Paternal deprivation prior to adolescence and vulnerability to pituitary adenomas.

It has been reported that women with prolactinoma were exposed, early in life, to an environment characterized by an absent or violent father. The present study was designed to evaluate whether paternal absence or violent paternal behavior were more prevalent in patients with pituitary adenomas (prolactinoma, acromegaly, non-secreting adenoma and Cushing's disease) compared to a control population. We conducted an observational case-control multicenter study. We interviewed 395 patients with prolactinoma (296 females and 99 males), 130 with acromegaly (87 females and 43 males), 237 with non-secreting adenoma (144 females and 93 males) and 68 with Cushing's disease (61 females and 7 males) and 365 patients from the same clinics with nodular thyroid disease or lymphocytic thyroiditis with euthyroidism as controls. Violent or absent fathers were significantly more prevalent in patients with prolactinoma or acromegaly than in controls (P = 0.001 and P = 0.002, respectively) but not in patients with non-secreting adenoma or corticotrophinoma. Absent fathers in prolactinoma and acromegaly versus controls: P = 0.001 and P = 0.119. Violent fathers in prolactinoma and acromegaly versus controls: P = 0.069 and P = 0.001. The prevalence of absent or violent fathers was also significantly higher in prolactinoma and acromegaly when compared to non-secreting adenoma (P = 0.039 and P = 0.033, respectively). Paternal deprivation before adolescence may be a risk factor for prolactinoma and acromegaly but not for non-secreting pituitary adenomas or Cushing's disease.

Underexpression of peroxisome proliferator-activated receptor (PPAR)gamma in PAX8/PPARgamma-negative thyroid tumours.

The expression of peroxisome proliferator-activated receptor (PPAR)gamma in thyroid neoplasias and in normal thyroid (NT) tissues has not been fully investigated. The objectives of the present work were: to study and compare the relative expression of PPARgamma in normal, benign and malignant thyroid tissues and to correlate PPARgamma immunostaining with clinical/pathological features of patients with thyroid cancer. We analysed the expression of PPARgamma in several types of thyroid tissues by reverse transcription-polymerase chain reaction (RT-PCR), interphase fluorescent in situ hybridisation, real-time RT-PCR and immunohistochemistry. We have demonstrated that NT tissues express PPARgamma both at mRNA and at protein level. PAX8-PPARgamma fusion gene expression was found in 25% (six of 24) of follicular thyroid carcinomas (FTCs) and in 17% (six of 36) of follicular thyroid adenomas, but in none of the 10 normal tissues, 28 nodular hyperplasias, 38 papillary thyroid carcinomas (PTCs) and 11 poorly differentiated thyroid carcinomas (PDTCs). By real-time RT-PCR, we observed that tumours negative for the PAX8-PPARgamma rearrangement expressed lower levels of PPARgamma mRNA than the NT. Overexpression of PPARgamma transcripts was detected in 80% (four of five) of translocation-positive tumours. Diffuse nuclear staining was significantly (P<0.05) less prevalent in FTCs (53%; 18 of 34), PTCs (49%; 19 of 39) and PDTCs (0%; zero of 13) than in normal tissue (77%; 36 of 47). Peroxisome proliferator-activated receptorgamma-negative FTCs were more likely to be locally invasive, to persist after surgery, to metastasise and to have poorly differentiated areas. Papillary thyroid carcinomas with a predominantly follicular pattern were more often PPARgamma negative than classic PTCs (80% vs 28%; P=0.01). Our results demonstrated that PPARgamma is underexpressed in translocation-negative thyroid tumours of follicular origin and that a further reduction of PPARgamma expression is associated with dedifferentiation at later stages of tumour development and progression.

Cortisol, prolactin, growth hormone and neurovegetative responses to emotions elicited during an hypnoidal state.

