OBJECTIVETo construct hu-PBL/SCID chimeras and to investigate the development of lymphoma and oncogenicity of the Epstein-Barr virus (EBV).

METHODSHuman peripheral blood lymphocytes (PBLs) were isolated from healthy adult donors and transplanted intraperitoneally into severe combined immunodeficient (SCID) mice. Mice with hu-PBL engraftment from healthy EBV seronegative donors were injected intraperitoneally with EBV-containing supernatant from suspension culture of B95-8 cell line (active infection), whereas mice receiving lymphocytes from healthy EBV seropositive donors were not re-infected with B95-8 derived EBV (latent infection). Pathological examination and molecular analysis were performed on experimental animals and induced neoplasms.

RESULTSIn the early stage of this experiment, 12 mice died of acute graft-versus-host disease, mortality was 34.3% (12/35 mice) with an average life span of 17.5 days. In 19 survival hu-PBL/SCID chimeric recipients from 12 healthy donors, tumor incidence was 84.2% (16/19 mice). The average survival time of tumor-bearing mice was 65.5 days. EBV-related neoplasms in SCID mice were nodular tumors with aggressive and fatal features. Histological morphology of tumors exhibited diffuse large cell lymphomas. Immunohistochemistry revealed that LCA (CD45) and L26 (CD20) were positive, but both PS1 (CD3) and UCHL-1 (CD45RO) were negative, and EBV products ZEBRA, LMP1, and EBNA2 were expressed in a small number of tumor cells. EB virus particles were seen in the nuclei of some tumor cells by electron microscopy, and EBV DNA could be amplified in the tumor tissues by PCR. In situ hybridization indicated that the nuclei of tumor cells contained human-specific Alu sequence.

CONCLUSIONSEBV-induced tumors were human B-cell malignant lymphomas. We obtained direct causative evidence dealing with EBV-associated tumor deriving from normal human cells.

Adult ; Animals ; Antigens, CD20 ; metabolism ; Chimera ; Epstein-Barr Virus Infections ; immunology ; virology ; Graft vs Host Disease ; prevention & control ; virology ; Herpesvirus 4, Human ; physiology ; Humans ; Leukocyte Common Antigens ; metabolism ; Leukocyte Transfusion ; methods ; Lymphoma, B-Cell ; immunology ; virology ; Lysosomal-Associated Membrane Protein 1 ; metabolism ; Mice ; Mice, SCID

OBJECTIVETo explorer the effectiveness of enriched bone marrow stem cells technique for lumbar fusion.

METHODSWith the randomization and control principles, 2 graft materials [Enrichment bone marrow mesenchymal stem cells hybridized with beta-tri calcium phosphate (composite graft group), autologous iliac crest bone graft (autograft group)] were compared in posterior lumbar fusion procedures. 56 patients with degenerative disc disease, lumbar instability or spinal stenosis, were included. The volume of cells suspension in pre- and post-enrichment and the number of nucleated cells (NCs) were identified. The number of osteoprogenitor cells was estimated by counting the colony-forming units which express alkaline phosphatase (CFUs/ALP+). Then the efficiency of the enrichment was evaluated. Clinical follow-up with roentgenogram and Oswestry scale scores was performed for outcome evaluation.

RESULTS(249 +/- 31) ml bone marrow per patient from bilateral iliac crests was aspirated peri-operatively. About (43 +/- 11) ml enriched bone marrow was collected. The number of NCs was concentrated from (15.9 +/- 3.3) x 10(6)/ml to (44.1 +/- 10.8) x 10(6)/ml, CFUs/ALP+ was significantly increased from (118 +/- 86)/ml to(486 +/- 305)/ml. The follow-up was about (26.3 +/- 7.5) months. There was no significant differences in age, gender, disease and fusion segments between the two groups. The fusion rate was 93.3% and 96.2% for composite graft group and autograft group, respectively (chi2 = 0.2146, P = 0.6432). There was no difference in operation time between the two group (t = 0.5243, P = 0.6022), but blood loss in composite graft group was more than that in autograft group (t = 6.4664, P < 0.01). Cell salvage for auto-transfusion could transfuse back half of the blood loss during operation. No hematoma or chronic soreness in the bone marrow donor sites of composite graft group occurred, but a little exudation or moderate swelling in the wound happened in 4 cases which disappeared under medical treatment. Meanwhile, 15.4% patients had hematoma in the iliac bone donor site and 26.9% patients had chronic soreness, but no case had wound problem in autograft group. As for Oswestry scale scores, there was no significant difference between the two groups.

CONCLUSIONSThe enrichment technique of autologous bone marrow stem cells can greatly increase the concentration of MSCs. It is a rapid and safe method used peri-operatively. The composite material of enriched MSCs and porous beta-TCP is a good bone substitute in posterior spinal fusion.

Adult ; Aged ; Bone Substitutes ; Bone Transplantation ; methods ; Calcium Phosphates ; Female ; Follow-Up Studies ; Humans ; Ilium ; transplantation ; Lumbar Vertebrae ; surgery ; Male ; Middle Aged ; Spinal Fusion ; instrumentation ; methods ; Stem Cell Transplantation ; Transplantation, Autologous ; Treatment Outcome

Objective To develop a biological canulated screw and evaluate its mechanical properties, so as to provide theoretical basis for the bio therapy of fracture with enrichment technique of bone marrow stem cells, which could be diffused through the canula, particularly for promoting bone union of femoral neck fracture and preventing avascular necrosis of femoral head. Methods TC4 titanium alloyed canulated cancellous screw (7.3 mm in diameter) commonly used for internal fixation of femoral neck fracture was improved by designing an end sealing plug and side holes in a 900 mm-long canulated screw. The side holes were arranged along the axis of screw, and the first side hole was 20 mm away from the screw tip. The distance between each side hole was 10 mm, and the number of side holes ranged from 0 to 6. The screws without holes were tested as control, and the rest were divided into two groups, i.e., group A: force direction parallel to the side hole, group B: force direction perpendicular to the side hole. Three point bending test on the screw was conducted by using Instron material testing machine, so as to study the relationship between the mechanical strength of the screw and the number of side holes, and the loading direction. Results (1) When the force direction was parallel to the side hole, which was confined within 0, 1 or 2, the bending deformation of the screw reached 3 mm, and no differences were found in the maximum loads and elastic modulus of the screw. However, when the number of side holes was increased to 3 or more, a significant reduction in the maximum load and elastic modulus of the screw was found (P＜0.05). (2) When the force direction was perpendicular to the side hole, which was confined within 0,1 or 2, and the bending deformation of the screw reached 3 mm, no significant differences were found in the maximum loads of the screw. While no significant difference was found in the elastic modulus of the screw when the side hole was 0, 1, 2 and 3. With an increase in the number of side holes, the maximum loads (≧ 3 side holes) and elastic modulus (≧ 4 side holes) were significantly reduced (P＜0.05). (3) When the force direction was perpendicular to the side hole and the number of side holes was 3 or more, the maximum loads and elastic modulus of the screw were all significantly higher than the screw with the same number of side hole under force direction parallel to the side hole. Conclusions (1) For achieving better mechanical properties of the screw, the number of side holes in titanium alloyed canulated cancellous screw (7.3 mm in diameter) should be within 2; (2) If the number of side holes was equal or over 3, screws under force direction perpendicular to the side hole could provide better mechanical properties than screws under force direction parallel to the side hole. This study may provide some theoretical evidence and support for future clinical development and practice of the biological canulated screw.