Hypertension in Pregnancy

ISSN: 1064-1955 (Print) 1525-6065 (Online) Journal homepage: http://www.tandfonline.com/loi/ihip20

Limited evidence for calcium supplementation in preeclampsia prevention: a meta-analysis and systematic review Reuben Tang, Ing Ching Tang, Amanda Henry & Alec Welsh To cite this article: Reuben Tang, Ing Ching Tang, Amanda Henry & Alec Welsh (2015) Limited evidence for calcium supplementation in preeclampsia prevention: a metaanalysis and systematic review, Hypertension in Pregnancy, 34:2, 181-203, DOI: 10.3109/10641955.2014.988353 To link to this article: http://dx.doi.org/10.3109/10641955.2014.988353

Published online: 30 Dec 2014.

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Date: 05 December 2016, At: 06:45

Hypertens Pregnancy, 2015; 34(2): 181–203 ! Informa Healthcare USA, Inc. ISSN: 1064-1955 print / 1525-6065 online DOI: 10.3109/10641955.2014.988353

Limited evidence for calcium supplementation in preeclampsia prevention: a meta-analysis and systematic review Reuben Tang,1 Ing Ching Tang,2 Amanda Henry,1,3,4 and Alec Welsh1,3,5 1

Department of Maternal-Fetal medicine, Royal Hospital for Women, Sydney, Australia, 2 Department of Obstetrics and Gynaecology, Rejang Medical Centre, Sibu, Malaysia, 3 School of Women’s and Children’s Health, UNSW Medicine, University of New South Wale, Randwick, NSW, Australia, 4 Department of Obstetrics & Gynaecology, St. George Public Hospital, Sydney, Australia, and 5 Australian Centre for Perinatal Science, University of New South Wales, Sydney, NSW, Australia Objective: This article synthesises evidence for calcium supplementation in preeclampsia prevention. Methods: Major databases and trial registries were searched, and comparisons were made against other meta-analyses. Results: Calcium supplementation reduced the overall risk of preeclampsia in 10 trials (n = 24 787; risk ratio (RR) 0.62; 95% confidence interval [CI] 0.47–0.81). Its effect was larger in two subgroups: lowbaseline calcium intake (RR 0.42 [0.23–0.76]) and increased risk of developing hypertensive disorders (RR 0.36 [0.10–0.98]). This effect was not significant amongst larger studies (RR 0.93 [0.83–1.04]). Funnel plotting suggested possible publication bias. Conclusion: Some evidence for calcium supplementation exists, but its utility is limited by the possibility of publication bias and a lack of large trials. Keywords

Preeclampsia, Calcium, Systematic review

Calcium

supplementation,

Meta-analysis,

INTRODUCTION Preeclampsia is a multisystem disorder usually characterized by de novo hypertension and proteinuria beyond 20 weeks of pregnancy. Preeclampsia falls under the broad spectrum of hypertensive disorders of pregnancy which complicate 2–8% of pregnancies, representing a major contribution to maternal mortality and morbidity worldwide (1,2). Preeclampsia’s precise aetiology remains unclear, although multiple factors including immunological, genetic, dietary and placental proteins have been implicated (3), suggesting that distinct clinical subtypes of preeclampsia may exist (4). Of particular interest in the prevention or management of preeclampsia is the administration of calcium, largely supported by Correspondence: Reuben Tang, Department of Maternal-Fetal medicine, Royal Hospital for Women, Sydney, Australia. E-mail: [email protected]

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epidemiological studies which have found an increased incidence of preeclampsia in regions with low-calcium intake (5–7) and biochemical studies demonstrating significantly depressed levels of serum calcium levels in preeclamptic versus normal women (8–10). Previous work regarding calcium’s role suggests that low-calcium intake may contribute to elevated blood pressures either via stimulation of parathyroid hormone or renin release, leading to vasoconstriction (11,12). Other studies propose that calcium supplementation reduces uterine smooth muscle contractility, possibly by increasing nitric oxide synthesis (via a calcium-dependant mechanism), leading to reduced vascular smooth muscle contractility and decreased resistance in both uterine and systemic vessels (4,13,14). Autoantibody induced Angiotensin-II receptor activation (which raises intracellular calcium via an IP3-dependant mechanism) has also been implicated (15–17). From a molecular perspective, recent in vivo studies assuming hypoxic conditions typical in preeclamptic syncytiotrophoblasts have found decreased expression of calcium transporters, and altered expression of the mRNA of certain calcium-related proteins (18–20). Therefore, impaired calcium metabolism found in preeclamptic syncytiotrophoblasts (due to hypoxic stress) may induce expression of calcium transport proteins in the placenta to maintain the balance between maternal and fetal calcium demand during pregnancy. Calcium supplementation for preeclampsia prevention remains controversial, with studies reaching differing conclusions regarding the role that calcium supplementation plays in improving patient outcomes in pregnancyrelated hypertension (21–24). Given the diversity of published data, this review aims to review the evidence for calcium supplementation in preeclampsia.

METHODS This paper consists of two parts, a meta-analysis to review data pertaining to the effectiveness of calcium in preventing preeclampsia and a systematic review of meta-analysis for this mode of treatment. For the purpose of this project, we accepted the RCOG clinical definition of preeclampsia that is the presence of gestational hypertension with significant proteinuria. To address both aspects of this paper, Pubmed/Medline, Embase, Cochrane, International Clinical Trials Registry Platform and ClinicalTrials.gov databases were searched under all subject headings using the terms: (preeclampsia [MeSH terms] OR (preeclamp*) OR (pre-eclamp*) OR (pregnancy-induced hypertension) OR (pregnancy induced hypertension) OR (gestational hypertension)) AND (calcium). Search terms were modified accordingly based on database operators. Only trials published in the English language and with an abstract were considered for review. Snowballing of trials based on references found in articles was also performed. Articles presented only as an abstract were included if there was sufficient detail to assess the final risk ratio. Only randomised trials comparing at least 1 g of calcium supplementation daily during pregnancy versus placebo were included. No limit on year of publication was set for the meta-analysis component of this paper, and only meta-analyses published after