The present study describes the responses of cortisol, prolactin and growth hormone (GH) to emotions elicited during sessions in which an hypnoidal state was induced. The purpose of the study was to provide answers for the following questions: 1) Do sessions with an emotional content have more hormonal surges than baseline, relaxation-only, sessions? 2) Does the induction of a fantasy of pregnancy and nursing elicit a prolactin response? 3) Are there any associations between surges of different hormones? 4) Are hormonal responses related to the intensity, type, or mode of expression of the emotions? For this purpose, thirteen volunteers and twelve patients with minor emotional difficulties were studied during sessions under hypnosis. The period of observation lasted for about three hours. Heart rate (HR), skin conductance (SC) and vagal tone (VT) were monitored. Serum cortisol, prolactin and growth hormone were sampled every 15 minutes. The volunteers had three types of sessions- "blank", consisting of relaxation only (12 sessions), "breast feeding", in which a fantasy of pregnancy and breast feeding was induced (12 sessions) and "free associations" in which the subjects were encouraged to evoke experiences or feelings (17 sessions). The patients had only sessions of free associations (38 sessions). Sessions of free associations had more hormonal surges than "blank" and "breast feeding" sessions. This was true for cortisol (8/17 v.3/24; p < 0.03), prolactin (7/17 v. 3/24; p < 0.05) and GH (9/17 v. 4/24; p < 0.02). During the 55 sessions of free associations (volunteers plus patients) there were 32 surges of cortisol, 18 of prolactin and 28 of GH. Cortisol and prolactin surges were negatively correlated (p < 0.03). GH had no significant association with either cortisol or prolactin. Visible emotions were positively associated with GH surges (p < 0.05). but not with cortisol or prolactin. Cortisol surges were correlated positively with evocations of real events (p < 0.01) and negatively with evocations containing defensive elements (p < 0.01). Cortisol correlated positively with shock and intimidation (p < 0.02) and negatively with rage (p < 0.04). The AUC of the cortisol peaks during shock and intimidation was significantly higher than that of the pool of all other cortisol peaks (12.4 micromol x min x l(-1) v. 7.1 micromol x min x l(-1); p < 0.005). Rage had a marginally significant positive association with prolactin surges (p=0.07). The distribution of GH surges did not show any significant association with types of emotions. The present study provides evidence that cortisol, prolactin and GH respond to psychological stress in humans. However, they are regulated differently from one another. Cortisol and prolactin surges appear to be alternative forms of response to specific emotions. GH surges depend on the intensity of the emotion, probably as a consequence of the associated muscular activity. The current paradigm of stress, implying corticotrophin-releasing hormone (CRH) as the initial step of a cascade of events, is insufficient to account for the diversity of hormonal changes observed in psychological stress in humans.

A minimal mathematical model of calcium homeostasis.

A mathematical model of calcium homeostasis is presented in which the controlling factors are the plasma concentrations of calcium, PTH, and calcitriol, and the effector organs are the parathyroids, bone, kidney, and intestine. Other factors can be added as the need arises. The model is aimed at simulating what happens in a single individual, but its parameters and variables were adjusted to the corresponding published average values. Simulations of published observations in humans undergoing the infusion of calcium or its chelators are presented. With a single exception, these simulations provided a good fit to the data. The response of the system to extrinsic perturbations was characterized by simulating chronic infusions of calcium, PTH, and calcitriol. Finally, the steady state response to perturbations in some of its parameters (the secretory mass of the parathyroids and the affinity and/or sensitivity of the calcium, PTH, and calcitriol receptors) and to renal failure were also investigated in an attempt to analyze the pathogenesis of clinical hypo- or hypercalcemias. In its present form the model cannot be used to base clinical decisions in individual cases. However, it requires modest computational resources, and clinicians with a modest mathematical background can manipulate it. It is a useful tool for the analysis of general mechanisms of the diseases of calcium metabolism and for the design of clinical experiments aimed at characterizing these diseases. The model can also be the core of future autoadaptive extensions to be used in individual patients.

Mutational analysis of Portuguese families with multiple endocrine neoplasia type 1 reveals large germline deletions.

