Biotin-deficient diet induces chromosome misalignment and spindle defects in mouse oocytes

Figures & data

Table 1. Diet compositions.

	Control diet (g/kg diet)	Biotin-deficient diet (g/kg diet)
Egg white solids	300	300
Gelatinized cornstarch	309.3	309.3
Sucrose	154.2	145.2
Corn oil	80	80
Dextrin	50	50
Cellulose	50	50
Choline bitartrate	2.5	2.5
Mineral mixture (AIN-93-G-MX)^*	42	42
Vitamin mixture (biotin free)^*	12	12
Biotin	0.004	–

^* Composition of mineral and vitamin mixtures formulated to meet AIN-76A^39–46).

Table 2. Effects of biotin deficiency.

	Control group	Biotin-deficient group
Body weight and food intake
Initial body weight at Day 1 (g)	12.0 ± 0.3	12.5 ± 0.4
Final body weight at Day 22 (g)	29.2 ± 0.7	27.4 ± 0.7
Body weight gain (g/21 days^†)	17.1 ± 0.9	14.9 ± 0.5^*
Total food intake (g/21 days^†)	90.5 ± 1.8	82.5 ± 2.2^**
Tissue weight
Liver (g)	1.50 ± 0.06	1.69 ± 0.10
(g/kg BW)	53.5 ± 1.3	63.0 ± 1.0^***
Uterus (g)	0.104 ± 0.024	0.087 ± 0.014
(g/kg BW)	3.66 ± 0.72	3.27 ± 0.53
Ovary (g)	0.0179 ± 0.0009	0.0157 ± 0.0009
(g/kg BW)	0.641 ± 0.024	0.594 ± 0.050
Oocyte
Total oocytes (number/mouse)	20 ± 4	25 ± 2
Dead oocytes (number/mouse)	1 ± 0	2 ± 1
Biotin
Urine (nmol/day)	14.6 ± 0.9	0.0743 ± 0.0105^***
Liver (nmol/g)	4.04 ± 0.16	0.84 ± 0.04^***
Uterus (pmol/g)	154 ± 11	23.9 ± 6.4^***
Ovary (pmol/g)	779 ± 93	138 ± 16^***
3-HIA
Urine (μmol/day)	N.D	7.81 ± 1.16
Steroid hormone
Estradiol (pg/mL plasma)	12.5 ± 2.6	12.8 ± 1.8

Notes: N.D, not detected; BW, body weight.

Values are mean ± standard error of mean; n = 5 except initial body weight, final body weight, and body weight gain, where n = 10.

^† The values were the summation of body weight gain or food intake from day 1 to day 22.

^* p <  0.05.

^** p  <  0.01.

^*** p  <  0.001 control vs. biotin-deficient groups.

Table 3. Effects of biotin restoration.

	Control group	Recovery group	Biotin-deficient group
Body weight and food intake
Initial body weight at Day 22 (g)	28.3 ± 0.7	26.7 ± 0.5	27.3 ± 0.5
Final body weight at Day 42 (g)	32.4 ± 0.9^a	31.8 ± 0.9^a	27.2 ± 0.7^b
Body weight gain (g/20 days^†)	4.0 ± 0.5^a	5.1 ± 0.5^a	−0.2 ± 0.9^b
Total food intake (g/20 days^†)	100.8 ± 4.2^a	99.0 ± 4.3^a	81.5 ± 3.9^b
Tissue weight
Liver (g)	1.52 ± 0.05	1.71 ± 0.12	1.51 ± 0.08
(g/kg BW)	47.3 ± 1.6^b	54.1 ± 2.0^a	55.9 ± 1.2^a
Uterus (g)	0.145 ± 0.019^a	0.118 ± 0.010^a	0.061 ± 0.014^b
(g/kg BW)	4.50 ± 0.50^a	3.80 ± 0.39^ab	2.20 ± 0.43^b
Ovary (g)	0.0167 ± 0.0026	0.0133 ± 0.0023	0.0112 ± 0.0010
(g/kg BW)	0.521 ± 0.074	0.416 ± 0.061	0.416 ± 0.037
Oocyte
Total oocyte (number/mouse)	40 ± 9	27 ± 3	27 ± 3
Dead oocyte (number/mouse)	5 ± 2	4 ± 1	2 ± 1
Biotin
Urine (nmol/day)	9.77 ± 0.78^a	11.6 ± 2.3^a	0.0370 ± 0.0053^b
Liver (nmol/g)	4.11 ± 0.24^a	4.39 ± 0.12^a	0.84 ± 0.06^b
Uterus (pmol/g)	171 ± 17^a	195 ± 17^a	13.7 ± 1.5^b
Ovary (pmol/g)	1302 ± 162^a	1165 ± 31^a	117 ± 35^b
3-HIA
Urine (μmol/day)	0.75 ± 0.75^b	N.D^b	24.7 ± 2.5^a
Steroid hormone
Estradiol (pg/mL plasma)	9.1 ± 2.1	6.4 ± 1.6	13.0 ± 2.5
Estrus
Estrus cycle (day/cycle)	4.7 ± 0.2^b	6.1 ± 1.0^ab	8.0 ± 0.7^a

Notes: N.D, not detected; BW, body weight.

Values are mean ± standard error of mean; n = 5 except initial body weight, final body weight, and body weight gain, where n = 10.

^† The values were the the summation of body weight gain or food intake from day 22 to day 42.

^a,b Different letters in the same row indicate significant differences (p <  0.05).

Fig. 1. Spindle abnormalities in oocytes induced by biotin deficiency.

Notes: Representative examples of meiotic spindles in oocytes from indicated mice (n  = 37–38 oocytes analyzed per group) after labeling α-tubulin antibody (green) and counter-staining DNA with DAPI (aqua blue). (A) control group, (B) biotin-deficient group.

Fig. 2. Higher frequency of abnormal oocytes induced by biotin deficiency not restored by re-feeding of biotin-containing diet.

Notes: Abnormal oocyte number is the summation of chromosome alignment and spindle defect in oocytes. (A) Day 22, (B) Day 42. (C) control; BD; biotin deficient group; R, recovery group. n = 37–54 oocytes analyzed per group.

Fig. 3. Meiosis aneuploidy induced by biotin deficiency, not restored by biotin re-feeding.

Notes: Representative examples of meiotic spindles in oocytes from indicated mice (n = 41–66 oocytes analyzed per group) after labeling α-tubulin antibody (green) and counter-staining DNA with DAPI (aqua blue). (A) control group, (B) biotin-deficient group, and, (C) recovery group.