Calcium for the prevention of preeclampsia

2008 were accepted for our systematic review. All searches were performed on the 29 March 2014. The primary outcome for our meta-analysis was the proportion of women developing preeclampsia in the calcium supplementation group versus placebo. No other outcomes such as preterm birth, maternal morbidity or mortality were reviewed. Data collected were presented as summary risk ratios with 95% confidence intervals. Statistical analysis for heterogeneity was performed using Cochran–Mantel–Haenszel’s test assuming a random effect model in Revman 5.2.11 (Cochrane Collaborative Group, Copenhagen, Denmark). Pre-specified subgroup analysis was performed based on: (1) Baseline dietary calcium intake, which if undefined by trial authors, was defined as low if 5840 mg/day, the estimated average requirement of pregnant women recommended by the National Health and Medical Research Council (25) (2) Baseline increased risk of hypertensive disorders of pregnancy, which included women at increased risk as determined by trial authors (teenagers, women with pre-existing hypertension or previous preeclampsia and positive angiotensin II sensitivity test) (26) As it was noted during data analysis that positive results (i.e. supporting calcium supplementation) appeared to be concentrated amongst smaller studies (51500 women), a post-hoc subgroup analysis by trial size was also performed. Studies were defined as large if the study sample was 1605 women in each arm. Critical appraisal of studies was conducted using Revman 5.2.11.

RESULTS The PRISMA process statement for the meta-analysis segment of this article is outlined in Figure 1. It encompassed 24 787 women across 10 trials. A summary of trials reviewed is presented in Table 1. Most studies indicated a low risk of bias (Appendix 2) though Nenad 2011, which was presented in abstract form only, was given an ‘‘unclear’’ rating. Pooling of the data demonstrated a statistically significant reduction in preeclampsia with calcium supplementation (692/12 399, 5.6%) versus a placebo (821/12 388, 6.6%), with RR 0.62, 95% CI 0.47–0.81 (Table 2). This was more pronounced in the low-baseline intake subgroup (n = 10534), with RR 0.42, 95% CI 0.23–0.76). In the adequate baseline calcium intake group (n = 5045), the risk reduction for preeclampsia with calcium supplementation did not reach statistical significance (RR 0.70; 95% CI 0.34–1.45; Table 3). Calcium supplementation in the high risk of hypertension subgroup (n = 5045) was associated with a significantly lower risk of preeclampsia versus placebo (RR 0.36; 95% CI 0.13–0.98). A lesser, but still statistically significant, decrease with calcium supplementation was seen in the subgroup with a normal risk of developing preeclampsia (n = 10534; RR 0.66; 95% CI 0.48–0.91; Table 4). Funnel plotting of studies based on subgroups demonstrated a lack of negative studies amongst trials with fewer recruits and a lack of positive studies amongst trials with a large number of recruits (Figure 2). The

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Records idenfied through database searching (n = 1589)

Records aer duplicates removed (n = 1535)

Title/abstract screen (n = 1535)

Records excluded (n = 1512)

Full-text arcles assessed for eligibility (n = 23)

Full-text arcles excluded (n = 13) Inappropriate outcome (n=4) Inappropriate intervenon (n=7) Inappropriate methodology (n=1) See appendix-1 for more details

Studies included in metaanalysis (n = 10)

Figure 1. PRISMA process statement.

resultant post-hoc subgroup analysis by trial size found that in large studies (3 studies, 22 079 patients), calcium supplementation did not significantly decrease preeclampsia risk (RR 0.93 [95% CI 0.83–1.04]). In contrast, a large and significant decrease in preeclampsia risk with calcium supplementation was found amongst small studies (7 studies, 2708 patients, RR 0.32 [95% CI 0.21–0.49]). Heterogeneity was considerable (I2 = 73%) amongst included studies, with subgroup analysis using the baseline calcium intake and preeclampsia risk failing to produce any substantially lower I2 statistic. Heterogeneity was associated with study size, with the I2 statistic dropping to 0 and 27% in the large study (defined as 1605 women in each arm) and small study subgroups (Table 5). This cut-off of 1605 in each arm was used to achieve a power of 0.8 using an alpha of 0.05, assuming a risk reduction of 0.4 and a preeclampsia incidence of 0.05, in line with our findings. Based on this finding, smaller studies seemed to indicate a more substantial benefit following calcium supplementation versus larger studies. In comparison to previous meta-analyses of the same topic, we found that our meta-analysis was the most stringent in defining preeclampsia, with most

Multicentre study, Argentina

Florida, USA

Nagpur, India

Multicentre study, USA

Quito, Ecuador

Sanchez-Ramos et al. (41)

Purwar et al. (30)

Levine et al. (23)a

Lopez-Jaramillo et al. (5)

Location

Belizan et al. (40)

Author

Table 1. Summary of trials.

Prospective, double-blind, randomised-controlled trial

Prospective, singleblind, randomised-controlled trial

Prospective, double-blind, randomised-controlled trial

Prospective, double-blind, placebo-controlled trial

Prospective, double-blind, randomised-controlled trial

Study type

260 nulliparas aged 517.5 years

4589 nulliparae

190 nulliparae

67 normotensive nulliparae

1167 nulliparae

Population

2 g oral calcium daily versus placebo

2000 mg oral calcium daily versus placebo

2 g oral calcium daily versus placebo

2 g oral calcium daily versus placebo

2 g oral calcium daily versus placebo

Intervention

Non-significantly lower rates of preeclampsia in intervention group (2.6 versus 3.9%; OR 0.65, 95% CI 0.35– 1.25; p = 0.01). Significantly lower indigence of preeclampsia in the intervention group (13.8 versus 44.1%; OR 0.31 [0.12, 0.84]). Significantly lower incidence of preeclampsia in the calcium group (2.06 versus 11.82%; RR 0.13; 95% CI 0.01–0.64). Non-significant reduction in preeclampsia incidence in intervention group (6.9 versus 7.3%; RR 0.94, 95% CI 0.76–1.16). Significant reduction in preeclampsia incidence in intervention group (3.2 versus 15.5%; RR 0.88; p50.001).

