Generation of suppressor cells in mice immunized with M locus-incompatible lymphocytes.

Alloimmunization of mice with M locus-incompatible lymphocytes resulted in the generation of suppressor cells in the immunized host. Lymph node cells from such alloimmunized mice suppressed the in vitro cytotoxic response of normal cells to H-2 alloantigens. The suppression generated was greater than could be accounted for by dilution of the prekiller cell population with cells possibly devoid of cytotoxic potential from M locus preimmunized mice. Using M locus pseudocongenic mice, the suppressive effect was shown to be largely attributable to M locus determinants; restimulation of suppressor cells in culture with the specific M locus was required for suppression of effector cell generation. The in vivo effect of suppressor cells was tested in a graft-versus-host reaction; injection of M locus preimmunized cells into footpads of F1 hybrid mice suppressed the popliteal lymph node enlargement compared with lymph node size after injection of control preimmunized cells. Although the suppressive effect is mainly attributable to M locus determinants, incompatibility for the DBA/2 antigen may add to the suppression. The study of inhibitory effects on T cell cytotoxicity because of serologically undetectable lymphocyte-activating determinants (Mls) could lead to the better understanding of suppressive mechanisms which may allow the growth of syngeneic tumours.     

Fast lymphoid reconstitution after vascularized bone marrow transplantation in lethally irradiated rats.

BACKGROUND: Bone marrow (BM) transplantation for treatment of hematological and solid malignancies is routinely carried out in conjunction with radio- and chemotherapy. Many patients achieve complete remission of the malignant process; however, their lymphohematopoietic recovery remains in most cases incomplete. This is probably due to the functional changes in the recipient BM stromal cells subsequent to myeloablative therapy. Transplantation of BM hematopoietic cells in a spatial relationship with stromal cells would give an insight into the kinetics of hematological repopulation of the recipient. The aim of this study was to investigate the lymphopoietic reconstitution of irradiated rats after vascularized bone marrow transplantation (VBMT) in comparison with i.v. bone marrow cell (BMC) infusion. METHODS: Lewis rats were totally irradiated with 8Gy and repopulated with syngeneic BMC introduced i.v. or in orthotopic hind limb graft. Ten days after irradiation and BMC graft BM, peripheral blood (PB) and mesenteric lymph nodes (MLN) were collected. The yield and the phenotype of cells were analyzed. RESULTS: VBMT brings much higher cell repopulation of BM cavities of lethally irradiated rats than BMC infusion. Orthotopic hind limb graft promotes also rapid lymphocyte replenishment of PB and MLN of lethally irradiated syngeneic recipients. The population rate of BMC, PB lymphocytes, and MLN lymphocytes was higher after VBMT than BMC injection in suspension. The percentage of T and B lymphocytes in PB and MLN on day 10 after VBMT was comparable with control values. Reconstituted PB lymphocytes showed two subsets of CD4+ cells: "bright" and "dull." All CD4+ cells in PB lymphocytes of i.v. BMC infused recipients expressed low level of these molecules ("dull" subset). CONCLUSIONS: The results of our studies indicate that the presence of stromal cells in their close relationship with stem cells is essential for the fast lyphohematopoietic repopulation of irradiated recipients. The population of CD4+dull cells may represent immature cells. These cells were not found in MLN of VBMT rats. All MLN CD4+ cells represented the "bright" subset, what suggests that the process of cell maturation may occur in the lymphoid organs.     

