Do We Still Need the TRH Stimulation Test?

Hulusi Atmaca,1 Fatih Tanriverdi,2 Cumali Gokce,2 Kursad Unluhizarci,2 and Fahrettin Kelestimur2

Objective: To evaluate the diagnostic value of the thyrotropin-releasing hormone (TRH) stimulation test in thediagnosis of central hypothyroidism in patients with Sheehan’s syndrome. Design: TRH stimulation test wasperformed in 72 patients with Sheehan’s syndrome. Basal free triiodothyronine (fT3) and free thyroxine (fT4)levels were measured. Serum thyrotropin (TSH) concentration was determined before and 30, 60, 90, and 120minutes after 200 mg TRH IV bolus injection. The peak TSH values <5.5 mIU=ml were defined as inadequateresponse. A peak TSH at 60 minutes or later was considered as delayed response. If TSH (60 minutes after peak),was more than 40% of the peak value it was considered as prolonged response. The diagnosis of central hypo-thyroidism (CH) was made if either serum fT4 concentration was subnormal with an inappropriately low serumTSH concentration or inadequate response to TRH stimulation test and=or a delayed or prolonged response toTRH stimulation test. Main outcome: Fifty-six (77.7%) of the patients had low serum fT4 and fT3 levels with aninappropriately low serum TSH levels were defined as CH (CH0 group). Ten (13.8%) patients with normal andlow-normal fT4 levels had no response and=or delayed or prolonged response to TRH stimulation test(CH1group). Six (8.3%) patients had fT3, fT4, and TSH levels within normal limits and peak TSH responses$5.5 mIU=ml consistent with euthyroidism (euthyroid group). Thus, 66 (91.6%) of 72 patients with Sheehan’ssyndrome had CH. Although fT4 levels were within normal reference range, TRH stimulation test revealed that10 (13.8%) of these had CH. Conclusion: TRH stimulation test is useful in the diagnosis of central hypothyroidism,especially in whom fT4 and=or TSH is low-normal and known to have hypothalamo-pituitary pathology.

Introduction

The thyrotropin-releasing hormone (TRH) stimula-tion test was a useful diagnostic test during the era of

first-generation thyrotropin (TSH) assays. The introductionof new generation assays for TSH and, to a lesser extent, im-provements in the assays for free thyroid hormones, havedramatically reduced the role for dynamic testing of thehypothalamo-pituitary-thyroid axis in most patients with sus-pected hormonal abnormalities of thyroid function (1). Thediagnosis of central hypothyroidism (CH) is often compli-cated because of varying clinical symptoms and hormonalprofiles. Basal serum TSH can be either low, normal, or even inthe high-normal range in patients with pituitary or hypo-thalamic disorders with secondary hypothyroidism. How-ever, some of the patients with CH have normal TSH valuesand thyroxine (T4) or free T4 (fT4) levels within the lower partof the normal range. Recently, it was reported that CH couldnot be detected only by measurement of peripheral thyroidhormones and TSH levels (2). In addition, CH may be asso-ciated with other pituitary hormone deficiencies, which mayhide the clinical manifestations of thyroid hormone insuffi-ciency. In this study we have investigated the usefulness of the

TRH stimulation test in the diagnosis of CH in patients withSheehan’s syndrome.

Materials and Methods

Seventy-two patients with Sheehan’s syndrome were in-cluded in the study. The study was approved by the EthicalCommittee of Erciyes University Medical School. The criteriafor the diagnosis of Sheehan’s syndrome included: (i) historyof postpartum hemorrhage and=or history of postpartumfailure of lactation and=or secondary amenorrhea; (ii) varyinggrades of loss of pituitary hormone reserve; (iii) good clinicalresponse to hormone replacement therapy; and (iv) exclusionof pituitary mass lesion and empty sella on magnetic reso-nance imaging (MRI) (3).

