Effects of parathyroid hormone-related protein and macrophage inflammatory protein-1α in Jurkat T-cells on tumor formation in vivo and expression of apoptosis regulatory genes in vitro - PubMed (original) (raw)

Sherry T Shu et al. Leuk Lymphoma. 2012 Apr.

Abstract

Parathyroid hormone-related protein (PTHrP) and macrophage inflammatory protein-1α (MIP-1α) have been implicated in the pathogenesis of adult T-cell leukemia/lymphoma, but their effects on T-cells have not been well studied. Here we analyzed the functions of PTHrP and MIP-1α on T-cell growth and death both in vitro and in vivo by overexpressing either factor in human Jurkat T-cells. PTHrP or MIP-1α did not affect Jurkat cell growth in vitro, but PTHrP increased their sensitivity to apoptosis. Importantly, PTHrP and MIP-1α decreased both tumor incidence and growth in vivo. To investigate possible mechanisms, polymerase chain reaction (PCR) arrays and real-time reverse transcription (RT)-PCR assays were performed. Both PTHrP and MIP-1α increased the expression of several factors including signal transducer and activator of transcription 4, tumor necrosis factor α, receptor activator of nuclear factor κB ligand and death-associated protein kinase 1, and decreased the expression of inhibitor of DNA binding 1, interferon γ and CD40 ligand in Jurkat cells. In addition, MIP-1α also increased the expression of transcription factor AP-2α and PTHrP increased expression of the vitamin D3 receptor. These data demonstrate that PTHrP and MIP-1α exert a profound antitumor effect presumably by increasing the sensitivity to apoptotic signals through modulation of transcription and apoptosis factors in T-cells.

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Conflict of interest statement

Potential conflict of interest: Disclosure forms provided by the authors are available with the full text of this article at www.informahealthcare.com/lal.

Figures

Figure 1

Figure 1

Overexpression of PTHrP and MIP-1α in Jurkat cells. Real-time RT-PCR of (A) PTHrP and (B) MIP-1α in Jurkat T-cells. Secretion of (C) PTHrP and (D) MIP-1α in Jurkat T-cells.

Figure 2

Figure 2

PTHrP and MIP-1α did not affect Jurkat cell growth. (A) Cell proliferation ratios of Jurkat-pcDNA-luc, Jurkat-PTHrP-luc and Jurkat-MIP-1α-luc cells and (B) cell viability ratios of Jurkat-pcDNA-luc, Jurkat-PTHrP-luc and Jurkat-MIP-1α-luc cells in 4-day cultures. Rates were normalized to the data from day 1.

Figure 3

Figure 3

PTHrP increased the sensitivity to an apoptotic agent (camptothecin) in Jurkat cells. Jurkat-pcDNA-luc and Jurkat-PTHrP-luc were treated with vehicle (black column), 2 µM of camptothecin (open column) or 10 µM of camptothecin (gray column) overnight and the percentages of apoptotic cells were detected by TUNEL assay using flow cytometry. Symbol * indicates p < 0.01 when compared to Jurkat-pcDNA-luc cells treated with 2 µM of camptothecin. Symbol ** indicates p < 0.001 when compared to Jurkat-pcDNA-luc cells treated with 10 µM of camptothecin.

Figure 4

Figure 4

PTHrP and MIP-1α decreased viable Jurkat tumor growth in vivo. (A) Tumor burden measured by bioluminescent imaging. Photon signals obtained at week 7 were normalized to day 1. There was a 16- and 66-fold decrease in viable Jurkat-PTHrP-luc and Jurkat-MIP-1α-luc cells, respectively. Symbol * indicates p < 0.05 when compared to Jurkat-pcDNA-luc group. (B) H&E images of Jurkat tumors at week 7. PTHrP and MIP-1α decreased the incidence and size of Jurkat tumors in vivo. Tumors were identified at postmortem in 10/10 Jurkat-pcDNA-luc mice, 10/10 Jurkat-PTHrP-luc mice and 3/10 Jurkat-MIP-1α -luc mice. Cross-sectional areas of tumors were significantly decreased (p < 0.05) seven- and 18-fold, respectively, in mice that received Jurkat-PTHrP-luc and Jurkat-MIP-1α-luc cells when compared to mice that received Jurkat-pcDNA-luc cells. Note that the pale regions of the tumors represent areas of necrosis. Bar = 10 mm.

Figure 5

Figure 5

PTHrP and MIP-1α regulated the expression of apoptosis and other factors in Jurkat cells in vitro. Relative gene expression of (A) CD40LG, (B) DAPK1, (C) TNFα, (D) IFNγ, (E) RANKL and (F) VDR in Jurkat-pcDNA-luc, Jurkat-PTHrP-luc and Jurkat-MIP-1α-luc cells was measured by real-time RT-PCR. Gene expression in Jurkat-pcDNA-luc cells was set as a basal level for comparison. Symbol * indicates p < 0.05 when compared to Jurkat-pcDNA-luc group.

Figure 6

Figure 6

PTHrP and MIP-1α regulated the expression of transcription factors in Jurkat cells in vitro. Relative gene expression of (A) ID1, (B) PAX6, (C) STAT4 and (D) TFAP2A in Jurkat-pcDNA-luc, Jurkat-PTHrP-luc and Jurkat-MIP-1α-luc cells was measured by real-time RT-PCR. Gene expression in Jurkat-pcDNA-luc cells was set as a basal level for comparison. Symbol * indicates p < 0.01 when compared to Jurkat-pcDNA-luc group.

Figure 7

Figure 7

HTLV-1-infected T-cells expressed the MIP-1α receptor CCR1. CCR1 expression of (1) Jurkat; (2) MT2; (3) SLB-1; (4) Hut102; (5) MET-1; (6) C8166; (7) RV-ATL; (8) HT-1RV; (9) positive control (PBMC-1); (10) positive control (PBMC-2); (11) negative control (water) was measured by RT-PCR.

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