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Departments of Medicine (Y.S., T.N.) and Clinical Pathology (H.D.), Tokyo Metropolitan Tama Geriatric Hospital, Tokyo; and the Departments of Medicine (Y.S., H.S., I.W.) and Neurosurgery (N.S.), Nippon Medical School, Tokyo 113-8603, Japan
Address all correspondence and requests for reprints to: Yujin Shuto, M.D., Department of Medicine, Nippon Medical School, 1–1-5 Sendagi, Bunkyo-ku, Tokyo 113-8603, Japan. E-mail: shuto{at}nms.ac.jp
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Introduction |
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TopAbstractIntroductionCase ReportMaterials and MethodsResultsDiscussionReferences |
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Case Report |
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TopAbstractIntroductionCase ReportMaterials and MethodsResultsDiscussionReferences |
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and 2.
The values in Table 1 were obtained at the time of admission, whereas
hormone levels were measured 2 days after.
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The patient became conscious after glucose administration. He had a diet consisting of 1600 Cal/day and consumed most of it. The levels of fasting plasma glucose never dropped below 50 mg/dL after he was admitted. During hypoglycemia, serum insulin and insulin-like growth factor II (IGF-II) were undetectable, whereas counterregulatory hormones, such as cortisol and adrenaline, were increased appropriately. Aspartate aminotransferase and alanine aminotransferase levels returned to normal in about 2 weeks. A needle biopsy of the liver was carried out after obtaining informed consent. Histological examination revealed no evidence of hepatitis or cirrhosis, suggesting that the liver enzyme abnormalities were a consequence of hypoglycemia (3). In periodic acid-Schiff-stained sections, the amount of glycogen was normal. Magnetic resonance imaging (MRI) of the head showed a round intrasellar mass without a suprasellar extension. The lesion was hyperintense on the T1-weighted image and isointense relative to the white matter on the T2-weighted image. Follow-up studies for hormone levels were performed every month. Serum IGF-I levels remained low, and GH levels were between 10–20 ng/mL. TSH levels remained elevated in the face of normal free T4 concentrations. The MRI findings continued to show a hyperintensity on the T1-weighted image. The patient was readmitted in November 1996 because of a hypoglycemic coma. As he could not consume a diet orally this time, iv high calorie infusion was performed. The patient developed pneumonia and died of acute sepsis on December 10, 1996. As it had been 9 months since the IGF-I generation test, it is highly improbable that the sepsis was induced by GH administration.
Autopsy
An autopsy revealed a cyst confined to the sella turcica. The cyst contained a yellowish mucoid material. The wall of the cyst consisted of a tall columnar epithelium with ciliated cells, confirming the diagnosis of a Rathke’s cleft cyst. The pathological examination of the thyroid tissue revealed no evidence of autoimmune thyroiditis. There was no recurrence of gastric cancer or occult malignancy.
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Materials and Methods |
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TopAbstractIntroductionCase ReportMaterials and MethodsResultsDiscussionReferences |
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GH levels were measured with an immunoradiometric assay (IRMA), and IGF-I levels were determined by RIA after extraction. Serum GHBP was measured by ligand immunofunctional assay. The concentration of serum IGF-binding protein 3 (IGFBP-3) was determined by RIA.
The ability of the patient to respond to GH was tested by injecting recombinant human GH on 4 consecutive days (IGF-I generation test). The patient received 4 IU recombinant human GH (provided by Novo Nordisk Pharma Ltd., Copenhagen, Denmark) by daily sc injection for 4 days. The glucose tolerance test, GHRH test, GnRH test, and TRH test were performed using standard methods.
GH bioassay
The patient’s serum was examined for bioactive GH with Nb2 rat lymphoma cells as described by Tanaka et al. (4). In this bioassay, the replication of cells was stimulated by lactogenic hormones, including human GH. To neutralize the growth-stimulating action of PRL, a rabbit antiserum to human PRL was added.
Immunohistochemistry
Immunohistochemical studies for GH and other pituitary hormones were performed on paraffin-embedded sections of the pituitary. The avidin-biotin complex method was followed as previously described (5).