Manipulation of graft-versus-host disease for a graft-versus-leukemia effect after allogeneic bone marrow transplantation in AKR mice with spontaneous leukemia/lymphoma

Graft-versus-host (GVH) disease can result in a beneficial graft-versus leukemia (GVL) effect after bone marrow transplantation in patients with malignant disease. In this report, we used bacteria-free AKR (H-2k) mice bearing advanced spontaneous T cell leukemia/lymphoma as a moel to evaluate the GVH and GVL effects of bone marrow transplantation using fully incompatible SJL (H-2s) donors. A therapeutic GVL effect, accompanied by increased leukemia-free survival, was obtained only when 0.5 X 10(6) allogeneic lymphocytes (lymph node cells) were added to the marrow inoculum. Transplantation of allogeneic bone marrow without added lymph node cells (or use of syngeneic cells) resulted in a significant increase in leukemia relapse; increasing the dose of allogeneic lymph node cells to 2.0 X 10(6) resulted in significantly higher GVH-associated mortality. Survival and therapeutic benefits were obtained only when the intensity of the GVH reaction was carefully controlled by manipulation of alloreactive lymphocytes present in the marrow. These results suggest, indirectly, that T cell depletion may abolish any GVL effect of marrow transplantation, even if the donor is mismatched with the host at the major histocompatibility complex. The frequency in the spleen of cytotoxic T lymphocytes (CTL) reactive against host alloantigens was estimated using limiting-dilution microcytotoxicity assays at various times after transplantation of allogeneic bone marrow with and without added lymph node cells. The average frequency of CTL was highest in mice that were given marrow plus lymph node cells and tested within the first four weeks after transplantation. The level of CTL activity measured in vitro was dependent on the dose of lymphocytes injected and correlated with both the GVL and GVH effects in vivo. Down-regulation of CTL activity against host, but not third-party, alloantigens in vitro was observed under limiting dilution assay conditions, leading to the suggestion that host-specific regulatory cells may be present in these allogeneic bone marrow chimeras.     

Murine autogeneic mixed lymphocyte culture. Role of culture conditions

In an attempt to determine whether culture conditions significantly influence autoreactivity, we tested the effects of fetal calf, syngeneic, and autogeneic serum with and without 2-mercaptoethanol on the murine self-reactive mixed lymphocyte culture. We used both unfractionated and T-enriched lymph node cells as responders and unfractionated and B-enriched irradiated spleen cells as stimulators in a primary mixed lymphocyte culture (MLC). Using unfractionated cells as responders to various stimulator cell populations, we found excellent allogeneic reactions in all media tested but minimal or no self-reactivity in syngeneic of autogeneic serum. When using T-enriched lymph node cells as responders, allogeneic reactivity was excellent but self-reactivity was present only in the cultures supplemented with fetal calf serum and 2-mercaptoethanol. The possibility that the substances in fetal calf serum and/or 2-mercaptoethanol may be needed to enhance minimal, but biologically relevant, self-reactivity is discussed, as well as the possibility that the culture supplements may be inducing "nonspecific" self-reactivity.     

Lymphocyte traffic within the bone marrow and selective retention of alloreactive cells.

Earlier studies showed that large numbers of isotopically labelled thoracic duct lymphocytes (TDLs) enter the bone marrow (BM) within hours of injection but depart equally as rapidly by 12 to 24 hr. The significance of this rapid flux was investigated further. Early (1/2 to 2 hr) after the i.v. injection of TDLs, BM was shown to contain T cells capable of initiating a graft-versus-host (GVH) reaction in F1 hybrids and in other experiments memory cells against human serum albumin (HSA). Both GVH and memory cell activity had markedly declined in the BM by 12 hr. In contrast to the rapid departure of TDLs from syngeneic BM, F1 hybrid BM retained parental lymphocytes with GVH activity for alloantigens of the opposite parent. F1 hybrid BM under these circumstances supported the transformation and proliferation of lymphocytes activated in situ by alloantigens. These selectively retained T cells also reacted to third-party alloantigens. In addition, TDLs with memory for HSA were retained in the BM of F1 hybrids. The BM is a site in which alloreactive immune responses may be initiated or sustained.     