Relevant findings

(Continued )

Low

Low

Low

Low

Low

Overall risk of bias

Calcium for the prevention of preeclampsia

185

Tehran, Iran

Multicentre study: Argentina, Egypt, India, Peru, Vietnam and South Africa New Delhi, India

Serbia

Niromanesh et al. (42)

Villar et al. (24)b

Kumar et al. (22)

Nenad et al. (37)

b

Prospective, double-blind, randomised-controlled trial

Prospective, double-blind, randomised-controlled trial

Prospective, double-blind, randomised-controlled trial Prospective, double-blind, randomised-controlled trial Prospective, double-blind, randomised-controlled trial

Study type

9178 nulliparous women

524 normotensive primigravidae

2 g oral calcium versus placebo

2 g oral calcium versus placebo

1.5 g oral calcium daily versus placebo

2 g oral calcium daily versus placebo

30 women at high risk of preeclampsia

8312 normotensive nulliparas from populations with 5600 mg/ day dietary calcium

1.8 g oral calcium daily versus placebo

Intervention

456 nulliparas

Population

Conducted on behalf of the Calcium for Pre-Eclampsia Prevention (CPEP) trial. Conducted on behalf of the World Health Organisation (WHO).

a

Multicentre study, Australia

Location

Crowther et al. (21)

Author

Table 1. Continued

Preeclampsia incidence was significantly lower in intervention group (4.0 versus 12.0%; OR = 0.31, 95% CI 0.15–0.63). Preeclampsia incidence was not significantly lower in the intervention group (6.9 versus 7.3%, RR 0.94, 95% CI 0.76–1.16).

Significant reduction in preeclampsia (4.4 versus 10%, RR 0.44, 95% CI 0.21–0.90, p = 0.02). Significant reduction in preeclampsia in intervention group (6.67 versus 46.67%; p = 0.014). Non-significant decrease in preeclampsia incidence (4.1 versus 4.5%; RR 0.91, 95% CI 0.69–1.19).

Relevant findings

Unclear

Low

Low

Low

Low

Overall risk of bias

186 R. Tang et al.

Total

12 388

Events

Total (95% CI) 12 399 Total events 692 821 Heterogeneity: Tau2 = 0.09; Chi2 = 33.26, df = 9 (p = 0.0001); I2 = 73% Test for overall effect: Z = 3.50 (p = 0.0005)

597 227 273 2295 125 4590 15 97 29 4151

Total 588 229 251 2294 135 4588 15 93 34 4161

15 10 11 158 4 316 1 2 4 171

Events

Control

23 23 30 168 21 336 7 11 15 187

Belizan (39) Crowther (21) Kumar (22) Levine (23) Lopez-Jaramillo (5) Nenad (27) Niromanesh (42) Purwar (30) Sanchez-Ramos (41) Villar (24)

Study or subgroup

Calcium supplementation

Table 2. Pooled data of trials.

100.0%

9.6% 8.4% 9.2% 18.8% 5.1% 19.9% 1.7% 2.9% 5.5% 18.9%

Weight [0.34, [0.21, [0.17, [0.76, [0.07, [0.81, [0.02, [0.04, [0.12, [0.75,

1.22] 0.90] 0.66] 1.16] 0.58] 1.09] 1.02] 0.77] 0.84] 1.12] 0.62 [0.47, 0.81]

0.64 0.44 0.34 0.94 0.21 0.94 0.14 0.17 0.31 0.92

Risk ratio M-H, Random, 95% CI

0.01 0.1 1 10 100 Favours calcium Favours control

Risk Ratio M-H, Random, 95% CI

Calcium for the prevention of preeclampsia

187

15 4 2 4 171 11

Events 597 29 97 125 4151 273 5272

Total

158 10

2295 227 2522

15 4590 4605 343

7 336

191

168 23

287

23 15 11 21 187 30

12 388

15 4588 4603

2294 229 2523

588 34 93 135 4161 251 5262

Total

Control Events

Total (95% CI) 12 399 Total events 692 821 Heterogeneity: Tau2 = 0.09; Chi2 = 33.26, df = 9 (p = 0.0001); I2 = 73% Test for overall effect: Z = 3.50 (p = 0.0005) Test for subgroup differences: Chi2 = 1.12, df = 2 (p = 0.57), I2 = 0%

Total events 317 Heterogeneity: Tau2 = 1.27; Chi2 = 3.50, df = 1 (p = 0.06); I2 = 71% Test for overall effect: Z = 0.82 (p = 0.41)

1.2.3 Unknown baseline calcium Niromanesh (42) 1 Nenad (27) 316 Subtotal (95% CI)

Total events 168 Heterogeneity: Tau2 = 0.22; Chi2 = 3.98, df = 1 (p = 0.05); I2 = 75% Test for overall effect: Z = 0.97 (p = 0.33)

1.2.2 High baseline calcium Levine (23) Crowther (21) Subtotal (95% CI)

Total events 207 Heterogeneity:Tau2 = 0.37; Chi2 = 22.31, df = 5(p = 0.0005);I2 = 78% Test for overall effect: Z = 2.88 (p = 0.004)

1.2.1 Low baseline calcium Belizan (39) Sanchez-Ramos (41) Purwar (30) Lopez-Jaramillo (5) Villar (24) Kumar (22) Subtotal (95% CI)

Study or subgroup

Calcium supplementation

Table 3. Subgroup analysis based on baseline calcium intake.

100.0%

1.7% 19.9% 21.6%

18.8% 8.4% 27.2%

9.6% 5.5% 2.9% 5.1% 18.9% 9.2% 51.2%

Weight [0.34, [0.12, [0.04, [0.07, [0.75, [0.17, [0.23,

1.22] 0.84] 0.77] 0.58] 1.12] 0.66] 0.76]

0.62 [0.47, 0.81]

0.14 [0.02, 1.02] 0.94 [0.81, 1.09] 0.48 [0.08, 2.82]

0.94 [0.76, 1.16] 0.44 [0.21, 0.90] 0.70 [0.34, 1.45]

0.64 0.31 0.17 0.21 0.92 0.34 0.42

Risk ratio M-H, Random, 95% CI

2001 2011

1997 1999

1991 1994 1996 1996 2006 2009

Year

0.01 0.1 1 100 10 Favours Calcium Favours Control

Risk Ratio M-H, Random, 95% CI

188 R. Tang et al.

Events 29 125 15 4151 4320

Total 15 21 7 187

Events

597 97 2295 227 273 4590 8079

23 11 168 23 30 336

Total (95% CI) 12 399 Total events 692 821 Heterogeneity: Tau2 = 0.09; Chi2 = 33.26, df = 9 (p = 0.0001); I2 = 73% Test for overall effect: Z = 3.50 (p = 0.0005) Test for subgroup differences: Chi2 = 1.30, df = 1 (p = 0.25), I2 = 22.8%

Total events 512 591 Heterogeneity: Tau2 = 0.08; Chi2 = 18.21, df = 5 (p = 0.003); I2 = 73% Test for overall effect: Z = 2.52 (p = 0.01)

1.4.2 Normal risk of preeclampsia Belizan (39) 15 Purwar (30) 2 Levine (23) 158 Crowther (21) 10 Kumar (22) 11 Nenad (27) 316 Subtotal (95% CI)

12 388

588 93 2294 229 251 4588 8043

34 135 15 4161 4345

Total

Control

Total events 180 230 Heterogeneity: Tau2 = 0.73; Chi2 = 14.75, df = 3 (p = 0.002); I2 = 80% Test for overall effect: Z = 2.00 (p = 0.04)

1.4.1 High risk of preeclampsia Sanchez-Ramos (41) 4 Lopez-Jaramillo (5) 4 Niromanesh (42) 1 Villar (24) 171 Subtotal (95% CI)

Study or subgroup

Calcium supplementation

Table 4. Subgroup analysis based on baseline hypertension risk.

