Potential for amelioration of aflatoxin B1-induced immunotoxic effects in progeny of white leghorn breeder hens co-exposed to vitamin E

Figures & data

Table 1. Anti-SRBC antibody titer in chicks obtained from layer breeders fed AFB₁ ± Vit E-containing feeds for 1 week.

Groups (AF mg/kg  + Vitamin E mg/kg	At day 7 post-primary injection		At day 14 post-primary injection		At day 7 post-booster injection		At day 14 post-booster injection
feed)	Total Ab	IgG	Total Ab	IgG	Total Ab	IgG	Total Ab	IgG
A (0 + 0)	5.00 ± 1.00^a	1.00 ± 0.70^abc	3.00 ± 1.00^abc	1.67 ± 0.58^bc	8.33 ± 0.58^a	5.67 ± 1.15^a	6.67 ± 0.58^a	5.33 ± 0.58^a
B (0 + 0.1)	5.33 ± 0.58^a	1.00 ± 1.00^abc	4.00 ± 0.10^a	2.67 ± 0.58^a	5.67 ± 2.52^abc	4.00 ± 2.00^ab	6.00 ± 1.00^ab	5.67 ± 0.58^a
C (0.1 + 0)	4.67 ± 0.58^ab	2.00 ± 1.00^a	3.00 ± 0.10^abc	2.00 ± 0.10^ab	8.67 ± 0.58^a	4.67 ± 1.53^ab	6.33 ± 0.58^ab	4.33 ± 0.58^ab
D (0.5 + 0)	4.33 ± 2.08^abc	1.66 ± 0.58^ab	3.33 ± 1.15^abc	1.67 ± 0.58^bc	5.67 ± 2.08^abc	4.33 ± 2.31^ab	4.67 ± 1.53^bcd	4.67 ± 1.53^ab
E (2.5 + 0)	2.33 ± 1.15^cd	0.67 ± 0.58^bc	2.00 ± 1.00^cd	1.00 ± 0.10^c	5.00 ± 2.00^bc	4.00 ± 2.65^ab	3.33 ± 0.58^de	3.33 ± 0.58^bc
F (5.0 + 0)	2.33 ± 0.58^cd	1.00 ± 0.40^abc	1.33 ± 0.58^d	0.67 ± 0.58^c	3.67 ± 0.58^c	3.67 ± 0.58^ab	2.33 ± 0.58^e	2.00 ± 1.00^cd
G (10.0 + 0)	2.00 ± 1.00^d	1.00 ± 1.00^abc	1.33 ± 0.58^d	1.00 ± 1.00^c	4.00 ± 1.00^c	2.67 ± 0.58^ab	2.00 ± 1.00^e	0.67 ± 0.58^d
H (0.1 + 0.1)	4.67 ± 1.15^ab	1.67 ± 0.58^ab	3.67 ± 1.53^ab	2.00 ± 0.10^ab	7.67 ± 1.53^ab	5.00 ± 1.00^ab	5.33 ± 1.53^abc	4.67 ± 1.15^ab
I (0.5 + 0.1)	3.67 ± 1.53^bcd	0.67 ± 0.58^bc	2.33 ± 1.15^bcd	1.00 ± 1.00^bc	5.67 ± 2.52^abc	4.67 ± 2.31^ab	4.67 ± 1.53^bcd	4.67 ± 1.53^ab
J (2.5 + 0.1)	2.67 ± 1.15^bcd	0.20 ± 0.10^c	2.33 ± 0.58^bcd	1.00 ± 0.10^bc	6.00 ± 2.00^abc	4.33 ± 1.15^ab	3.67 ± 1.15^cde	3.33 ± 0.58^bc
K (5.0 + 0.1)	2.33 ± 0.58^c	1.00 ± 0.10^abc	1.33 ± 0.58^d	1.00 ± 0.10^c	4.67 ± 1.53^bc	2.33 ± 0.58^b	2.33 ± 0.58^e	1.00 ± 0.30^d
L (10.0 + 0.1)	2.00 ± 1.00^d	0.67 ± 0.58^bc	1.33 ± 0.58^d	1.00 ± 0.10^bc	4.00 ± 1.00^c	2.67 ± 0.58^ab	3.00 ± 1.00^de	3.00 ± 1.00^bc

Values shown are mean ± SD. n = 8/group/time-point.