OBJECTIVE: To determine the spectrum of MEN1 mutations in Portuguese kindreds, and identify mutation-carriers. PATIENTS, DESIGN AND RESULTS: Six unrelated MEN1 families were studied for MEN1 gene mutations by single-strand conformational polymorphism (SSCP) and DNA sequence analysis of the coding region and exon-intron boundaries of the MEN1 gene. These methods identified 4 different heterozygous mutations in four families: two mutations are novel (mt 1539 delG and mt 655 ims 11 bp) and two have been previously observed (mt 735 del 46p and mt 1656 del C) all resulting in a premature stop codon. In the remaining two families, in whom no mutations or abnormal MEN1 transcripts were detected, segregation studies of the 5' intragenic marker D11S4946 and codon 418 polymorphism in exon 9 revealed two large germline deletions of the MEN1 gene. Southern blot and tumour loss of heterozygosity analysis confirmed and refined the limits of these deletions, which spanned the MEN1 gene at least from: exon 7 to the 3' untranslated region, in one family, and the 5' polymorphic site D11S4946 to exon 9 (obliterating the initiation codon), in the other family. Twenty-six mutant-gene carriers were identified, 6 of which were asymptomatic. CONCLUSIONS: These results emphasize the importance of the detection of MEN1 germline deletions in patients who do not have mutations of the coding region. Important clues indicating the presence of such deletions may be obtained by segregation studies using the intragenic polymorphisms D11S4946 and at codon 418. The detection of these mutations will help in the genetic counselling of clinical management of the MEN1 families in Portugal.

The hyperparathyroidism-jaw tumour syndrome in a Portuguese kindred.

The hyperparathyroidism-jaw tumour (HPT-JT) syndrome is an autosomal dominant disease characterized by the occurrence of parathyroid tumours and fibro-osseous tumours of the jaw bones. Some HPT-JT patients may also develop renal abnormalities, which include Wilms' tumours, hamartomas and polycystic disease. The HPT-JT gene has been mapped to chromosome 1q25-q31, and we report the clinical and genetic findings in a kindred from central Portugal. HPT-JT was observed in six members from three generations; all had primary hyperparathyroidism (five had parathyroid adenomas, one a parathyroid carcinoma). Ossifying jaw fibromas affecting the maxilla and/or mandible were observed in 5/6. Renal cysts (<2.5 cm) were observed in four. Genetic studies using 18 polymorphic loci from chromosome 1q25-q31, together with leukocyte DNA from 11 family members and tumour DNA from three parathyroids (two adenomas and one carcinoma), revealed loss of tumour heterozygosity in the parathyroid carcinoma only, and the retained haplotype was found to cosegregate with the disease in the six affected members. A new Portuguese kindred with the HPT-JT syndrome that maps to chromosome 1q25-q31 has been identified, and these findings will help in the further characterization of this inherited disorder.

Medullary thyroid carcinoma: an accurate pre-operative diagnosis by reverse transcription-PCR.

OBJECTIVE: To study the expression of calcitonin (CT) and thyroglobulin mRNA in samples of leftover cells in needles used for fine-needle aspiration biopsy either from thyroid tumours or cervical lymph nodes. PATIENTS AND METHODS: Specimens were analysed using reverse transcription-polymerase chain reaction; 12 samples from 11 patients were included and molecular diagnosis was compared with cytological or histological diagnosis and serum CT measurements. RESULTS: Transcripts of the CT gene were detected in all six patients with medullary thyroid carcinoma (MTC) but in none of the other patients. CONCLUSIONS: Present data reinforce this technique as a reliable and alternative tool to establish the pre-operative diagnosis of MTC, especially when cytological examination is not conclusive or when cytological information is not in agreement with clinical data. Furthermore, it may be clinically useful to identify those conditions in which increased serum CT in the presence of a thyroid nodule is not due to MTC.

Somatic trinucleotide change encompassing codons 882 and 883 of the RET proto-oncogene in a patient with sporadic medullary thyroid carcinoma.

