Egg white composition, antioxidant capacity, serum and yolk lipids and oxidative damage of the oviduct magnum in laying hens fed diets contaminated with different concentrations of cadmium

Figures & data

Table 1. Ingredient compositions and nutrient levels of basal diet for hens.^a

Basal ingredients	Value	Nutrient level	Value
Corn, %	65	Metabolism energy, MJ/kg	2.65
Soybean meal (42.0% CP)	21	Crude protein (CP), %	15.73
Feather meal, %	1	Ether extract, %	6.32
Fish meal, %	1	Lysine, %	0.78
Calcium carbonate, %	7	Methionine, %	0.34
Premix^b, %	5	Cysteine, %	0.32
		Total phosphorus, %	0.61
Total, %	100	Calcium, %	3.45

^aEstimated from Chinese feed database provided with tables of feed composition and nutritive values in China (2015 26th edition).

^bThe premix provided following per kilogram of diet: vitamin A, 7000 IU; vitamin D₃, 2500 IU; vitamin E, 49.5 mg; vitamin K₃, 1 mg; vitamin B₁, 1.5 mg; vitamin B₂, 4 mg; vitamin B₆, 2 mg; vitamin B₁₂, 0.02 mg; niacin, 30 mg; folic acid, 0.55 mg; pantothenic acid, 10 mg; biotin, 0.16 mg; chloride becholine, 400 mg; Cu, 20 mg; Fe, 70 mg; Mn, 100 mg; Zn, 70 mg; I, 0.4 mg; Se, 0.5 mg.

Table 2. Primer used for quantitative real-time PCR.

