Propanil exposure induces delayed but sustained abrogation of cell-mediated immunity through direct interference with cytotoxic T-lymphocyte effectors

Abstract

The postemergent herbicide propanil (PRN ; also known as 3,4-dichloropropionanilide) is used on rice and wheat crops and has well-known immunotoxic effects on various compartments of the immune system, including T-helper lymphocytes, B lymphocytes, and macrophages. It is unclear, however, whether PRN also adversely affects cytotoxic T lymphocytes (CTLs) , the primary (1 degrees ) effectors of cell-mediated immunity. In this study we examined both the direct and indirect effects of PRN exposure on CTL activation and effector cell function to gauge its likely impact on cell-mediated immunity. Initial experiments addressed whether PRN alters the class I major histocompatibility complex (MHC) pathway for antigen processing and presentation by antigen-presenting cells (APCs) , thereby indirectly affecting effector function. These experiments demonstrated that PRN does not impair the activation of CTLs by PRN-treated APCs. Subsequent experiments addressed whether PRN treatment of CTLs directly inhibits their activation and revealed that 1 degrees alloreactive CTLs exposed to PRN are unimpaired in their proliferative response and only marginally inhibited in their lytic activity. Surprisingly, secondary stimulation of these alloreactive CTL effectors, however, even in the absence of further PRN exposure, resulted in complete abrogation of CTL lytic function and a delayed but significant long-term effect on CTL responsiveness. These findings may have important implications for the diagnosis and clinical management of anomalies of cell-mediated immunity resulting from environmental exposure to various herbicides and other pesticides.

Figure 1

CTL clone 33 lytic response in a 4-hr ⁵¹Cr-release assay against N1 targets: EtOH treated, exposed to PRN for 18 hr, or exposed to PRN for 2 hr. The x-axis indicates the ratio of effector T cells per target cell added to each well in the ⁵¹Cr-release assay; ratios range from 30:1 to 0.1:1.

Figure 2

Clone 33 stimulated with EtOH-treated N1 (A) or PRN-treated N1 (B) stimulators, tested against N1 targets: EtOH treated, exposed to PRN for 18 hr, or exposed to PRN for 2 hr. Results shown are representative of three experiments; all sample points were run in triplicate. Effector:target ratios range from 30:1 to 0.1:1.

Figure 3

Peptide titration of clone 33 lytic response to VSV-N p52–59 tested on EL4 targets: EtOH treated, exposed to PRN for 18 hr, or exposed to PRN for 2 hr. A constant 3:1 effector:target ratio was used in this experiment. The x-axis indicates the peptide concentrations are titrated from 32 μM to 5 pM. Results shown are representative of three experiments; all sample points were run in triplicate.

Figure 4

Primary B6 anti-BALB/c MLC-derived alloreactive effector CTLs tested against P815 (H-2^d) target cells. MLC-derived effectors are from cultures with the following added on day 0: EtOH control, 16 μM PRN, 33 μM PRN, or 66 μM PRN. (A) Proliferation of MLC-derived T lymphocytes. The column designated “0” indicates that no CTLs were added to this group. (B) ⁵¹Cr-release assay results depicted as percent specific lysis by the following treatment groups: 0 PRN, 16 μM PRN, 33 μM PRN, or 66 μM). Results shown are representative of three experiments; all sample points were run in triplicate and represent mean ±SE.

Figure 5

Secondary B6 anti-BALB/c MLC-derived effectors are tested for ³H-TdR uptake to measure the proliferative response (A) and ⁵¹Cr release as a measure of CTL-mediated lytic activity (B). Treatment groups in A are as follows: Un/Un, 1° EtOH-treated/2° EtOH-treated; Un/66, 1° EtOH-treated/2° 66 μM; Un/165, 1° EtOH-treated/2° 165 μM PRN; 66/Un, 1° 66 μM PRN/2° EtOH-treated; 66/66, 1° 66 μM PRN/2° 66 μM PRN; 165/Un, 1° 165 PRN/2° EtOH-treated. (B) ⁵¹Cr-release as a measure of CTL-mediated lytic activity. Treatment groups in B are as follows: 0 PRN; 2° 66 μM PRN; 2° 165 μM PRN. Results shown represent three experiments; all sample points were run in triplicate and represent mean ± SE.