RNA extraction and RT-PCR
Liver tissue was obtained with a needle biopsy and immediately frozen in liquid nitrogen. Polyadenylated RNA was extracted using a Micro Fasttrak kit (Invitrogen, San Diego, CA). Complementary DNA (cDNA) was obtained from 100 ng of the poly(A) RNA prepared, using avian myeloblastosis virus reverse transcriptase (Takara Biomedicals, Tokyo, Japan) in a 20-µL reaction solution containing 50 mmol/L Tris-HCl (pH 8.3), 75 mmol/L KCl, 5 mmol/L MgCl2, 1 mmol/L deoxy-NTPs, 2.5 mmol/L random hexamers, and 1 U of a ribonuclease inhibitor. Then, the reaction mixtures were incubated for 30 min at 42 C and inactivated for 5 min at 95 C. The synthesized cDNA was amplified by PCR with primers (sense, 5'-GGATAAGGAATATGAAGTGC-3'; antisense, 5'-GATTTCTCATGGTCACTGC-3') as described by Ross et al. (6). The amplification by PCR involved an initial period of denaturation at 94 C for 3 min, followed by 30 cycles consisting of 30 s of denaturation at 94 C, 1 min of annealing at 56 C, and 2 min of extension at 72 C, and a final period of extension at 72 C for 10 min. The PCR products were separated on a 2% agarose gel, and the bands were visualized with ethidium bromide staining. For a positive control, poly(A) RNA extracted from IM-9 cells (7) was also reverse transcribed as described above, and the cDNA was amplified with PCR. Also, as an internal control, the expression of ß-actin mRNA in the liver tissue and IM-9 cells was examined by RT-PCR.
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Results |
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TopAbstractIntroductionCase ReportMaterials and MethodsResultsDiscussionReferences |
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GHBP was undetectable (<15.6 pmol/L; normal range, 65.0–408). IGFBP-3, which is the quantitatively dominant IGFBP in the blood and is regulated by GH, was decreased to 1.05 µg/mL (normal range, 2.59–4.26). Even though the levels of these binding proteins decline with age (8, 9, 10), it is highly likely that these levels in the patient were below the age-appropriate normal range. Follow-up studies for GHBP and IGFBP-3 were performed twice. There was no significant change in the levels of these binding proteins. Serum GH levels remained stable between 12.1–19.9 ng/mL after ingestion of 75 g glucose. Serum GH increased from 13.4 to 33.5 ng/mL after iv administration of 100 µg human GHRH. Serum GH levels did not change in response to GnRH or TRH administration. Administration of GH for 4 consecutive days produced a slight increment of IGF-I from an undetectable baseline level (<6.3 ng/mL) to 16.1 ng/mL, whereas there was no increase in the concentration of IGFBP-3. No side-effects were observed after GH administration.
A GnRH stimulation test showed that FSH levels remained stable between 68.8–85.8 mIU/mL. LH measured with a time-resolved fluoroimmunoassay (TR-FIA) remained below 0.2 mIU/mL at 0–90 min, and LH measured with an IRMA remained stable between 29.0–37.0 mIU/mL. Administration of 500 µg TRH produced a rise in serum TSH levels from a baseline level of 18.0 µIU/mL to 66.2 after 60 min.
GH bioactivity
The bioactivity of the patient’s GH was 21.0 ng/mL. GH immunoreactivity measured in the same serum using an IRMA was 19.2 ng/mL. These data confirm that the GH in the patient’s serum was biologically active.
Immunohistochemistry for pituitary hormones
Immunohistochemistry revealed that the number of GH-immunopositive
cells was normal (not shown), suggesting that there was no evidence of
a tumor overproducing GH. There was a cluster of cells with strong
immunoreactivity for the ß-subunit of TSH. The immunostaining for
PRL, ACTH, FSH ß-subunit, LH ß-subunit, and the common 
-subunit
of FSH and LH was normal in amount and distribution.
