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Summary

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References

Objective

To investigate whether the live birth rate following in vitro fertilization (IVF) is affected by thyroid autoimmunity (TAI) and/or subclinical hypothyroidism in subfertile women.

Design and setting

Retrospective study in a university infertility clinic.

Patients

A total of 627 women without past or current history of thyroid disorder undergoing their first IVF cycle.

Intervention

Pre-IVF archived blood serum samples were tested for TAI and thyroid function tests.

Main outcome measure

Live birth rate.

Results

The clinical pregnancy rate, live birth rate and miscarriage rate were similar among women with or without TAI and/or subclinical hypothyroidism using a TSH threshold 4·5 mIU/l. Thyroid autoantibody level did not affect these IVF outcomes.

Conclusion

The live birth rate and miscarriage rate of women with TAI and/or subclinical hypothyroidism following IVF were not impaired.


Introduction

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References

Hypothyroidism is associated with a broad spectrum of reproductive problems, including menstrual irregularities, impaired ovulation, miscarriage and late pregnancy complications.[1, 2] Because of these potential consequences, subclinical hypothyroidism and thyroid autoimmunity (TAI), which are closely related to hypothyroidism, have been a topic of active investigation and controversy.

Thyroid autoimmunity, characterized by the presence of antithyroglobulin and/or antithyroid peroxidase antibodies, affects 5–20% of women of childbearing age. It is the main cause of hypothyroidism/subclinical hypothyroidism although it can be present without thyroid dysfunction.[3] Subclinical hypothyroidism occurs when serum TSH concentrations are raised and serum T4 concentrations are normal. There is no consensus among endocrinologists regarding the cut-off value for TSH in subclinical hypothyroidism, although the Endocrine Society suggests that TSH levels should be <2·5 mIU/l in the first trimester of pregnancy to avoid adverse outcomes.[4]

Both TAI and subclinical hypothyroidism have independently been associated with adverse pregnancy outcomes in different trimesters including miscarriage, placental abruption, preterm birth, pre-eclampsia and an increased risk of perinatal mortality.[4-6] Women with TAI with or without subclinical hypothyroidism were more frequently found to have subfertility problems than controls.[7] When these women undergo assisted reproductive technologies (ART), the rapid rise in oestradiol concentrations may stress the hypothalamic–pituitary–thyroid axis.[8] A recent review found that the evidence regarding the effect of ovarian stimulation on thyroid function or TAI was inconclusive.[9] TSH levels were found to be inversely proportional to the fertilization rate at ART,[10] and women with TAI undergoing in vitro fertilization (IVF) had poorer pregnancy outcomes when compared with controls.[11] However, Michalakis et al.[12] found that preconception subclinical hypothyroidism was not associated with adverse ART or pregnancy outcomes. Negro et al.[13] showed that it was the TSH level rather than the presence of thyroid antibodies that affected the ART/pregnancy outcomes while Kim et al.[14] found evidence that the level of thyroid antibodies might also play a role.

The relevance of preconception TAI and subclinical hypothyroidism in infertile patients undergoing ART remains controversial. Currently, there is a paucity of studies looking into the possible association between these conditions and live birth rates, which is considered to be the ultimate aim of IVF treatment. The objective of this retrospective study was to examine the possible relationship between pre-IVF TAI and/or subclinical hypothyroidism and the live birth rates following IVF treatment.

Materials and methods

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References

This was a retrospective study carried out at the Centre of Assisted Reproduction and Embryology, The University of Hong Kong – Queen Mary Hospital, Hong Kong. Clinical details of all treatment cycles were prospectively entered into a computerized database, which were checked for accuracy and completeness on a regular basis and were retrieved for analysis. Ethical approval was obtained from the Institutional Review Board of the University of Hong Kong/Hospital Authority Hong Kong West Cluster for this retrospective study.

Patients

We analysed data of infertile women who underwent first IVF/ICSI cycles between February 2007 and December 2009. Only those women who had archived serum samples that were taken 6 months prior to ovarian stimulation were included. Women with a known history of thyroid disease (whether or not on medication) and cycles carried out for pre-implantation genetic diagnosis or those using donor oocytes were excluded. The archived serum samples of the included women were retrieved and assayed for serum antithyroid peroxidase antibodies, antithyroglobulin antibodies, TSH and free T4 concentrations.

