Effects of Sensitization to Dinitrochlorobenzene (DNCB) on Clinical Pathology Parameters and Mitogen-Mediated Blastogenesis in Cynomolgus Monkeys (Macaca fascicularis)

Abstract

Immunotoxicologic testing of drug candidates and environmental contaminants is of growing importance. Cutaneous delayed type hypersensitivity (DTH) is a convenient way of testing immune function in vivo. However, DTH testing must not interfere with interpretation of other relevant parameters. We have evaluated the effects of sensitization and challenge with dinitrochlorobenzene (DNCB) on clinical parameters routinely evaluated in toxicity testing and on lectin-mediated blastogenesis. Female cynomolgus monkeys were sensitized to DNCB with 4 daily applications of DNCB in acetone to the skin of the axilla. Fifteen days later, the monkeys were challenged for DTH by applying DNCB to the antecubital skin. Skin fold thickness was measured and the macroscopic appearance of the challenge site was scored 24 and 48 hr after challenge. All 5 monkeys were successfully sensitized to DNCB. There was a significant increase in the mean skin fold thickness (compared to pre-challenge thickness) of 2 mm at 24 hr and 1 mm at 48 hr (p < 0.001). The clinical score of the challenge site was also increased. Histologic examination of the sensitization and challenge sites from a second group of monkeys exposed to DNCB in an identical manner showed the perivascular inflammatory cell infiltrate typical of DTH. Evaluation of hematologic parameters at days 7, 14, and 21 revealed no change in the erythron at any interval and a mild decrease in total WBC, neutrophil and lymphocyte counts on day 7 in 4/5 monkeys. The WBC parameters remained within the normal range and returned to pre-sensitization values at the later intervals. Clinical biochemical parameters related to liver and kidney function were evaluated at the same intervals and no changes were found. The blastogenic response of peripheral blood leukocytes to concanavalin A, phytohemagglutinin and pokeweed mitogen was mildly but not significantly decreased at day 7 but not on day 14 or 21. This study has shown that sensitization to DNCB in cynomolgus monkeys is a reliable means of evoking a DTH response which is quantifiable. It has also shown that sensitization and challenge with DNCB can be incorporated into toxicity studies without confounding interpretation of other toxicity data.

Keywords

Lectin mediated blastogenesis delayed type JryDersensitivity contact hypersensitivity im-munotoxicity non-human primate

References

Bleumink E

Nater JP

Schraffordt-Koops, H

, and

The TH

(1974). A standard method for DNCB sensitization testing in patients with neoplasms. Cancer33:911–915.

Botham PA

Rattray NJ

Woodcock DR

Walsh ST

, and

Hext PM

(1989). The induction of respiratory allergy in guinea-pigs following intradermal injection of trimellitic anhydride: A comparison with the response to 2,4-dinitrochlorobenzene. Toxicol. Lett.47(1): 25–39.

Bronaugh RL

and

Maibach HI

(1985). Percutaneous absorption of nitroaromatic compounds, in vivo and in vitro studies in human and monkey. J. Invest. Dermatol.84:180–183.

Byers NE

and

Allison AC

(1987). Adjuvant formulation for use in vaccines to elicit both cell mediated and humoral immunity. Vaccine5:223–228.

Chalifoux LV

Schlossman SF

, and

Letvin NK

(1984). Delineation of lymphocyte subsets in lymph nodes of nonhuman primates. Clin. Immunol. Immunopathol.31:96–101.

Cohen DJ

Loertscher R

Rubin MF

Tilney NL

Carpenter CB

, and

Strom TB

(1984). Cyclosporine: A new immunosuppressive agent for organ transplantation. Ann. Intern. Med.101:667–682.

David JR

Goetzl EJ

, and

Austen KF

(1981). Immunology. In: Pathophysiology, The Biological Principles of Disease,

LH Smith

and

SO Thier

(eds). W.B. Saunders Company, Philadelphia, pp. 165–267.

Ferraresi RW

Byers NE

, and

Shott LD

(1981). Induction of contact sensitivity to dinitrochlorobenzene in the cynomolgus monkey. J. Med. Primatol.10:26–32.