Development of a new method in promoting fracture healing: multiple cryopreserved bone marrow injections using a rabbit model

Introduction It was hypothesized that multiple injections of cryopreserved bone marrow (BM) would promote fracture healing.Materials and methods Experiments were performed to select a method for freezing BM nucleated cells based on the recovery rate, and to investigate the effects of frozen BM nucleated cells on fracture healing in a rabbit fracture nonunion model. Radiographic and histological evidences of bone union and new bone growth were compared between the experimental group and the control.Results Storage of BM nucleated cells in 10% DMSO plus 90% autologous serum showed the optimal rate of cell recovery, cell viability, and osteoprogenitor cell recovery. Consecutive injections directly to the bone defect site with cryopreserved BM nucleated cells significantly increased the incidence of union, the radiographic volume and bone mineral density of repaired bone as compared with the control (p<0.05).Conclusion A new method was developed to efficiently freeze BM nucleated cells, and cells cryopreserved with this method were found to promote fracture healing by multiple injections. It is suggested that cryopreserved BM injection provides an alternative approach for grafting in delayed union or nonunion.     

Preservation of lymphokine-activated killer activity following T cell depletion of human bone marrow.

T cell depletion has decreased the incidence and severity of graft-versus-host disease following transplantation of allogeneic bone marrow. In the treatment of leukemia, decreased GVHD has often been associated with diminished antileukemia or graft-versus-leukemia (GVL) reactivity resulting in higher relapse rates. However, we have not seen a loss of the GVL effect following transplantation of marrow grafts depleted of CD3+ T cells. This suggests that non-T-cell effectors may play a role in preventing leukemic relapse. To study whether natural killer and lymphokine-activated killer (LAK) activity in BM was compromised by T cell depletion, the effect of T-cell-specific monoclonal antibodies against CD3 and CD6 determinants alone, or in combination, on the generation and expansion of NK/LAK cells was examined in vitro and compared to the effect of T depletion on mitogen-driven T cell proliferation. Limiting dilution analysis revealed that T depletion with CD3 and/or CD6 specific antibodies significantly reduced the number of proliferating T lymphocytes but did not significantly affect the frequency of cells able to expand and mediate LAK activity. Bone marrow, depleted of CD3+ or CD6+ T cells, generated levels of LAK activity equivalent to non-T-cell-depleted bone marrow following long-term culture in recombinant interleukin 2. CD3- NKH-1+ cells were the predominant population in rIL-2 expanded marrow cultures prior to transplant and in the peripheral blood of patients who had received a CD3-depleted marrow graft 21-65 days earlier. These studies show that it is possible to selectively reduce GVH-reactive T cells in allogeneic bone marrow while retaining non-T-effector cells with potential to mediate an antileukemia effect in vivo.     

Age-related osteogenic potential of mesenchymal stromal stem cells from human vertebral bone marrow.

Mesenchymal stem cells (MSCs) residing in bone marrow (BM) are the progenitors for osteoblasts and for several other cell types. In humans, the age-related decrease in bone mass could reflect decreased osteoblasts secondary to an age-related loss of osteoprogenitors. To test this hypothesis, BM cells were isolated from vertebral bodies of thoracic and lumbar spine (T1-L5) from 41 donors (16 women and 25 men) of various ages (3-70 years old) after death from traumatic injury. Primary cultures were grown in alpha modified essential medium with fetal bovine serum for 13 days until adherent cells formed colonies (CFU-Fs). Colonies that stained positive for alkaline phosphatase activity (CFU-F/ALP+) were considered to have osteogenic potential. BM nucleated cells were plated (0.5, 1, 2.5, 5, or 10 x 106 cells/10-cm dish) and grown in dexamethasone (Dex), which promotes osteoblastic differentiation. The optimal plating efficiency using BM-derived cells from donors of various ages was 5 x 106 cells/10-cm dish. BM-derived cells were also grown in the absence of Dex at this plating density. At the optimal plating density, in the presence of Dex, the number of CFU-F/ALP+ present in the BM of the younger donors (3-36 years old) was 66.2 +/- 9.6 per 106 cells (mean +/- SEM), but only 14.7 +/- 2.6 per 106 cells in the older donors (41-70 years old). With longer-term culture (4-5 weeks) of these BM cells in medium containing 10 mM beta-glycerophosphate and 100 microg/ml ascorbic acid, the extracellular matrix mineralized, a result consistent with mature osteoblastic function. These results demonstrate that the number of MSCs with osteogenic potential (CFU-F/ALP+) decreases early during aging in humans and may be responsible for the age-related reduction in osteoblast number. Our results are particularly important in that the vertebrae are a site of high turnover osteoporosis and, possibly, the earliest site of bone loss in age-related osteoporosis.     