The mean age of the patients was 53.6� 10.7 (range 29–75)years and mean duration of the disease was 17.8� 8.7 (range4–39) years. Laboratory evaluation included levels of basalhormones (free triiodothyronine [fT3], fT4, TSH, prolactin,cortisol, estradiol, follicle-stimulating hormone, luteinizinghormone, and insulin-like growth factor-1) and dynamic pi-tuitary function tests including; TSH and prolactin responsesto TRH, follicle-stimulating hormone, and luteinizing

1Department of Endocrinology, Zonguldak Karaelmas University Medical School, Zonguldak, Turkey.2Department of Endocrinology, Erciyes University Medical School, Kayseri, Turkey.

THYROIDVolume 17, Number 6, 2007ª Mary Ann Liebert, Inc.DOI: 10.1089=thy.2006.0311

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hormone responses to gonadotropin-releasing hormone, cor-tisol responses to corticotropin (ACTH), and growth hor-mone (GH) and cortisol responses to insulin tolerance test.None of them reported either transient or permanent poly-uria or polydipsia. Other causes of hypopituitarism wereruled out by the typical history, physical examination, resultsof laboratory findings, and radiological investigations. Allthe tests including TRH stimulation test have been carriedout during the first admission of the patients in the course ofusual clinic care.

All the patients were clinically stable and without any in-tercurrent illness, such as infectious disease, coronary arterydisease, gastrointestinal system disorder, etc. The patients whohad secondary adrenal failure were on glucocorticoid treat-ment (61 patients) during TRH stimulation test. None ofthem were on estrogen, GH, or thyroid replacement therapy.TRH stimulation tests were performed in the fasting state(8.00–10.00 hours). All patients were given 200mg TRH (pro-tirelin, Ferring Laboratory, Kiel, Germany) intravenously asa bolus injection. Blood samples were obtained basally and30, 60, 90, and 120 minutes for TSH analysis after TRHstimulation. In a previously performed study, the TSH peakafter TRH was between 5.5 and 14 mIU=ml in our 10 healthyeuthyroid control women (4). The lowest peak TSH afterTRH has been used as a cut-off value to define TSH responseto TRH. In that study, serum TSH concentration was deter-mined before and 30 and 60 minutes after 200 mg TRH in-travenous (IV) bolus injection. The peak TSH value $5.5 mIU=ml was accepted as a normal response. The value below5.5 mIU=ml was accepted as inadequate response. A peakTSH response at 60 minutes or later was considered as de-layed response. If TSH (60 minutes after peak), was morethan 40% of the peak value it was considered as prolongedresponse (5). An inadequate response and=or a delayed orprolonged response(s) was defined as abnormal response toTRH stimulation test. The diagnosis of CH was made if theserum fT4 concentration was subnormal with an inappropri-ately low serum TSH concentration. According to the TRHstimulation test, the diagnosis of central hypothyroidism re-quired an abnormal response. The TRH-stimulated serumTSH response was also calculated in terms of both an abso-lute response (peak minus basal TSH) as well as a fold re-sponse (peak=basal TSH).

Serum TSH levels were measured by IRMA method (Bio-source,Nivelles,Belgium,functionalassaysensitivity,0.025mIU=ml; reference range 0.20–4.5mIU=ml). The reference range offT3 was 2.2–4.7 pg=ml and of fT4 was 8–20 pg=ml fT3 and fT4

were assayed by using the kits of Amersham (London, UK).Both inter- and intraassay coefficients of variation for TSH were4.1%, 6%; fT3, 9.8%, 5.8%; and fT4, 7.5%, 6.5%, respectively.Serum samples were stored at �208C until assayed.

Results are expressed as mean� SD in the text and tables.Two groups were compared by the nonparametric Mann–Whitney U test. Correlations were evaluated by Spearman’srank correlation test. P value less than 0.05 was considered asstatistical significance.

Results

Fifty-one (70.8 %) of the patients had complete and 21(29.2 %) had partial empty sella on pituitary MRI. Both go-nadotropins and GH were deficient in 97%, and ACTH and

prolactin were deficient in 84.7% and 80.5% of cases, re-spectively.

The patients were evaluated according to the number of an-terior pituitary hormone deficiencies. Only one patient (1.4%)had TSH deficiency plus one other hormone (GH) deficiency.Two (2.8%), 22 (30.6%), and 47 (65.3%) of the patients hadTSH deficiencies plus two, three, and four other hormones,respectively. Fifty-six (77.7%) of the patients having low se-rum fT4 (range, 0.10–7.80 pg=ml) and fT3 levels in conjunc-tion with an inappropriately low serum TSH levels weredefined as CH (CH0). Of these, the TRH stimulation testrevealed peak TSH response <5.5 mIU=ml in 42 (75%) pa-tients and $5.5 mIU=ml in 14 (25%) patients.

