Harry Segall

Tolerance induction by megadose hematopoietic transplants : Donor-type human cd34 stem cells induce potent specific reduction of host anti-donor cytotoxic T lymphocyte precursors in mixed lymphocyte culture'

BACKGROUND Recently, the use of megadoses of CD34+ hematopoietic progenitors has been reported to abrogate resistance to engraftment, thus overcoming major histocompatibility barriers in bone marrow transplantation in leukemia patients. METHODS The ability of human CD34+ cells to possess potent tolerizing activity was studied by limiting dilution analysis of cytotoxic T lymphocyte (CTL) precursors (CTL-p) in human peripheral blood lymphocytes after addition of purified CD34+ cells. RESULTS The addition of purified human CD34+ cells to primary mixed lymphocyte culture led to a marked reduction of antiallogeneic CTL-p frequency against stimulator cells of the same origin, compared with the response against cells of third-party origin. The CD34+ cells caused a marked inhibition of the CTL activity, when added at an equal number with the responder T cells, and they were still present after the mixed lymphocyte culture, which suggests that no significant killing of CD34+ cells had occurred. The tolerizing activity is abrogated by irradiation and requires cell contact. This pattern of tolerization most closely resembles what has been ascribed to veto cells in other systems. Phenotypic analysis of the purified CD34+ cells showed that they express MHC class I and class II antigens, but do not express costimulatory molecules of the B7 family. CONCLUSIONS It is possible, that CD34+ cells in the megadose transplants-perhaps by their inability to provide costimulatory molecules-are actively reducing the frequency of CTL-p directed against their antigens, and thereby help to overcome allogeneic rejection, and enhance their own engraftment.

Conversion of normal rats into scid-like animals by means of bone marrow transplantation from scid donors allows engraftment of human peripheral blood mononuclear cells

We have recently shown that lethally irradiated normal strains of mice, radioprotected with SCID bone marrow, can be engrafted with human peripheral blood mononuclear cells (PBMC). We now demonstrate that lethally irradiated Lewis rats can also be radioprotected with a transplant of SCID bone marrow cells, administered 1 day after total body irradiation. Split chimerism was found in PBMC, 30 days after transplantation, with predominance of SCID donor-type cells. The average percentages of CD4 and CD8 T cells, of mouse or rat origin, were < 1%. This chimerism status could be maintained for over 3 months. When human PBMC (300-1000 x 10(6) cells) were transplanted intraperitoneally 1 day after the administration of SCID bone marrow, prompt engraftment of human CD4 and human CD8 T cells, as well as human CD20 B cells, was found in the peritoneum and in internal organ (such as liver, lung, spleen, thymus, and lymph nodes). T cell activation was high: about 50% of the cells expressed HLA-DR and almost all expressed CD45RO. High titers of human Ig (> 1 mg/ml) were initially found after 2 weeks; these levels were similar to those found in the irradiated mouse model and in the SCID model. Likewise, marked human anti-tetanus response, predominantly of the IgG type, was recorded 2 weeks after the immunization, reaching maximal levels at 4 weeks. The triple-chimeric SCID-like rats, which accept as much as 1000 x 10(6) human PBMC, can potentially be used to elicit both antibody responses and T cell responses against specific antigens, with the advantages of a larger animal.

Hematopoietic stem cell transplantation for cancer therapy

Bone marrow transplantation has become well established in the treatment of malignant disorders. High-dose chemotherapy with hematopoietic stem cell support is widely used for most hematological malignancies, as well as for some solid tumors. In the light of recent developments in blood progenitor cell harvest, there have been clinical trials with autologous and allogeneic transplants. In particular, the availability of large numbers of blood stem cells, mobilized by granulocyte colony-stimulating factor and collected by leukapheresis, has made it possible to overcome histocompatibility barriers in HLA-mismatched leukemia patients. Other recent developments include new methods for blood progenitor cells mobilization and ex vivo expansion, the use of umbilical cord blood as an alternative source of stem cells, and molecular techniques that may, in the future, provide other modalities of purging tumor cells from autologous grafts.

Human bone marrow-derived mitogenic stimulation selective for breast carcinoma and neuroblastoma cells.

In patients with neuroblastoma (NB) or breast carcinoma (BC), metastatic disease in the bone marrow (BM) is observed more frequently than at any other site, and a high incidence of BM metastases in these patients is associated with advanced disease and poor prognosis. These observations suggest the presence of BM micro‐environmental elements that are favorable for NB and BC tumor cell growth. The influence of normal human BM cell‐derived conditioned medium (CM) on clonogenic growth of BC and NB cell lines was investigated in vitro. The effects obtained were compared with those on tumor cells with a lower potential for BM metastasis. CM from unstimulated cultures of normal, healthy, low‐density BM cells reproducibly and markedly augmented clonogenic growth of 3 BC and 3 NB cell lines. In contrast, growth of cell lines established from human tumors with differing metastatic propensity was unaffected by BM CM. Initial characterization, using crude BM CM, indicated that mitogenic activity (i) is mediated by peptides released by the non‐adherent fraction of low‐density BM cells and (ii) is not abolished by neutralizing anibodies against various cytokines known to be produced by BM cells and to regulate hematopoietic cell growth. Our observations suggest that certain specific peptides in the BM micro‐environment may be responsible for the preferential growth of NB and BC metastases in BM. Int. J. Cancer 78:624–628, 1998.

T cell control of staphylococcal enterotoxin B (SEB) lethal sensitivity in mice: CD4+ CD45RBbright / CD4+ CD45RBdim balance defines susceptibility to SEB toxicity

Radiation chimeras, generated by transplantation of SCID bone marrow into C3H/HeJ mice, show lethal susceptibility to staphylococcal enterotoxin B (SEB), thus constituting a valid murine model for SEB shock. This SEB sensitivity is due to the ability of the irradiated host to restore residual T cell populations, since the SCID donor bone marrow is unable to generate T cells. SCID bone marrow transplanted into irradiated nude mice does not generate SEB‐sensitive chimeras, as a consequence of the inability of the recipient nude mice to develop mature T cells. Thymectomy of normal recipient mice prior to bone marrow transplantation does not affect the development of susceptibility to SEB, suggesting that post‐thymic, residual T cells of the host probably mediate this SEB sensitivity. In vivo depletion experiments show that CD4+ T cells are required for the SEB‐triggered shock, while CD8+ cells suppress it. A further examination of the T helper subpopulations in the SEB‐sensitive mice reveals a prevalence of CD4+ CD45RBdim cells over CD4+ CD45RBbright cells. This T helper balance was statistically significant when correlated with SEB‐induced mortality. Our model provides a possible explanation for the SEB resistance of normal mice: they have a prevalence of CD4+ CD45RBdim over CD4+ CD45RBbright cells.