Factors affecting the transduction of pluripotent hematopoietic stem cells: long-term expression of a human adenosine deaminase gene in mice

An amphotropic retroviral vector, LgAL(delta Mo + PyF101) containing a human adenosine deaminase (ADA) cDNA was used to optimize procedures for the lasting genetic modification of the hematopoietic system of mice. The highest number of retrovirally infected cells in the hematopoietic tissues of long-term reconstituted mice was observed after transplantation of bone marrow (BM) cells that had been cocultured in the presence of both interleukin-1 alpha (IL-1 alpha) and IL-3. A significantly lower number was detected when IL-1 alpha was omitted from such cocultures. The yield of cells that generate spleen colony-forming cells (CFU-S) in the BM of lethally irradiated recipients (MRA-CFU-S) significantly improved on inclusion of the adherent cell fraction of cocultures in the transplant. Retroviral integration patterns in MRA-CFU-S-derived spleen colonies showed that an MRA-CFU-S can produce many CFU-S during BM regeneration. Expression of hADA was detected in the circulating white blood cells of long-term reconstituted animals, demonstrating that the LgAL(delta Mo + PyF101) vector is capable of directing the sustained expression of hADA, and in approximately 35% of the transduced MRA-CFU-S-derived spleen colonies. These results should facilitate the development of gene therapy protocols for the treatment of severe combined immunodeficiency caused by a lack of functional ADA.     

五种布鲁氏菌核酸检测试剂盒检测性能的评价北大核心CSCD

目的比较5种布鲁氏菌核酸实时荧光PCR检测试剂盒的一致性和检出能力,为临床实验室选择检测方法和布鲁氏菌的诊断提供参考依据。方法选用经病原学检测确定为布鲁氏菌阳性的血液样本38份,健康人的血液样本24份,潘氏变形杆菌、溶藻弧菌、河弧菌、铜绿假单胞菌、肺炎克雷伯菌DNA各1份,使用5种试剂盒(编号A-E)分别进行核酸检测,比较5种试剂盒临床样本检测的一致性;选择1份阳性样本核酸用无RNA酶水梯度稀释得到5个浓度(浓度1:4453.13 fg/μL,浓度2:1113.28 fg/μL,浓度3:278.32 fg/μL,浓度4:69.58 fg/μL,浓度5:17.40 fg/μL),每个浓度使用5种试剂盒(编号A-E)分别进行3次检测,比较5种试剂盒的阳性检出率及批内重复性。结果5种试剂盒检测67份DNA样品的符合率稍有不同,试剂盒ABDE的符合率均为100%,试剂盒C的符合率为98.51%。批内重复性显示5种试剂盒在浓度1、浓度2、浓度3水平重复检测DNA的Ct值变异系数均<5%;在浓度1与浓度4梯度区间,试剂盒的阳性检出能力比较显示试剂盒A、B、D较高,为11/12,试剂盒C和E较低,为8/12。结论5种试剂盒的真实性和可靠性较好,灵敏度和符合率稍有差别,特异度均为100%;重复性较好,检测性能良好。部分试剂盒对弱阳性样本的检出能力不强,该类样本可使用多种试剂盒复核,以保障结果的准确性。     

Epstein-Barr virus (EBV)-associated lymphoproliferations in severe combined immunodeficient mice transplanted with Hodgkin's disease lymph nodes: implications of EBV-positive bystander B lymphocytes rather than EBV-infected Reed-Sternberg cells

To establish an in vivo model for the study of Hodgkin's disease and Reed-Sternberg (RS) cells, 25 lymph node tissue samples involved by Hodgkin's disease were grafted into severe combined immunodeficiency (SCID) mice. Ten Epstein-Barr virus (EBV)-associated tumors were obtained in SCID mice. EBV-positive tumors growing in SCID mice were correlated with the presence of EBV-positive nonneoplastic B cells in patient tumors (90% v 26.6%; P<.01) and was independent of the EBV status of RS cells. Our results suggested that EBV-positive tumors growing in SCID mice originated from normal EBV-positive small lymphocytes (bystander B lymphocytes). We also compared the characteristics of these tumors with those obtained after transplantation of 15 non-Hodgkin's lymphoma and four reactive lymph nodes. The latent period to observe a growing tumor in SCID mice was similar between the two groups (12.86 +/- 5.59 weeks for Hodgkin's disease v 13.6 +/- 5.36 weeks for non-Hodgkin's lymphoma and reactive lymph nodes). The relatively high number of EBV-positive small lymphocytes detected in Hodgkin's disease and T-cell lymphoma compared with B-cell lymphoma may account for the greater percentage of EBV- positive tumors obtained in SCID mice. Our results show that SCID mice do not provide the growth conditions that are required for in vivo growth of RS cells. We noted in some SCID tumors, the presence of binucleated and/or multinucleated giant cells resembling RS cells. However, the presence of such cells was not restricted to mice grafted with lymph nodes involved by Hodgkin's disease. We postulate that in previous reports, cells resembling RS cells were just binucleated EBV- positive lymphoma blastoid cells rather than actual RS cells.     

