To investigate the influence of recombinant adenovirus carrying tissue inhibitor of metalloproteinase-3 (RAdTIMP-3) on the main compositions of rabbits intervertebral discs and to assess its potential in treatment for intervertebral disc degeneration.[Method]RadTIMP-3 and empty adenovims vector with Lac-Z gene (Rad66) was propagated in 293 Cells and was purified, identified and tittered. Thirty Japanese white rabbits were randomly divided into 5 groups. And 25 μl of various reagents were injected to the L4、5 and L5、6 intervertebral discs of the rabbits as follows:normal saline in group 1, 1.0×1010 OPU/ml of RAd66 in Group 2, and 1.0×1010 OPU/ml of RAdTIMP-3 in group 3, 4 and 5. The intervertebral discs of each group were collected after 2, 2, 1, 2 and 4 weeks after injection respectively.Then X-gal staining, And Group 1, RT-PCR for TIMP-3 and aggrecan core protein,TUNEL staining, immunohistochemical staining for TIMP-3 and type I! Collagen and Safranin O-Fast green staining was carried out to assess the effects of RadTIMP-3 transfection.[Result](1)concentration of RAdTIMP-3 reached 1.9×1012 OPU/ml after propagation and purification. (2)RT-PCR shows that the expression of TIMP-3 was significantly raised in group 3, 4, 5, as compared with group 1 or 2. And the expression of core protein gene in group 3, 4, 5 increased slightly than in group 1 and 2. (3) TUNEL staining revealed that there was not significant difference between the positive-staining rates of any two of the groups. (4)TIMP-3 staining exhibited an obvious increase of positive-staining rates in group 3, 4 and 5 as compared with groupi or 2. The staining density of Safranin O-Fast Green staining and immunohistochemical staining for type II collagen of group 5 was obviously higher than that of group 1 or 2.[Conclusion]RAdTIMP-3 can express widely and safely in rabbit intervertebral discs, and improve the quantity and quality of matrix. It has the potential to be used in treatment for intervertabral disc degeneration.

In order to investigate the apoptotic pathway of rabbit annulus fibrosus (AF) cells induced by mechanical overload, an experimental air-pressure model was established in this study to pressurize the rabbit AF cells in vitro. Cells were randomly divided into five groups in which the cells were exposed to a continuous pressure of 1.1 MPa for different lengths of time (0, 5, 12, 24 and 36 h). The cell proliferation and apoptosis were detected by cell counting kit-8 (CCK-8) assay and flow cytometry; the alterations in mitochondrial membrane potential were measured by fluorescence microscopy and fluorescence spectrophotometer; the activities of caspase-8 and 9 were determined by spectrophotometry. The results showed that after the cells were subjected to the pressure for 24 or 36 h, the cell proliferation was inhibited; the ratio of cell apoptosis was increased; the mitochondrial membrane potential was decreased; the activity of caspase-9 was enhanced; no activity changes were observed in caspase-8. The results suggested that treatment with a pressure of 1.1 MPa for more than 24 h can lead to the proliferation inhibition and the apoptosis of rabbit AF cells in vitro, and the mitochondrial-dependent pathway is implicated in the pressure-induced AF cell apoptosis.

The present study examined the role of Wnt/β-catenin signaling pathway in the degeneration of nucleus pulposus cells and the protective effect of DKK1 on nucleus pulposus cells. The model of nucleus pulposus cell degeneration was induced by intra-disc injection of TNF-α, and the expression of β-catenin protein was detected by Western blotting. The cultured rabbit nucleus pulposus cells were divided into 4 groups. In group A, the cells were cultured with normal medium and served as control group. In group B, the cells were cultured with TNF-α and acted as degeneration group. In group C, the cells were cultured with TNF-α and transfected with Adv-eGFP and was used as fluorescence control group. In group D, the cells were cultured with TNF-α and transfected with Adv-hDKK1-eGFP, serving as intervention group. The expression of type II collagen, proteoglycan, β-catenin, and MMP-13 in each group was detected by immunocytochemistry and RT-PCR. The result showed that TNF-α increased the expression of β-catenin and MMP-13, and significantly inhibited the synthesis of type II collagen and proteoglycan, which resulted in the degeneration of nucleus pulposus cells. This effect could be obviously reversed by DKK1. We are led to concluded that TNF-α could activate the Wnt/β-catenin signaling pathway, and increase the expression of MMP-13, thereby resulting in disc degeneration. Specifically blocking Wnt/β-catenin signaling pathway by DKK-1 could protect the normal metabolism of intervertebral disc tissue. The Wnt pathway plays an important role in the progression of the intervertebral disc degeneration.