OBJECTIVE: Restriction analysis is a straightforward procedure for mutational analysis. It is commonly used for screening RET mutations. Incomplete digestion is a well-known cause of false results. Herein, we report another limitation of the method. DESIGN AND METHODS: Screening for somatic mutations in RET exons 16, 13 and 15 was performed in a patient with a sporadic medullary thyroid carcinoma. Genetic study was carried out by both restriction analysis and direct sequencing. RESULTS: A somatic trinucleotide change encompassing codons 882 and 883 of the RET proto-oncogene (GTA GCT to GTT TTT) was documented. Particular to this case is the silent mutation (GTA-->GTT) at codon 882. Independently, both the novel silent mutation and the missense mutation at codon 883 may disrupt the same AluI restriction site. Based on the restriction pattern we were able to say that both mutations occurred in the same allele. CONCLUSIONS: Restriction analysis is an easy approach for screening RET mutations; however, it is not enough to assign a final diagnosis.

Presence of a Gs alpha mutation in an adrenal tumor expressing LH/hCG receptors and clinically associated with Cushing's syndrome.

We describe the case of a patient with Cushing's syndrome due to a functioning adrenal adenoma. There was a pronounced increase in serum and urinary cortisol after administration of human chorionic gonadotropin. Immunocytochemistry revealed positive immunostaining for LH/hCG receptors. Molecular analysis documented the presence of a gsp mutation at codon 201 (CGT to TGT). The existence of this type of hCG-responsive adrenal tumor may help explain the higher prevalence of cortisol-secreting adrenal tumors versus pituitary-dependent disease in pregnant women with Cushing's syndrome as well as some reported cases of remission following delivery.

Hyperprolactinemia due to big big prolactin is differently detected by commercially available immunoassays.

Macroprolactinemia, i.e. sustained hyperprolactinemia where the predominant circulating form of prolactin (PRL) is of large molecular weight, is a common phenomenon comprising up to one-fourth of all cases of hyperprolactinemia. We measured serum PRL levels by four different immunoassay systems (PROL-CTK, RIAgnost, Delfia, ACS 180) and by the Nb2 bioassay in patients with prolactinomas/idiopathic hyperprolactinemias in whom monomeric PRL was the major species of PRL (n=11, group 1) and in patients with macroprolactinemia (n=12, group 2). In group 1, the results obtained with the different immunoassays and with the Nb2 assay were highly correlated (r=0.945-0.982). On the other hand, big big-PRL (bb-PRL) was differently recognized by the immunoassays, since measured serum PRL values from each patient were highly variable in group 2. RIA-gnost Prolactin and Delfia Prolactin detected bb-PRL similarly and they were highly correlated with each other (r=0.937, p<0.0001). ACS 180 detected bb-PRL somewhat differently from the RIA-gnost and Delfia systems, but likewise most of the patients of group 2 had PRL values above normal. PROL-CTK was the method less influenced by the presence of bb-PRL since most of the subjects with macroprolactinemia had PRL levels either within the normal range or only marginally elevated. From the immunoassays tested, PROL-CTK was the system which was less correlated with the Nb2 bioassay in group 2 (r=0.252; NS). Our experience is that macroprolactinemia is an asymptomatic condition in most of the cases. Therefore, we suggest that the routine measurement of PRL should be done with methods that are only minimally affected by the presence of macroprolactin. Such an approach would obviate the use of extensive, frequently expensive and ultimately useless diagnostic tests that are needed to determine the cause of the hyperprolactinemia.

Mapping the gene causing hereditary primary hyperparathyroidism in a Portuguese kindred to chromosome 1q22-q31.

A Portuguese kindred with autosomal dominant isolated primary hyperparathyroidism (HPT) that was associated with parathyroid adenomas and carcinomas was investigated with the aim of determining the chromosomal location of this gene, designated HPTPort. Leukocyte DNA from 9 affected and 16 unaffected members and 7 parathyroid tumors from 4 patients was used in comparative genomic hybridization (CGH), tumor loss of heterozygosity (LOH), and family linkage studies. The CGH studies revealed abnormalities of chromosomes 1 and 13, and the results of LOH studies were consistent with the involvements of tumor suppressor genes from these regions. Family segregation studies mapped HPTPort to chromosome 1q22-q31 by establishing linkage with eight loci (D1S254, D1S222, D1S202, D1S238, D1S428, D1S2877, D1S422, and D1S412) (peak two-point LOD scores = 3. 46-5.14 at 0% recombination), and defined the location of HPT Port to a 21 cM region flanked centromerically by D1S215 and telomerically by D1S306. Thus, HPTPort has been mapped to chromosome 1q22-q31, and a characterization of this gene will help to elucidate further the mechanisms that are involved in the development of parathyroid tumors.