Ovarian stimulation and embryo transfer

All women were treated either with the long GnRH agonist protocol or the GnRH antagonist protocol for pituitary down-regulation. The details of the long protocol of ovarian stimulation regimen, gamete handling, standard insemination and intracytoplasmic sperm injection (ICSI) were as previously described.[15] In short, women received buserelin (Suprecur®; Hoechst, Frankfurt, Germany) nasal spray 150 μg four times a day starting from the mid-luteal phase of the cycle preceding the treatment cycle and received hMG or recombinant FSH for ovarian stimulation following the menstrual period. In the GnRH antagonist protocol, after confirming a basal serum oestradiol level, ovarian stimulation was started with either hMG or recombinant FSH, and ganirelix 250 μg was started from the sixth day of stimulation. The doses used for stimulation were adjusted according to the baseline antral follicle count.

Transvaginal ultrasonography together with measurement of blood oestradiol was used to assess the ovarian response. When the mean diameter of the leading follicle reached 18 mm and there were at least three follicles reaching a mean diameter of 16 mm or more, HCG [Pregnyl® (Organon, Oss, Holland) 5000 or 10000 units or Ovidrel® (Merck Sernono S.p.A., Modugno, Italy) 250 μg] was injected on the same day and oocytes were collected about 36 h later. Fertilization was carried out in vitro either by conventional insemination or by ICSI depending on semen parameters. Women were allowed to have replacement of at most two embryos 2 days after oocyte retrieval. Progesterone pessaries (Endometrin 100 mg twice per day; Ferring Pharmaceuticals, Parsippany, NJ, USA) were administered from the day of embryo transfer for 2 weeks for luteal support. Pregnancies were confirmed by positive urine HCG tests and transvaginal ultrasonographic evidence of a gestational sac.

Collection of clinical information

Clinical information including age, body mass index, basal serum levels of follicle-stimulating hormone and anti-Mullerian hormone were collected. During the IVF treatment, data including days of stimulation, total dosage of gonadotrophin, oestradiol level on day of hCG, number of oocytes retrieved, fertilization rate, number of available embryos, pregnancy rate and miscarriage rate were recorded.

Clinical pregnancy was defined as the presence of a gestational sac by ultrasonography, whereas miscarriage rate per clinical pregnancy was defined as the proportion of patients who failed to continue development to 20 weeks of gestation in all clinical pregnancies. Pregnancy outcome was collected from all pregnant women by postal questionnaire or by phone. Live birth was defined as the delivery of a foetus with signs of life after 24 completed weeks of gestational age.

Determination of thyroid antibodies and hormone levels

Thyroid antibodies were tested using the Access TPO Antibody Reagent Pack and Access Thyroglobulin Antibody II Reagent Pack (Beckman Coulter Inc., Brea, CA, USA). Serum TSH and free T4 were measured using the Access HYPERsensitive hTSH Reagent Pack and Access Free T4 Reagent Pack (USG; Beckman Coulter), respectively. The reference ranges and intra- and interassay coefficients of variation were as follows: TPO antibodies <9 IU/ml (intra-assay CV 5·7% and interassay CV 5·2%); Tg antibodies <4 IU/ml (intra-assay CV 6·5% and interassay CV 4·4%); TSH 0·34–5·60 mIU/l (intra-assay CV 2·49% and interassay CV 2·76%); free T4 7·86–14·41 pm (intra-assay CV 1·82% and interassay CV 4·71%). TAI was defined as having either positive antithyroid peroxidase antibody or antithyroglobulin antibody.

Statistical analysis

Women with overt hypothyroidism, overt hyperthyroidism and subclinical hyperthyroidism were excluded for analysis. Subclinical hypothyroidism was defined as an elevated serum TSH level ≥2·5 mIU/l with a normal free T4 level.[4]

The primary outcome measure was the live birth rate in the first cycle. Statistical analysis for the comparisons of mean values was performed using Mann–Whitney test, Kruskal–Wallis test or Student's t-test, as appropriate. The chi-squared test and Fisher's exact test were used for the comparisons of categorical variables. Statistical analysis was carried out using the Statistical Program for Social Sciences (Version 20.0; SPSS Inc., Chicago, IL, USA). The two-tailed value of P < 0·05 was considered statistically significant.

Results

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References

A total of 656 women underwent their first IVF cycle during the study period, and 20 women were excluded because of past or current history of thyroid disorders.

The mean age was 35·3 ± 3·2 years (mean ± SD) with a TSH level of 1·84 ± 1·3 mIU/l (median, 1·59 mIU/l). The prevalence of TAI and subclinical hypothyroidism was 19·7% (125/636) and 18·7% (119/636). Twenty-six per cent of TAI-positive women had coexisting subclinical hypothyroidism when compared with 16·8% of TAI-negative women. Table 1 showed the prevalence of each thyroid status.

Table 1. Prevalence of thyroid disorders
Thyroid statusTAI positive (n = 125) (%)TAI negative (n = 511) (%)
  1. TAI, thyroid autoimmunity.