The recommended dietary allowance for protein may not be adequate for older people to maintain skeletal muscle

BACKGROUND: Inadequate dietary protein intake results in loss of skeletal muscle mass. Some shorter-term nitrogen balance studies suggest that the Recommended Dietary Allowance (RDA) of protein may not be adequate for older people. The aim of this study was to assess the adequacy of the RDA of protein for older people by examining longer-term responses in urinary nitrogen excretion, whole-body protein metabolism, whole-body composition, and mid-thigh muscle area. METHODS: This was a 14-week precisely controlled diet study. Ten healthy, ambulatory men and women, aged 55 to 77 years, were provided eucaloric diets that contained 0.8 g protein.kg(-1).day(-1). The study was conducted at a General Clinical Research Center using an outpatient setting for 11 weeks and an inpatient setting for 3 weeks. The main outcome measures included urinary nitrogen excretion, postabsorptive and postprandial whole-body leucine kinetics via infusion of L-[1-(13)C]-leucine, whole-body density via hydrostatic weighing, total body water via deuterium oxide dilution, and mid-thigh muscle area via computed tomography scans. RESULTS: Mean urinary nitrogen excretion decreased over time from Weeks 2 to 8 to 14 (p =.025). At Week 14, compared with Week 2, there were no changes in postabsorptive or postprandial leucine kinetics (turnover, oxidation, incorporation into protein via synthesis, release via breakdown, or balance). Whole-body composition (% body fat, fat-free mass, and protein + mineral mass) did not change over time in these weight-stable subjects. Mid-thigh muscle area was decreased by -1.7 +/- 0.6 cm(2) (p =.019) at Week 14 compared with Week 2. The loss of mid-thigh muscle area was associated with the decrease in urinary nitrogen excretion (Spearman r =.83, p =.010). CONCLUSIONS: The maintenance of whole-body leucine metabolism and whole-body composition is generally consistent with a successful adaptation to the RDA for protein. However, the decrease in mid-thigh muscle area and the association with decreased urinary nitrogen excretion are consistent with a metabolic accommodation. These results suggest that the RDA for protein may not be adequate to completely meet the metabolic and physiological needs of virtually all older people.     

Acute simultaneous proximal occlusion of two major coronary arteries in acute myocardial infarction: successful treatment with percutaneous coronary intervention

Background: Acute myocardial infarction (AMI) due to acute simultaneous proximal occlusion of two major coronary arteries (ASOMC) is a rare but life‐threatening situation. Most patients die suddenly or go into cardiogenic shock (CS). In patients with AMI due to ASOMC identified by coronary angiography (CA), percutaneous coronary intervention (PCI) performed in both infarct‐related arteries (IRAs) at the same time as diagnostic CA is the fastest option to complete revascularization. However, no prospective studies regarding the outcome of such procedures have been published so far. Methods: In this prospective single‐center study, between October 2004 and March 2007, consecutive patients with acute coronary syndrome (ACS) reporting to our university hospital and regional referral center were evaluated for ASOMC by means of emergent CA. When diagnosed with ASOMC, PCI of the IRAs was performed. Clinical data were obtained at baseline, discharge, after 6 months, and after 1 year. Results: Out of 417 patients with ACS, 379 patients (90.9%) suffered an AMI. In 5 patients CA revealed an ASOMC. PCI was performed in 4 patients. One patient with severe triple‐vessel disease was referred for emergent coronary artery bypass graft (CABG) surgery after conventional PCI of one IRA. One patient died in‐hospital due to early in‐stent thrombosis after PCI. At 6‐month follow‐up and at 1‐year follow‐up, 4 patients were alive. Conclusion: In spite of the complex interventions, PCI patients had low in‐hospital mortality and good clinical results at 1‐year follow‐up. Our observations are important in the clinical decision‐making process of AMI due to ASOMC.     

Positionspapier zur „Automatisierten Externen Defibrillation“

Ohne ZusammenfassungDiese Leitlinie ist ursprünglich erschienen in: Z Kardiol 94:287–295 (2005)Herausgegeben vom Vorstand der Deutschen Gesellschaft für Kardiologie Herz- und Kreislaufforschung—in Zusammenarbeit mit der Deutschen Gesellschaft für Internistische Intensivmedizin und Notfallmedizin Bearbeitet im Auftrag der Kommission für klinische Kardiologie (R. H. Strasser, D. Andresen, G. Ertl, F. de Haan, H. Mudra, A. Osterspey, H.-J. Trappe, K. Werdan, außerdem G. Arnold, E. Fleck, H. M. Hoffmeister)     

Prognosis and follow-up of patients with ventricular tachycardias or ventricular fibrillation without coronary heart disease

We studied the follow-up of 49 patients (pts), mean age 34 +/- 9 years, without coronary artery disease who had sustained (duration greater than 30 s) monomorphic ventricular tachycardia (smvt) (n = 42) or ventricular fibrillation (vf) (n = 7). There were 9/49 pts (18%) with smvt who had right ventricular dysplasia (RVD) and 32/49 pts (65%) without structural heart disease ("idiopathic" ventricular arrhythmia) (26/32 pts with smvt and 6/32 pts with vf). There were 6/49 pts (12%) with congestive (COCM) and 2/49 pts (4%) with hypertrophic (HOCM) cardiomyopathy. Mean follow-up was 49 +/- 13 months. During the follow-up 1/9 pts (11%) with RVD died postoperatively from heart failure, 1/26 pts (4%) with idiopathic smvt from cancer and 2/6 pts (33%) with COCM from heart failure. There were no deaths in pts with idiopathic vf. Recurrent smvt occurred in 5/9 pts (56%) with RVD, in 10/26 pts (39%) with idiopathic smvt, in 2/6 pts (33%) with idiopathic vf, in 3/6 pts (50%) with COCM and in 1/2 pts (50%) with HOCM. Our data show that pts with smvt or vf without coronary artery disease have a good prognosis. However, there is a high incidence of recurrent ventricular arrhythmia in these patients.