Target gene	Primer	Primer Sequence (5'-3')	Accession no.
β-actin	Forward	TCCCTGGAGAAGAGCTATGAA	NM_205518.1
	Reverse	CAGGACTCCATACCCAAGAAAG
OV	Forward	GCTGTGTGCTTTACCTCTTTG	XM_040673607.1
	Reverse	GAACTGGGTCCTTCTGTCATC
OVA	Forward	GAGAGCAGATCACGGCATATT	MH360742.1
	Reverse	GCTCATCAATTCCTGGGTAGAA
LYZ	Forward	TACGACACTGGCAACATGAG	KU933268.1
	Reverse	CGGCTGTTGATCTGTAGGATT
OVM	Forward	TGCCTGCACGATGGAATACA	XM_035559733.1
	Reverse	TCAGCATTCTCCAAAATGGCTTA
AVD	Forward	CCTACATCACAGCCGTAACA	AM779412.1
	Reverse	GCTTTCCAGTCATCACCAATG
Nrf2	Forward	CTGCCCAAAACTGCCGTA	NM_205117.1
	Reverse	TCAAATCTTGCTCCAGTTCCA
NQO1	Forward	CTCCGAGTGCTTTGTCTACG	XM_015874307.1
	Reverse	AATGGCTGGCATCTCAAACC
HO-1	Forward	GCTGAAGAAAATCGCCCAA	NM_205344.1
	Reverse	ATCTCAAGGGCATTCATTCGG
SOD1	Forward	TGTGCATGAATTTGGAGACAAC	NM_205064.1
	Reverse	TTGCAGTCACATTGCCGAG
SOD2	Forward	TGCACTGAAATTCAATGGT	NM_204211.1
	Reverse	GTTTCTCCTTGAAGTTTGCG
SOD3	Forward	TTTTCTCCTAAAGATGGCAAG	XM_420760.3
	Reverse	CTTCCTGCTCATGGATCACAA
GST	Forward	GGAAGCCATTTTAATGACAGA	XM_046913335.1
	Reverse	TCCTTTAAAAGCCTGTAGCAGA
GCLC	Forward	TCTGTAGATGATCGAACGC	XM_419910.4
	Reverse	TCCTTTATTAGGTGCTCGTAG
GCLM	Forward	GCTGCTAACTCACAATGACC	NM_001007953.1
	Reverse	TGCATGATATAGCCTTTGGAC
IL1β	Forward	GCCTCTGCTCCCATTTAGTT	XM_015297469.1
	Reverse	CTCACAGTCCTTCGACATCTTC
IL-6	Forward	CGGTACATACGAGATGGAAACC	XM_015281283.2
	Reverse	GATCCGGCAGATGGTGATAAA
TNFα	Forward	GAGGATTGTGCCCGAACTAAA	AY765397.1
	Reverse	GACAGCCTATGCCAACAAGTA
NF-κB	Forward	TCCACATCTTTCAGAGCATCAA	XM_015856764.1
	Reverse	CTCCTCAACCTCACTTCCTTAC
MFN1	Forward	GTTGTTGGCGGAGTGATTTG	XM_046923917.1
	Reverse	CTTGGAGTGGCTCTGTATGTT
MFN2	Forward	GACAGGTTGCCTTGTGAGATAG	XM_040689233.2
	Reverse	CCCATTCTTACCCTGGCATTAG
PINK1	Forward	AGCAGCGATTCCTTCATCTC	NM_001389481.2
	Reverse	CTTCAGGTCTCTGTGTGCTATC
Parkin	Forward	GTCCAGCAAAGCATCGTTCA	XM_046914604.1
	Reverse	CAACGATGGAAGGATGCTGG
Fundc1	Forward	GTTGTGCTGGGTTCTCATTTAC	NM_001276363.2
	Reverse	TGTTCCTCGGCGTTTCTT
GRP75	Forward	AAGAGGCAGGCAGTAACTAATC	NM_001006147.2
	Reverse	CTTCAGACTTGTCCAGACCATAG
VDAC1	Forward	GCCTGAAGCTGACTTTTGACTCC	XM_046927011.1
	Reverse	GATGTGCTCCCTTTTGTATCCTGT
MCU	Forward	TTGGCAGAGTGTGAGAGTGG	XM_046920626.1
	Reverse	AATTCCTCGGTCCTCTGCTT
LC3I	Forward	TTACACCCATATCAGATTCTTG	XM_040688401.2
	Reverse	ATTCCAACCTGTCCCTCA
LC3II	Forward	AGTGAAGTGTAGCAGGATGA	NM_001031461.2
	Reverse	AAGCCTTGTGAACGAGAT
MFF	Forward	ACTCAAAGTGGCTCCTCA	XM_040679333.2
	Reverse	CCTGCATAGTTACACTGG
PGC1α	Forward	TACAGCAATGAGCCTGCCAA	XM_046916274.1
	Reverse	AGGCAATCCATCCTCATCCAC
Chop	Forward	GCACAGCCCATTTCTGTTTC	XM_015273173.2
	Reverse	TGCCATCCCATTCTGCTAAG
GRP78	Forward	GTTACTGTGCCAGCCTACTT	NM_205491.1
	Reverse	CCGCTTCGCTTTCTCTACTT
ATF4	Forward	TCACCCAATGACAACCCG	NM_204880.3
	Reverse	TCACCTTTGCTGACGCTACC
ATF6	Forward	CGTCGTCTGAACCACTTACTGA	XM_040677276.2
	Reverse	CCTTCTTTCCTAACAGCCACAC
IRE1α	Forward	CTACAGGTCGCTCCTCACATC	XM_046927561.1
	Reverse	ATCAGTCCTTCTGCTCCCATCT
GRP94	Forward	CAAAGACATGCTGAGGCGAGT	NM_204289.2
	Reverse	TCCACCTTTGCATCCAGGTCA

Figure 1. Effects of dietary Cd contamination on Cd levels in egg white and magnum of laying hens. Data are presented as mean ± standard error of means (SEM). Different letters (a, b, c, d) indicate significant differences between groups (p < .05). Linear regression analysis was used to assess the dose-response relationship between Cd exposure and Cd contents in albumen and magnum.