Comparative analysis of immunological reconstitution induced by vascularized bone marrow versus bone marrow cell transplantation

Abstract  We have reported previously that vascularized bone marrow transplantation (VBMT) in an orthotopic hind limb graft brings about complete repopulation of bone marrow cavities in lethally irradiated syngeneic recipients within 10 days. Intravenous infusion of an equivalent volume of bone marrow cell suspension was evidently less effective. The purpose of this study was to investigate the reconstitution of immunocompetent compartments of lethally irradiated syngeneic rats after VBMT. Lewis rat hind limbs were transplanted orthotopically into irradiated recipients. Ten days after irradiation and bone marrow transplantation, bone marrow, mesenteric lymph nodes, and sera from rats were harvested. Mesenteric lymph node lymphocytes were analyzed. The responsiveness fomesenteric lymph node lymphocytes (MLNL) to mitogens and cell proliferation in the presence of sera and bone marrow cell (BMC) culture supernatants were measured. Our studies have shown that vascularized bone marrow transplantation brings about rapid replenishment of lymphoid organs of lethally irradiated syngeneic recipients. The re-populating subsets are fully responsive to mitogens. Sera from reconstituting rats had no effect on the proliferation of mature lymphocytes. Intravenous infusion of a number of BMC in suspension equivalent to that grafted in hind limb transplant was less efficient in reconstitution of lymphoid tissue.     

Changes in mixed lymphocyte culture-reactive lymphocytes following alloimmunization of single lymph nodes in sheep.

When an "isolated" single lymph node is challenged with irradiated allogeneic lymphocytes, there is a change in the reactivity of the lymphocytes flowing out of the node when they are cultured in vitro in unidirectional mixed lymphocyte cultures (MLC) against the immunizing lymphocytes. These changes in the reactivity of the recipient lymphocytes are shortlived, follow a set time sequence in relation to the cell traffic changes accompanying the immune response, are the property of small lymphocytes and not blast cells, are exhibited by surface Ig-negative cells, and they are specific for the donor lymphocytes. It is suggested that antigen causes the selective retention of antigen-specific lymphocytes within the stimulated node followed by the proliferation and differentiation of large numbers of antigen-specific cells, which then leave the lymph node as small lymphocytes.     

Development of a simplified counterflow centrifugation elutriation procedure for depletion of lymphocytes from human bone marrow

Graft-versus-host disease (GVHD) remains a major complication of allogeneic bone marrow (BM) transplantation. Techniques that effectively purge BM of mature T lymphocytes should reduce the incidence of GVHD and improve survival. We have developed a simplified, two-flow rate, fixed rotor speed counterflow centrifugation-elutriation (CCE) procedure that reproducibly depletes 99% of lymphocytes from Ficoll-Hypaque(F/H)-separated BM or BM buffy-coat. Two predetermined flow rates (24 and 28 ml/min) were used to purge small and intermediate-to-large lymphocytes, respectively, whereas faster sedimenting cells were recovered at the termination of the run. Lymphocyte depletion was substantiated by pan-T monoclonal antibody analysis as well as by complete loss of responsiveness to alloantigens and mitogens. Despite the lack of mature T cells, the depleted marrow fraction retained lymphoid colony-forming ability. Lymphocyte-purged marrow was obtained in high yield (72%), and retained high viability (greater than 97%) and hematopoietic colony-forming ability (greater than 99%). The ratio of total myeloid/erythroid colony-forming cells to T lymphocytes was 73-fold higher in the lymphocyte-depleted fraction than in unseparated BM. We concluded that a two-step CCE procedure can be used to rapidly deplete lymphocytes from both F/H-separated and buffy-coat BM inocula without altering hematopoietic capacity as measured by the in vitro clonogenic assays. It may be possible to adapt this procedure to the separation of the large number of marrow cells required for human BM transplantation.     