Euthyroid89 (71·2)419 (82·0)
Subclinical hypothyroidism33 (26·4)86 (16·8)
Hypothyroidism0 (0)0 (0)
Subclinical hyperthyroidism1 (0·8)2 (0·4)
Hyperthyroidism2 (1·6)4 (0·8)

Only euthyroid women and those with subclinical hypothyroidism were included for analysis (n = 627), and these were divided into four different groups according to their thyroid and TAI status. They were comparable with respect to patient characteristics, such as age, body mass index, basal FSH and anti-Mullerian hormone levels (Table 2). TSH levels, as expected, were significantly higher in women with subclinical hypothyroidism when compared with those who were euthyroid. TSH levels ranged from 2·52 to 16·82 mIU/l for patients with subclinical hypothyroidism. There were no significant differences among the four groups with respect to the cause of infertility, the duration of infertility, the type of protocol used and the requirement for ICSI. The response of ovarian stimulation results in terms of the duration of stimulation, the total dosage of gonadotrophins, the oestradiol level on the day on HCG administration, the numbers of oocytes retrieved and fertilization rate was similar among the four groups (data not shown). Clinical pregnancy, miscarriage rate and live birth rates were also similar among the groups (Table 3). Clinical outcomes were analysed again using different TSH thresholds and showed that there were no differences in the clinical pregnancy, live birth or miscarriage rates using thresholds of 2·5, 3·5 and 4·5 mIU/l (Table 4).

Table 2. Patient characteristics
 TAI positiveTAI negativeP-value
EuthyroidSubclinicalEuthyroidSubclinical
  1. TAI, thyroid autoimmunity.

  2. Data expressed as mean ± SD or mean (range) or number (%).

  3. a

    TSH level was significantly higher in subclinical than in euthyroid group.

Patients, n893341986 
Age (year)35·3 ± 3·134·5 ± 3·835·4 ± 3·334·9 ± 3·0NS
BMI (kg/m2)21·1 ± 3·021·4 ± 2·721·6 ± 2·721·7 ± 2·7NS
Basal FSH (IU/l)8·7 ± 3·611·7 ± 15·58·7 ± 4·08·2 ± 3·1NS
AMH (μg/l)3·9 ± 3·04·9 ± 3·63·6 ± 3·44·2 ± 3·5NS
TSH (mIU/l)1·5 (0·35–2·47)4·0 (2·53–10·71)1·4 (0·32–2·50)3·6 (2·52–16·82)<0·01a
Free T4 (pm)14·1 ± 2·113·2 ± 2·014·2 ± 1·914·1 ± 2·1NS
Indication for in vitro fertilization
Endometriosis5 (5·6)0 (0)14 (3·3)5 (5·8)NS
Tuboperitoneal factor13 (14·6)7 (21·2)53 (12·6)13 (15·1)
Male factor48 (53·9)19 (57·6)249 (59·4)51 (59·3)
Anovulation2 (2·2)1 (3·0)3 (0·7)0 (0)
Unexplained4 (4·5)3 (9·1)17 (4·1)1 (1·2)
Mixed factors17 (19·1)3 (9·1)83 (19·8)16 (18·6)
Duration of infertility (years)4·2 ± 2·34·3 ± 2·84·4 ± 2·74·4 ± 2·8NS
GnRH agonist:antagonist77:1230:3385:3479:7NS
Percentage of ICSI (%)32·530·332·731·4NS
Table 3. Outcomes of in vitro fertilization
 TAI positiveTAI negativeP-value
EuthyroidSubclinicalEuthyroidSubclinical
  1. TAI, thyroid autoimmunity.

Clinical pregnancy rate per cycle initiated, % (n)44·9 (40/89)42·4 (14/33)45·8 (192/419)41·9 (36/86)NS
Miscarriage rate, % (n)20·0 (8/40)14·3 (2/14)19·3 (37/192)8·3 (3/36)NS
Live birth rate per fresh cycle, % (n)32·6 (29/89)33·3 (11/33)36·0 (151/419)36·0 (31/86)NS
Table 4. Clinical outcomes by different TSH thresholds
 TSH < 2·5 mIU/l (n = 508)TSH ≥ 2·5 mIU/l (n = 119)P-valueTSH < 3·5 mIU/l (n = 586)TSH ≥ 3·5 mIU/l (n = 41)P-valueTSH < 4·5 mIU/l (n = 602)TSH ≥ 4·5 mIU/l (n = 25)P-value
Clinical pregnancy rate per cycle initiated, %45·742·0NS45·143·9NS45·240·0NS
Miscarriage rate, %19·410·0NS18·211·1NS17·620·0NS
Live birth rate per fresh cycle, %35·435·3NS35·239·0NS35·532·0NS

We further divided the TAI-positive women based on their TPO antibody and Tg antibody levels and looked at their clinical outcomes. Clinical pregnancy, miscarriage and live birth rates were comparable among the groups, regardless of the levels of antibodies (Figs 1 and 2).

image

Figure 1. Clinical outcomes with different anti-TPO antibodies level.

