3.1 Population characteristics
Mothers with probable postpartum depression were more likely to belong to middle socioeconomic group (p = 0.002), have had more than one child (p=0.002), were dissatisfied with their marriage (p < 0.001) & the gender of the child (p=0.004), have had an unplanned pregnancy (p < 0.001), and have had higher rates of cesarean section (p = 0.014). They also reported lower intake of milk (p < 0.001), meat (p = 0.012) and eggs (p = 0.002) during the postpartum period. Age, BMI, other maternity stressor (e.g., lack of sleep) did not differ between those with and without postpartum depression (Table 1). All the women included in the study reported intake of folate supplementation during pregnancy, which was supplied by the hospital as a routine care.
Table 1: Maternal anthropometric, social, obstetrical and psychological characteristics in 434 women with and without probable PPD according to EPDS score cut-off of >10 (N, %, mean and SD)
Variable
|
Group
|
P-value
|
Women with probable postpartum
depression (n=217)
|
Women without probable postpartum depression (n=217)
|
Anthropometric characteristics
|
Age(years)
Mean (SD)
Min – Max
|
26.22(4.30)
18 – 40
|
25.56(4.08)
19 – 36
|
0.102^
|
Age
<=25yrs
26 – 30 yrs
>30 yrs
|
111(51.2%)
77(35.5%)
29(13.4%)
|
118(54.4%)
73(33.6%)
26(12.0%)
|
0.785@
|
BMI (kg/m2)
Mean(SD)
Min – Max
|
25.08(4.55)
13.33 – 39.12
|
25.61(4.57)
13.90 – 40.50
|
0.225^
|
BMI (kg/m2)
<=18.5
>18.5 – 24.9
>24.9 – 29.9
>30
|
12(5.5%)
109(50.2%)
60(27.7%)
36(16.6%)
|
6(2.8%)
109(50.2%)
62(28.6%)
40(18.4%)
|
0.523@
|
Socio-demographic characteristics
Family structure
Extended
Nuclear
|
149(68.7%)
68(31.3%)
|
150(69.1%)
67(30.8%)
|
0.917@
|
SES$
Lower
Upper lower
Lower middle
Upper middle
Upper
|
4(1.8%)
112(51.6%)
54(24.9%)
47(21.7%)
-
|
9(4.2%)
142(65.4%)
31(14.3%)
32(14.8%)
3(1.4%)
|
0.002@*
|
Total children
1
>=2
|
116(53.5%)
101(46.5%)
|
148(68.2%)
69(31.8%)
|
0.002@*
|
Marital dissatisfaction
No
Yes
|
153(70.5%)
64(29.5%)
|
213(98.2%)
4(1.8%)
|
<0.001@*
|
Any death in the family in the last one year
No
Yes
|
213(98.2)
4(1.8)
|
215(99.1)
2(0.9)
|
0.411@
|
Breast feeding
Yes
No
|
214(98.6)
3(1.4)
|
215(99.1)
2(0.9)
|
0.653@
|
Sleeping hours after delivery
<6hrs/day
>6hrs/day
|
100(46.1)
117(53.9)
|
98(45.2)
119(54.8)
|
0.847@
|
Are you happy with the baby gender
Yes
No
|
46(21.2)
171(78.8)
|
24(11.1)
193(88.9)
|
0.004@*
|
Family members happy with the sex of the child
No
Yes
|
55(25.4)
162(74.6)
|
49(22.6)
168(77.4)
|
0.500@
|
Obstetrics characteristics
Parity
Primi
Multi
|
102(47.0)
115(53.0)
|
120(55.3)
97(44.7)
|
0.084@
|
History of miscarriage/still birth
No
Miscarriage
Still birth
|
165(76.0)
51(23.5)
1(0.5)
|
165(76.0)
45(20.7)
7(3.2)
|
0.087@
|
Pregnancy planning
Unplanned
Planned
|
108(49.8)
109(50.2)
|
66(30.4)
151(69.6)
|
<0.001@*
|
Delivery type
Vaginal
Vaginal assisted
Cesarian
|
135(62.2)
17(7.8)
65(30.0)
|
163(75.1)
10(4.6)
44(20.3)
|
0.014@*
|
Difficulty in delivery
Yes
No
|
86(39.6)
131(60.4)
|
77(35.5)
140(64.5)
|
0.372@
|
Perinatal complications
Yes
No
|
18(8.3)
199(91.7)
|
18(8.3)
199(91.7)
|
0.999@
|
Nutritional characteristics
Diet
Veg
Mixed diet
|
11(5.1)
206(94.9)
|
13(6.0)
204(94.0)
|
0.674@
|
Milk
Seldom
Once per day
2-3 times per day
>4 times per day
|
7(3.2)
86(39.8)
109(50.5)
14(6.5)
|