Spontaneously occurring anti-PTH autoantibodies must be considered in the differential diagnosis of patients with elevated serum PTH levels.

We investigated the cause of elevated immunoreactive circulating parathyroid hormone (PTH) levels in two females of 41 (case 1) and 39 (case 2) years of age with low/normal serum calcium levels and hypocalcemia, respectively, and, in the latter case, hyperprolactinemia. Serum samples from both patients were fractionated by Sephadex G-100 Superfine chromatography. Fractions were assayed for PTH and prolactin (PRL) by immunoradiometric assays (IRMA) and for immunoglobulin G (IgG) by radial immunodiffusion. Sera from both patients were incubated with protein A and protein G Sepharose, centrifugated and the supernatant was assayed for PTH by IRMA. Sera were also subjected to affinity chromatography with an anti-human-IgG-agarose column. IgG and PTH or PRL were measured in the fractions by radial immunodiffusion and IRMA, respectively. In both cases the majority of serum PTH immunoreactivity eluted in the same fractions of IgG after gel filtration and was precipitated by protein A (89% in patient 1 and 96% in patient 2) and protein G (83% in patient 1 and 100% in patient 2), thus, behaving as IgG. In case 1, 79% of PTH was also retained by an anti-hIgG agarose column. High PRL levels in patient 2 were due to macroprolactinemia since most of PRL eluted as big,big (40%) and big-PRL (45%) after gel filtration. Forty-eight percent of PRL from patient 2 was retained by the anti-hIgG column indicating the presence of an anti-PRL autoantibody. These data suggest that spontaneously occurring anti-PTH autoantibodies must be considered in the differential diagnosis of patients with elevated serum PTH levels.

Emotional aspects of hyperprolactinemia.

Patients with hyperprolactinemia often present with emotional difficulties. These occasionally persist even after successful treatment. Insight into the roots of their diseased state makes a difference in the handling of all cases, but becomes crucial in the not-so-rare situations in which the normalization of hormonal levels is not followed by a feeling of cure. This chapter attempts to provide details, discuss and situate in context the following blocks of pertinent information: (1) prolactin acts upon the central nervous system and variations in its concentrations do affect mood, emotions and behavior; (2) most actions of prolactin are directed to metabolical and behavioral adaptation to pregnancy and the care of the young; (3) even in the absence of pregnancy prolactin secretion responds to environmental stimuli under specific conditions. Whether adaptive, as in the case of surrogate maternity, or pathological, as in the case of pseudopregnancy, prolactin responds to a perceived need to take care of a child; (4) the facts that the clinical onset of prolactinomas often follows life-events and that these tumors occur preferentially in women brought up under specific conditions suggest the possibility that psychological factors may predispose to prolactinomas; (5) dealing with individual cases requires the perception that the relations between prolactin, emotions and feelings are circular, i.e., prolactin affects the brain and mood but, on the other hand, personality traits and environmental factors may stimulate the secretion of prolactin and may play a role in the genesis of the disease.

Rapid weight gain, at least in some women, is an expression of a neuroendocrine state characterized by reduced hypothalamic dopaminergic tone.

