Doses Of Spleen Cells Research Articles

Human Immune Reconstitution With Spleen Cells in SCID/Beige Mice

Background We developed an original experimental model to study chronic vascular rejection (CVR) consisting of a graft of human mesenteric artery followed by human immune reconstitution into CB.17 SCID/Beige mice. Human immune reconstitution achieved after human PBMC injection has often been variable and incomplete. The aim of this work was to develop an alternative method to achieve a complete, functional human immune reconstitution. Method After institutional authorizations, spleen cells were recovered from cadaveric organ donors. Single intraperitoneal injections of various doses of spleen cells were made into 70 CB.17 SCID/Beige mice. Reconstitution of the human immune system was monitored by flow cytometry (circulating human cells) and ELISA (human IgG). Colonization of murine lymphoid organs by human cells was studied by immunohistochemistry and flow cytometry. Evaluation of the immune function consisted of examination of CVR lesions in human arterial grafts. The animals were humanely killed at day 28. Results After injection of 30 to 40 × 10 6 spleen cells, the mice showed significant human CD3 +, CD19 +, and CD56 + populations in peripheral blood. The mean human cells levels were, respectively, 8.2% ± 5.4%, 2.9% ± 1.2%, and 5.3% ± 5.1%. Murine spleen and mesenteric lymph nodes were colonized by human T and B cells, while the murine thymus was only colonized by human T cells. Human IgG was detected in murine serum (65.9 ± 63.3 mg/L) and typical CVR lesions were observed within the allogeneic grafts. Conclusion Intraperitoneal injection of 30 to 40 × 10 6 human spleen cells into CB.17 SCID/Beige mice induces complete and functional human immune reconstitution allowing the study of CVR under human allogeneic conditions.

Inhibition of lung metastasis by adoptive immunotherapy using iscador

Adoptive immunotherapy using spleen cells activated with Iscador was found to inhibit tumour growth significantly (P<0.001). Metastatic B16F10 tumour bearing animals treated with a single dose of spleen cells activated with Iscador (in vitro) showed 100% inhibition of tumour nodule formation on 21st day. Single injection of splenocytes isolated from mice treated with Iscador inhibited the tumour nodule formation by 93.8%. Animals treated with in vivo and in vitro activated spleen cells along with Iscador had an increase in life span of 119% and 81% respectively. Treatment of animals with low dose of Iscador after adoptive immunotherapy further augmented the life span. Animals which underwent adoptive immunotherapy showed significantly reduced lung collagen hydroxyproline (9.8μg/mg protein) and serum sialicacid (24 6μg/ml serum) levels compared to control tumour bearing animals with increased levels of lung hydroxyproline (26.95μg/mg protein) and serum sialic acid levels (151.3μg/ml serum) Serum γ -glutamyl transpeptidase levels were found to be significantly reduced in the group of animals treated with Iscador and Iscador activated splenocytes (16.6 ± 2.3 nmol P-nitroaniline released/ml serum) Group of animals treated with Iscador activated splenocytes alone also showed significantly reduced serum γ -glutamyl transpeptidase levels (17.3 ± 10 nmol P-nitroaniline released/ml serum) compared to control tumour bearing animals (91 • 12 nmol P-nitroaniline released /ml serum)

Effect of granulocyte-macrophage colony-stimulating factor on the induction of unresponsiveness by lymphoid cells.

Skin grafts can be significantly prolonged in ALS-treated mice by the injection of 25 x 10(6) donor bone marrow cells or 50 x 10(6) spleen cells. Lymph node cells and thymocytes are only minimally effective in prolonging grafts. The effect of a hematopoietic growth factor, granulocyte-macrophage colony stimulating factor (GM-CSF) was studied in this model of unresponsiveness. C3H/He lymphoid cell donors were treated with GM-CSF. Either normal or GM-CSF-treated cells were injected into ALS-treated B6AF1 mice grafted with C3H/He skin. GM-CSF treatment significantly augmented the effect of marrow in prolonging graft survival at doses of 1 to 25 x 10(6) cells. In contrast, GM-CSF had no effect on the graft-prolonging effect of spleen cells when 50 x 10(6) cells were given. When the dose of cells was reduced to 25 x 10(6), graft survival in the group given GM-CSF-treated cells was prolonged compared with survival in the group given normal cells. Grafts in the group given GM-CSF-treated lymph node cells were rejected in sensitized fashion. When marrow and spleen are separated on a Percoll gradient, the cell active in promoting graft survival is recovered primarily in the 52.5% fraction. The graft-prolonging effect of the 52.5% marrow fraction was not affected by GM-CSF treatment. In contrast, GM-CSF-treated marrow cells in the 60% fraction significantly prolonged graft survival, while normal marrow cells in this fraction had no effect on graft survival. GM-CSF-treated spleen cells in the 52.5% and 60% fractions significantly decreased graft survival compared with normal cells when given at a dose equal to the number of cells recovered from 50 x 10(6) cells. When the dose of fractionated spleen cells was reduced, GM-CSF-treated spleen cells were more effective than normal cells in prolonging graft survival. These results indicate that GM-CSF activates a cell in marrow that promotes graft survival. This cell is recovered in the 60% Percoll fraction. In contrast, GM-CSF appears to affect two cell populations in spleen, one beneficial and one detrimental to graft survival. The predominant effect depends on the dose of spleen cells that is given.

Further evidence for H-2-unrestricted induction of minor histocompatibility antigens-specific T cell immunity in vivo.

