SARS-CoV GDH株N蛋白全长基因克隆及其可溶性表达

目的构建SARS冠状病毒N蛋白的原核表达质粒,诱导重组蛋白表达并纯化,鉴定其抗原性。方法以我国SARS冠状病毒GDH株总RNA为模板,采用RT-PCR技术扩增N蛋白的全长基因,TA克隆后测序。构建pET-23d的N基因表达载体,用IPTG诱导目的蛋白表达,利用硫酸铵沉淀、分子筛层析及离子交换层析纯化重组蛋白,免疫印迹鉴定重组蛋白。结果RT-PCR扩增出1269bpSARS冠状病毒N蛋白的基因片段,其序列分析结果与SARS-CoVGD01、BJ01株的同源性为99.92%;该基因在大肠杆菌表达系统中高效表达,占可溶性蛋白的33.57%,表达产物为非融合的可溶性蛋白,Westernblot结果显示重组N蛋白具有良好的抗原性;纯化后重组N蛋白纯度为92.9%。结论成功构建了SARS冠状病毒N蛋白的重组表达质粒,并在大肠杆菌中以非融合蛋白的形式得到高效的可溶性表达,为SRAS的诊断和疫苗的研制奠定了基础。     

诺氟沙星缓释片的初步研究

本文初步研究了诺氟沙星缓释片的处方组成。并对不同处方进行了体外溶出试验,实验结果表明：本品的体外释药可维持12h以上且体外溶出符合一级动力学过程。     

Autologous transplantation for diffuse aggressive non-Hodgkin's lymphoma in patients never achieving remission: a report from the Autologous Blood and Marrow Transplant Registry.

PURPOSE: To evaluate the results of high-dose chemotherapy and autologous hematopoietic stem-cell transplantation (autotransplants) in patients with diffuse aggressive non-Hodgkin's lymphoma (NHL) who never achieve a complete remission with conventional chemotherapy. PATIENTS AND METHODS: Detailed records from the Autologous Blood and Marrow Transplant Registry (ABMTR) on 184 patients with diffuse aggressive NHL who never achieved a complete remission with conventional chemotherapy and subsequently received an autotransplant were evaluated. Transplants were performed between 1989 and 1995 and were reported to the ABMTR by 48 centers in North and South America. RESULTS: Seventy-nine (44%) of 184 patients achieved a complete remission or a complete remission with residual imaging abnormalities of unknown significance after autotransplantation. Thirty-four (19%) of 184 had a partial remission and 55 (31%) of 184 had no response or progressive disease. Eleven patients (6%) were not assessable for response because of early death. The probabilities of progression-free and overall survival at 5 years after transplantation were 31% (95% confidence interval [CI], 24% to 38%) and 37% (95% CI, 30% to 45%), respectively. In multivariate analysis, chemotherapy resistance, Karnofsky performance status score less than 80 at transplantation, age > or = 55 years at transplantation, receiving three or more prior chemotherapy regimens, and not receiving pre- or posttransplant involved-field irradiation therapy were adverse prognostic factors for overall survival. CONCLUSION: High-dose chemotherapy and autologous hematopoietic stem-cell transplantation should be considered for patients with diffuse aggressive NHL who never achieve a complete remission but who are still chemotherapy-sensitive and are otherwise transplant candidates.     

HPLC法测定5-单硝酸异山梨酯缓释胶囊含量

建立了高效液相色谱法测定5-单硝酸异山梨酯缓释胶囊中5-单硝酸异山梨酯的含量方法.采用十八烷基键合硅胶填充柱(250×4.6mm,10μm),以咖啡因为内标,甲醇-水(2476)为流动相,流速为1.0ml/min,检测波长为210nm,柱温为室温.线性范围为80～600μg/ml,平均回收率为99.93%(RSD=0.61%),辅料和内标物对测定无干扰.     

Outcome of lower-intensity allogeneic transplantation in non-Hodgkin lymphoma after autologous transplantation failure

We studied the outcome of allogeneic hematopoietic stem cell transplantation after lower-intensity conditioning regimens (reduced-intensity conditioning and nonmyeloablative) in patients with non-Hodgkin lymphoma who relapsed after autologous hematopoietic stem cell transplantation. Nonrelapse mortality, lymphoma progression/relapse, progression-free survival (PFS), and overall survival were analyzed in 263 patients with non-Hodgkin lymphoma. All 263 patients had relapsed after a previous autologous hematopoietic stem cell transplantation and then had undergone allogeneic hematopoietic stem cell transplantation from a related (n = 26) or unrelated (n = 237) donor after reduced-intensity conditioning (n = 128) or nonmyeloablative (n = 135) and were reported to the Center for International Blood and Marrow Transplant Research between 1996 and 2006. The median follow-up of survivors was 68?months (range, 3-111 months). Three-year nonrelapse mortality was 44% (95% confidence interval [CI], 37%-50%). Lymphoma progression/relapse at 3 years was 35% (95% CI, 29%-41%). Three-year probabilities of PFS and overall survival were 21% (95% CI, 16%-27%) and 32% (95% CI, 27%-38%), respectively. Superior Karnofsky Performance Score, longer interval between transplantations, total body irradiation-based conditioning regimen, and lymphoma remission at transplantation were correlated with improved PFS. Allogeneic hematopoietic stem cell transplantation after lower-intensity conditioning is associated with significant nonrelapse mortality but can result in long-term PFS. We describe a quantitative risk model based on pretransplantation risk factors to identify those patients likely to benefit from this approach.     

