Objective: To study the characteristics of mouse embryonic stem (ES) cell line R1 expressing green fluorescent protein (GFP) efficiently and stably. Methods: Four kinds of biological characteristics: growth curve, express level of alkaline phosphatase, karyotype and pluripotentiality of the subclones of R1 ES cell expressing GFP were observed and analysed. Results: No obvious differences between R1 ES cells and the 4 ES cell clones expressing GFP［ESG（+）］ on the proliferation speed, differentiation state and the ratio of normal karyotype were observed. Teratocarcinoma concluded 3 germinal layers could form after inoculating ESG(+) cells to nude mice. Conclusion: The expression of GFP may not make any detectable effect on the proliferation speed, differentiation state and the pluripotentiality of R1 ES cells. This research work ensures efficient utilization of GFP as a reporter gene tracing ES cell in vivo.

Objective To observe and evaluate the effect of ultrasound guided percutaneous ethanol injection for hyperparathyroidism secondary to chronic renal failure. Methods Fifty-three patients with secondary hyperparathyroidism in Nanjing General Hospital Military Command from January 2011 to May 2013 were treated by percutaneous ethanol injection. The echogenicity, size and blood supply of parathyroid glands were observed by color Doppler ultrasound before and after treatment. Parathyroid hormone (PTH) and volumes of parathyroid glands were analyzed and compared with non-paired t test. Results Before treatment, the parathyroid gland volume was (5.28±0.84) cm3. The enlarged parathyroid glands had homogeneous hypoechogenicity and abundant blood supply, and the PTH was (1041.6±37.1) ng/L. After treatment, the size of solitary parathyroid gland was (3.93±0.67) cm3 which did not change significantly compared with that before treatment. The echogenicity of parathyroid glands enhanced and blood supply decreased after injection ethanol into solitary parathyroid. PTH [(509.2±27.6) ng/L] decreased obviously (t=3.792, P < 0.05). Conclusion Color Doppler ultrasound guided percutaneous injection of ethanol has an important role in the treating of secondary hyperparathyroidism and the curative effect can be observed timely.

Objective To investigate the clinicopathologic and immunohistochemical characteristics of breast carcinosarcoma with osteosarcoma components and its differential diagnosis. Methods The pathologic features and clinical manifestations of the two patients were retrospectively reviewed. Immunohistochemistry was performed and the literature was reviewed. Results Histopathologically, the neoplasm consisted of invasive ductal carcinoma of no special type and poorly ( case 1 ) or well (case 2) differentiated osteosarcomatoid elements. The morphological transition from carcinoma to sarcomatoid elements was seen. Immunohistochemically,the carcinoma cells were positive for CK and EMA , the sarcomatoid elements were stained positive for vimentin, and a few cells of two elements were positive for S-100 protein. Ki-67 and VEGF were over expressed in both elements of Case 1.In case2 Ki-67 expression was low in both elements and VEGF over expression was only seen in sarcomatoid elements. Some of the carcinoma cells were positive for ER. Conclusions Carcinosarcoma of the breast with osteosarcomatoid elements is a rare type of mixed epithelial/mesenchymal metaplastic breast carcinoma. The types and proportion of carcinoma and sarcomatoid elements determine the diagnosis.

Objective To analyze the causes for failed treatments of proximal femoral fractures and to evaluate the curative results of total hip replacement(THR)to salvage the failures.Methods A retrospective study was conducted for 23 consecutive cases of THR to salvage failed treatments of proximal femoral fractures from February 2002 to April 2005.There were 17 cases of femoral neck fracture,five cases of intertrochanteric frac- ture,and one case of femoral head fracture.Results Inadequate reduction after inadequate fixation,incorrect materials or technical errors in internal fixation were the most significant reasons for the failure of early treatment. Twenty-one of the 23 patients were available for fullow-up that ranged from 8 to 46 months(mean,32 months). No such complications as wound infection,hip dislocation,prosthesis breakage or deep venous thrombosis were found.Their mean Harris score increased from 48.3 preoperatively to 87.6 postoperatively.Condusions Re- duction,internal fixation,and fixation technique are very important to treatments of proximal femoral fractures. High energy trauma in young patients and osteoporosis in old patients make treatment of proximal femoral fractures even more difficult.THR is a satisfactory and safe way to salvage a failed treatment of proximal femoral fracture.

Adolescent ; Adult ; Aged ; China ; epidemiology ; Humans ; Male ; Middle Aged ; Motor Activity ; Urban Population ; Young Adult

Objective: To study the factors effecting the expression of the reporter green fluorescent protein (GFP) gene in the mouse embryo stem cell line R1. Methods: Three different kinds of GFP euko-expression vectors were constructed, and the expression efficiency was contrasted both at mRNA and protein levels after they were integrated into the chromosomes of host cells. Results: At protein level, the GFP expression level of the colonies transfected by the expression vector-pEF-GFP with the promoter of the peptide elongation factor (EF) were significantly higher than that of the colonies transfected by pCMV-GFP with CMV promoter and by pdCMV-GFP with double copies of CMV-GFP expression unit. There was no significant difference between the colonies transfected by pCMV-GFP and pdCMV-GFP. The detection results on mRNA level of GFP had the same tendency as that at protein level. Conclusion: (1) GFP gene expression efficiency controlled by EF promoter is distinctly higher than that by CMV promoter in NIH3T3 and R1 ES cell line.(2) A slight increase of the copy number of the foreign gene expression units in the host chromosome can not make obvious increase of its expression efficiency. (3) The vector express GFP in R1 ES cell line efficiently and stablely is obtained.