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image

Figure 2. Clinical outcomes with different anti-Tg antibodies level.

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Discussion

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References

The aim of this study was to evaluate whether TAI, with or without subclinical hypothyroidism, has an impact on IVF/ICSI outcomes, in particular the live birth rate, which is the most relevant information for women undergoing treatment and their clinicians. Our study showed that TAI and subclinical hypothyroidism were common among women pursuing ART (19·7% and 18·7%, respectively). The clinical pregnancy, live birth and miscarriage rates were similar among women with or without TAI and/or subclinical hypothyroidism using a TSH threshold of between 2·5 and 4·5 mIU/l. Thyroid autoantibody level did not affect these IVF outcomes.

The relevance of TAI and preconception subclinical hypothyroidism in infertile patients undergoing ART has been a topic of debate. Singh et al.[16] in 1995 reported that women who successfully conceived with IVF had a higher incidence of miscarriage if they were positive for TAI, and Poppe et al.[17] showed similar findings. However, such an association was not reproduced by other researchers.[18, 19] Negro et al.[13] found no difference in pregnancy and miscarriage rates in women with or without thyroid antibodies, but within the group of antibody-positive women, high-normal TSH values before ART were associated with an increased risk of unsuccessful pregnancy and miscarriage. Our study took both TAI status and TSH level into account and showed no difference in miscarriage rates between TAI-positive and TAI-negative women. We failed to demonstrate an effect on the live birth rates in women with TSH ≥ 4·5 mIU/l or who were TAI positive. For TAI-positive women, the level of anti-Tg and anti-TPO antibodies did not affect the clinical outcomes including the miscarriage rates.

Our study is limited by its retrospective design and lack of data regarding antenatal complications or neonatal outcomes. Another concern of (sub)clinical hypothyroidism is altered neurological development in the foetus. Impaired mental development has been reported in children born to women who were inadequately treated for subclinical hypothyroidism, but not in children born to women who were adequately treated.[20] Our study failed to address this concern. Also, the limitations of assays, particularly with regard to assay bias, should be taken into account.[21] We used a TSH level of 2·5 mIU/l as the cut-off value as suggested by the Endocrine Society, which advised that TSH level should be kept below 2·5 mIU/l prior to pregnancy in women diagnosed with overt hypothyroidism. However, no consensus has been reached by endocrinologists regarding the appropriate cut-off value of TSH for diagnosis of subclinical hypothyroidism. No difference in the rates of clinical pregnancy, miscarriage or live birth were observed using the 2·5 mIU/l cut-off value, and our findings were consistent with those of Reh et al.[22] We therefore further examined the effect of different TSH cut-off values on IVF outcomes, and we found that the live birth rate was not affected by TSH level, up to 4·5 mIU/l, which is considered to be more clinically applicable, but with the trade-off of a much smaller sample size.

Women with overt hypothyroidism should be treated with an adequate dose of levothyroxine (LT4) before contemplating pregnancy to avoid maternal and foetal adverse effects.[4] However, in infertile women with subclinical hypothyroidism, the effect of LT4 supplementation in infertility treatment including IVF remains unclear. A prospective randomized study by Kim et al.[14] showed that LT4 therapy in patients with subclinical hypothyroidism improved embryo quality and enhanced embryo implantation after IVF/ICSI, but the result was limited by the small sample size. Nonetheless, given that LT4 is safe to use in pregnancy and its potential benefits outweigh the potential risks, the Endocrine Society recommends its use in women with subclinical hypothyroidism.

The NICE guideline in 2012 stated that routine measurement of thyroid function should not be offered to women with infertility and should be confined to women with symptoms of thyroid disease.[23] However, the American Association of Clinical Endocrinologists and the Endocrine Society advised TSH measurement in any women before pregnancy and those who present with infertility problems.[4, 24] The need for screening remains a topic of debate, and until more robust evidence is available, the decision to perform screening is made on the basis of individual choice and finding a balance between the available evidence and personal experience.

In conclusion, our retrospective data suggested that live birth and miscarriage rates were not impaired among women who were thyroid autoimmunity positive and/or had preconception subclinical hypothyroidism with TSH > 2·5 mIU/l.

Acknowledgement

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References

The authors would like to thank Dr Matthew Yeung and Miss Wong Po Chau Benancy for their input and technical assistance.

References

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgement
  8. Financial disclosure
  9. References
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