Distinct role of antigen-specific T helper type 1 (Th1) and Th2 cells in tumor eradication in vivo - PubMed (original) (raw)

Distinct role of antigen-specific T helper type 1 (Th1) and Th2 cells in tumor eradication in vivo

T Nishimura et al. J Exp Med. 1999.

Abstract

The role of T helper type 1 (Th1) and Th2 cells in tumor immunity was investigated using Th cells induced from ovalbumin (OVA)-specific T cell receptor transgenic mice. Although Th1 cells exhibited stronger cytotoxicity than Th2 cells, both cell types completely eradicated tumors when transferred into mice bearing A20 tumor cells transfected with the OVA gene (A20-OVA). Th1 cells eradicated the tumor mass by inducing cellular immunity, whereas Th2 cells destroyed the tumor by inducing tumor necrosis. Both Th1 and Th2 cells required CD8(+) T cells to eliminate tumors, and neither of these cells were able to completely eliminate A20-OVA tumors from T and B cell-deficient RAG2(-/-) mice. Mice cured from tumors by Th1 and Th2 cell therapy rejected A20-OVA upon rechallenge, but CD8(+) cytotoxic T lymphocytes were induced only from spleen cells prepared from cured mice by Th1 cell therapy. Moreover, we demonstrated that Th1 and Th2 cells used distinct adhesion mechanisms during tumor eradication: the leukocyte function-associated antigen (LFA)-1-dependent cell-cell adhesion step was essential for Th1 cell therapy, but not for Th2 cell therapy. These findings demonstrated for the first time the distinct role of antigen-specific Th1 and Th2 cells during eradication of established tumors in vivo.

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Figures

Figure 1

Figure 1

Characteristics of Th1 and Th2 cells from DO11.10 TCR-Tg mice and their activation by A20-OVA tumor cells. Th1 and Th2 cells were induced from CD4+CD45RB+ naive Th cells obtained from DO11.10 TCR-Tg mouse spleen cells under the conditions described in Materials and Methods. The expression of CD4/CD8 antigen (A and C) and cytoplasmic cytokine (IL-4/IFN-γ) expression (B and D) in Th1 (A and B) and Th2 cells (C and D) was determined by flow cytometric analysis. The cytokine-producing ability of Th1 (hatched bars) and Th2 cells (black bars) was determined by measuring the amounts of IFN-γ (E) or IL-4 (F) in culture supernatants after coculture with A20 tumor cells, A20 tumor cells pulsed with OVA-pep (A20-pep), or A20-OVA tumor cells, for 12 h. ND, not detected. The cytotoxic activity of Th1 (G) and Th2 cells (H) against A20 tumor cells (▴), A20 tumor cells pulsed with OVA-pep (A20-pep) (•), or A20-OVA tumor cells (○) was measured by 4-h 51Cr-release assays. The bars represent mean ± SE of triplicate samples.

Figure 3

Figure 3

The distinct role of Th1 and Th2 cells for eradication of established tumors in vivo. (A) OVA-specific Th1 cells (•), OVA-specific Th2 cells (○), or saline (▴) were intravenously injected into BALB/c mice bearing A20-OVA tumors. The antitumor activity of Th1 and Th2 cells was determined by measuring changes over time of the means of two perpendicular diameters of the tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d. (B) Typical tumor growth or regression pattern in saline- (a), Th1- (b), or Th2-treated mice (c). Similar results were obtained in >10 independent experiments. The arrow indicates the site of tumor inoculation in Th1-treated mice.

Figure 3

Figure 3

The distinct role of Th1 and Th2 cells for eradication of established tumors in vivo. (A) OVA-specific Th1 cells (•), OVA-specific Th2 cells (○), or saline (▴) were intravenously injected into BALB/c mice bearing A20-OVA tumors. The antitumor activity of Th1 and Th2 cells was determined by measuring changes over time of the means of two perpendicular diameters of the tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d. (B) Typical tumor growth or regression pattern in saline- (a), Th1- (b), or Th2-treated mice (c). Similar results were obtained in >10 independent experiments. The arrow indicates the site of tumor inoculation in Th1-treated mice.

Figure 2

Figure 2

Establishment of an adoptive tumor immunotherapy model using antigen-specific Th cells. (A) The protocol for adoptive tumor immunotherapy using OVA-specific Th1 or Th2 cells. A20-OVA tumor cells were intradermally inoculated into wild-type BALB/c mice. When the tumor formed a mass of 6–8 mm in diameter, 2 × 107 OVA-specific Th1 or Th2 cells, induced from DO11.10 TCR-Tg mice, were transferred into the tumor-bearing mice. The therapeutic effect of Th1 or Th2 cell transfer was determined by measuring tumor size. According to the above protocol (A), OVA-specific Th1 cells (OVA-Th1, •), antigen-nonspecific 2C11-activated Th1 cells (2C11-Th1, ▴), or saline (None, ○) were intravenously injected into A20-OVA–bearing BALB/c mice (B) or parental A20-bearing BALB/c mice (C). The antitumor activity of Th1 cells was determined by measuring changes over time of the means of two perpendicular diameters of tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d.

