Human CD34+ stem cells produce bone nodules in vivo.

OBJECTIVES: The aim of this study was to select and provide enough stem cells for quick transplantation in bone engineering procedures, avoiding any in vitro expansion step. MATERIALS AND METHODS: Dental germ pulp, collected from 25 healthy subjects aged 13-20 years, were subjected to magnetic-activated cell sorting to select a CD34(+) stem cell population capable of differentiating into pre-osteoblasts. These cells were allowed to adhere to an absorbable polylactic-coglycolic acid scaffold for 30 min, without any prior expansion, and the CD34(+) cell-colonized scaffolds were then transplanted into immunocompromised rats, subcutaneously. RESULTS: After 60 days, analysis of recovered transplants revealed that they were formed of nodules of bone, of the same dimensions as the original scaffold. Bone-specific proteins were detected by immunofluorescence, within the nodules, and X-ray diffraction patterns revealed characteristic features of bone. In addition, presence of platelet endothelial cell adhesion molecule and von Willebrand factor immunoreactivity were suggestive of neo-angiogenesis and neovasculogenesis taking place within nodules. Importantly, these vessels were HLA-1(+) and, thus, clearly human in origin. CONCLUSIONS: This study indicates that CD34(+) cells obtained from dental pulp can be used for engineering bone, without the need for prior culture expanding procedures. Using autologous stem cells, this schedule could be used to provide the basis for bone regenerative surgery, with limited sacrifice of tissue, low morbidity at the collection site, and significant reduction in time needed for clinical recovery.

[Home care for patients with ictus cerebri. An observational study in the metropolitan area of Milano]. / L'assistenza domiciliare ai pazienti colpiti da ictus cerebrale. Studio osservazionale nell'ambito della città di Milano.

Health care assessment is a very relevant process in health services organization. Despite home care appears to be a very important tool in health care delivery system, its impact has been only rarely evaluated in this country. The observational study we performed in the ASL Città di Milano on patients affected by ictus cerebri was aimed at addressing some questions in order to assess whether services can be delivered more cost effectively. We chose to look mainly at the effectiveness and cost of services for the people in need and their caregivers selected in two years time. The results are showed according to different patients' profiles and precocity of recruitment. Their analysis shows that further improvements can still be made in order to achieve a better tailored profile of delivery. Moreover it can be seen how the burden of costs still falls largely on families and caregivers. The study can be seen as premises for further analyses as well as a follow up intervention.

Excimer Laser Assisted Non-occlusive Anastomosis (ELANA). Our experience with a training model in vivo.

AIM: In this study the ELANA Technique has been reproduced in our experimental laboratory in order to verify its feasibility and reproducibility, the percentage of patent anastomosis in acute at different steps along the learning curve of the surgical team, specific problems related to the surgical technique. METHODS: In 20 rabbits New Zealand 4kg body weight the training model in vivo proposed by Tulleken and coworkers has been reproduced, realizing 40 ELANA anastomosis. The model consists in the realization of two different end-to-side anastomosis on the abdominal aorta of each experimental animal, assisted by a special designed suction/excimer laser catheter, then connected by an end-to-end suture. After a few hours the animals are sacrificed and the by-pass site withdrawn and examined in order to verify the percentage of patency in acute. RESULTS: In the first 5 animals (group A), the anastomosis were realized using a jugular vein graft and the procedure results successful in only 3 cases out of ten (30%). For the following experiments - groups B, C and D where an aorta artery graft was used, the percentage of arterial flap retrieval was respectively 50%, 60% and 80%. CONCLUSIONS: ELANA is a feasible fascinating microsurgical technique for the realization of high flow, non-occlusive anastomosis. The rate of success results progressively higher along the learning curve of the surgical team. In our opinion, before the application of the ELANA technique on humans, a period of propaedeutic training in vivo on laboratory animals is essential for the dedicated team.

Limited transgene immune response and long-term expression of human alpha-L-iduronidase in young adult mice with mucopolysaccharidosis type I by liver-directed gene therapy.

Mucopolysaccharidosis type I (MPS I) due to deficient alpha-L-iduronidase (IDUA) activity results in the accumulation of glycosaminoglycans (GAGs) in many of the cells of affected patients. Stable gene replacement by in vivo administration of lentiviral vectors (LVs) has therapeutic potential for metabolic disorders and other systemic diseases. We have previously shown in a murine model the therapeutic potential of lentiviral IDUA vector-mediated gene therapy, in which human IDUA cDNA was driven by the cytomegalovirus promoter. However, the major limitation of this approach was the induction of an immune response against the therapeutic protein, which limited the efficacy and long-term duration of treatment. In this study, we evaluate the potential of liver-directed gene therapy, that is, programming of murine hepatocytes to secrete the enzyme with mannose 6-phosphate (M6P), which can be taken up by distant cells. Eight- to 10-week-old mice were injected via the tail vein with a lentiviral vector expressing human IDUA cDNA driven by the albumin gene promoter selectively expressed in hepatocytes. One month after treatment, IDUA activity was present in the liver and spleen of treated mice; an expression level of 1% normal IDUA activity was sufficient to reduce the GAG level in liver, spleen, kidney, heart, and lung. Interestingly, 6 months after a single injection of this vector, IDUA activity was detectable in several murine tissues; the level of enzyme activity was low but sufficient to maintain the decrease in GAG levels in liver, spleen, kidney, heart, and lung. Also, the level of enzyme-specific antibodies reached at 6 months postinjection was nearly null, and real-time polymerase chain reaction analysis showed high levels of vector DNA content in liver and spleen. Thus, these results show that the use of LV with the albumin gene promoter selectively expressed in hepatocytes limited the immune response to the transgene and allowed stable and prolonged expression of the IDUA enzyme and a partial correction of the pathology.

Evaluation of the 1982 ARA lupus criteria data set in pediatric patients. Committees of Pediatric Rheumatology of the Brazilian Society of Pediatrics and the Brazilian Society of Rheumatology.

OBJECTIVE: Although commonly used, the 1982 revised criteria for the classification of Systemic Lupus Erythematosus (SLE) have not been completely evaluated in pediatric patients. This study was aimed at evaluating the sensitivity and specificity of the 1982 revised criteria when applied to pediatric patients. METHODS: One hundred and three children with SLE and 101 children with other rheumatic diseases were selected from 5 rheumatology centers in Brazil. Diagnosis of SLE by the 1982 criteria were compared with our clinical diagnosis. The diagnosis of other diseases was made according to internationally accepted classification criteria or, when these were not available, according to the physician's own experienced judgement. RESULTS: The median number of criteria fulfilled by the patients with SLE and the controls were 6 and 1, respectively. The most common criteria observed in children with SLE were: abnormal antinuclear antibody titers (94%), arthritis (83%), immunologic disorder (83%), hematologic disorder (70%), malar rash (67%), and photosensitivity (58%). When the immunologic disorder was broken down into its constituent elements, antibodies to dsDNA and Sm were observed in 73.0% (65/89) and 31.4% (15/48), respectively. The sensitivity and specificity observed were 96% and 100%, respectively. CONCLUSION: The 1982 classification criteria can be successfully applied to children with SLE. These criteria may serve as a basis for multi-center collaborative studies on children with SLE.