 6 months after their stroke.Results The scores of the experimental groups were higher than those of the controls.The experimental groups scores were 26.10% of normal at the time of the enrollment,and improved to 42.52%,65.62% and 83.71% by the end of the 1st,3rd and 6th month,respectively.The control groups started at 18.51%,and progressed to 24.85% ,37.24% and 45.84% over the same interval.Conclusion STR was associated with improved LMF scores of stroke pa- tients.

OBJECTIVETo investigate the expression features of glypican-3 (GPC-3) and its diagnostic and differential values in hepatocellular carcinoma (HCC).

METHODSRat hepatoma models were made and the dynamic expression features of GPC-3 protein and its gene were investigated by Western blotting and RT-PCR respectively. Liver specimens from 36 HCC patients were collected by self-control method and the expression and clinicopathological features of GPC-3 were analyzed by immunohistochemistry. Serum GPC-3 levels were quantitatively detected by ELISA and its efficiency for HCC diagnosis was evaluated in patients with liver diseases.

RESULTSThe incidence of GPC-3 was 0% in control, 83.3% in degeneration, 100% in precancerosis and 100% in canceration during dynamic formation of rat hepatoma, respectively. The positive GPC-3 was brown granule- like staining localized in membrane and cytoplasm in human HCC.

CONCLUSIONSThe GPC-3 positive rates were 80.6% in HCC, 41.7% in surrounding tissues and none in distal tissues (P < 0.01), respectively. No positive relationship presented between GPC-3 and differentiation grade or the number of tumor except of tumor size (Z = 2.941, P < 0.01). The incidence of serum GPC-3 was 52.8% in HCC patients except of one patient with cirrhosis. No significant differences were found between GPC-3 and sex, age, AFP, tumor number, Child classification or extrahepatic metastasis except of tumor size (χ² = 6.318, P < 0.05) and HBV infection (χ² = 23.362, P < 0.01). Combined detection of GPC-3 and AFP could rise up diagnosis of HCC. GPC-3 expression closely associated with HCC and might be useful for early diagnosis of HCC.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Animals ; Carcinoma, Hepatocellular ; diagnosis ; metabolism ; pathology ; Diagnosis, Differential ; Female ; Glypicans ; metabolism ; Humans ; Liver ; pathology ; Liver Neoplasms ; diagnosis ; metabolism ; pathology ; Male ; Middle Aged ; Rats ; Rats, Sprague-Dawley ; Young Adult

OBJECTIVEUse sagittal reconstruction CT to verify the surgical strategy for cervical ossification of the posterior longitudinal ligament (OPLL).

METHODSA retrospective study of 161 patients (106 males and 55 females) who had undergone surgery for OPLL from July 2007 to November 2010 was performed. The mean age at surgery was 54.5 years (range from 26 to 77 years). The mean follow-up period was 28 months (12 - 54 months). There were 40 patients accept anterior approach surgeries (anterior group) which include 14 cases of anterior cervical corpectomy and fusion and 26 cases of anterior cervical discectomy and fusion. There were 120 patients accept posterior approach surgeries (posterior group) which was spinous process-splitting laminoplasty for cervical myelopathy using coralline hydroxyapatite. One patient accepted combined anterior and posterior approach. According to the sagittal reconstruction CT, the main reason for spinal cord compression was cervical disc herniation in anterior group, and OPLL in posterior group. The level of spinal cord compression was 1 to 2 levels in anterior group, and 1 to 5 levels in posterior group with a major of 2 to 4 levels. As the classification of OPLL, segmental type and circumscribed type were major of segmental type in anterior group and all of the four types were in posterior group, the distribution of each type was average. The patients of posterior group were classified into two groups according to the modified K-line classification, and clinical results were compared between the two groups. The modified K-line was defined as a line that connects the midpoints of the spinal canal at C(2) and C(7) on sagittal CT myelography. Compression to the spinal cord did not exceed the K-line in the modified K-line(+) group and did exceed it in the modified K-line(-) group. Clinical data were compared using t-test or χ(2) test. Correlation analysis was used to determine the relationships of C(2)-C(7) angulation between sagittal reconstruction CT and neutral position X-ray.

