Immunotoxic effects of cyclophosphamide and cyclosporine in the dog

Figures & data

Table 1.  Male (n = 3) and female (n = 3) Beagle dogs treated orally with water (controls), 25 mg/kg cyclosporine daily, or 2 mg/kg cyclophosphamide 4 days/week from Day 1 of a 4-week treatment period. WBC, neutrophil, and lymphocytes were counted by ADVIA120 and lymphocyte subset counts (B-cells, total T-cells, as well as CD4⁺ and CD8+ T-cells) were analyzed by flow cytometry, before treatment, and on Days 11, 18, and 28 of this 4-week treatment period.

	Controls	Cyclosporine	Cyclophosphamide
Males
Pre-Test
WBC	10.70 ± 1.44	11.99 ± 2.10	10.82 ± 0.40
Neutrophils	6.59 ± 0.76	8.16 ± 1.97	7.10 ± 0.95
Lymphocytes	3.17 ± 0.46	2.70 ± 0.84	2.88 ± 0.57
B-cells	0.49 ± 0.07	0.51 ± 0.26	0.50 ± 0.07
Total T-cells	1.78 ± 0.07	1.72 ± 0.50	1.81 ± 0.46
CD4^+ T-cells	1.25 ± 0.14	1.15 ± 0.28	1.17 ± 0.43
CD8^+ T-cells	0.40 ± 0.07	0.35 ± 0.11	0.41 ± 0.15
Day 11
WBC	10.07 ± 0.91	11.34 ± 1.18	5.41 ± 0.88
Neutrophils	5.79 ± 0.87	6.72 ± 1.36	2.53 ± 0.87
Lymphocytes	3.43 ± 0.44	3.72 ± 0.56	2.40 ± 0.12
B-cells	0.69 ± 0.08	0.65 ± 0.13	0.34 ± 0.09
Total T-cells	2.02 ± 0.38	2.64 ± 0.48	1.71 ± 0.05
CD4^+ T-cells	1.28 ± 0.24	1.76 ± 0.30	1.12 ± 0.16
CD8^+ T-cells	0.42 ± 0.06	0.51 ± 0.10	0.40 ± 0.15
Day 18
WBC	10.70 ± 0.86	10.98 ± 2.19	5.66 ± 0.80
Neutrophils	6.04 ± 0.70	6.78 ± 1.70	3.05 ± 0.23
Lymphocytes	3.72 ± 0.70	3.29 ± 0.87	2.21 ± 0.56
B-cells	0.69 ± 0.15	0.53 ± 0.14	0.24 ± 0.06*
Total T-cells	2.21 ± 0.54	2.34 ± 0.72	1.62 ± 0.42
CD4^+ T-cells	1.27 ± 0.46	1.66 ± 0.52	1.06 ± 0.36
CD8^+ T-cells	0.47 ± 0.14	0.40 ± 0.13	0.37 ± 0.08
Day 28
WBC	8.55 ± 1.57	12.12 ± 1.94	4.82 ± 1.06
Neutrophils	5.34 ± 0.91	8.28 ± 2.04	2.58 ± 0.45
Lymphocytes	2.52 ± 0.58	2.81 ± 0.48	1.81 ± 0.52
B-cells	0.38 ± 0.12	0.45 ± 0.17	0.23 ± 0.06
Total T-cells	1.51 ± 0.41	1.94 ± 0.27	1.21 ± 0.39
CD4^+ T-cells	1.01 ± 0.26	1.31 ± 0.18	0.82 ± 0.31
CD8^+ T-cells	0.35 ± 0.09	0.37 ± 0.05	0.31 ± 0.07
Females
Pre-Test
WBC	11.27 ± 1.58	9.10 ± 2.12	8.71 ± 1.04
Neutrophils	7.33 ± 1.71	5.62 ± 2.09	5.27 ± 0.80
Lymphocytes	2.91 ± 0.58	2.76 ± 0.03	2.62 ± 0.14
B-cells	0.52 ± 0.18	0.46 ± 0.09	0.37 ± 0.11
Total T-cells	1.91 ± 0.42	1.68 ± 0.09	1.72 ± 0.14
CD4^+ T-cells	1.28 ± 0.38	1.13 ± 0.09	1.20 ± 0.17
CD8^+ T-cells	0.39 ± 0.01	0.36 ± 0.09	0.32 ± 0.04
Day 11
WBC	10.69 ± 0.81	9.86 ± 2.44	6.73 ± 0.22
Neutrophils	6.08 ± 0.35	5.14 ± 2.00	3.46 ± 0.43
Lymphocytes	3.51 ± 0.84	3.99 ± 0.33	2.60 ± 0.29
B-cells	0.69 ± 0.20	0.63 ± 0.09	0.27 ± 0.15*
Total T-cells	2.19 ± 0.69	2.71 ± 0.33	1.90 ± 0.23
CD4^+ T-cells	1.43 ± 0.58	1.80 ± 0.25	1.28 ± 0.17
CD8^+ T-cells	0.49 ± 0.05	0.57 ± 0.07	0.38 ± 0.10
Day 18
WBC	10.99 ± 1.63	10.63 ± 1.25	6.13 ± 1.39
Neutrophils	5.87 ± 1.01	5.42 ± 1.19	3.65 ± 1.06*
Lymphocytes	3.95 ± 0.99	4.42 ± 0.04	2.07 ± 0.30
B-cells	0.73 ± 0.27	0.65 ± 0.03	0.17 ± 0.04*
Total T-cells	2.61 ± 0.75	2.91 ± 0.22	1.42 ± 0.27
CD4^+ T-cells	1.71 ± 0.52	1.95 ± 0.17	1.03 ± 0.19
CD8^+ T-cells	0.56 ± 0.10	0.61 ± 0.09	0.30 ± 0.04
Day 28
WBC	9.22 ± 1.43	9.57 ± 1.47	5.37 ± 0.58
Neutrophils	5.30 ± 0.99	5.93 ± 1.80	3.07 ± 0.70
Lymphocytes	3.10 ± 0.79	2.94 ± 0.46	1.75 ± 0.21
B-cells	0.51 ± 0.18	0.38 ± 0.12	0.15 ± 0.06*
Total T-cells	2.09 ± 0.58	1.98 ± 0.16	1.20 ± 0.25
CD4^+ T-cells	1.40 ± 0.51	1.35 ± 0.12	0.85 ± 0.16
CD8^+ T-cells	0.46 ± 0.04	0.41 ± 0.10	0.28 ± 0.07*