Promotion of pancreatic islet allograft survival by intrathymic transplantation of bone marrow.

An important goal in the treatment of insulin-dependent diabetes by pancreatic islet transplantation is the development of strategies that allow permanent survival of islet allografts without continuous host immunosuppression. In this study, we demonstrate that inoculation of allogeneic bone marrow into the thymus of adult rats treated with a single dose of anti-lymphocyte serum induces an unresponsive state that permits survival of subsequent pancreatic islet allografts transplanted to an extrathymic site. This effect is donor specific, cannot be reproduced by systemic administration of bone marrow, and is associated with persistence of chimeric cells in the thymus of the recipient. In addition, lymph node cells from long-term recipients of intrathymic bone marrow display markedly reduced proliferative responses to donor alloantigens in mixed lymphocyte culture. Interaction of maturing thymocytes with foreign alloantigens may produce the unresponsiveness. This model offers a potential approach for establishing donor-specific allograft acceptance in adult recipients.     

Transplantation tolerance in primates after total lymphoid irradiation and allogeneic bone marrow injection. III. Lymphocyte responsiveness and suppressor cell activity

After total lymphoid irradiation (TLI), allogeneic bone marrow (BM) injection, and organ transplantation in baboons, there is a prolonged period of reduced lymphocyte proliferative responsiveness to polyclonal mitogens and allogeneic lymphocytes. The effect observed is greater with the use of fractionated TLI than after single doses of irradiation. Suppressor cell activity can be demonstrated in vitro in most animals by inhibition of mixed lymphocyte reactivity (MLR) by mitomycin-treated recipient lymphocytes harvested after TLI, with or without allogeneic BM injection, and organ transplantation. Preliminary data suggest the presence of both donor-specific and nondonor-specific suppression, although other interpretations are possible, and suppressor phenomena may not be responsible for the transplantation tolerance observed.     

Transplantation of allogeneic or xenogeneic bone marrow within the donor stromal microenvironment

Successful engraftment of hematopoietic stem cells requires a supportive hematopoietic stromal microenvironment (HSM). Defects in the HSM associated with aplastic anemia, myelofibrosis, or caused by intensive ionizing radiation and chemotherapy generally result in failure of bone marrow (BM) engraftment. Transplantation of donor BM within donor HSM may therefore provide optimal conditions for allogeneic BM transplantation. We have transplanted donor hematopoietic cells together with their own HSM to improve acceptance of allogeneic or xenogeneic BM. The non-myeloablative treatment used induced tolerance to murine allografts and provided conditions for the life-long acceptance of allogeneic HSM. Allogeneic BM transplanted within it's own HSM under the kidney capsule caused less graft-versus-host disease than BM transplanted i.v. Tolerance in mice to xenogeneic (rat) HSM was less complete. Ectopic ossicles were small and contained fewer hematopoietic cells. However, simultaneous transplantation of rat BM and HSM to preconditioned mice improved engraftment of rat BM compared with transplantation of BM alone. Donor hematopoietic cells survived longer on their own HSM than on HSM of recipients.     

