Phase-partitioning with Triton X-114 (TX114) was applied to the TSP antigen, which may be preferentially associated with the cell wall of M. tuberculosis. The hydrophilic protein components of the TSP antigen were successfully separated from integral hydrophobic macromolecules. To further characterize and examine the cellular immune response of the aqueous fraction of the TSP antigen (TSPa), the in vitro properties of the antigen were measured by lymphoproliferation; surface expression of IL-2 Ra on T lymphocytes was analyzed by flow cytometry; and the cytokine mRNA expression pattern was determined by RT-PCR. Significant lymphoproliferative responses to the TSPa antigen were observed in healthy tuberculin reactive donors after a 5 day in vitro stimulation. TSPa treatment of PBMCs from healthy tuberculin positive subjects for 5 days resulted in progressive augmentation of IFN-r, II 2, and IL-2Ra mRNA expression, as measured by RT-PCR, but considerably reduced IL-4 mRNA expression. In addition, the TSPa antigen stimulated more IL-12 p40 mRNA production than did the PPD antigen, and graduaBy suppressed IL- 10 mRNA expression. Moreover, the CD3' T cells of tuberculin positive subjects displayed a profound increase in their expression of the II 2Ru protein (39.0%) in response to the TSPa antigen. Proliferation was correlated with IL-2 and IL-2Ra mRNAs, but not correlated with distinct IFN-r or IL-12 p40 mRNA production. These findings strongly suggest that the TSPa antigen preferentially evokes the generation of a Thl-like immune response in healthy tuberculin reactors.
Cell Wall ; Flow Cytometry ; Humans* ; Immunity, Cellular* ; Interleukin-12 ; Interleukin-2 ; Interleukin-4 ; Mycobacterium tuberculosis* ; Mycobacterium* ; Neptune ; RNA, Messenger ; T-Lymphocytes ; Tissue Donors ; Tuberculin ; Tuberculosis

Tumor-draining lymph node mononuclear (TDLMN) cells are specifically sensitized to the growing tumor but such cells are deficient for mediating an antitumor response. In this study, we examined the feasibility of using mycobacterial 30 kDa or Triton X-100 solubilized protein (TSP) antigen to stimulate mononuclear cells of colon cancer-draining lymph node for the generation of cell mediated immune effector cells. The proliferative response of TDLMN cells stimulated with mycobacterial 30 kDa or TSP antigen was determined by H-thymidine incorporation assay. The proliferation of TDLMN cells to mycobacterial 30 kDa or TSP antigen was significantly increased in PPD (+) patients, but a poor response to the 30 kDa or TSP antigen was observed in PPD (-). The expression on ro T cells to mycobacterial 30 kDa or TSP antigen was assessed by flow cytometry. The ro T cells from PPD (+) patient responded only to 30 kDa antigen but to TSP antigen. An investigation of cytokine mRNA expression was undertaken using reverse transcription- polymerase chain reaction (RT-PCR) to follow TDLMN cells stimulated with the 30 kDa or TSP antigens for 5 days. The IFN-r and TNF-a mRNA expression was only induced in TDLMN cells of PPD (+) patient in response to the 30 kDa or TSP antigen. The IL-2 mRNA expression was induced in both PPD (+) and PPD (-) in response to the 30 kDa or TSP antigen. But the IL-4 mRNA expression was not induced in response to the 30 kDa or TSP antigen. These results suggest that the 30 kDa and TSP antigens may serve as biologic response modifier for the generation of cell mediated immune effector cells.
Colon* ; Flow Cytometry ; Humans ; Interleukin-2 ; Interleukin-4 ; Lymph Nodes* ; Mycobacterium tuberculosis* ; Mycobacterium* ; Negotiating ; Neptune* ; Octoxynol* ; Polymerase Chain Reaction ; RNA, Messenger ; T-Lymphocytes

No abstract available.
Introns* ; Point Mutation* ; Protein-Tyrosine Kinases* ; Tyrosine*

Present study was aimed to investigate the immunological activities of the 47 kDa glycoprotein antigen from Treponema pallidum and conducted on 24 patients with syphilis, (early, late, spontaneously healed, congenital and treated patients) and on 17 normal healthy controls. Two opposite lymphoproliferative manifestations to the 47 kDa antigen were observed in syphilis patients by H-thymidine incorporation assay. Ten responders (stimulation index [Sl] >4) showed a 3-fold-higher proliferation than the nonresponders, and four of those responders were spontaneously healed patients. Furthermore, analysis by flow cytometry indicated a preferential expansion of CD4' T lymphocytes by the 47 kDa antigen in the spontaneously healed syphilis patients. Stimulation of PBMCs of spontaneously healed syphilis patients with the 47 kDa antigen for greater than 72 hrs resulted in piogressive augmentation of IFN-r, IL-2Ra and IL-2 mRNA measured by RT-PCR, but considerably reduced IL-4 and IL-10 mRNA expression. However, patients with late latent syphilis exhibited more increased IL-4 and IL-10 mRNA expressions in response to the 47 kDa antigen than spontaneously healed syphilis group. In contrast to other groups, when cultured with the 47 kDa antigen very low IFN-#y, IL-2Ra and IL-2 mRNA expressions were shown in early syphilis group. These data suggest that the Th1-predominant cellular responses induced by the 47 kDa antigen may be involved in the clinical outcome of syphilis and provide the immunologic basis for further functional studies regarding the role of the 47 kDa in the immunopathogenesis of syphilis.