Objective To evaluate the clinical effects and experiences of short stem ceramic-on-ceramic total hip arthroplasty in the treatment of young and middle-aged patients with femoral head necrosis.Methods From December 2009 to March 2011,40 patients with femoral head necrosis were treated primarily with short stem ceramic-on-ceramic hip arthroplasty (43 hips) through posterior-lateral or anterior-lateral approach.There were 23 males,17 females； with the mean age of 40.5 years (range,27-55 years).All cases were stage Ⅳ or more according Steinberg stage.The patients were followed up at the 3,6,12 months after operation,and then once each year.Standard X-ray films were taken at the follow-ups.Femoral prosthesis subsidence was assessed through X-ray films.Harris scores system and WOMAC Osteoarthritis Index were recorded before and after operation to evaluate clinical effects of surgery.Results Thirty-eight patients (41 hips) were followed up for 21-36 months,with an average of 31.3 months.The femoral stem subsided 0.68±2.90 mm at the last follow-up.After operation,hip pain of patients was relieved obviously.The Harris score improved from 38.7±12.1 (15-66) preoperatively to 95.2±4.49 (83-100) postoperatively,WOMAC Index from 66.7±12.0 (49-94) to 12.8±5.6 (0-24).During follow-up period,there was no serious complication such as femoral neck fracture,infection of wound,pulmonary embolism,et al.No dislocation,implant prosthesis failure and infection were observed.Conclusion The short stem hip arthroplasty preserves bone stock to a great extent.The short stem implant showed a good stability and osseous integration.The wear rate of ceramic-on-ceramic bearing surface is extremely low.Short stem hip arthroplasty plus ceramic-on-ceramic bearing surface might be a good option for the young and middle-aged patients with femoral head necrosis.

ObjectiveTo explore the expression of novel protein kinase C ε (PKCε) in normal prostate (NP) tissue, benign prostate hyperplasia(BPH), peficancerous (PC) tissue and prostate cancer (Pca), and study its correlation with the grade and stage of Pca.MethodsTen NP slides, ten BPH slides, ten PC slides and 43 Pca slides were collected from our hospital. These slides were routinely proceased and analyzed according to the requirement of immunohistochemical staining. Tumors were classified according to the 2002 TNM staging system. The grading system used in the study was based on the Gleason grade.ResultsWe was found that the expression of PKCεs in Pca (27/43) were significantly higher than those in NP(1/10), BPH (0/10) and PC (2/10) tissue, and the difference was statistically significant ( P ＜0.05 ). With regard to grade of prostate cancer, the expression of PKCε in Pca with Gleason score ≥8 group (12/13) was higher than the Gleason score 2 -4 group (4/10) and the Gleason score 5 -7 group (11/20). The difference was statistically significant (P ＜ 0.05 ). Moreover, the T3 and T4 stages had a more positive rate (10/12 & 9/10) than the T1 and T2 stages( 1/6 &7/15). There is statistically significant difference between early and advanced stage of prostate cancer ( P ＜ 0. 05 ). Furthermore, the positive expression of PKCε in prostatic carcinoma samples increased significantly in the metastasis group (9/10)compared to the non-metastasis group ( 18/33 ) ( P ＜ 0. 05 ), but the difference was not statistically significant between the concentration of prostate-specific antigen in blood serum ( P ＞ 0. 05 ).Conclusions PKCε is expressed in prostate cancer, and it correlates with the grade and stage of prostate cancer. PKCε may be related to the origin and the development of Pca, and it may be used as a prognostic factor for Pca.

