Object To establish and optimize the technology of clonig rapid propagation by tissue culture in Scutellaria baicalensis Georgi for the protection and development of nature resource as well as seeking for new breeding way of S. baicalensis by biotechnology. Methods Research on the optimized technology of tissue culture including callus inducing, differentiation, plantlet propagation, and rooting were designed. Results Callus could be induced from nodes and internodes easily with 100% inducing rate compared with that from leaves. PP 333 has significant stocky effect to the plantlets. Conclusion The nodes are good explant source in inducing callus. PP 333 in certain concentration which is added to culture media could improve and regulate the differentiation of callus and growth of plantlets as well as increase the survival rate of cultivated seeding.

ObjectiveTo evaluate the inhibiting effects of mitomycin C（MMC） on fibroblasts of human pterygium （HPF） in vitro．MethodsThe fibroblasts derived from the 5 th to 8 th generation of the cultured tissues in human pterygium．These fibroblasts were exposed to culture plates with MMC at 6 gradient concentrations，adjusting the concentration value of the culture medium to 105，104，103，102，10 and 1 μg/L．The cells had been treated with PBS in the control group．After 48 h of incubation，the absorbency had been determined individually by MTT［3-（4，5-dimethylthiazol-2-yl）-2，5-diphenyl-tetrazolium bromide］ method.The inhibitory rates were calculated accordingly．All the data were analyzed with student t test and linear regression．ResultsThe extensive breakage of fibroblasts was observed．There was a statistical difference between the tested group and control group in absorbency (P＜0.05)，especially at the concentrations of more than 100 μg/L (P＜0.01)．The degree of growth inhibition increased with exposure of cells to MMC over 48 h．It showed that there is a positive correlation between the doses of MMC and the antiproliferative effects on HPF．ConclusionMMC significantly inhibites the proliferation of HPF in a dose-dependent manner．

Objective To discuss the application value of CT angiography (CTA) in evaluating renal artery anatomy of laparoscopic nephron-sparing surgery (LNSS) before operation. Methods Retroperitoneal LNSS was performed in 87 patients with renal tumors. Forty cases underwent CTA before LNSS, volume rendering and maximum intensity projection of renal artery were used as three-dimensional reconstruction. The other 47 without CTA were compared as control. Treatment out-comes of the 2 groups were compared, including operation time, warm ischemia time, blood loss, con-version to open surgery, hospitalization stay after operation, and complications after operation. Re-sults All the procedures were completed laparoscopically with no conversion to open surgery in CTA group. One patient underwent conversion to open surgery in the control group. In CTA and control group, the operation time was 94 min(range, 76-118)and 115 min(90-190, P<0.05). The warm ischemia time was 23 min(12-39)and 29 min(18-40, P<0.05). Blood loss was 90 ml(9.0-160) and 130 m1(90-600, P<0.01). Hospitalization stay after operation was 4.1 d(3-5) and 5.5 d(3-9), respectively(P<0.05). The pathologic examination showed negative surgical margin in all cases of renal cell carcinoma. The patients were followed up for 4 months to 32 months. Neither distant nor local recurrences were observed, and the renal function was normal in all cases. Urinary leakage oc-curred in 1 patient of each group. Conclusions CTA could be an effective method in evaluating the supply blood vessels of kidney before LNSS. And it can shorten the operation time and warm ischemia time, reduce the blood loss, as the result of helping the operaters deal with renal artery more quickly and accurately.