Figure 5

Figure 5

Tumor eradication mechanisms mediated by Th1 and Th2 cells. Th1 cell therapy was carried out using wild-type BALB/c mice and DO11.10 TCR-Tg mice according to the protocol described in the legend to Fig. 2 A. (A) At days −1 and 0 before Th1 cell transfer, either saline (○), anti-CD4 mAb (▵), anti-CD8 mAb (▴), anti–IFN-γ mAb (▪), or rat IgG (•) was intravenously injected into tumor-bearing BALB/c mice. (B) A20-OVA tumor cells were intradermally inoculated into wild-type BALB/c mice (○, •) or BALB/c background TCR-Tg mice (▵, ▴). When the tumor mass became palpable, Th1 cells (•, ▴) were transferred into the tumor bearing mice. As a control, mice were treated with saline (○, ▵). The antitumor activity of Th1 cells was determined by measuring changes over time of the means of two perpendicular diameters of the tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d.

Figure 4

Figure 4

Histochemical analysis of the cellular mechanisms underlying Th1- and Th2-mediated tumor eradication in vivo. Tumor tissues were removed from tumor-bearing mice 7 d after treatment with saline (A), Th1-cell transfer (B), or Th2-cell transfer (C). The tissues were stained with hematoxylin and eosin.

Figure 8

Figure 8

Th1 and Th2 cells use distinct cell adhesion interactions during eradication of tumors in vivo. (A) Expression of LFA-1 antigen on Th1 and Th2 cells. (B) Expression of ICAM-1 on Th1 and Th2 cells. Dotted line, unstained control cells. (C and D) LFA-1/ICAM-1–dependent homotypic cell–cell aggregation. Th1 (C) and Th2 cells (D) (2 × 106 cells) were stimulated with 20 ng/ml of PMA. After culture for 1 h, homotypic aggregates were formed. Typical cell–cell aggregates were photographed. We confirmed that the homotypic aggregation was strongly blocked by addition of either anti–LFA-1 mAb or anti–ICAM-1 mAb (data not shown). (E and F) The effect of anti–LFA-1 mAb administration in vivo on the therapeutic ability of Th1 (E) or Th2 cells (F) was examined using the protocol described in the legend to Fig. 2 A. (E) When the A20-OVA tumor mass became palpable, the mice were treated with saline (○), Th1 cell transfer (•), or Th1 cell transfer after anti–LFA-1 mAb administration (▵). (F) When the A20-OVA tumor mass became palpable, the mice were treated with saline (○), Th2 cell transfer (•), or Th2 cell transfer after anti–LFA-1 mAb administration (▵). The anti–LFA-1 mAb (500 μg/mouse) was intravenously injected into the mice at days −1 and 0 before cell transfer. The antitumor activity of Th1 and Th2 cells was determined by measuring changes over time of the means of two perpendicular diameters of the tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d.

Figure 6

Figure 6

Requirement for CD8+ T cells in inducing complete elimination of transplanted tumors in response to Th1 or Th2 cell therapy. The tumor therapy model was carried out according to the legend to Fig. 2 A except that aliquots (2 × 105/mouse) of CD8+ T cells from immunized mice (i-CD8, A) or unprimed mice (ni-CD8, B) were transferred into the mice 7 d before the inoculation of A20-OVA tumor cells. CD8+ T cells were isolated by cell sorting from A20-OVA–immunized BALB/c mice or untreated BALB/c mice. (A) When the tumor mass became palpable, the mice were treated with intravenous injection of saline (A20-OVA, •), Th1 cells (▴), or Th2 cells (▪). The tumor-bearing mice transferred with CD8+ T cells from tumor-immunized mice (i-CD8) were also treated with intravenous injection of saline (○), Th1 cells (▵), or Th2 cells (□). (B) When the tumor mass became palpable, the mice were treated with intravenous injection of saline (A20-OVA, •), Th1 cells (▴), or Th2 cells (▪). The tumor-bearing mice transferred with CD8+ T cells from unprimed mice (ni-CD8) were also treated with intravenous injection of saline (○), Th1 cells (▵), or Th2 cells (□). The antitumor activity of Th1 and Th2 cells was determined by measuring changes over time of the means of two perpendicular diameters of the tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d.

Figure 7

Figure 7

Adoptive transfer of Th1 cells but not Th2 cells induces cellular immunological memory. (A) Mice cured from A20-OVA tumors after Th1 (•) or Th2 cell therapy (○) were rechallenged by intradermal injection of 2 × 106 viable A20-OVA tumor cells. Untreated mice (▴) were used as controls. Tumor growth was determined by measuring changes over time of the means of two perpendicular diameters of the tumor mass. Results are presented as mean ± SE of six mice. The tumor-free mice were followed for >90 d. (B) Mice cured from A20-OVA tumors after Th1 or Th2 cell therapy were killed, and their spleen cells were restimulated in vitro with mitomycin C–treated A20-OVA by mixed lymphocyte tumor reaction (MLTR). As control, untreated BALB/c mice (None) were used for MLTR. After culture for 2 d, the levels of IFN-γ (black bars) and IL-4 (hatched bars) in culture supernatants were measured by ELISA. The bars represent mean ± SE of three mice. (C) The cells were harvested from MLTR cultures from mice that had received Th1 cells (•, ○), Th2 cells (▴, ▵), or no cells (None, ▪, □), and their cytotoxicity was measured using 51Cr-labeled A20-OVA (•, ▴, ▪) or A20 tumor cells (○, ▵, □).The results are presented as mean ± SE of three mice.

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