Objective To explore the effectiveness and compliance of ketogenic-diet(KD) treatment for infantile spasm(IS).Methods Ninety-eight IS patients who were treated with KD in Wuhan Children's Hospital from March 2009 to June 2015 were analyzed by using retrospective case-control study,the patients were divided into 4 groups:newly diagnosed IS patients group (group A,including 24 patients),one antiepileptic drug (AEDs) failure IS patients (group B,including 28 patients),two and more AEDs failure IS patients (group C,including 29 patients),and two or more AEDs combined with ACTH failure IS patients(group D,including 17 patients).The spasm-free andretention rates after 3,6 and 12 months KD treatment were compared among these groups.Results Overall retention rate was 80.6％ (79/98 cases),69.4％ (68/98 cases),and 42.9％ (42/98 cases)at 3,6,12 months,respectively.The 3-month retention rate in group A,B,C and D was 83.3 ％ (20/24 cases),78.6％ (22/28 cases),82.7％ (24/29 cases) and 76.4％ (13/17 cases) respectively,and there was no significant difference among these groups (P ＞ 0.05).The 6-month retention rates in each group was 75.0％ (18/24 cases),67.9％ (19/28 cases),68.8％ (20/29 cases) and 65.0％ (11/17 cases) in sequence,and there was also no significant difference among these groups(P ＞0.05).The 12-month retention rate was 54.2％ (13/24 cases),21.4％ (6/28 cases),48.3％ (14/29 cases) and 52.9％ (9/17 cases) in group A,B,C and D in sequence,the 12-month retention rate of group B was significantly lower than that of other 3 groups,and the differences were statistically significant(x2 =5.973,4.508,4.727,all P ＜ 0.05),and there was no significant difference among the A,C,D groups (all P ＞ 0.05).The spasm-free rate at 3,6,12 months of KD treatment was 19.4％ (19/98 cases),20.4％ (20/98 cases),30.6％ (30/98 cases).The 3-month spasm-free rate in A,B,C,D groups were as follow:41.7％ (10/24 cases),14.3％ (4/28 cases),10.3％ (3/29 cases),11.8％ (2/17 cases),respectively.The 3-month spasm-free rate in group A was significantly higher than that of other 3 groups,and the differences were statistically significant (x2 =10.238,9.219,6.697,all P ＜ 0.05),but there was no significant difference among the B,C,D groups (all P ＞ 0.05).The 6-month spasm-free rates were 41.7％ (10/24 cases),14.3％ (4/28 cases),13.8％ (4/29 cases),and 11.8％ (2/17 cases) in group A,B,C and D in order,and the spasm-free rate in group A was significantly higher than that of other 3 groups,and the differences were statistically significant(x2 =4.924,5.249,4.298,all P ＜ 0.05),but there was no significant difference among the A,C,D groups (all P ＞ 0.05).The 12-month spasm-free rates were 54.2％ (13/24 cases),21.4％ (6/28 cases),24.1％ (7/29 cases),and 23.5 ％ (4/17 cases) in group A,B,C and D,and the spasm-free rate in group A was significantly higher than that in other 3 groups,and the differences were statistically significant(x2 =8.354,7.923,4.364,all P ＜ 0.05),but there was no significant difference among the A,C,D groups (all P ＞ 0.05).Conclusions The spasm-free rate of KD therapy for newly-diagnosed IS is higher than that of IS patients whose drug-therapy failed.KD therapy may be the top priority for IS patients and part of those patients whose drug-therapy failed can still get seizure-free with KD diet.

This study sought to construct recombinant eukaryotic plasmid pcDNA3. 1-sFlt-1 and observe its effect on proliferation of vascular endothelial cells. Total RNA was extracted from human umbilical vein endothelial cells (HUVECs) firstly. The 1st-3rd Ig-like domains of Flt were amplified by polymerase chain reaction (PCR) from the full-length cDNA. Subsequently, the PCR product was cloned into the eukaryotic plasmid pcDNA3.l(+)/myc-His. The constructed recombinant plasmid pcDNA3. 1-sFlt-1 was sequenced. Then recombinant plasmid was transfected into Lewis lung cancer cells. RT-PCR and SDS-PAGE were used to detect the expression of soluble vascular endothelial growth factor (VEGF) receptor mRNA and protein, respectively. MTT method was used to evaluate the effect of sFlt-1 protein on proliferation of HUVECs induced by VEGF. The results showed: (1) The sequence of inserted fragment was correct. (2) Lewis lung cancer cells with recombinant plasmid transfection could express the soluble VEGF receptor mRNA and protein stably. (3) Culture supernatant of Lewis lung cancer cells with sFlt-1 could significantly inhibit the proliferation of HUVECs induced by VEGF. These data suggested that recombinant eukaryotic plasmids pcDNA3. 1-sFlt-1 was constructed successfully, sFlt-1 mRNA and protein were expressed in eukaryotic system stably and sFlt-1 protein could significantly inhibit the proliferaton of endothelial cells induced by VEGF.
Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cells, Cultured ; Genetic Vectors ; genetics ; Human Umbilical Vein Endothelial Cells ; cytology ; Humans ; Transfection ; Vascular Endothelial Growth Factor A ; pharmacology ; Vascular Endothelial Growth Factor Receptor-1 ; biosynthesis ; genetics ; pharmacology