Objective To investigate the cytokine spectrum and henlatopoiesis-supportive function of umbilical cord derived mesdnchymal stem cells(UC-MSC),and compare with those of normal adult bone marrow derived mesenchymal stem cells(BM-MSC).Methods The mRNA of cytokine production of UC-MSC and BM-MSC were determined by reverse transcriptasc polymerase chain reaction(RT-PCR)analysis.To evaluate hematopoiesis supporting activity,cord blood(CB)CD+34 cells were co-cultured with UC-MSC or BM-MSC.Colony-forming cells(CFC)were determined after 5 weeks of culture.Results RT-PCR assay showed that UC-MSC had a cytokine spectrum very similar to that of BM-MSC.including expression of the mRNA ofstem cell factor,leukemia inhibitor factor,macrophage colony stimulating factor,Flt3-ligand,interleukin-6,vascular endothelial growth factor and stromal derived factor-1.but UC-MCS additionally expressed mRNA of granulocyte macrophage and granulocyte colony-stimulating factors.After co-culture with CD+34 cord blood cells for 5 weeks,no significant difference in CFC was observed between the CD+34 cells/UC-MSC and CD+34 cells/BM-MSC co-cultures (P＞0.05). Conclusion The cytokine spectrum and hematopoiesis-supponive function of UC-MSC ale similar with that of BM-MSC.

BACKGROUND:Pax6 gene plays an important role in eye development and differentiation, and to study how it regulates the differentiation of human bone marrow mesenchymal stem cel s (BMSCs), gaining the BMSCs stably over-expressing Pax6 is crucial, which is also the basis of stem cel replacement therapy. OBJECTIVE:To construct a lentivirus vector containing Pax6 and detect the expression of Pax6 in transfected human BMSCs.
METHODS:Pax6 gene was extracted using PCR. After its connection with lentivirus vector pHIV-EGFP, it was then packaged by 293T cel s. The human BMSCs were transfected with recombinant lentivirus Pax6-EGFP as wel as lentivirus vector pHIV-EGFP, which was considered negative control group. The cel ular morphology was observed by a fluorescence microscope, and the mRNA expression of Pax6 was detected by real-time PCR.
RESULTS AND CONCLUSION:The recombinant lentivirus Pax6-EGFP was constructed successful y with a titer of 3×109 pfu/L. After the transfection, both the green fluorescent protein and Pax6 gene were expressed detected using fluorescence microscope and real-time PCR, showing that the method of lentiviral transfection is a safe and effective way to modify BMSCs.

Objective To study the natural history of renal cell carcinomas associated with von Hippel-Lindau disease (VHL). Methods An active surveillance strategy was carried out on 11 VHL cases(5 males and 6 females with average age of 45 years)with 32 renal masses. The mean maximum diameter of these masses at initial diagnosis was 2. 5 cm ranging from 0. 5 to 6.2 cm. Clinical materials, radiographic, and pathologic records were reviewed to determine tumor growth rate, subsequent interventions and outcome of follow-up. Results During a median follow-up of 70 months (range 32 to 258 months), bilateral solid renal tumors developed in 6 patients;13 surgical interventions were performed in 8 cases;tumor local recurrence occurred in 4 cases;3 cases died (2 of metastasis diseases and 1 of an unrelated disease) ;8 cases survived with 1 case taking regular hemodialysis.The median follow-up duration for 32 renal masses was 51 months (range 19 to 106 months). The mean tumor growth rate observed were 0. 55 cm/year (range 0. 04 to 1.75 em/year). Three tumors grew faster (1.13-1.75 cm/year), and the other 29 tumors grew slower (0. 17-0.88 cm/year).Among the 3 tumors, 1 was found to be grade Ⅱ at pathologic examination and another developed metastasis. Progression to metastatic disease was found in 2 patients. At the last follow-up, 27 (84 %)tumors were larger than 3 cm and no metastasis disease developed among tumors less than 4 cm.Conclusions The majority of enhanced renal masses with VHL disease may still be indolent and do not metastasize during long period of follow-up even in tumors larger than 3 cm. Active surveillance appears to be a reasonable option for VHL patients with enhanced renal masses less than 4 cm.