RESULTSThe patient of anterior group had better recovery rate of the JOA score (72% ± 27%) than the posterior group (59% ± 35%) at the latest follow-up (t = 2.238, P = 0.027). In posterior group, the patients of modified K-line(+) group had better recovery rate of the JOA score (63% ± 37%) than the K-line(-) group (49% ± 30%) at the latest follow up (t = 2.150, P = 0.034). The C(2)-C(7) angulation on sagittal reconstruction CT was 11° ± 9° which has significantly correlated with the C(2)-C(7) angulation on neutral position X-ray which was 10° ± 10° (r = 0.947, P < 0.01).

CONCLUSIONSConsidering the selection of surgical approach, it should be combined with the main clinical diagnosis for spinal cord compression, the level of compression, the classification of OPLL and the kyphotic alignment of the cervical spine. The modified K-line is a simple and practical tool for making decisions regarding the surgical strategy for cervical OPLL patients.

Adult ; Aged ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Ossification of Posterior Longitudinal Ligament ; diagnostic imaging ; surgery ; Retrospective Studies ; Tomography, X-Ray Computed

OBJECTIVEThe aim of this study was to evaluate the correlation of protein expressions of CXC chemokine receptor 4 (CXCR4), vascular endothelial growth factor-C (VEGF-C) and cytokeratin 19 (CK-19) with lymph node metastasis (LNM) in patients with hepatocellular carcinoma (HCC), and their survival.

METHODSThe expressions of CXCR4, VEGF-C and CK-19 in HCC patients with (n = 123) or without (n = 145) LNM were determined using tissue microarray and immunohistochemical staining. The relationship between clinicopathological features and CXCR4, VEGF-C and CK-19 were analyzed. Evaluation of immunostaining was performed semiquantitatively by visual assessment.

RESULTSThe UICC T stage, and expressions of nuclear CXCR4, VEGF-C and CK-19 were independent risk factors for LNM. Nuclear CXCR4, VEGF-C and CK-19 expression were predictive factors for LNM in HCC patients. In patients with LNM, the median survival time was 15.1 months for patients with high nuclear CXCR4 expression and 24.5 months for those with low nuclear CXCR4 expression. The median survival time was 15.1 months for patients with high tumor VEGF-C expression and 31.1 months for those with low tumor VEGF-C expression. The median survival time was 12.0 months for patients with positive CK-19 expression and 19.2 months for patients with negative CK-19 expression. Patients with high nuclear CXCR4, VEGF-C or CK-19 expression had significantly poorer prognosis than those with low expression (all P < 0.05). PVT, UICC T stage and expressions of nuclear CXCR4, VEGF-C, and CK-19 were independent prognostic factors.

CONCLUSIONIncreased protein expressions of nuclear CXCR4, VEGF-C, and CK-19 are independent risk factors for developing lymph node metastasis, and they are significantly correlated with LNM and poor outcome in HCC patients.

Carcinoma, Hepatocellular ; metabolism ; pathology ; Cell Nucleus ; metabolism ; Female ; Follow-Up Studies ; Humans ; Keratin-19 ; metabolism ; Liver Neoplasms ; metabolism ; pathology ; Lymphatic Metastasis ; pathology ; Male ; Middle Aged ; Neoplasm Staging ; Proportional Hazards Models ; Receptors, CXCR4 ; metabolism ; Risk Factors ; Survival Rate ; Vascular Endothelial Growth Factor C ; metabolism

OBJECTIVETo study the correlations between clinical features and liver pathohistological changes of chronic hepatitis B virus (HBV) carriers and to discuss the factors which may influence the prognosis.

METHODSNinety HBV carriers who had liver biopsies were enrolled in this study.