In order to investigate the apoptotic pathway of rabbit annulus fibrosus(AF)cells induced by mechanical overload,an experimental air-pressure model was established in this study to pressurize the rabbit AF cells in vitro.Cells were randomly divided into five groups in which the cells were exposed to a continuous pressure of 1.1 MPa for different lengths of time(0,5,12,24 and 36 h).The cell proliferation and apoptosis were detected by cell counting kit-8(CCK-8)assay and flow cytometry; thealterations in mitochondrial membrane potential were measured by fluorescence microscopy and fluorescence spectrophotometer; the activities of caspase-8 and 9 were determined by spectrophotometry.The results showed that after the cells were subjected to the pressure for 24 or 36 h,the cell proliferation was inhibited; the ratio of cell apoptosis was increased; the mitochondrial membrane potential was decreased;the activity of caspase-9 was enhanced; no activity changes were observed in caspase-8.The results suggested that treatment with a pressure of 1.1 MPa for more than 24 h can lead to the proliferation inhibition and the apoptosis of rabbit AF cells in vitro,and the mitochondrial-dependent pathway is implicated in the pressure-induced AF cell apoptosis.

Objective To assess the effectiveness and safety of dexmedetomidine (Dex) in anesthesia for end-stage renal disease (ESRD)recipients in living related renal transplantation.Methods Forty ESRD patients undergoing living related renal transplantation in the Affiliated Provincial Hospital of Anhui Medical University from October 201 3 to December 201 4 were randomized into the Dex group and the control group,20 patients in each group.In the Dex group,the patients were pumped with the loading dose of Dex at 0.6 μg/kg before anesthesia induction and the procedure was completed within 20 min.Then,the patients were pumped at 0.2 μg/(kg·h)for 1 h and underwent general anesthesia induction.In the control group,the patients were pumped with equivalent normal saline.The anesthesia induction and the maintenance drug of the two groups were the same.The mean arterial pressure (MAP),heart rate (HR)and bispectral index (BIS) before administration (T0 ),20 min after administration (T1 ),before laryngeal mask placement (T2 )and after laryngeal mask placement (T3 )of the two groups were observed and recorded.Delayed recovery and emergence delirium were also observed.Urine output during transplantation,4,8,1 2,24 h after transplantation,as well as endogenous creatinine clearance rate (Ccr)before transplantation and 1 2,24 h after transplantation were recorded.Results Compared with T0 ,the MAP,HR and BIS of the two groups at T2 decreased significantly (all in P <0.05).As for the comparison between two groups,the HR and BIS of the Dex group decreased significantly (both in P <0.05 ),but the MAP didn't decrease significantly (P >0.05 ).The emergence delirium in the Dex group was all mild with the incidence of 1 5% (3 /20),which significantly deceased in comparison with that in the control group (30%,6 /20)(P <0.05).The urine outputs of Dex group during transplantation,and 4 h and 8 h after transplantation were significantly higher than those of the control group (all in P <0.05).The Ccr of the two groups at 1 2 h and 24 h after transplantation significantly increased, compared with that before transplantation (all in P <0.01 ).Conclusions Dex may reduce the incidence of emergence delirium of recipient in living related renal transplantation,increase urine output after transplantation and cause no delayed recovery,which may be used in ESRD patients safely.

OBJECTIVETo evaluate the clinical efficacy of a new formulation of albendazole emulsion (AbzE) in cases of liver cystic hydatidosis.

METHODSTwo regimens of AbzE (10 mg.kg(-1).d(-1) and 12.5 mg.kg(-1).d(-1)) were given to 212 patients with liver cystic hydatidosis in courses ranging from 3 months to more than one year. Assessment of drug efficacy was essentially based on imaging signs with ultrasonography as the main tool. Assessments were performed at the end of different courses and in the follow-up study of 1 - 4 years after the cessation of therapy.

RESULTSAt the end of therapeutic courses, the overall cure rate of the 212 cases was 74.5%, with a 99.1% effective rate. In the follow-up study, the cure rate was 83.1%, effective rate was 89.3%, ineffective rate was 0.6%, and recurrence rate was 10.2%. The highest cure rate was observed in cases receiving AbzE 12.5 mg.kg(-1).d(-1) for 9 months. Retreatment of recurrent cases with AbzE obtained satisfactory results.

CONCLUSIONSAbzE surpassed other currently used antihydatidosis drugs or formulations with its promising efficacy and mild side effects, and could be recommended as a drug of choice in the treatment of cystic hydatidosis.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Albendazole ; administration & dosage ; adverse effects ; Child ; Child, Preschool ; Echinococcosis, Hepatic ; drug therapy ; Emulsions ; Female ; Humans ; Male ; Middle Aged