Ten patients with normal and low-normal fT4 levelsshowed peak TSH response <5.5 mIU=ml. Of these, eight hadnormal TSH levels and two had low TSH levels. Eight hadfT4 in the lower portion of the normal range (range, 8.0–12.5 pg=ml), and only two had fT4 in the upper portion of thenormal range (14.9 and 16 pg=ml) (Fig. 1). Nine of thesepatients (90%) had delayed or prolonged response after TRHstimulation test. Therefore, 10 patients with abnormal TRHstimulation test were considered as having CH (CH1). Inother words, 10 (13.8%) of the diagnoses of CH would havebeen missed by reliance on fT4 and basal TSH values alone.

Six (8.3%) of 72 patients had normal fT3, fT4, and TSHlevels and peak TSH response $5.5 mIU=ml consistent witheuthyroidism (euthyroid group). Five of euthyroid patientshad fT4 in the lower portion of the normal range (range, 8.3–12.5 pg=ml) and the remaining one had fT4 in the upperportion of the normal range (19.7 pg=ml). As a result, 10 outof 16 Sheehan’s syndrome patients with normal FT4 levelswere diagnosed CH by TRH stimulation test. On the con-trary of euthyroid patients, CH1 patients were treated withL-thyroxine.

The results of TRH stimulation test are shown in Figure 2,and peak TSH responses of each groups, including healthy

20

18

16

14

FT

4 (

pg/m

l)

12

10

8

6

-,5 0,0 ,5 1,0 1,5

TSH (µlU/ml)

2,0 2,5 3,0 3,5 4,0 4,5

FIG. 1. Serum free thyroxine (fT4) and thyrotropin (TSH)levels in patients with central hypothyroidism in whom fT4

levels were within normal range. Horizontal broken line in-dicates lower limit of normal fT4 range; vertical broken lineindicates lower limit of normal TSH range.

530 ATMACA ET AL.

euthyroid control group, are shown in Figure 3. In thismanner, a total of 66 (91.6%) patients with Sheehan’s syn-drome were suggested to have CH.

Patients with CH were evaluated according to basal TSHlevels; 56 (85%) had normal, four (6%) had low, and six (9%)had mildly elevated TSH levels. A significant correlation(r¼ 0.85, p< 0.001) was found between basal TSH and peakTSH levels in patients with CH (Fig. 4), but not in euthyroidpatients. Patients with CH (group CH0 and CH1) did notdiffer from the euthyroid group in terms of the duration ofthe disease and the age (Table 1).

fT3 ( p< 0.005) and fT4 ( p< 0.001) levels were lower in theCH0 group than in the CH1 and euthyroid groups. As theSheehan’s patients have potential hypopituitary findings inother axes, we also compared the data with the age-matched

control subjects. Although the mean basal TSH levels weresimilar between the groups, peak TSH ( p< 0.005), abso-lute TSH ( p< 0.005) and fold TSH ( p< 0.05) responses werehigher in the euthyroid and control groups compared to theCH0 and CH1 patients. CH0 and CH1 groups were similar interms of absolute and fold TSH responses. Only fold TSHresponse was higher ( p< 0.01) in control group than in eu-thyroid group, whereas basal TSH, peak and absolute TSHresponses were similar between euthyroid and control groups(Table 1). Individual absolute and fold TSH responses areshown in Figure 5 and Figure 6, respectively.

Discussion

TRH secreted from the hypothalamus regulates the syn-thesis and secretion of TSH. It is also important in the post-translational processing of the oligosaccharide moieties onTSH which are necessary for biological activity (6). The TRHstimulation test was established after the introduction of syn-thetic TRH in 1969 as a simple and safe diagnostic tool forthe evaluation of pituitary-thyroidal functions (5).