Objective To explore the safety and efficacy of laparoscopic nephron sparing nephrectomy for renal tumor. Methods The data of 72 patients diagnosed as renal rumor and treated with laparoscopic nephron sparing nephrectomy were retrospectively analyzed.Procedure detailed as following:firstly,the renal tumor was exposed completely after routine institution of 4 passages;secondly,the tissue 3 cm around the tumor was labeled and then incised using cool scissors after Bulldog occlusion of the renal artery;thirdly,the lesion was covered with anti-bleeding gauze and then intermittently sutured using 1-0 absorbable Dixon suture and crossing Hem-o-lok. Results Operation time was 90-190 min,with warm ischemia time 12-40 min.Bleeding volume was 20-600 m1.with one intra-operative infusion and 4 post-operative infusions.Pathological diagnosis was clear cell RCC in 52 cases,angiomyolipoma in 14,chromophobe RCC in 5,and collective duct cancer in 1 case.Hospitalization time was 5-12 days.With 1-52 months follow-up,no tumor recurrence and metastasis was observed. Conclusions Treatment by laparoscopic nephron sparing nephrectomy for renal tumor could be safe and efficient,but operative experiences and skills are needed.The method of intermittent suture and crossing Hem-o-lok could be useful to shorten the operation time and reduce the complications.

Objective To discuss the management of renal cell carcinoma(RCC) associated with von Hippel-Lindau(VHL) disease. Methods Clinical data were analyzed retrospectively from 28 ca-ses ( 16 males and 12 females, with a mean age of 45 years), of whom 15 had bilateral RCC and 13 had unilateral RCC. VHL germline mutation was analyzed in 25 cases. Nephron sparing surgery (NSS) or radical nephrectomy was performed in 24 cases. Results VHL germline mutations were detected in 25 cases including 14 asymptomatic patients. Among 29 solid renal tumors in 9 cases observed for a mean time of 44 months (range 12 to 86), the mean increase in tumor size was 0. 531 cm/year. There were 19(65.5%) tumors>3 cm at the end of follow-up but only 1 developed retroperitoneum lymph nodes metastasis. A total of 87 solid tumors were removed and 62 (71.3%) solid tumors were man-aged by NSS. Pathological results showed 86 clear cell carcinomas (73 Fuhrman Ⅰ and 12 Fuhrman Ⅱ ) and 1 calcified lesion. During mean follow-up of 50(5-237) months, local recurrence occurred in 4 cases treated with NSS; 26 patients were alive at the end of follow-up. Conclusions DNA testing might be helpful in the earlier detection of asymptomatic VHL patients. Most solid renal tumors in VHL disease grow slowly. The majority of the tumors >3 cm may still be indolent and do not metas-tasize during longer follow-up and can be observed. NSS is effective and safe for RCC in VHL disease.