Fuller TC

Trevithick JE

Fuller AA

Colvin RB

Cosimi AB

, and

Kung PC

(1984). Antigenic polymorphism of the T4 differentiation antigen expressed on human T helper/inducer lymphocytes. Hum. Immunol.9:89–102.

10.

Harves JS

Felsburg PJ

Heberling RL

Kniker WT

, and

Kalter SS

(1974). Immunological competence in non-human primates: Differences observed in four species. Clin. Exp. Immunol.16:267–278.

11.

Humber DP

, and

Hetherington CM

(1981). A micro-method for the evaluation of PHA-responsive T cells in Callithrix jacchus and Macaca fascicularis. J. Med. Primatol.10: 170–174.

12.

Jonker M

Neuhaus P

Zurcer C

Fucello A

, and

Goldstein G

(1985). OKT4 and OKT4A antibody treatment as immunosuppression for kidney transplantation in rhesus monkeys. Transplantation39(3): 247–253.

13.

Letvin NL

King NW

Reinherz EL

Hunt RD

Lane H

, and

Schlossman SF

(1983). T-lymphocyte surface antigens in primates. Eur. J. Immunol.13:345–347.

14.

Maddison SE

Hicklin MD

Conway BP

, and

Kagan IG

(1972). Transfer factor: Delayed hypersensitivity to Schistosoma mansoni and tuberculin in Macaca mulatta. Science178: 757–759.

15.

Martin LN

Gormus BJ

, and

Bozelka BE

(1983). Functional analysis of monkey lymphocytes subjects defined by OKT4 and OKT8 monoclonal antibodies. Cell Immunol.77:338–347.

16.

McDevitt HO

Perry R

, and

Steinman LA

(1987). Monoclonal anti-la antibody therapy in animal models of autoimmune disease. Ciba Found. Symp.129:184–193.

17.

Muller AE

and

Levis WR

(1973). Lymphocyte transformation during dinitrochlorobenzene contact sensitization. In vitro and in vivo evaluation of the primary immune response in man. J. Clin. Invest.52:1925–1930.

18.

Murayama Y

Fukao K

Noguchi A

, and

Takenaka O

(1986). Epitope expression on primate lymphocyte surface antigens. J. Med. Primatol.15:215–226.

19.

Nakagawa S

Amos HE

Gotoh M

, and

Tanioku K

(1979). The distribution of 2,4-dinitrophenyl groups in lymphoid tissue of guinea pigs following skin painting with 2,4-dinitrochlorobenzene. Immunology36:851–857.

20.

Nakagawa S

Takei Y

Bang D

, and

Ueki H

(1987). Delayed-type skin reaction to 2,4-dinotrophenylated epidermal cells in guinea pigs with contact sensitivity to 2,4-dinitrochlorobenzene. Arch. Dermatol. Res.279 (4): 236–240.

21.

Roth JA

Morton DL

, and

Holmes EC

(1978). Rejection of dinitrochlorobenzene-conjugated syngeneic tumor cells by dinitrochlorobenzene-sensitized guinea pigs. J. Surg. Res.25:1–7.

22.

Tatsumi M

Kohase M

, and

S Yamazaki

(1983). Inhibitory effect of human interferons on lymphocyte proliferation induced by phytomitogens in cynomolgus monkeys (Macaca fascicularis). Int. Arch. Allergy Appl. Immunol.72:316–324.

23.

Unanue ER

(1980). Cooperation between mononuclear phagocytes and lymphocytes in immunity. N. Eng. J. Med.303:977–985.

24.

Whittum-Hudson, JA

Taylor HR

Farazdaghi M

, and

Prendergast RA

(1986). Immunohistochemical study of the local inflammatory response to chlamydial ocular infection. Invest. Ophthalmol. Vis. Sci.27:64–69.

25.

Zapata-Sirvent RL

and

Hansbrough JF

(1985). Post-burn immunosuppression in an animal model. III. Maintenance of normal splenic helper and suppressor lymphocyte subpopulations by immunomodulating drugs. Surgery97:721–727.