Autologous Transplantation in Follicular Lymphoma with Early Therapy Failure: A National LymphoCare Study and Center for International Blood and Marrow Transplant Research Analysis

Patients with follicular lymphoma (FL) experiencing early therapy failure (ETF) within 2 years of frontline chemoimmunotherapy have poor overall survival (OS). We analyzed data from the Center for International Blood and Marrow Transplant Research (CIBMTR) and the National LymphoCare Study (NLCS) to determine whether autologous hematopoietic cell transplant (autoHCT) can improve outcomes in this high-risk FL subgroup. ETF was defined as failure to achieve at least partial response after frontline chemoimmunotherapy or lymphoma progression within 2 years of frontline chemoimmunotherapy. We identified 2 groups: the non-autoHCT cohort (patients from the NLCS with ETF not undergoing autoHCT) and the autoHCT cohort (CIBMTR patients with ETF undergoing autoHCT). All patients received rituximab-based chemotherapy as frontline treatment; 174 non-autoHCT patients and 175 autoHCT patients were identified and analyzed. There was no difference in 5-year OS between the 2 groups (60% versus 67%, respectively; P?=?.16). A planned subgroup analysis showed that patients with ETF receiving autoHCT soon after treatment failure (≤1 year of ETF; n?=?123) had higher 5-year OS than those without autoHCT (73% versus 60%, P?=?.05). On multivariate analysis, early use of autoHCT was associated with significantly reduced mortality (hazard ratio, .63; 95% confidence interval, .42 to .94; P?=?.02). Patients with FL experiencing ETF after frontline chemoimmunotherapy lack optimal therapy. We demonstrate improved OS when receiving autoHCT within 1 year of treatment failure. Results from this unique collaboration between the NLCS and CIBMTR support consideration of early consolidation with autoHCT in select FL patients experiencing ETF.     

Degradation products of extracellular matrix affect cell migration and proliferation

Biologic scaffolds composed of extracellular matrix (ECM) are utilized in numerous regenerative medicine applications to facilitate the constructive remodeling of tissues and organs. The mechanisms by which the host remodeling response occurs are not fully understood, but recent studies suggest that both constituent growth factors and biologically active degradation products derived from ECM play important roles. The objective of the present study was to determine if degradation of ECM scaffold materials in vitro by methods that are biochemically and physiologically relevant can yield products that possess chemotactic and/or mitogenic activities for fully differentiated mammalian endothelial cells and undifferentiated multipotential progenitor cells. ECM harvested from porcine urinary bladder was degraded enzymatically with pepsin/hydrochloric acid or papain. The ECM degradation products were tested for chemoattractant properties utilizing either 48-well chemotaxis filter migration microchambers or fluorescence-based filter migration assays, and were tested for mitogenic properties in cell proliferation assays. Results showed that ECM degradation products possessed chemotactic and mitogenic activities for multipotential progenitor cells and that the same degradation products inhibited both chemotaxis and proliferation of differentiated endothelial cells. These findings support the concept that degradation products of ECM bioscaffolds are important modulators of the recruitment and proliferation of appropriate cell types during the process of ECM scaffold remodeling.     

Improved mobilization of peripheral blood CD34+ cells and dendritic cells by AMD3100 plus granulocyte-colony-stimulating factor in non-Hodgkin's lymphoma patients

AMD3100 is a drug capable of mobilizing peripheral blood stem cells (PBSCs) in donors and in cancer patients as a single agent or in combination with granulocyte-colony-stimulating factor (G-CSF). We initiated a phase II study of 11 refractory or relapsed non-Hodgkin's lymphoma (NHL) patients, receiving 16 microg/kg daily of G-CSF for 4 days followed by 240 microg/kg of AMD3100 given subcutaneously on a new schedule of 9-10 h before apheresis collection on day 5. Our aims were to assess the effect of AMD3100 on the mobilization of CD34+ cells, dendritic cells (DCs) and lymphoma cells. Administration of G-CSF and AMD3100 were continued daily until >or=2 x 10(6) CD34+ cells/kg were collected. Adequate collection of the target of CD34+ cells was achieved in all but 1 patient within 2 days, and 10/11 patients were transplanted within 2 months. All transplanted patients engrafted with a mean of 10 and 12 days for neutrophils and platelets, respectively. Addition of AMD3100 to G-CSF resulted with >2.5-fold increase in CD34+ cells/microl (p = 0.0001) and in a >2-fold increase in pDC1 and pDC2 cells/microl (p = 0.003). Adverse events related to AMD3100 were minimal. AMD3100 was generally safe and improved PBSC and DC cell mobilization with no apparent contamination of lymphoma cells.