100.0%

9.6% 2.9% 18.8% 8.4% 9.2% 19.9% 68.8%

5.5% 5.1% 1.7% 18.9% 31.2%

Weight

1.22] 0.77] 1.16] 0.90] 0.66] 1.09] 0.91]

0.84] 0.58] 1.02] 1.12] 0.98]

0.62[0.47, 0.81]

0.64[0.34, 0.17[0.04, 0.94[0.76, 0.44[0.21, 0.34[0.17, 0.94[0.81, 0.66[0.48,

0.31[0.12, 0.21[0.07, 0.14[0.02, 0.92[0.75, 0.36[0.13,

Risk ratio M-H, Random, 95% CI

1991 1996 1997 1999 2009 2011

1994 1997 2001 2006

Year

0.01 0.1 1 10 100 Favours calcium Favours control

Risk ratio M-H, Random, 95% CI

Calcium for the prevention of preeclampsia

189

190

R. Tang et al.

Figure 2. Funnel plot of trials.

other meta-analyses accepting pregnancy-induced hypertension as being synonymous with preeclampsia. A comparison of other studies including our own is made in Table 6.

DISCUSSION While preeclampsia remains the most common medical disorder of pregnancy, its pathophysiology remains unclear, further confounding attempts to fully understand the role calcium supplementation may play in its prevention. In this meta-analysis, we found that calcium supplementation of at least 1 g/day during pregnancy versus placebo was associated with an almost 40% decrease (RR 0.62) in the risk of preeclampsia. These benefits appeared most striking in the group of women with either low-baseline calcium intake (RR 0.42) or highbaseline risk of preeclampsia (RR 0.36). However, the beneficial effects of calcium supplementation appear concentrated amongst the seven smaller studies, which all enrolled 51500 women each. When the three large trials (41605 women in each) were analysed, the risk reduction for preeclampsia is both modest (RR 0.93) and not statistically significant. Lack of demonstrated benefit of calcium supplementation in the larger trials [Levine et al.(23), Villar et al. (24), Nenad et al. (27)] is one reason why this cheap and uncomplicated measure has not been widely adopted for preeclampsia prophylaxis. Levine et al.’s (23) conclusions particularly shaped the conclusions of Trumbo & Ellwood (28) (on behalf of the United States Food and Drug Administration), that insufficient evidence exists for calcium supplementation in a US population (28). Several possibilities exist as to why larger studies have reached differing conclusions from smaller studies. Firstly, as postulated by Hofmeyr et al. (7) populations might differ regarding baseline calcium intake and/or baseline preeclampsia risk, with smaller studies recruiting higher risk populations in which calcium supplementation has greater effect (7). Given epidemiological studies that found increased preeclampsia incidence in populations with lowcalcium intake, and that the US population mean dietary calcium intake in women aged 19–30 was adequate at 1367 mg in 2011 (29), it is likely that

15 158 316 171

1.3.1 Large Belizan (39) Levine (23) Nenad (27) Villar (24) Subtotal (95% CI)

597 2295 4590 4151 11 036

Total

23 168 336 187

events

15 10 11 4 1 2 4

597 227 273 125 15 97 29 1363

23 23 30 21 7 11 15

588 229 251 135 15 93 34 1345

9.6% 8.4% 9.2% 5.1% 1.7% 2.9% 5.5% 42.5%

0.0% 18.8% 19.9% 18.9% 57.5%

[0.34, [0.21, [0.17, [0.07, [0.02, [0.04, [0.12, [0.26,

[0.33, [0.76, [0.81, [0.75, [0.84,

1.22] 0.90] 0.66] 0.58] 1.02] 0.77] 0.84] 0.52]

1.23] 1.16] 1.09] 1.12] 1.04]

0.62 [0.47, 0.81]

0.64 0.44 0.34 0.21 0.14 0.17 0.31 0.37

0.63 0.94 0.94 0.92 0.93

Risk ratio Weight M-H, Random, 95% CI

Total (95% CI) 12 399 12 388 100.0% Total events 692 821 Heterogeneity: Tau2 = 0.09; Chi2 = 33.26, df = 9 (p = 0.0001); I2 = 73% Test for overall effect: Z = 3.50 (p = 0.0005) Test for subgroup differences: Chi2 = 25.21, df = 1 (p50.00001), I2 = 96.0%

Total events 47 130 Heterogeneity: Tau2 = 0.01; Chi2 = 6.41, df = 6 (p = 0.38); I2 = 6% Test for overall effect: Z = 5.63 (p = 0.00001)

1.3.2 Small Belizan (39) Crowther (21) Kumar (22) Lopez-Jaramillo (5) Niromanesh (42) Purwar (30) Sanchez-Ramos (41) Subtotal (95% CI)

588 2294 4588 4161 11 043

Total

Control

Total events 645 691 Heterogeneity: Tau2 = 0.00; Chi2 = 0.04, df = 2 (p = 0.98); I2 = 0% Test for overall effect: Z = 1.29 (p = 0.20)

Events

Study or subgroup

Calcium supplementation

Table 5. Subgroup analysis based on study size.

Risk ratio M-H, Random, 95% CI

Calcium for the prevention of preeclampsia

191

10 (11 405 women)

16 (21 079 women)

10 (24 787 women)

Imdad et al. (35)

Patrelli et al. (36)

This review

Databases

Medline/Pubmed, Cochrane, Embase

Medline/Pubmed

Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, handsearch, weekly current awareness of selected journals Pubmed, Cochrane Library, WHO regional databases

PubmedElsevier ScienceSCI (1991– 2011)

Assuming definition used in this meta-analysis.