Objective To explore the diagnostic values of vascular endothelial growth factor (VEGF) and p -human chorionic gonadotropin( β -hCG) for early ectopic pregnancy. Methods The serum levels of VEGF and β-hCG were measured in 109 patients with suspected ectopic pregnancy by ELISA and RIA respectively, receiver operating characteristic (ROC) curvers were established, the diagnostic values of VEGF and β -hCG for ectopic pregnancy were analyzed. Results One hundred and nine patients with suspected ectopic pregnancy were diagnosed in the end: 62 cases with ectopic pregnancy and 47 cases with intrauterine pregnancy. The levels of VEGF in ectopic pregnancy were significantly higher than those in intrauterine pregnancy (P ＜ 0.05), but the levels of β -hCG were significantly lower than those in intrauterine pregnancy (P ＜ 0.05). The areas under ROC curves of VEGF were 0.87, the areas under ROC curves of β -hCG were 0.71. In VEGF ＞ 140 ng/L for diagnosis of ectopic pregnancy, the sensitivity and specificity were 75.8% (47/62) and 80.9% (38/47) respectively. In β-hCG ＜ 1200 U/L for diagnosis of ectopic pregnancy, the sensitivity and specificity were 64.5%(40/62) and 61.7%(29/47) respectively. The sensitivity and specificity of combined detection of VEGF and β -hCG were 80.6%(50/62) and 57.4%( 27/47) respectively. Conclusions The diagnostic values of VEGF are better than those of β -hCG. The combined detection of VEGF and β -hCG can increase the rate of early diagnosis and shorten observation time.

This study was based on live yeast cell derivative (LYCD), which was produced by live yeast cell stressed with high temperature and H 2O 2. The results showed that pretreating of low dose(37℃and 0.2mmol/L H 2O 2) could increase the content of GSH and the activity of SOD and CAT. These pretreatment could induce the resistance to lethal concentration of H 2O 2. LYCD was produced by yeast treated with 37℃ and 0.2mmol/L H 2O 2. And it was found that the survival of yeast treated with lethal concentration of H 2O 2 obviously increased, while LYCD was added in yeast culture. It indicated that LYCD could have resistance to oxidative condition.

Objective To investigate effects of different doses of recombinant human bone morphogenetic protein-2 (rhBMP-2) on vascular endothelial growth factor (VEGF) expression in fetal mouse osteoblasts.Methods Calvaria osteoblasts of fetal mice of 19 days were cultured.The effects of rhBMP-2 at different doses and different action times on VEGF expression patterns in fetal mouse osteoblasts were observed with reverse transcrip- tion-polymerase chain reaction (RT-PCR) and Western blot staining.Results In the present study,RT-PCR detected a steady expression of VEGF mRNA in the control fetal mouse osteoblasts.The levels of VEGF mRNA increased in an apparent biphasic manner with a maximum stimulation (about 2-fold above the control,P＜0.05 ) in both VEGF mRNA species observed at 300 ng/mL of rbBMP-2.After 48 h of rhBMP-2 treatment,the VEGF mRNA levels approached those in the control.The VEGF mRNA levels appeared to be biphasic in rhBMP-2-treated cultures,showing peak induction at 3 and 24 h and remaining elevated at 48 b.Compared with the individual control value at each time point,an apparent maximum increase (about 2.5-fold above the control,P＜0.05) occurred at 6 h.The second peak induction,about 2-fold above that in the control,occurred at about 36 h.Conclusions The expression of VEGF mRNA is steady in the control fetal mouse osteoblasts.RhBMP-2 can promote the expression of VEGF in dose-dependent and time-dependent manners.

Objective: To study the construction and application of a vector with Cre recombinase recognition site lox66 for mouse HPRT gene targeting in embryonic stem(ES) cells. Methods: Using the HPRT genomic DNA fragment and synthesized oligonucleotides, pSP HPRT lox66 Neo was designed and constructed as a replacement vector by common molecular cloning techniques. After the linearized pSP HPRT lox66 Neo DNA was electroporated into ES cells, and transfected cells were cultured in G418/6 TG drug selection medium. The recombination efficiency of this vector was tested. Results: The main components of pSP HPRT lox66 Neo were a positive selection gene Neo, lox66, long and short homologous fragments of mouse HPRT gene and plasmid backbone. Twenty ES cell clones with HPRT gene inactivated were obtained. Conclusion: An effective replacement vector with Cre recombinase recognition site lox66 is constructed and applied to HPRT gene targeting in ES cells.