Values that are followed by different superscript letters in a column are significantly different from one another (p < 0.05).

Table 2. Anti-SRBC antibody titer in chicks obtained from layer breeders fed AFB₁ ± Vit E-containing feeds for 3 week.

Groups (AF mg/kg  + Vitamin E mg/kg	At day 7 post-primary injection		At day 14 post-primary injection		At day 7 post-booster injection		At day 14 post-booster injection
feed)	Total Ab	IgG	Total Ab	IgG	Total Ab	IgG	Total Ab	IgG
A (0 + 0)	4.67 ± 0.58^ab	1.33 ± 1.15	2.67 ± 0.58^abc	1.67 ± 0.58^abc	8.33 ± 0.58^a	5.67 ± 0.58^a	6.33 ± 0.58^ab	4.67 ± 0.58^ab
B (0 + 0.1)	5.33 ± 0.58 ^a	1.67 ± 0.58	2.67 ± 0.58^ab	1.67 ± 0.58^abc	8.00 ± 1.00^a	5.33 ± 1.53^a	6.67 ± 0.58^a	4.67 ± 0.58^ab
C (0.1 + 0)	4.00 ± 1.00^abc	0.67 ± 0.58	2.67 ± 0.58^ab	2.00 ± 0.30^ab	7.00 ± 1.00^abc	5.00 ± 1.00^ab	5.33 ± 0.58^abc	4.00 ± 1.00^ab
D (0.5 + 0)	4.33 ± 1.53^ab	1.00 ± 0.50	3.00 ± 1.00^a	1.67 ± 0.58^abc	6.67 ± 1.53^abcd	5.67 ± 1.53^a	5.00 ± 1.00^bc	4.00 ± 1.00^ab
E (2.5 + 0)	2.33 ± 0.58 ^c	0.67 ± 0.58	2.00 ± 1.00^bc	1.00 ± 0.00^cd	4.67 ± 1.15^d	3.33 ± 0.58^bc	3.33 ± 0.58^d	3.33 ± 0.58^bc
F (5.0 + 0)	2.33 ± 0.58 ^c	0.67 ± 0.58	1.33 ± 0.58^c	0.67 ± 0.58^d	5.00 ± 1.00^cd	3.00 ± 1.00^ce	3.33 ± 0.58^d	2.00 ± 1.00^c
G (10.0 + 0)	*	*	*	*	*	*	*	*
H (0.1 + 0.1)	5.33 ± 0.58 ^a	1.33 ± 0.58	3.67 ± 0.58^a	2.33 ± 0.58^a	8.33 ± 1.15^a	5.67 ± 0.58^a	6.00 ± 1.00^ab	5.33 ± 1.15^a
I (0.5 + 0.1)	4.33 ± 1.15^ab	1.00 ± 0.60	2.33 ± 1.15^abc	1.33 ± 0.58^bcd	7.33 ± 1.53^ab	4.67 ± 0.58^abc	5.33 ± 1.15^abcd	4.33 ± 1.53^ab
J (2.5 + 0.1)	3.00 ± 1.00^bc	1.00 ± 0.20	2.33 ± 0.58^abc	1.00 ± 0.20^cd	5.67 ± 1.53^bcd	4.33 ± 1.53^abc	4.00 ± 1.00^cd	3.33 ± 0.58^bc
K (5.0 + 0.1)	3.00 ± 1.00^bc	0.67 ± 0.58	1.67 ± 1.15^bc	1.33 ± 0.58^bcd	5.00 ± 1.00^cd	3.33 ± 0.58^bc	4.00 ± 1.00^cd	3.00 ± 1.00^bc
L (10.0 + 0.1)	*	*	*	*	*	*	*	*

Values shown are mean ± SD. n = 8/group/time-point (except G and L). * Progenies in Groups G and L were not available due to limitations on egg production (and thus, progeny available to examine) by dams in these groups.

Values that are followed by different superscript letters in a column are significantly different from one another (p < 0.05).