13 (15 730 women)

Hofmeyr et al. (7)

a

4 (14 537 women)

Studies included

An et al. (38)

Study

0.88 [0.77–1.02] in women with adequate calcium intake 0.73 [0.61–0.87] in women with low calcium intake 0.62 [0.47, 0.81]

0.41 [0.24–0.69]

0.45 [0.31–0.65]

0.90 [0.79–1.04]

Risk ratio of calcium versus placebo [95% CI]

Table 6. Overview of contemporary meta-analyses with regards to calcium in preventing preeclampsia.

Inclusion of two studies with inappropriate interventions and one with an inappropriate outcomea. Inclusion of three studies with inappropriate interventions and two with inappropriate outcomesa.

Included criteria were nulliparous women without hypertension, diabetes mellitus or renal disease. Inclusion of three studies with inappropriate outcomesa.

Additional notes

192 R. Tang et al.

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calcium supplementation had a much smaller effect in at least the Levine et al. population, where other processes may have been the primary cause of preeclampsia. No data concerning dietary calcium intake of Slovakian women, whom were studied by Nenad and colleagues, could be found, though it is possible that a similar process resulted in non-significant reduction of preeclampsia following calcium supplementation. In contrast, the third large multicentre trial (24), on behalf of the WHO in populations with a low-baseline calcium intake, demonstrated a non-significant decrease in preeclampsia following calcium supplementation. The results from this study contrast the findings of the smaller studies of Kumar et al. (22) and Purwar et al. (30), who both demonstrated that a more substantial decrease in preeclampsia incidence in an Indian population is characterised by low-baseline intake that was also studied by the WHO study (22,24,30). However, as acknowledged by Villar and colleagues, women studied in their trial were assumed to have low-calcium intakes, rather than measuring their respective dietary intakes. This may be significant as the urban location (and therefore overall nutritional status of the women studied) of these six hospitals may have played a role. The other major explanation for why large studies do not demonstrate significant benefit of calcium supplementation, while smaller studies do relates to publication bias. The bias towards publication of studies with positive results is well documented (31–33). Our meta-analysis did not identify any negative small studies or positive large studies, leading to an asymmetrical funnel plot (Figure 2), which is highly suspicious for publication bias (34). While it is recognised that trial registries make it easier to screen for publication bias, most small included studies were published more than 10 years ago, prior to many universal trial registration systems, so ‘‘missing’’ small studies with negative results dating from this time period would likely not be registered. Search using ClinicalTrial.gov (by the US National Institutes of Health) and International Clinical Trials Registry Platform (by the WHO) failed to find any small negative studies. With regards to prior contemporary reviews of preeclampsia incidence following calcium supplementation, most have also found overall significant reductions when calcium supplementation is used (7,35,36). This review includes slightly fewer studies, but includes the largest total number of women amongst meta-analyses published to date due to the large recent study of Nenad and colleagues 27) which, to the authors’ knowledge, has not been previously included in any other meta-analysis (27). However, due to the significant difference noted between the large and small trial results, which has not been discussed previously, we would caution against accepting the pooled trials as definitive evidence of benefit of calcium supplementation in preeclampsia. This review also differs from previous meta-analyses by not accepting pregnancy-induced hypertension as being synonymous with preeclampsia, believing the two to be fundamentally different pathological processes. Using the RCOG’s definition of preeclampsia and requiring calcium be compared with a placebo, careful evaluation of Patrelli et al. (36) and Imdad et al. (35) found that the respective authors likely erred on several minor points, with Patrelli et al. (36) merging data of pregnancy-induced hypertension with

193

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preeclampsia and Imdad et al. (35) including studies with inappropriate controls. How this review compares to other reviews is briefly summarised in Table 6. This difference in the definition of preeclampsia is probably the basis for the differing results in our meta-analysis. With regards to subgroup analysis, our review concurs with Patrelli et al. (36), Imdad et al. (35) and Hofmeyr et al. (7), in finding a decreased risk ratio of preeclampsia following calcium supplementation in high-risk women and in women with low-calcium intake (RR 0.49 [95% CI 0.25–0.93] and 0.36 [0.10–1.35] respectively) (7,35,36). This supports the notion that calcium supplementation should be offered to women at high risk of preeclampsia or has a low-baseline calcium intake. A final meta-analysis to consider is one conducted by An et al. (37) who reported a smaller risk reduction following calcium administration, using extremely stringent criteria by only accepting trials that studied nulliparous women without hypertension, diabetes mellitus or renal disease. Given that the majority of trials found in this review tended not to report these conditions, only four studies (n = 14 537) were included (37). However, given that this analysis was presented as a poster abstract only, additional analysis of this article proved difficult. While the overall quality of this meta-analysis is likely to be high, it possibly occurs at the cost of external validity in reporting findings substantially different from other meta-analyses. Finally, Simon and colleagues have recently analysed preeclampsia trial data, finding that the majority of randomised controlled trials mixed multiparous women with primiparous individuals, in turn suggesting a major confounding factor may have been overlooked (38). All trials included in this study required that women be nulliparous or primigravid, largely circumventing this problem. Pregnancies that were terminated after 20 weeks may represent a small confounding factor, though this problem is likely to be small. Use of the cut-off n = 1605 in each arm lead to the exclusion of a positive large study conducted by Belizan et al. (39) from the large study subgroup. Its addition, however, is of little significance, reducing the RR of the large study subgroup from 0.93 [95% CI 0.83–1.04] to 0.92 [95% CI 0.82–1.03] in a separate analysis using the methods outlined earlier. Strengths of this review include a standardised methodology using Revman, inclusion of recent data, and use of a rigorous definition of preeclampsia. A key limitation of this review is that secondary outcomes were not studied in this meta-analysis. Previous work by Hofmeyr et al. (7) and Imdad et al. (35) found that calcium supplementation in pregnancy-reduced maternal mortality and morbidity; and preterm delivery. Incidence of admission to the neonatal ICU, placental abruption, caesarean section, neonates being small-for-gestational age and HELLP syndrome were not, however, reduced.