9(4.2)
41(18.9)
148(68.2)
19(8.8)
|
<0.001@*
|
Curd/Butter milk
Seldom
1-2 times per week
3-4 times per week
Once a day
|
178(82.0)
11(5.1)
11(5.1)
17(7.8)
|
196(90.3)
13(6.0)
6(2.8)
2(0.9)
|
0.002@*
|
Fishes
Seldom
1-2 times per week
3-4 times per week
Once a day
|
65(30.4)
83(38.8)
54(25.2)
12(5.6)
|
64(29.6)
87(40.3)
57(26.4)
8(3.7)
|
0.808@
|
Meat
Seldom
1-2 times per week
3-4 times per week
Once a day
|
85(39.7)
88(41.1)
25(11.7)
16(7.5)
|
74(34.3)
88(40.7)
47(21.8)
7(3.2)
|
0.012*
|
Egg
Seldom
1-2 times per week
3-4 times per week
Once a day
|
38(17.7)
36(16.7)
55(25.6)
86(40.0)
|
32(14.8)
100(46.3)
68(31.5)
16(7.4)
|
<0.001@*
|
Term baby
Pre-term
Term
|
34(15.7)
183(84.3)
|
24(11.1)
193(88.9)
|
0.158@
|
Gender of baby
Male
Female
|
121(55.8)
96(44.2)
|
133(61.3)
84(38.7)
|
0.242@
|
$ Based on Kuppusamy’s Socioeconomic scale (21)
@ P-value based on Chi-square test
^ P-value based on Mann-Whitney U test
3.2. Biochemical analysis
The median total vitamin B12 levels in cases were significantly lower than controls with no difference in its active form, holotc. MMA, a marker of functional deficiency of vitamin B12 was significantly elevated in depressed women compared those without depression (p = 0.002) (Table 2). Though hcy, a non-specific marker of vitamin B12 deficiency was notably increased in cases than control, the difference did not achieve a statistical significance (p = 0.057). Also, 5-methyl THF was increased in depressed women (p < 0.0001) signifying a ‘folate trap’ due to existing B12 deficiency. A significantly lower value of cB12 was observed in women with probable depression as compared to controls (p < 0.001).
Table 2: Plasma levels of vitamin B12, and other metabolites in study population
Characteristics
|
Control
Median (IQR)
|
Cases
Median (IQR)
|
P value
|
Total vitamin B12 (pg/ml)
|
353.9
(284.4-415.9)
|
294
(247.7-365.8)
|
<0.001
|
Holotranscobalamin
(pmol/L)
|
57.4
(49.4-86.7)
|
60.9
(52.3-74.4)
|
0.510
|
Homocysteine
(micromol/L)
|
3.02
(1.9-4.52)
|
2.99
(2.39-5.52)
|
0.057
|
MMA
(micromol/L)
|
19.1
(16.61-23.05)
|
22.09
(14.27 – 124.49)
|
0.002
|
cB12
|
1.59
(-0.94-2.69)
|
1.25
(-0.79-2.7)
|
<0.001
|
SAM (ng/ml)
|
32.89
(20.01-73.02)
|
38.47
(22.54 – 64)
|
0.373
|
5 methyl THF (ng/ml)
|
2.52
(2.08-3.25)
|
2.93
(2.39-3.67)
|
<0.001
|
Women in the study were divided into 3 groups based on clinical decision limits of total vitamin B12: Deficient (less than 150 pmol/L), low normal (190-300 pmol/L) and Sufficient (>300 pmol/L). In the whole cohort (n = 434), 58% of the women had sufficient vitamin B12, 38.5% had low normal levels and 3.5% women were overtly deficient. Among cases and controls, significantly higher number of women with PPD had low normal vitamin B12 (49.8%) compared to women without PPD (27.2%). The cutoff limit for holotranscobalamin is 35pmol/L, to define sufficient and deficient status. There was no significant difference among women with and without depression for clinical decision limits of holotranscobalamin.
3.3 Association of Vitamin B12 and its deficiency indicators with probable postpartum depression
Regression analyses showed negative association of vitamin B12, cB12 and serotonin with probable PPD, whereas a positive association of MMA, hcy and 5-methyl THF was observed with depressive symptoms (table 3).