Figure 2. Effects of Cd on gene expressions in magnum (A) and protein levels in egg white (B, C) of laying hens. Data are presented as mean ± standard error of means (SEM) (n = 6). Different letters (a, b, c) above the histogram indicate significant differences between groups (p < .05). OV: ovomucin; OVA: ovalbumin; LYZ: lysozyme; OVM: ovomucoid; AVD: avidin.

Figure 3. Effect of Cd on the antioxidant ability of the oviduct magnum (A) and egg white (B) of laying hens. Data are presented as mean ± standard error of means (SEM) (n = 6). Different letters (a, b, c) above the histogram indicate significant differences between groups (p < .05). GSH: glutathione; GSH-Px: glutathione peroxidase; T-SOD: total superoxide dismutase; CAT: catalase; MDA: malondialdehyde.

Table 3. Effect of Cd on the serum and yolk lipids in laying hens.

Items	Dietary Cd dosage, mg/kg				SEM	p Value
	0	15	30	60
SERUM, mmol/L
T-CHO	3.25^b	4.16^ab	3.81^ab	5.01^a	0.442	.013
TG	17.12	19.26	15.28	10.24	3.491	.116
HDLC	0.51^b	1.31^ab	2.02^ab	4.55^a	0.806	.005
LDLC	0.99^b	1.16^b	1.40^b	2.71^a	0.393	.005
EGG YOLK, mg/g
T-CHO	19.11^b	20.62^ab	24.01^a	24.68^a	1.472	.005
TG	53.43	49.82	61.14	60.73	6.073	.223
HDLC	2.61	3.10	3.45	3.00	0.596	.580
LDLC	10.36^b	13.23^ab	13.23^ab	17.01^a	2.097	.055

Notes: SEM: standard error of the means; T-CHO: total cholesterol; TG: triglyceride; HDLC: high density lipid-cholesterol; LDLC: low density lipid-cholesterol. Values are presented as the mean and SEM (n = 12).

^a–bMeans within a row with different superscripts are significantly different (p < .05).

Figure 4. Effects of Cd on the Nrf2 signalling pathway in the oviduct magnum of laying hens. (A) Nrf2 pathway related gene expression; (B) Nrf2 and Keap1 protein expression levels. Data are presented as mean ± standard error of means (SEM) (n = 6). Different letters (a, b, c) above the histogram indicate significant differences between groups (p < .05). **p < .01 indicates a significant difference compared to the control group. Nrf2: nuclear factor-erythroid 2 p45-related factor 2; Keap1: kelch-like ECH-associated protein 1; NQO1: NAD(P)H quinone dehydrogenase 1; HO-1: haem oxygenase 1; GCLC: glutamate-cysteine ligase catalytic subunit; GCLM: glutamate-cysteine ligase modified subunit.

Figure 5. Effects of Cd on the expression of inflammatory factors in the oviduct magnum of laying hens. Data are presented as mean ± standard error of means (SEM) (n = 6). Different letters (a, b, c) above the histogram indicate significant differences between groups (p < .05). **p < .01 indicates a significant difference compared to the control group. IL: interleukin; TNFα: tumour necrosis factor α; NF-κB: nuclear factor kappa-B.

Figure 6. Effects of Cd on endoplasmic reticulum-stress in the oviduct magnum of laying hens. Data are presented as mean ± standard error of means (SEM) (n = 6). Different letters (a, b, c) above the histogram indicate significant differences between groups (p < .05). *p < .05 and **p < .01 indicates a significant difference compared to the control group.

Figure 7. Effects of Cd on mitochondrial function and dynamics in the oviduct magnum of laying hens. Data are presented as mean ± standard error of means (SEM) (n = 6). Different letters (a, b, c) above the histogram indicate significant differences between groups (p < .05).

Data availability statement

The data that support the findings of this study are available from the corresponding author, upon reasonable request.