IMPLICATIONS FOR FUTURE RESEARCH Although calcium supplementation during pregnancy appears effective for preeclampsia prevention, particularly in women with high-baseline preeclampsia risk or low-baseline calcium intake, the relative lack of effect seen in large versus small clinical trials means caution should be applied regarding recommending calcium supplementation for all pregnant women. Identification of trials performed but not published through trial registries, grey literature and

Calcium for the prevention of preeclampsia

enquiries of authors of existing trials regarding their knowledge of nonpublished work would be helpful in resolving concerns around publication bias. If further large trials do occur, then analysis with regards to confounding factors, particularly the relative contributions to preeclampsia prevention of aspirin versus calcium, should occur. As calcium supplementation appears more effective in high-risk subgroups, future research should also focus on determining the aetiology of the disease within different population subgroups to allow better targeting of prophylactic therapy. From a molecular standpoint, biochemical studies by Hache et al. (20), Jeung et al. (18) and Yang et al. (19) suggest that calcium metabolism is impaired in preeclamptic syncytiotrophoblast, and correction or amelioration of this defect may underlie calcium supplementation’s beneficial effect (18–20). Finally, future trials may aim to quantify calcium intake versus preeclampsia risk, as while some studies have shown retrospectively depressed serum calcium levels in women in preeclampsia, no trials to the authors’ knowledge have yet prospectively demonstrated this (13,40).

CONCLUSION Calcium appears effective at reducing the incidence of preeclampsia. Its effect is of particular use in women at high risk of developing preeclampsia and women with low-baseline calcium intake. Its utility in the general pregnant population remains less clear, given that the largest randomised trials do not show clear evidence of benefit, and meta-analysis suggests likely small-trial publication bias.

DECLARATION OF INTEREST No potential conflicts of interest are declared.

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10. Gabbay A, Tzur T, Weintraub AY, et al. Calcium level during the first trimester of pregnancy as a predictor of preeclampsia. Hypertens Pregnancy 2014;33:311–21. 11. Belizan JM, Campodonico L. Calcium supplementation and toxaemia of pregnancy. Gynecol Obstet Invest 1988;26:341–2. 12. Beliza´n J, Villar J, Repke J. The relationship between calcium intake and pregnancyinduced hypertension: up-to-date evidence. Am J Obstet Gynecol 1988;158:898–902. 13. Repke JT, Villar J, Anderson C, et al. Biochemical changes associated with blood pressure reduction induced by calcium supplementation during pregnancy. Am J Obstet Gynecol 1989;160:684–90. 14. Carroli G, Merialdi M, Wojdyla D, et al. Effects of calcium supplementation on uteroplacental and fetoplacental blood flow in low-calcium-intake mothers: a randomized controlled trial. Am J Obstet Gynecol 2010;202:45.e1–e9. 15. Sumners C, Zhu M, Gelband CH, Posner P. Angiotensin II type 1 receptor modulation of neuronal K+ and Ca2+ currents: intracellular mechanisms. Am J Physiol Cell Physiol 1996;271:C154–63. 16. Zhou CC, Zhang Y, Irani RA, et al. Angiotensin receptor agonistic autoantibodies induce pre-eclampsia in pregnant mice. Nat Med 2008;14:855–62. 17. Siddiqui AH, Irani RA, Blackwell SC, et al. Angiotensin receptor agonistic autoantibody is highly prevalent in preeclampsia correlation with disease severity. Hypertens 2010;55:386–93. 18. Jeung EB, Yang H. Membrane and cytosolic calcium proteins, TRPV6, PMCA1, NCKX3, NCX1 and CaBP-28k, appear to be distinctly regulated in human choriocarcinoma and placental cells. Reprod Fertil Dev 2012;24:153. 19. Yang H, Kim TH, An BS, et al. Differential expression of calcium transport channels in placenta primary cells and tissues derived from preeclamptic placenta. Mol Cell Endocrinol 2013;367:21–30. 20. Hache S, Takser L, Lebellego F, et al. Alteration of calcium homeostasis in primary preeclamptic syncytiotrophoblasts: effect on calcium exchange in placenta. J Cell Mol Med 2011;15:654–67. 21. Crowther CA, Hiller JE, Pridmore B, et al. Calcium supplementation in nulliparous women for the prevention of pregnancy-induced hypertension, preeclampsia and preterm birth: an Australian randomized trial. Aust N Z J Obstet Gynaecol 1999; 39:12–18. 22. Kumar A, Devi SG, Batra S, et al. Calcium supplementation for the prevention of pre-eclampsia. Int J Gynaecol Obstet 2009;104:32–6. 23. Levine RJ, Hauth JC, Curet LB, et al. Trial of calcium to prevent preeclampsia. N Engl J Med 1997;337:69–76. 24. Villar J, Abdel-Aleem H, Merialdi M, et al. World Health Organization randomized trial of calcium supplementation among low calcium intake pregnant women. Am J Obstet Gynecol 2006;194:639–49. 25. NHMRC. Nutrient reference values for Australia and New Zealand including recommended dietary intakes. Canberra: Australian Government: Department of Health and Ageing; 2005. 26. Myatt L, Redman CW, Staff AC, et al. Strategy for standardization of preeclampsia research study design. Hypertension 2014;63:1293–301. 27. Nenad S, Olivera KV, Goran R, Ljiljana S. Did calcium management prevent preeclampsia? Pregnancy Hypertens 2011;1:287. 28. Trumbo PR, Ellwood KC. Supplemental calcium and risk reduction of hypertension, pregnancy-induced hypertension, and preeclampsia: an evidence-based review by the US Food and Drug Administration. Nutr Rev 2007;65:78–87. 29. Mangano KM, Walsh SJ, Insogna KL, et al. Calcium intake in the United States from dietary and supplemental sources across adult age groups: new estimates from the National Health and Nutrition Examination Survey 2003–2006. J Am Diet Assoc 2011;111:687–95. 30. Purwar M, Kulkarni H, Motghare V, Dhole S. Calcium supplementation and prevention of pregnancy induced hypertension. J Obstet Gynaecol Res 1996;22: 425–30.