Antigenic requirements for inducing minor histocompatibility antigens (MIHA)-specific T cell immunity for second set rejection (SSR) of a MIHA-allogeneic tumor were studied. An intravenous injection of surprisingly small numbers (10(4)-10(5] of live allogenetic spleen cells (SC) effectively primed mice for SSR of the allogeneic tumor, and this immunity was developed as early as 2-3 days after injection of the SC. In contrast, sonication-disrupted allogeneic SC, which should be readily processed by host antigen presenting cells (APC), were not active as immunogens, even at a dose 1000 times higher than the minimum effective dose of live SC. The possibility that host APC preferentially receive MIHA antigens shed by live allogeneic SC for T cell activation was ruled out. These results demonstrated that antigen processing via conventional pathways is very little involved in the mechanism of T cell activation. Under such restricted experimental conditions, the induction phase but not the effector phase of the MIHA-specific T cell immunity was shown to be H-2-unrestricted.

Differences in the effect of indomethacin and preincubation on the lymphoproliferative response to concanavalin A of spleen cells from low responder C57BL/6 and high responder BALB/c mice

In vivo treatment with 100 μg of indomethacin each 48 h for 2 weeks enhanced the proliferative response to concanavalin A (Con A) of spleen cells from mice of the C57BL/6 (B6) strain, low responder to T cell mitogens, but did not modify the response of spleen cells from mice of the high responder strain BALB/c (C). The enhancing effect of in vivo indomethacin treatment was more marked in cultures of B6 splenocytes stimulated with high, moderately supraoptimal doses of Con A than in cultures stimulated with optimal mitogen doses. Addition of indomethacin to cultures of spleen cells from untreated donors induced greater increase of the lymphoproliferative response of cells from low responder B6 than from high responder C mice. The enhancing effect of indomethacin added in vitro was observed in cultures stimulated by optimal but not by supraoptimal doses of Con A. The addition of indomethacin did not enhance the response of B6 spleen lymphocytes depleted of adherent cells. Preincubation for 24 h prior to mitogen stimulation increased the response to high Con A doses of spleen cells from low responder B6 mice whereas this procedure did not enhance lymphocyte proliferation in cultures of spleen cells from high responder C mice. Supplementation with indomethacin in vitro combined with preincubation induced additive enhancing effects on DNA synthesis by B6 spleen lymphocytes, suggesting that each treatment acts through different mechanism(s). The results indicated that spleen cells from low responder B6 strain mice are more sensitive than cells from high responder C mice to the potentiating effect of indomethacin and preincubation on the proliferative response to Con A. These observations suggest that mechanisms sensitive to indomethacin and to preincubation contribute to the depression of mitogen induced DNA synthesis in low responder B6 mice.

T and B Lymphocytes in the Regulation of Delayed Hypersensitivity

A correlation was demonstrated between the transient nature of a) delayed intradermal responses of guinea pigs sensitized to hen egg albumin in incomplete Freund's adjuvant and b) the proliferative response of sensitized lymphocytes to the specific antigen. Spleen cells from sensitized animals suppressed the proliferative response of lymph node cells to specific antigen. This suppression was dependent on the dose of spleen cells and the time of their removal after sensitization. Thymus cells were suppressive to a lesser extent, and their activity was not correlated with the time of removal after sensitization. Separation of spleen and thymus cells into T and B populations indicated that the B cell was the major suppressor cell in the spleen, while the T cell in the thymus had a similar but less pronounced action.

Heterotransplantation of mouse tumors to immunologically paralyzed rats

A comparative study of the critical factors involved in the induction of tolerance to mouse tumors in rats treated after birth by intraperitoneal injections of mouse spleen, liver, and tumor cell suspensions has revealed that the induction and maintenance of tolerance is essentially conditioned by repeated inoculation of the antigen. Tolerance to mouse tumors was conferred on rats by spleen cells, provided the latter were administered in daily inoculations the first 10 days of life. The incidence rate of actively developing tumors and the duration of tolerance were directly related to the total dose of postnatally administered spleen cells. By the administration of a second course of large doses of spleen cells, tolerance could be maintained after the onset of recovery from paralysis. Similarly, tolerance could be reinduced by repeated inoculations of tumor cells in rats that had been sensitized by a mouse skin heterograft during the period of recovery from paralysis. Actively growing tumors developed in rats treated with nonviable liver cells during the first week of life and thereafter repeatedly reinoculated with tumor cells. Tolerance was induced by repeated inoculations of tumor cells administered during the first 3 to 5 days of life; a similar dose proved inefficient when administered in a single inoculation during the neonatal period. The theoretical implications of these results are discussed.

Induction of homograft tolerance in adult mice by sublethal x-irradiation and injection of homologous spleen cells.

Homograft tolerance was induced in adult DBA/2 mice by sublethal x irradiation and an injection of a suspension of adult BALB/c spleen cells. The optimal conditions for thls procedure were determined by varying the dose of x irradiation (200 to 400 r), the time of splenic injection (1 to 72 hours after irradiation), the time of skin grafting (1 to 17 days after irradiation), and the dose of spleen cells (0.01 to 1.0, expressed as a fraction of one spleen). The first 3 factors were found to be of maximal importance. The mortality attributable to the ertire procedure was 2%. The procedure failed to induce tolerance across an H-2 barrier. The possible use of this system as a model for studying tolerance induction and maintenance was emphasized.