同种异体血清在体外培养骨髓间充质干细胞的作用研究

[目的]适当的血清浓度可维持骨髓间充质干细胞(MSCs)的体外培养,血清浓度过低不利于细胞的生长,而高浓度血清则易引起细胞分化。而血清的不同来源对MSCs的培养同样产生影响。传统的培养方法用胎牛血清作为营养支持,临床应用时存在潜在的风险。本研究探讨同种异体血清在体外培养人骨髓间充质干细胞(hBM-SCs)的可行性。[方法]无菌条件下收集人骨髓悬液,分离hBMSCs,分别用含胎牛血清和人血清的培养基培养。分别测定2种方法培养的hBMSCs细胞生长曲线;采用流式细胞仪检测分析培养的hBMSCs表面抗原类型;并将培养的hBMSCs诱导分化为软骨细胞及神经细胞,诱导后的软骨细胞及神经细胞采用免疫组化染色分析。[结果]2种方法培养的间充质干细胞表面抗原CD29、CD44、CD105阳性,CD34、CD45、CD106以及HLA-DR阴性,表达强度无显著性差异;人血清培养的hBMSCs细胞生长速度快于胎牛血清培养组,但分化效率低于后者。[结论]通过生长特性、表面抗原表达以及分化潜能等方面的对比研究,人血清培养hBMSCs与胎牛血清培养差别不大,可以作为一种适合临床的安全培养方法。     

Vascularized bone marrow transplanted in orthotopic hind‐limb stimulates hematopoietic recovery in total‐body‐irradiated rats

Abstract Hematopoietic recovery after bone marrow transplantation (BMT) is reported to be slow with long‐lasting immune deficiency. This may be attributable to lack of a proper microenvironment for hematopoietic cell proliferation and differentiation. We have designed a model in which complete hematopoietic reconstitution of lethally irradiated rats can be achieved by vascularized bone marrow transplantation (VBMT) in an orthotopic hind‐limb graft. The aim of the study was to investigate the process of repopulation of bone marrow cavities and peripheral blood of irradiated rats after VBMT and, in particular, to follow the contribution of grafted BM cells and residual recipient BM cells in hematopoietic regeneration. Lewis hind‐limbs were transplanted orthotopically to totally irradiated (8 Gy) syngeneic sex‐mismatched recipients (VBMT). In the control group 8 × 10⁷ BM cells in suspension were injected intravenously (BMCT). After 10 days BM and peripheral blood (PB) cells were collected from the recipient. For cell subset analysis cytomorphological evaluation of BM smears and flow cytometry of PB cells were performed. Additionally, PCR was performed using specific primers for rat Y chromosome (sex‐determining region Y‐Sry) to detect male (donor or recipient) cells in sex‐mismatched BM graft recipients and the products were analyzed by electrophoresis. VBMT brought about much faster replenishment of nucleated cells in BM and PB than did BMCT. Cytometry analysis of PB cells revealed more lymphocytes in VBMT than in BMCT recipients. The amount of donor DNA of bands corresponding to Y‐Sry was also higher in PB cells of VBMT than of BMCT recipients. The presence of host DNA was observed in PB cells of VBMT rats but was not detected in PB population of BMCT recipients. VBMT is highly effective in hematopoietic reconstitution of irradiated recipients. The fast cell maturation and repopulation may be due to the presence of stromal cells transplanted in a normal spatial relationship with donor hematopoietic cells in hind‐limb graft. Self renewal of radioresistant host cells was seen after VBMT but not after BMCT.     

Hybrid resistance to parental bone marrow grafts in nonlethally irradiated mice

Resistance to parental bone marrow (BM) grafts in F1 hybrid recipients is due to natural killer (NK) cell–mediated rejection triggered through “missing self” recognition. “Hybrid resistance” has usually been investigated in lethally irradiated F1 recipients in conjunction with pharmacological activation of NK cells. Here, we investigated BM‐directed NK‐cell alloreactivity in settings of reduced conditioning. Nonlethally irradiated (1‐3 Gy) or nonirradiated F1 (C57BL6 × BALB/c) recipient mice received titrated doses (5‐20 x 10⁶) of unseparated parental BALB/c BM without pharmacological NK cell activation. BM successfully engrafted in all mice and multilineage donor chimerism persisted long‐term (24 weeks), even in the absence of irradiation. Chimerism was associated with the rearrangement of the NK‐cell receptor repertoire suggestive of reduced reactivity to BALB/c. Chimerism levels were lower after transplantation with parental BALB/c than with syngeneic F1 BM, indicating partial NK‐mediated rejection of parental BM. Activation of NK cells with polyinosinic–polycytidylic acid sodium salt poly(I:C), reduced parental chimerism in nonirradiated BM recipients but did not prevent hematopoietic stem cell engraftment. In contrast, equal numbers of parental lymph node cells were completely rejected. Hence, hybrid resistance leads to incomplete rejection of parental BM under reduced conditioning settings.     