RESULTS(1) The mean follow-up period of the patients was 118 weeks. (2) Fifty-four patients (60.0%) had G1 hepatitis and 21 (23.3%) had G2 hepatitis. The fibrosis stages were graded as S1(42) and S2(21). (3) There were significant age differences among S0, S1 and S2. (4) There were significant differences in aminotransferase levels between patients who had a normal liver histology and those who had mild hepatitis. (5) The grades of liver inflammation were not correlated with the titers of HBeAg and HBV DNA in sera. The stages of liver fibrosis were not correlated with the titers of HBVDNA in sera. Most of the HBeAg negative patients progressed to S2. (6) There were significant differences in spleen dimensions measured by ultrasonography between S0, S1 and S2 patients. (7) During the follow-up period serum aminotransferase (ALT) levels remained normal in 60 patients (group A); 22 patients had transient elevations (group B), and 8 patients had persistent increases (group C). There were significant differences of the ratios of S0 and S2 cases among patients in groups A, B and C. (8) Age and fibrosis stages were predictive factors of liver cirrhosis.

CONCLUSIONSMost chronic HBV carriers had mild inflammatory histological changes in their livers and also had different degrees of liver fibrosis. This follow-up study shows that some of those carriers should have had antiviral therapy.

Adult ; Carrier State ; diagnosis ; pathology ; virology ; Female ; Hepatitis B virus ; Hepatitis B, Chronic ; diagnosis ; pathology ; Humans ; Liver Cirrhosis ; diagnosis ; pathology ; virology ; Male ; Middle Aged ; Prognosis

[Objective] To investigate the level of wild -measles neutralization antibody of both mother and infant , and its correlation between the paired mother and infant . [ Methods] The wild-mea-sles neutralization antibody in the serum of the women and their infants were detected directly by a neutral -ization test (NT) methods. [ Results] The positivity rates of neutralization antibody in mothers and their infants were 91.52%and 88.57%respectively, geometric mean title of neutralization antiboby (GMT) being 61.32 and 58.17 respectively.And the titer of neutralization antibody was highly correlated ( r=0.899, P<0.01)between mother and infant in pairs.When the GMT of mother was ≥1∶16,the positivity rate of neutralization antibody in their infants might reach 100.00%. [ Conclusion] It is important to increase the maternal measles antibody level in order to prevent infants from measles .

This study was purposed to investigate the effect of a hypoxia-inducible factor inhibitor (YC-1) on expression of hypoxia-inducible factor 1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) as well as induction of apoptosis in leukemic cell lines. RT-PCR was used to determine the levels of HIF-1alpha mRNA and VEGF mRNA in K562, U937 and Jurkat cells. After treatment of U937 cell with 4 micromol/L YC-1, cell apoptosis was assayed by DAPI staining under fluorescent microscope and flow cytometry with Annexin V-FITC/PI staining; the expression levels of HIF-1alpha mRNA and VEGF mRNA were measured with RT-PCR; the expression levels of HIF-1alpha, VEGF, BAX, BCL-2 and caspase-3 proteins were measured by Western blot. The results showed that HIF-1alpha mRNA and VEGF mRNA were expressed in all three leukemia cell lines. After treatment of U937 cell with 4 micromol/L YC-1 for 0, 8, 16 and 24 hours, the changes of morphologic features of U937 cells could be observed under fluorescent microscope and the apoptotic rates significantly increased in time-dependent manner, they were (4.87 +/- 0.70)%, (27.27 +/- 2.00)%, (51.53 +/- 2.81) and (60.5 +/- 3.20)% respectively, the expression levels of VEGF mRNA reduced, while the expression levels of HIF-1alpha mRNA had no obviously changes.Furthermore, the expression of HIF-1alpha, VEGF and BCL-2 decreased, while the expression of BAX and caspase-3 increased, the ratio of BAX/BCL-2 increased in time-dependent manner (r = 0.973, p < 0.01). It is concluded that HIF-1alpha mRNA and VEGF mRNA are all expressed in in K562, U937 and Jurkat cells, YC-1 has significant effect on down-regulating the protein expression of HIF-1alpha and VEGF, and induces the apoptosis in U937. The mechanism of apoptosis in leukemic cells may involve in up-regulating BAX/BCL-2 ratio and expression of protein caspase-3.
Apoptosis ; drug effects ; Cell Hypoxia ; Gene Expression Regulation, Leukemic ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; antagonists & inhibitors ; metabolism ; Indazoles ; pharmacology ; Jurkat Cells ; K562 Cells ; U937 Cells ; Vascular Endothelial Growth Factor A ; metabolism