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31. Stern JM, Simes RJ. Publication bias: evidence of delayed publication in a cohort study of clinical research projects. BMJ 1997;315:640–5. 32. Dwan K, Altman DG, Arnaiz JA, et al. Systematic review of the empirical evidence of study publication bias and outcome reporting bias. PLoS One 2008;3:e3081. 33. Callaham ML, Wears RL, Weber EJ, et al. Positive-outcome bias and other limitations in the outcome of research abstracts submitted to a scientific meeting. JAMA 1998;280:254–7. 34. Egger M, Smith GD, Schneider M, Minder C. Bias in meta-analysis detected by a simple, graphical test. BMJ 1997;315:629–34. 35. Imdad A, Jabeen A, Bhutta ZA. Role of calcium supplementation during pregnancy in reducing risk of developing gestational hypertensive disorders: a meta-analysis of studies from developing countries. BMC Public Health 2011;11:S18. 36. Patrelli TS, Dall’Asta A, Gizzo S, et al. Calcium supplementation and prevention of preeclampsia: a meta-analysis. J Matern Fetal Neonatal Med 2012;25:2570–4. 37. An L, Li W, Xie S, Xu J. Calcium supplementation reducing risk of hypertensive disorders complicating pregnancy: a meta analysis of multi-center RCTs. Circulation 2012;125:e714. 38. Simon E, Caille A, Perrotin F, Giraudeau B. Mixing nulliparous and multiparous women in randomised controlled trials of preeclampsia prevention is debatable: evidence from a systematic review. PLoS One 2013;8:e66677. 39. Beliza´n JM, Villar J, Gonzalez L, et al. Calcium supplementation to prevent hypertensive disorders of pregnancy. N Engl J Med 1991;325:1399–405. 40. Proverbio F, Chiarello DI, Pinero S, et al. Changes in the Ca2+ or the Mg2+ content of basal and microvillous membranes of the human placenta syncytiotrophoblast and their impact on levels of lipid peroxidation and Ca-ATPase activity. Placenta 2013;34:A41. 41. Sanchez-Ramos L, Briones DK, Kaunitz AM, et al. Prevention of pregnancyinduced hypertension by calcium supplementation in angiotensin II-sensitive patients. Obstet Gynecol 1994;84:349–53. 42. Niromanesh S, Laghaii S, Mosavi-Jarrahi A. Supplementary calcium in prevention of pre-eclampsia. Int J Gynaecol Obstet 2001;74:17–21.

APPENDIX 1. TRIALS NOT INCLUDED IN ANALYSIS Cong 1995

Cong K, Chi S, Liu G. Calcium supplementation during pregnancy for reducing pregnancy induced hypertension. Chin Med J 1995;108:57–9. Only studied pregnancy-induced hypertension without investigating preeclampsia as an outcome. [Inappropriate outcome]. Hall 2000 Hall DR, Odendaal HJ, Steyn DW, Grove D. Expectant management of early onset, severe preeclampsia: maternal outcome. Br J Obstet Gynaecol 2000;107:1252–7. Did not study the role of calcium supplementation in the study. [Inappropriate intervention]. Herrera 1993 Herrera JA. Nutritional factors and rest reduce pregnancy-induced hypertension and pre-eclampsia in positive roll-over test primigravidas. Int J Gynaecol Obstet 1993;41:31–5. Studied calcium supplementation and conjugated linoleic acid. [Inappropriate intervention]. Herrera 2006 Herrera JA, Arevalo-Herrera M, Shahabuddin AK, et al. Calcium and conjugated linoleic acid reduces pregnancy-induced hypertension and decreases intracellular calcium in lymphocytes. Am J Hypertens 2006;19:381–7. Studied calcium supplementation (300 mg/day) in addition to soy proteins (25 g/day) and linoleic acid (300 mg/day). [Inappropriate intervention].

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Hiller 2007 Hiller JE, Crowther CA, Moore VA, et al. Calcium supplementation in pregnancy and its impact on blood pressure in children and women: follow up of a randomised controlled trial. Aust N Z J Obstet Gynaecol 2007;47:115–21. Only studied the effects of calcium supplementation on maternal and child blood pressure. [Inappropriate outcome]. Ito 1994 Ito M, Koyama H, Ohshige A, et al. Prevention of preeclampsia with calcium supplementation and vitamin D3 in an antenatal protocol. Int J Gynaecol Obstet 1994;47:115–20. Studied calcium supplementation (156 or 312 mg/day) and vitamin D3. [Inappropriate intervention]. Lopez-Jaramillo 1989 Lopez-Jaramillo P, Narvaez M, Weigel RM, Yepez R. Calcium supplementation reduces the risk of pregnancy-induced hypertension in an Andes population. Br J Obstet Gynaecol 1989;96:648–55. This study only studied the effects of calcium supplementation on the effects of pregnancy-induced hypertension, without investigating preeclampsia as an outcome. [Inappropriate outcome]. Marcoux 1991 Marcoux S, Brisson J, Fabia J. Calcium intake from dairy products and supplements and the risks of preeclampsia and gestational hypertension. Am J Epidemiol 1991;133:1266–72. This was a case–control study. [Inappropriate methodology]. Rogers 1999 Rogers MS, Fung HY, Hung CY. Calcium and low-dose aspirin prophylaxis in women at high risk of pregnancy-induced hypertension. Hypertens Pregnancy 1999;18:165–72. This trial investigated the role of calcium supplementation and aspirin prophylaxis. [Inappropriate intervention] Taherian 2002 Taherian AA, Taherian, A Shirvani A. Trial of aspirin and calcium on prevention of preeclampsia. J Res Med Sci 2002;7:136–40. This trial investigated the role of calcium supplementation and aspirin prophylaxis. [Inappropriate intervention]. Villar 1987 Villar J, Repke J, Belizan JM Pareja G. Calcium supplementation reduces blood pressure during pregnancy: results of a randomized controlled clinical trial. Obstet Gynecol 1987;70:317–22. This trial only evaluated the effects of calcium supplementation on blood pressure during pregnancy without studying the effects on preeclampsia. [Inappropriate outcome]. Villar 1990 Villar J, Repke JT. Calcium supplementation during pregnancy may reduce preterm delivery in high-risk populations. Am J Obstet and Gynecol 1990;163:1124–31. This trial only evaluated the effects of calcium supplementation on blood pressure and other selected outcomes during pregnancy without studying the effects of preeclampsia. [Inappropriate outcome]. Wanchu 2001 Wanchu M, Malhotra S, Khullar M. Calcium supplementation in pre-eclampsia. J Assoc Physicians India 2001;49:795–8. This trial investigated the role of calcium supplementation in preeclampsia without comparison to a placebo. [In appropriate intervention]

Calcium for the prevention of preeclampsia APPENDIX 2. Critical appraisal of selected trials.

Bias Belizan 1991 Random sequence generation (selection bias)

Authors’ judgement

Allocation concealment (selection bias)

Low risk

Blinding of participants and personnel (performance bias)

Low risk

Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias) Selective reporting (reporting bias) Other bias

Low risk

Randomisation was performed at each hospital by a random-sample generator program provided by the Epistat Statistical Package. Randomisation codes were sent to each hospital in numbered, sealed, opaque envelopes. Women were allocated a randomisation number such that they would only receive medication from a central unit. Personnel involved in her treatment were thus unaware of the woman’s treatment status. As above.