GH receptor mRNA expression in the liver
As shown in Fig. 1, a 453-bp band
was detected in IM-9 cells, whereas there was no detectable band in the
sample from the patient’s liver. The amplified products of ß-actin
mRNA were equivalent in both samples. These data imply that the
expression of the GH receptor gene in the liver was markedly decreased
in this patient.
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Discussion |
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TopAbstractIntroductionCase ReportMaterials and MethodsResultsDiscussionReferences |
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The mechanism of GH resistance in malnutrition is not established. In animals, protein restriction can result in both GH receptor and postreceptor defects with decreased IGF-I mRNA transcription (11). A 3-day fast caused a loss of high affinity GH receptors in the rat liver (12). In the patient described here, serum GHBP, which is derived proteolytically from the receptor, was undetectable. In addition, we could not detect GH receptor mRNA in the liver with RT-PCR. Therefore, the decreased expression of GH receptor mRNA in the liver may explain the GH resistance in this patient.
GH plays an important role in glucose counterregulation and has a diabetogenic effect. Therefore, a lack of GH function induces hypoglycemia. Clinically important hypoglycemia can occur in young children with GH deficiency, whereas most adults with GH deficiency do not experience hypoglycemia (13, 14). Subjects with GH deficiency become obese with increasing age and develop hyperinsulinemia and even glucose intolerance (13). It is considered that GH deficiency in adults may be offset by the development of insulin resistance caused by increased abdominal fat. A clinical manifestation of this patient was hypoglycemia in the presence of undetectable levels of serum insulin and IGF-II. The patient was extremely emaciated and had very little abdominal and visceral fat. These factors may have augmented the insulin sensitivity further and made him supersensitive to insulin. It is, thus, possible that the GH resistance and the shortage of fat tissue acted synergistically on serum glucose regulation and provoked refractory hypoglycemia in this patient. Furthermore, it is noteworthy that the amount of glycogen in the patient’s liver appeared to be normal. GH resistance might prevent mobilization and utilization of these glycogen stores.
Chronic malnutrition is usually associated with decreased serum T3 and T4 concentrations with preserved TSH levels. Even though the patient was severely malnourished, serum TSH levels were elevated in the presence of normal thyroid hormone levels. A cluster of cells showing strong immunohistochemical staining for the TSH ß-subunit was present in the pituitary. The secretion from these cells may be autonomous and resistant to feedback regulation by thyroid hormone.
In this patient, LH was undetectable using an IRMA, whereas with a TR-FIA the level of serum LH was about 38 mIU/mL. The difference in LH levels between the two methods could be explained by an immunologically anomalous LH variant (15, 16). Others have described point mutations in the LH ß-subunit gene in patients with anomalous LH variants (16). Their serum LH levels were undetectable with an IRMA, but were detected normally with a TR-FIA. Some patients with anomalous LH variants suffered from infertility or menstrual disorders, whereas others had no apparent abnormalities. The bioactivity of their mutant LH was normal. The patient described here had three daughters. The low level of serum testosterone may not be abnormal considering that he was 87 yr old. Although the LH ß-subunit gene remains to be analyzed, the patient might have a LH ß-subunit gene with a point mutation.
Rathke’s cleft cysts are nonneoplastic lesions of the sellar region derived from the foregut (17, 18). They are usually asymptomatic, and in 32% of the cases, the pituitary mass was an incidental finding. Pituitary function was intact in half of these patients, whereas varying degrees of anterior pituitary malfunction were evident in the reminder. The patient had a Rathke’s cleft cyst that was found incidentally by MRI. Although there were abnormalities in pituitary hormone levels, these may be independent of the Rathke’s cleft cyst.
In summary, we have described acquired GH resistance in a malnourished elderly man with multiple endocrine abnormalities. GH resistance in the patient may be due to malnutrition. Decreased expression of GH receptor mRNA in the liver may be responsible for the GH resistance.
Received November 2, 1998.
Revised January 12, 1999.
Revised March 24, 1999.
Accepted March 29, 1999.
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