A group of 17 consecutive regularly menstruating women who gained at least 5 kg the previous year (Group 1) was compared with a control group of similar age, parity and social class (Group 2). Galactorrhea was observed in 6/17 women from group 1 and in 1/16 women from group 2 (chi 2 4.571; p < .05). Average morning prolactin levels were higher in group 1 (8.15 +/- 4.92 micrograms/l) than in group 2 (5.29 +/- 2.48 micrograms/l; p < .05). The two groups were similar in their morning thyroxin, triiodothyronine, TSH, estradiol, cortisol, gastrin, cholecystokinin, somatostatin, oxytocin, insulin and IGF-1 levels. Leptin levels were significantly higher in group 1 than in group 2 (18.85 +/- 10.63 micrograms/l vs. 10.15 +/- 6.38 micrograms/l; p < .02) but this difference could be attributed exclusively to the higher body mass index (BMI) of group 1 (MANCOVA). Analysis of the distribution of basal prolactin levels in group 1 revealed a skewed distribution due to the presence of six outliers (Barnett and Lewis test associated with Mahalanobis distance) whose values were higher than the highest value found in group 2. These outliers were henceforth considered as subgroup 1a, and the remnant patients in group 1 as subgroup 1b. Besides the expected difference in basal prolactin levels between subgroups 1a and 1b (13.72 +/- 3.69 and 5.12 +/- 1.81 micrograms/l, respectively) and the higher frequency of galactorrhea in group 1a (4/6 vs. 2/11; p < .05) no other differences were observed in clinical or basal biochemical parameters. Following domperidone (10 mg, i.v.) the percentual increase in prolactin (delta Prl 20'/Prl 0') was significantly lower in group 1 than in group 2 (23.9 + 15.2 vs. 37.0 +/- 21.2; p < .05). In absolute values, the prolactin rise in subgroup 1a (100.7 +/- 45.5 micrograms/l) was significantly lower (p < .02) than that of subgroup 1b (157.3 +/- 50.3 micrograms/l) and group 2 (152.7 +/- 34.5 micrograms/l). Group 1 (and each one of its two sub-groups) also differed from group 2 in a higher incidence of meaningful life-events the year preceding the study. This study confirms previous observations that recent weight gain in women is preceded by important life-events and is associated with galactorrhea and increased prolactin levels in a number of them. Besides, it provides evidence that the increased prolactin levels are due to reduced hypothalamic dopaminergic tone.

An unusual case of papillary carcinoma of the thyroid with cutaneous and breast metastases only.

Cutaneous metastases of thyroid carcinoma are infrequent and, when present, are usually located in the vicinity of a widespread primary tumor. Breast metastases from these tumors are even less common. We report the case of a 64-year-old female with a toxic multinodular goiter in whom a fine-needle biopsy, performed in 1985 at the age of 52, was suggestive of papillary carcinoma of the thyroid. Total thyroidectomy for a papillary carcinoma, follicular variant, was performed in 1988. Four months after surgery, a cutaneous metastasis was discovered in the right thigh. Surgical excision of the lesion followed by treatment with radioactive iodine decreased serum Tg levels from 7495 to 3.3 micrograms/l. Under suppressive therapy with L-thyroxine, serum Tg remained undetectable for the next 4 years. Then, serum Tg levels rose to 3.9-5.6 micrograms/l and a second cutaneous metastasis was removed from the abdominal wall. The patient was again treated with radioactive iodine and the post-treatment whole-body scan did not show any area of increased uptake of the radionuclide. However, serum Tg levels under suppression with L-thyroxine remained elevated at 4-20 micrograms/l for the next 2 years. In August 1995, a 1.5 cm nodule was found in the right breast. Cytological examination was suggestive of a breast metastasis from thyroid carcinoma and the lesion was removed by enucleation. This proved to be a metastasis from a papillary carcinoma of the thyroid. Elevated (19-44 micrograms/l) serum Tg levels persisted postoperatively. A third cutaneous metastasis was revealed by 131I scintigraphy in the right buttock and surgically removed in December 1996. Serum Tg levels have remained undetectable since then. To the best of our knowledge, this is a unique case of a papillary carcinoma of the thyroid with a propensity to metastasize only to the skin and breast during a follow-up of 11 years.

ACTH-producing carcinoma of the pituitary with haematogenic metastases.