Table 3: Association of Edinburg Postpartum Depression Scale (EPDS) scores with maternal circulating parameters (log transformed)
Model
|
Vitamin B12 metabolites
|
Β
|
p-value
|
Unadjusted
|
Total B12
|
-0.103
|
<0.027*
|
MMA
|
0.263
|
<0.001*
|
5-methyl THF
|
0.157
|
0.001*
|
Hcy
|
0.140
|
0.015*
|
holotc
|
0.979
|
0.922
|
SAM
|
-0.098
|
0.085
|
Serotonin
|
-0.124
|
<0.040*
|
cB12
|
-0.276
|
<0.0001*
|
Adjusted$
|
Total B12
|
-0.064
|
0.146
|
MMA
|
0.161
|
0.001*
|
5-methyl THF
|
0.118
|
0.010*
|
Hcy
|
0.155
|
0.005*
|
holotc
|
-0.051
|
0.260
|
SAM
|
-0.067
|
0.218
|
Serotonin
|
-0.161
|
0.005*
|
cB12
|
0.169
|
<0.001*
|
* p-value<0.05 using linear regression
$Adjusted for socioeconomic status, marital dissatisfaction, unplanned pregnancy and type of delivery
Regression model after adjusting for significant socio-demographic and nutritional characteristics demonstrated that the likelihood of probable postpartum depression decrease by a factor of 0.39 for every unit increase in total vitamin B12 (OR = 0.394; 95% CI: 0.189-0.822) and by a factor of 0.29(OR=0.293; 95% CI:0182-0.470) for cB12. Both MMA and 5-methyl THF were found be significant predictors of probable postpartum depression (table 4). SAM and hcy although were associated with probable PPD in unadjusted model, on subsequent adjustment with nutritional and socio-demographic covariates, lost that significance (table 4).
Table 4: Logistic regression model of the association of maternal circulating vitamin B12 deficiency markers with probable postpartum depression based on Edinburg Postpartum Depression Scale (EPDS) cut-off of >10
Model
|
Vitamin B12 metabolites
|
Odds ratio
|
95% CI
|
p-value
|
Predictive value of the model
|
Unadjusted
|
Total B12
|
0.355
|
0.175-0.718
|
0.004*
|
76.5%
|
MMA
|
1.99
|
1.53-2.59
|
<0.001*
|
5-methyl THF
|
2.96
|
1.46-6.01
|
0.003*
|
Hcy
|
1.45
|
1.00-2.09
|
0.048*
|
holotc
|
0.979
|
0.641-1.497
|
0.922
|
SAM
|
0.74
|
0.573-0.967
|
0.027*
|
Serotonin
|
0.844
|
0.581-1.22
|
0.372
|
cB12
|
0.357
|
0.239-0.532
|
<0.0001*
|
Adjusted$
|
Total B12
|
0.514
|
0.289-0.912
|
0.009*
|
78.6%
|
MMA
|
2.04
|
1.53-2.11
|
<0.001*
|
5-methyl THF
|
3.18
|
1.42-6.08
|
0.001*
|
Hcy
|
1.42
|
0.959-2.1
|
0.079
|
holotc
|
0.968
|
0.61-1.49
|
0.893
|
SAM
|
0.762
|
0.59-1.01
|
0.063
|
Serotonin
|
0.832
|
0.573-1.21
|
0.309
|
cB12
|
0.312
|
0.192-0.501
|
<0.001*
|
*p-value <0.05
$Adjusted for socioeconomic status, marital dissatisfaction, unplanned pregnancy and type of delivery
In quartile analysis, decreasing total vitamin B12 levels were associated with increasing odds of probable postpartum depression. With the highest vitamin B12 quartile as reference, those in the lowest vitamin B12 quartile had 4.53 times higher likelihood of probable postpartum depression (p < 0.001). Increasing MMA and 5-methyl THF levels were also significantly associated with the risk of PPD (OR = 3.26 and 2.97, respectively). No associations were observed between quartiles levels of hcy, SAM and probable postpartum depression.
On path analysis, the model with total vitamin B12 as the predictor, MMA the mediator and EPDS score as the dependent variable was statistically significant (p < 0.001). This indicates the role of MMA as a potential mediator in appearance of depressive symptoms in postpartum period. On the other hand, 5-methyl THF as a mediator for the effect of vitamin B12 deficiency on EPDS score was not found to be significant. Mediation analysis with serotonin and SAM was not statistically significant.