Although TRH has many routes of administration such assubcutaneous, intramuscular, oral, rectal, etc., it is widely rec-ognized that the rapid IV injection of TRH is the best standard-ized and most reliable way to investigate the TSH response(6–8). Although the recent introduction of an ultrasensitiveTSH assay has rendered the use of TRH stimulation test inmany clinical conditions such as subclinical hyperthyroidism,it is still useful in the differential diagnosis of resistance to thy-roid hormones and TSHoma. Patients with central hypothy-roidism may show different patterns of basal TSH levels whichmay lead to problems in the diagnosis of secondary hypothy-roidism (1). Patients with central hypothyroidism may show

FIG. 2. The mean (� standard error of the mean [SEM])thyrotropin (TSH) responses of the study groups obtainedduring thyrotropin-releasing hormone (TRH) stimulation test.

FIG. 3. Peak thyrotropin (TSH) responses of patients (CH0¼patients diagnosed central hypothyroidism according to basalthyroid hormone levels only; CH1¼patients diagnosed cen-tral hypothyroidism in whom free thyroxine [fT4] levels werewithin normal range) and control groups.

FIG. 4. There was a significant correlation in patients withcentral hypothyroidism between basal thyrotropin (TSH) lev-els and peak TSH levels (r¼ 0.85, p< 0.001).

TRH TEST AND CENTRAL HYPOTHYROIDISM 531

different patterns of TSH response to TRH, changing fromabsent to exaggerated responses and from delayed to pro-longed response curve. Absent or reduced responses are infavor of pituitary hypothyroidism while positive responses,often delayed, exaggerated response indicate hypothalamicorigin of hypothyroidism (1).

The standard TRH stimulation test is performed by giving200–500 mg TRH IV and measuring the TSH level at 0, 20 (or30), and 60 minutes. There is no ‘‘gold standard’’ rangeof TSH values to develop for TRH stimulation test, but the

TSH levels rise at 20–30 minutes to around 10mIU=ml andfall slightly to around 8mIU=ml may be considered as nor-mal (2). In our healthy population we found that peakTSH response higher than 5.5 mIU=ml indicates adequateresponse.

There were two main indications for TRH stimulation testused to be undertaken. The first is to confirm the diagnosis ofthyrotoxicosis which became reluctant after the introductionof sensitive TSH assays. The second one is to investigatesuspected secondary and tertiary hypothyroidism. It has

Table 1. Comparisons of Characteristics of the Groupsa

GroupsCH0

(n¼ 56)CH1

(n¼ 10)Euthyroid(n¼ 6)

Control(n¼ 10)

Age (yr) 54.3� 10.9 (29–75) 52.2� 8.4 (38–65) 50.02� 12.3 (31–63) 49.0� 9.1 (32–67)Duration of the

disease (yr)17.2� 8.4 (4–36) 21.0� 9.4 (5–39) 18.6� 10.5 (6–30)

fT3 (N; 2.2–4.7 pg=ml) 0.96� 0.77 (0.03–3.05) 1.81� 0.81 (0.24–2.90) 2.61� 1.00b (0.86–3.30)fT4 (N; 8–20 pg=ml) 3.10� 2.30 (0.10–7.80) 10.3� 3.02 (8–16) 11.6� 4.30b (8.30–19.7)TSH (N; 0.2–4.5 mIU=ml) 2.09� 1.71 (0.10–6.80) 0.96� 1.03 (0.01–2.90) 2.43� 1.81 (0.29–4.79) 1.33� 0.53 (0.35–2.0)Peak TSH (mIU=ml) 4.67� 4.37 (0.12–23.5) 2.29� 2.24 (0.02–5.47) 11.5� 5.13 (5.92–19.2) 11.2� 2.43c (5.50–14.0)Absolute TSH response

(peak TSH minusbasal TSH)

2.58� 3.04 (0.04–16.7) 1.20� 1.23 (0.03–3.83) 8.46� 5.03 (3.17–16.2) 9.92� 2.41)c (5.22–13.1)

Fold TSH response(peak TSH=basal TSH)