Depletion of T cells from human bone marrow using monoclonal antibodies and rabbit complement. A quantitative and functional analysis

Graft-versus-host-disease (GVHD) remains the principal complication of allogeneic bone marrow transplantation. In animal models mature T lymphocytes have been shown to be responsible for GVHD and, therefore, in vitro treatment of donor bone marrow using monoclonal T cell specific antibodies and complement is currently being investigated as a strategy for the prevention of GVHD. In the present studies anti-T12 and anti-T11 monoclonal antibodies and rabbit complement were used to remove T lymphocytes from normal bone marrow. The efficacy of depletion was investigated by immunofluorescence assays and by in vitro culture of the residual cells using nonspecific mitogens or allogeneic B cells as the proliferative stimulus in the presence of lymphocyte-conditioned medium containing interleukin 2 (IL-2). Immunofluorescence analysis showed complete depletion of T12+ and T11+ cells after treatment with the respective antibodies and with the combination. Nevertheless, culture of treated bone marrow with phytohemagglutinin (PHA) or concanavalin A (Con A) and conditioned media containing IL-2 resulted in the proliferation of mature T cells (T3+, T4+ or T8+, T11+). Stimulation of treated marrow with allogeneic cells (Laz 388) resulted in the growth of a population with natural killer (NK) cell phenotype (T3-, T11+, NKH1+). The latter population was found to be strongly cytotoxic against K562 cells, a standard NK target. As expected, NK cells that are T11+ and T12- appeared to be more effected by in vitro treatment with anti-T11 than with anti-T12. A clonogenic assay was then used to quantitate the efficacy of target cell depletion in vitro. Three sequential incubations of bone marrow with either anti-T12 or anti-T11 plus complement resulted in depletion of 1-2 logs of clonogenic cells. Treatment with both antibodies concurrently resulted in elimination of 2-3 logs of clonogenic target cells. Although multiple treatments with both anti-T12 and anti-T11 were more effective than similar treatment with only one antibody, it remains to be established whether such combinations will be necessary in the clinical setting or whether more selective depletion of T cells without removal of NK cells might be optimal.     

Elevation of serum albumin levels in nagase analbuminemic rats by allogeneic bone marrow cell transplantation

We investigated the feasibility of correcting the congenital absence of albumin in Nagase analbuminemic rats (NARs) by allogeneic bone marrow cell transplantation (BMT). Seven-week-old male NARs were used as recipients, and 6- to 8-week-old male Sprague-Dawley (SD) rats were used as allograft donors. NARs were divided into three groups: a BMT group (n=10) in which bone marrow cells were infused into the liver; a hepatocyte transplantation (HCT) group (n=8) in which hepatocytes were transplanted into the liver, and a control group (n=8) in which PBS was injected into the portal vein. Serum albumin levels were measured as an indicator of the function of the grafted cells, and the phenotypic characteristics of the engrafted cells in the recipient's liver were assessed with immunohistochemical and immunofluorescence techniques. At 8 weeks after cell transplantation, the serum albumin levels of the BMT group and HCT group were significantly higher than in the control group. The hepatocyte-like cells derived from bone marrow cells expressed albumin in liver of the NARs. According to this result, bone marrow cells can differentiate into hepatocyte-like cells in vivo. The results show that BMT is an effective treatment for congenital analbuminemia in a rat model and suggest that allogeneic BMT can be used as an efficient therapy for hereditary metabolic diseases.