The case report is presented of a 47-year-old white woman with Cushing's disease treated by bilateral adrenalectomy in June 1981. A first computed tomography (CT) scan in September 1984 showed a voluminous pituitary adenoma with invasion of the sphenoid sinus and left parasellar extension. The tumour increased progressively in size, a suprasellar extension developed and the optic chiasm was eventually affected. In March 1988 and June 1989 the patient underwent two surgeries for the pituitary tumour, the second followed by radiotherapy. During this period, the ACTH values varied between 100 pmol/l and 403 pmol/l (normal: < 13 pmol/l). After radiotherapy, a progressive shrinking of the tumour was observed and the ACTH concentrations decreased to a lowest value of 27.5 pmol/l. The patient was clinically well until September 1993 when, suddenly, the plasma ACTH concentration increased to very high levels (greater than 965 pmol/l). There was no evidence of tumour growth on the sellar CT scan. In January 1995, an ACTH-producing pituitary carcinoma was diagnosed, based on the presence of bone metastases. The patient died in May 1995.

A possible role for D8/PSF-A-like sequences in lactotroph versus somatotroph expression of the human prolactin gene.

The transcription factor GHF-1/Pit-1 is essential for the expression of GH and prolactin (PRL) by somatotrophs and lactotrophs respectively. However, PRL is not expressed in mature somatotrophs despite the presence of GHF-1/Pit-1. A possible mechanism is the presence of a somatotroph-specific repressor in the 5'-flanking sequences of the PRL gene. The region -3500/-1750 of the human (h) PRL gene is associated with negative regulatory activity and contains an element, designated D8, that resembles repressor PSF-A sequences which are located in the distal upstream region of placental members of the human GH family. An internal deletion of D8 sequences resulted in a significant stimulation of promoter activity in somatotroph GC (P < 0.005) and somatolactotroph-like GH3 and GH4C1 cells (P < 0.05), but not lactotroph-like 235-1 cells after gene transfer. However, D8 binding was observed by nuclease protection with lactotroph- as well as somatotroph-like cell nuclear protein. Although proteins that bind to the D8 element appear ubiquitous, this element does yield tissue-specific complexes in mobility shift assays. Further, competition studies do not suggest an interaction between GHF-1/Pit-1 and D8 proteins. The hPRL D8 element was inserted upstream of a thymidine kinase promoter and used to transfect pituitary and non-pituitary HeLa cells, to assess intrinsic repressor activity and/or promoter specificity. Although no repression was observed, a significant ninefold increase in expression was observed in HeLa cells (P < 0.001) which was at least twofold greater than observed in any of the pituitary cell lines tested. These results implicate D8 in the somatotroph-specific repression of hPRL; however, they also suggest that D8 can act as a stimulator as well as a repressor, depending on the interaction of a ubiquitous D8 factor forming promoter and cell-specific complexes with other elements/factors.

Some forms of big big prolactin behave as a complex of monomeric prolactin with an immunoglobulin G in patients with macroprolactinemia or prolactinoma.

The purpose of this study was to characterize the structure of big big PRL (bb-PRL) in patients with macroprolactinemia or prolactinomas. Serum samples from these patients were fractionated by Sephadex G-100 chromatography, and PRL was measured in the eluate by an immunoradiometric assay. The fractions containing bb-PRL were subjected to affinity chromatography with an antihuman immunoglobulin G (IgG) agarose column. PRL and IgG were assayed in the fractions obtained after affinity chromatography by immunoradiometric assay and radial immunodiffusion, respectively. bb-PRL was also immunoprecipitated with an antihuman PRL antibody, followed by polyacrylamide gel electrophoresis and Western blotting. We found that an average of 60% (range, 27-87%) of bb-PRL from patients with macroprolactinemia was retained by the agarose, indicating that this form of PRL contains an IgG. In one of the patients with prolactinoma, the big big form constituted 76% of the total PRL immunoreactivity. Most (75%) of the bb-PRL from this patient behaved as an IgG after affinity chromatography. In the two other patients with prolactinoma, we found that 22% and 25% of the bb-PRL, which represented only 2% and 3% of the total PRL in the serum, reacted as an IgG. In both groups of patients, the 23-kilodalton form of PRL was detected after the immunoprecipitation of bb-PRL. These results show that bb-PRL is in part a complex of 23-kilodalton PRL with an IgG and not an antibody mimicking the actions of PRL, as has been demonstrated for some large forms of growth-hormone.