Figure 1. Lymphoproliferative response against phytohemagglutinin (PHA-P) in chicks from hens fed the test feeds for 1 or 3 weeks. At 14 days-of-age, 10 chicks/group were injected with PHA-P (100 μg PHA-P in 0.1 ml normal saline) into the intra-digital space between digits 3 and 4 of the right foot; 0.1 ml normal saline was injected similarly in the left foot. Skin thickness was then measured 24- and 48-h post-injection. The thickness response was then calculated and expressed (mm) as: cutaneous basophilic hypersensitivity response = (right foot skin thickness – left foot skin thickness), at each time-point. Dam on diets for (a) 1 week and (b) 3 weeks. Values shown are mean ( ± SD); n = 10/group. All groups (A–L) were fed the diets containing various amounts of AFB₁ in the presence/absence of Vit E; specific exposure levels for each group are indicated in tables in this paper. Values significantly different from control (Group A chicks) values at *p ≤ 0.05 are indicated. Note: Due to limitations on overall egg production by hens in the 3-week Groups G and L, eggs (and thus progeny) were not available for this particular study.

Figure 2. Phagocytic activity of peritoneal macrophages recovered from chicks of hens fed AFB₁ ± Vit E for 1, 2, or 3 weeks. At 29 days-of-age, peritoneal macrophages were isolated from the chicks (n = 10/group). The cells were then cultured for 1 h to permit adherence and subsequently provided SRBC to engulf in a 1-h period. The cells were then stained and a total of 200 macrophages/chick scored for phagocytosis and number of SRBC engulfed/macrophage. Dam on diets for (a) 1 week, (b) 2 weeks, and (c) 3 weeks. Values shown are mean percentage (%) of phagocytically-active cells present (±SD); n = 10/group. All groups (A–L) were fed the diets containing various amounts of AFB₁ in the presence/absence of Vit E; specific exposure levels for each group are indicated in tables in this paper. Values significantly different from control (Group A chicks) values at *p ≤ 0.05 are indicated.

Figure 3. Number of SRBC engulfed/macrophage obtained from progeny chicks of hens fed AFB₁ ± Vit E for 1, 2, or 3 weeks. Cells were prepared and treated as outlined in the legend for Figure 2. Dam on diets for (a) 1 week, (b) 2 weeks, and (c) 3 weeks. Values shown are mean number of SRBC engulfed per phagocytically-active cell (±SD); n = 10/group. All groups (A–L) were fed the diets containing various amounts of AFB₁ in the presence/absence of Vit E; specific exposure levels for each group are indicated in tables in this paper. Values significantly different from control (Group A chicks) values at *p ≤ 0.05 are indicated.

Figure 4. LPS-induced nitrite production by peritoneal macrophages recovered from chicks of hens fed AFB₁ ± Vit E for 1, 2, or 3 weeks. Cells were prepared for culture as outlined in the legend for Figure 2. Lipopolysaccharide (LPS, 1 μg/well) was then added to each well to induce nitrite production over a 24-h period. Culture supernatants were then collected and assessed for nitrite using Griess reagent. Dam on diets for (a) 1 week, (b) 2 weeks, and (c) 3 weeks. Values shown are mean µM nitrite/chick cell culture (±SD); n = 10/group. All groups (A–L) were fed the diets containing various amounts of AFB₁ in the presence/absence of Vit E; specific exposure levels for each group are indicated in tables in this paper. Values significantly different from control (Group A chicks) values at *p ≤ 0.05 are indicated.

Figure 5. Comparison of rate of hemolysis values. Values shown are for the tim-epoint at which 50% hemolysis was observed using a free radical generator AAPH. All values shown are with RBC obtained from progenies of breeder hens fed AFB₁ ± Vit E for (a) 1 week, (b) 2 weeks, or (c) 3 weeks. Values shown are mean ± SD; n = 8/group. All groups (A–L) were fed the diets containing various amounts of AFB₁ in the presence/absence of Vit E; specific exposure levels associated with each group are indicated in each table in this paper. Values significantly different from control (Group A chicks) values at *p ≤ 0.05 or **p ≤ 0.01 are indicated. Note: Due to limitations on overall egg production by hens in the 3-week Groups G and L, eggs (and thus progeny) were not available for this particular study.