Objective To study the histological classification,clinical stage,histological grade and prognosis of renal cell carcinoma by analyzing the records of the patients in Shanghai Renji hospital. Methods A consecutive series of 435 patients with renal cell carcinoma between 2003 and 2005derived from the renal cancer database were reviewed clinically and pathologically.The 1997 version of WHO histological classification for renal epithelial tumor,the 2002 version of AJCC clinical TNM staging system and the 1982 version of Fuhrmaffs system for nuclear grade were used.By survival analysis of 57 cases with advanced renal cell carcinoma using Kaplan-Meier method prognostic factors were confirmed using logrank test. Results Of a total 435 patients,cases were classified into 10(accounting for 2.4%of renal cell tumors)hereditary renal cancer in VHL disease,372(85.5%)clear cell renal cell carcinoma(CCRCC),13(3.0%)papillary renal cell carcinoma(PRCC),18(4.1%)chromophobe renal cell carcinoma(CRCC),4(0.9%)oncocytoma,4(0.9%)carcinoma of the collecting ducts of Bellini(CCDB),and 14(3.2%)renal cell carcinoma unclassified.There were 335(77%)patients undergone radical nephrectomy,74(17%)nephron sparing surgery and 26(6%)others,such as palliative nephrectomy.The patients with VHL disease come from 5 Chinese kindred and all had bilateral clear cell renal cell carcinomas and multifocal renal cysts.There were 7 paients of stage Ⅰ and 3 cases of stage Ⅱ and 6 cases of grade Ⅰ and 4 cases of grade Ⅱ.Genetic test revealed that all patients had VHL gene mutation.4 patients had recurrence while no evidence of local advance and distant metastasis were found during a mean of 28.6 months.Patients with chromophobe RCC are all of stage Ⅰ and 5 cases of grade Ⅰ and 13 cases of gradeⅡ.All patients are alive without recurrence or metastasis during a mean of 19.8 months.Collecting ducts RCC all presented with stage Ⅰ but grade Ⅲand with the median survival only 11.3 Months.Of clear cell and papillary RCC,260(67.6%),64(16.6%),32(8.3 %),29(7.5%)were stage Ⅰ,Ⅱ,Ⅲand Ⅳ,and of stage Ⅰ patients 147(38.2%),113(29.4%)were T1a and T1b respectively.124(32.2%),219(56.9%),40(10.4)and 2(0.5%)were grade Ⅰ,Ⅱ,Ⅲ,Ⅳ,respectively.Median survival of 57 advanced RCC is 16.0±1.3months,1-year survival is 55%,and 2-year survival is 31%,respectively.By using logrank test,clinical stage(＜0.01),tumor size(＜0.01),lymphadenopathy(＜0.01),metastasis(＜0.01)and tumor grade(＜0.01)were anatomical and histological prognostic factors for advanced RCC. Coneluslons Different RCC subtypes have different clinical course.The RCC patients in VHL disease have VHL gene mutation and the tumors are often multifocal,bilateral,clear cell type with a low stage and grade which often recurrence but without metastasis.Chromophobe RCC may have a favorable prognosis but collecting duct RCC poor prognosis.In anatomical and histological level,clinical stage,tumor size,lymphadenopathy,metastasis and tumor grade are prognostic factors of survival for advanced RCC.

Objective To study the evolution of renal cell carcinoma treated in a single institution over the past 12 years.Methods A total of 1923 patients with renal cell carcinoma surgically treated from 1999 to 2010 were included in this study.Age at diagnosis,gender,symptoms,tumor size,TNM stage,histological subtype,Fuhrman grade and type of treatment were compared among 3 periods (Period 1:1999-2002,Period 2:2003-2006,Period 3:2007-2010).Results The number of patients grew fasty from 267 cases in the Period 1 to 1092 cases in the Period 3.The incidentally diagnosed cases increased significantly from 49.8％ to 73.6％ (P ＜0.01),while the mean age of patients at diagnosis,the male to female ratio and the proportion of young (＜40 years old) patients were not statistically different.The tumor size at diagnosis gradually decreased from 6.1 cm to 4.8 cm (P ＜ 0.01),and the proportion of small tumors less than 4 cm increased remarkably from 30.3％ to 54.4％ (P ＜ 0.01).Concerning the surgical approach,open surgery decreased from 98.5％ to 73.8％ (P ＜ 0.01),and laparoscopic surgery increased from 1.50％ to 26.2％ (P ＜ 0.01).Overall,the rate of nephron-sparing surgery increased from 5.2％ to 34.7％ (P ＜0.01) in all cases.The use of nephron-sparing surgery increased significantly from 17.9％ to 54.7％ in T1a subset and from 0.00％ to 14.8％ in T1b subset respectively (P ＜ 0.01).Furthermore,radiofrequency ablation was applied and the percent of this approach reached 2.47％ in Period 3.Conclusions The evolution of clinical-pathological characteristics and surgical treatment for renal cell carcinoma in a single institution is apparent over the last 12 years.With the increase of incidentally diagnosed cases and small renal tumors,nephron-sparing surgery has been widely performed in T1 subset instead of traditional radical nephrectomy.