Results are presented as absolute cell numbers in Giga/L (± SD).

*p < 0.05 (Dunn test).

Table 2.  Anti-KLH IgM and IgG mean responses. Male (n = 3) and female (n = 3) Beagle dogs treated orally with water (controls), 25 mg/kg cyclosporine daily, or 2 mg/kg cyclophosphamide 4 days/week from Day 1 of a 4-week treatment period, were injected intramuscularly with 5 mg KLH on Day +11. Serum anti-KLH IgM and IgG were measured using a sandwich ELISA before treatment and subsequently on Days +18 and +23 (to measure IgM anti-KLH antibody response) and on Days +23 and +28 (to measure IgG anti-KLH antibody response).

	Controls	Cyclosporine	Cyclophosphamide
IgM
Males
Pre-test	blq^a	blq^b	blq
Day 18	609 ± 458	381 ± 184	804 ± 645
Day 23	442 ± 322	342 ± 196	572 ± 461
Females
Pre-test	blq	blq	blq
Day 18	666 ± 208	213 ± 97*	726 ± 227
Day 23	484 ± 211	141 ± 73*	462 ± 178
IgG
Males
Pre-test	blq	blq	blq
Day 23	677 ± 474	220 ± 123	196 ± 145
Day 28	971 ± 493	196 ± 145	1565 ± 837
Females
Pre-test	blq	blq	blq
Day 23	1112 ± 482	72 ± 37*	1059 ± 228
Day 28	1490 ± 742	52 ± 27*	1378 ± 312

Results are presented as µg/ml, Lower Limit Of Quantification = 20 µg/mL; blq, below limit of quantification.

^aOne male out of three had 21.39 µg/ml. ^bOne male out of three had 42.61 µg/ml.

*p < 0.05.

Figure 1.  Anti-KLH IgM and IgG individual responses. Male (n = 3) and female (n = 3) Beagle dogs treated orally with water (controls), 25 mg/kg cyclosporine daily, or 2 mg/kg cyclophosphamide 4 days/week from Day 1 of a 4-week treatment period, were injected intramuscularly with 5 mg KLH on Day +11. Serum anti-KLH IgM and IgG were measured using a sandwich ELISA before treatment and subsequently on Days +18 and +23 (to measure IgM anti-KLH antibody response) and on Days +23 and +28 (to measure IgG anti-KLH antibody response).