Low risk

All or partial data available for 544/593.

Low risk

All primary outcomes addressed.

Low risk

None detected.

Low risk

Randomisation was performed by the drug company with stratification made by centre using variable blocks. Identically sealed treatment packs were prepared by the drug company. Each treatment pack contained identically labelled drug containers of either 600 mg tablets of calcium or placebo tablets of lactose, identical in size, colour and consistency. The code was held centrally and only broken after the analysis had ended.

Crowther 1999 Random sequence generation (selection bias) Allocation concealment (selection bias) Blinding of participants and personnel (performance bias)

Low risk

Support for judgement

Low risk Low risk

Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias)

Low risk

Selective reporting (reporting bias) Other bias

Low risk

Kumar 2009 Random sequence generation (selection bias) Allocation concealment (selection bias)

Low risk

Low risk

Recruitment resulted in only 456 of 948 needed due to lack of funds, potentially affecting the power of this study. Intention-to-treat analysis 227 in calcium group versus 229 in placebo group. None detected

Low risk

Simple randomization sequence developed manually.

Low risk

Treatment packets were then distributed to the participants using the random numbers in sequence. (Continued )

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Bias

Authors’ judgement

Blinding of participants and personnel (performance bias)

Low risk

Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias)

Low risk

Selective reporting (reporting bias)

Unclear risk

Other bias

Low risk

Low risk

Levine 1997 (on behalf of CPEP) Random sequence Low risk generation (selection bias)

Allocation concealment (selection bias)

Low risk

Blinding of participants and personnel (performance bias) Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias) Selective reporting (reporting bias)

Low risk

Other bias

Unclear risk

Lopez-Jaramillo 1997 Random sequence generation (selection bias) Allocation concealment (selection bias) Blinding of participants and personnel (performance bias)

Support for judgement Double blind. Tablets appeared identical in appearance. The randomization code was disclosed only after completion of the study. As above.

273/290 women were analysed in the calcium group, and 251/262 women were analysed in the placebo group. An imbalance between groups was noted at 273 in the intervention group versus 251 in the placebo group. None detected. Packages of study tablets were prepared and numbered by the pharmaceutical manufacturer according to a computergenerated simple randomization sequence developed by the study statisticians. Each woman was assigned the next numbered package of medication, contained in identically looking blister packs. The study was ‘‘double-blinded’’ with all codes held centrally.

Low risk

As above.

Low risk

Compliance was measured when women returned dispensed blister packs. The authors failed to report the total number of women enrolled in the study. Some discrepancies in the overall number between the control and intervention group were noted, but were unlikely to cause significant distortion to the authors’ results. None detected.

Low risk

Low risk

Low risk

A random number table was used to assign each patient independently in sequence. Tablets were identical in appearance.

Low risk

Double-blind.

(Continued )

Calcium for the prevention of preeclampsia APPENDIX 2. Continued

Bias

Authors’ judgement

Support for judgement

Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias)

Low risk

As above.

Low risk

Selective reporting (reporting bias) Other bias

Low risk

260 of 274 recruited were analysed. Only women with no missing values for any of the covariate and outcome variables were included. As above.

Low risk

None detected.

Unclear risk

‘‘Randomly assigned’’, no other details provided.

Unclear risk

Not stated.

Low risk

Low risk

Surveillance for preeclampsia was conducted by personnel unaware of treatment-group assignments. As above.

Unclear risk

Attrition was not reported.

Unclear risk

As above.

Unclear risk

This trial was presented as a poster abstract. A large amount of data relating to study design was not reported.

Unclear risk

Women were ‘‘randomly assigned’’

Low risk

Low risk

Tablets had identical packaging and physical characteristics. All pills were dispensed by the pharmacy. Double-blind

Low risk

As above.

Low risk

No incomplete outcome data.

Low risk

As above.

Low risk

None detected.

Nenad 2011 Random sequence generation (selection bias) Allocation concealment (selection bias) Blinding of participants and personnel (performance bias) Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias) Selective reporting (reporting bias) Other bias

Niromanesh 2001 Random sequence generation (selection bias) Allocation concealment (selection bias) Blinding of participants and personnel (performance bias) Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias) Selective reporting (reporting bias) Other bias

(Continued )

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Bias Purwar 1996 Random sequence generation (selection bias) Allocation concealment (selection bias) Blinding of participants and personnel (performance bias) Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias) Selective reporting (reporting bias) Other bias Sanchez-Ramos 1994 Random sequence generation (selection bias) Allocation concealment (selection bias) Blinding of participants and personnel (performance bias) Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias) Selective reporting (reporting bias) Other bias

Authors’ judgement

Support for judgement

Low risk

Women were assigned using a computer generated randomization list.

Low risk Low risk

Tablets had identical physical characteristics. Double-blind.

Low risk

As above.

Low risk

No incomplete outcome data.

Low risk

As above.

Low risk

None detected.

Low risk

Randomisation by computer-generated list.

Low risk

‘‘Double-blind’’.

Low risk

Double blind. Tablets were physically identical.

Low risk

As above.

Low risk Low risk

4/33 individuals in the calcium group were lost to follow up. As above.

Low risk

None detected.

Villar 2006 (on behalf of the WHO) Random sequence Low risk generation (selection bias)

Randomisation was performed independently for each study sight by the Clinical Trials Unit at WHO/HRP Geneva, Switzerland, using computer-generated random number blocking with randomly varying groups of 6 to 8 women. Randomisation within strata was also restricted via the allocation of consecutively numbered treatment boxes for each woman. (Continued )

Calcium for the prevention of preeclampsia APPENDIX 2. Continued

Bias

Authors’ judgement

Support for judgement

Allocation concealment (selection bias)

Low risk

Blinding of participants and personnel (performance bias)

Low risk

Blinding of outcome assessment (detection bias) Incomplete outcome data (attrition bias)

Low risk

Selective reporting (reporting bias) Other bias

Low risk

Delivery data were missing for 143 women in the calcium group and 155 women in the placebo group. None detected.

Low risk

None detected.

Low risk

Boxes and tablet bottles were prepared and numbered. Complete sets, all of which were identical, were shipped by WHO to centers and kept in a designated area with a locker. Individual treatment boxes were provided consecutively to the randomized subject. Randomization codes remained at the WHO Clinical Trial Unit until the time of analysis and were not available to any person until the analyses were completed. As above.

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