2.71� 2.31 (1.06–15.3) 2.67� 1.06 (1.11–4.03) 4.06� 1.77d (1.86–6.41) 9.98� 4.42c (4.78–15.9)

aRanges are given in parenthesis. TSH, thyrotropin; fT3, free triiodothyronine; fT4, free thyroxine; CH0, patients diagnosed central hypo-thyroidism according to basal thyroid hormone levels only; CH1, patients diagnosed central hypothyroidism in whom fT4 levels were withinnormal range; euthyroid, patients with both normal basal thyroid hormone levels and with normal TRH test.

bCH0 vs. CH1 and Euthyroid. Significant difference ( p< 0.05) between pairs.cEuthyroid and control vs. CH0 and CH1. Significant difference ( p< 0.05) between pairs.dEuthyroid vs. control. Significant difference ( p< 0.05) between pairs.

FIG. 5. Absolute thyrotropin (TSH; peak TSH minus basalTSH) responses of each group.

FIG. 6. Fold thyrotropin (TSH; peak TSH=basal TSH) res-ponses of each group.

532 ATMACA ET AL.

been suggested that TRH stimulation test has a low sensi-tivity to be used in routine clinical practice (8,9). However, in10 patients with Sheehan’s syndrome fT4 levels were normalor at a low-normal level associated with normal TSH levels(in eight patients) and low TSH levels (in two patients).Those patients might be considered as normal hypothalamo-pituitary thyroid axis unless a TRH stimulation test had beenperformed. Basal TSH values may be normal or even mildlyelevated in patients with Sheehan’s syndrome which is ex-plained by the presence of TSH isoforms, especially second-ary to differences in oligosaccharide content, which havevariable bioactivity (2,10).

Hartoft-Nielsen et al. (5) evaluated the TRH stimulationtest in the diagnostic work-up of the thyroid function inpatients with pituitary pathology. They compared the TSHresponse in 35 patients with pituitary pathology and com-pared the results with 26 healthy subjects. The authors foundthat the absolute response to TRH and the fold response de-fined as stimulated TSH response divided by basal TSH weresimilar. They concluded that the use of the TRH stimulationtest as a routine examination in the diagnostic work-up of thepatients with central hypothyroidism is questionable (5). Inthat study the patients had pituitary adenomas, which is verydifferent from Sheehan’s syndrome in terms of pathogenesis.In pituitary tumors, the cells may exhibit hypofunction whichmight be due to mass effect of the tumor and may not bedestroyed extensively. In contrast Sheehan’s syndrome is dueto postpartum necrosis of the pituitary gland which leads toextensive and mostly irreversible damage of the pituitary cells.We agree with the authors that TRH stimulation test shouldnot be used indiscriminately in the evaluation of central hypo-thyroidism but it may be useful in patients with low-normalthyroid hormone levels associated with low-normal TSHlevels.

Circulating TSH has multiple molecular forms or isoformsdue to variations in the oligosaccharide structures (11,12). Inseveral thyroid disorders, different biological activities hasbeen reported. Additionally, it has been shown that patientswith Sheehan’s syndrome may have unexpectedly normal orelevated TSH level without any pulsality (13,14). Due to theimmunoreactive, but less bioactive TSH, patients with Shee-han’s syndrome may mimic normal thyroid axis albeit withsecondary hypothyroidism. In addition, low levels of cortisolas well as possible decrease in hypothalamic somatostatindue to decreased GH secretion could contribute to furtherincrease TSH release in these patients (15).

In conclusion the diagnosis of central hypothyroidism isoften complicated because of varying clinical symptoms andbiochemical profiles. The associated hormone deficiencies inpatients with Sheehan’s syndrome even complicates the clin-ical picture. Thus, in selected patients TRH stimulation test isuseful in the diagnosis of central hypothyroidism, especiallywith patients in whom fT4 is low-normal or within normallimits and who are known to have hypothalamo-pituitarypathology.

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13. Abucham J, Castro V, Maccagnan P, Vieira JG 1997 Increasedthyrotropin levels and loss of nocturnal thyrotropin surge inSheehan’s syndrome. Clin Endocrinol 47:515–522.

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Address reprint requests to:Prof. Dr. Fahrettin KelestimurDepartment of Endocrinology

Erciyes University Medical School38039, Kayseri

Turkey

E-mail: fktimur@erciyes.edu.tr

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