Effect of vasoactive intestinal polypeptide (VIP) on growth hormone (GH) and prolactin (PRL) release and cell morphology in human pituitary adenoma cell cultures.

Six GH adenomas and three prolactinomas were investigated by light- and electron-microscopic morphological and immunocytochemical methods and the effect of vasoactive intestinal polypeptide (VIP) on growth hormone (GH) and prolactin (PRL) secretion was tested in vitro. The tumour cells of the acromegalic patients revealed both GH and PRL immunoreactivity while prolactinomas showed only PRL activity. All the adenomas stained immunocytochemically also for VIP. By electron microscopy, the tumours included two densely and two sparsely granulated GH, two mixed GH/PRL, and three sparsely granulated PRL adenomas. The dissociated cells were explanted, and cultured in vitro. The cultures in micro test plates were treated with VIP at different concentrations between 10(-5)-10(-12) M. GH and PRL contents in the culture media were measured by radioimmunoassay. GH release was significantly stimulated by VIP in a dose-dependent manner over the whole concentration range, while VIP was effective on the PRL release only at 10(-6)-10(-7) M concentration. The cells of a mixed adenoma were grown in Petri dishes and used for ultrastructural and immunocytochemical studies. The cytoplasmic structure of the cells treated with VIP corresponded to that of active hormone-secreting cells with large ergastoplasmic fields and Golgi zones containing secretory granules. Massive exocytotic events were encountered mainly in the GH-type cells. GH and PRL double immunocytochemistry showed the predominance of GH cells, many of them containing low amounts of PRL as well. Cells predominantly containing PRL were spread among them, they also might contain GH as well. Some of the cells contained only a single immunoreactive hormone. The intensity of gold labelling of the secretory granules appeared higher in the VIP-treated cells than in the untreated control ones which showed a cytoplasmic structure characteristic of glandular cells with low secretory activity. As all the adenoma cells both contained and reacted to VIP, our results are in agreement with an autocrine or paracrine effect of this peptide. The fine structure of the cells in the cultures treated with VIP supply an additional argument to the assumption that VIP may serve as a growth factor for these cell types.

Locomotion and proliferation of glioblastoma cells in vitro: statistical evaluation of videomicroscopic observations.

OBJECT: The motility and doubling of human glioblastoma cells were investigated by means of statistical evaluation of large sets of data obtained using computer-aided videomicroscopy. METHODS: Data were obtained on cells in four established glioblastoma cell lines and also on primary tumor cells cultured from fresh surgical samples. Growth rates and cell cycle times were measured in individual microscopic fields. The averages of cell cycle time and the duplication time for the recorded cell populations were 26.2 +/- 5.6 hours and 38 +/- 4 hours, respectively. With these parameters, no significant differences among the cell lines were revealed. Also, there was no correlation in the cell cycle time of a parent cell and its progeny in any of the cultures. Statistical analysis of cell locomotion revealed an exponential distribution of cell velocities and strong fluctuations in individual cell velocities across time. The average velocity values ranged from 4.2 to 27.9 micro/hour. In spite of the uniform histopathological classification of the four tumors, each cell line produced by these tumors displayed distinct velocity distribution profiles and characteristic average velocity values. A comparison of recently established primary cultures with cell lines that had propagated multiple times indicated that cells derived from different tumors sustain their characteristic locomotor activity after several passages. CONCLUSIONS: It can be inferred from the data that statistical evaluation of physical parameters of cell locomotion can provide additional tools for tumor diagnosis.

Transformation of a human mixed GH-PRL adenoma cell population in response to long-term bromocriptine treatment.

A child was operated 3 times because of a recurrent growth hormone- and prolactin-producing pituitary adenoma. Between the operations she was treated for five years with bromocriptine. The characteristics of the tumour cell population collected after the last operation was now examined by electron microscopy, immunocytochemistry, and in tissue culture and compared to those of the primary tumour cells reported earlier. A prominent change was the reversal of the proportion of the densely and sparsely granulated cells in favour of the GH-type, densely granulated cells. These cells, some of them coexpressing PRL, did not essentially change their characteristics either in vivo or in culture. On the other hand, pleomorphous cells with smaller cytoplasmic area and prominent lysosomal structures represented the sparsely granulated population containing PRL or both PRL and GH. The morphological alteration of the PRL-type cells was also reflected in vitro. Hence, while GH-type cells prevail, at least a sub-population of PRL-type cells survives long-term bromocriptine administration. A shift in the incidence of the two cell types in favour of the GH-type cells explains the change in the endocrine status of the patient.

The role of psychosocial and biological variables in separating chronic and non-chronic major depression and early-late-onset dysthymia.

Psychosocial (sociodemographic characteristics, loss and separation and family atmosphere in childhood, recent life events) and biological (family history, DST, TRH-test) variables were investigated in 180 patients with Major Depression (MD) and Dysthymic Disorder (DD). The aim of the study was to reveal certain differences between the chronic and non-chronic course of MD and the early- and late-onset subtypes of dysthymia. When comparing the two course patterns of MD, a higher rate of malignant tumours among first-degree relatives, a greater number of long-lasting stress situations before the index depressive episode, longer duration of the previous episodes, less frequent DST nonsuppression, and a blunted TSH response to TRH were found in patients with a chronic course of MD. Several factors seem to influence the course pattern of MD, or else the chronic form represents a subgroup within MD. The late-onset dysthymics were mainly women with a low level of education, a lower suicidal tendency, normal suppression in DST, and a lack of blunted TSH responses to TRH administration during the period of double depression. The early-onset dysthymics showed a higher number of persons who had never married, who presented a more traumatic and frustrating childhood background, and who had a higher rate of DST non-suppressors and blunted TSH responses after TRH administration during the period of their double depression. Our data suggest that late-onset dysthymia might be a biologically distinct subgroup of chronic depression.

[Reducing the incidence of fatal pulmonary embolism by preventive administration of heparin]. / Halálos tüdöembóliák számának csökkenése heparin preventióra.

The authors had introduced the heparin prevention routinely from 01 January 1985 on the patients with predisposing factors in consideration of the contraindications. The death of pulmonary embolism was compared retrospectively with the pre- and post-prevention five years period in an autopsy material. During 10 years 340 patients died in pulmonary embolism. During the first period the total mortality was 24.6% because of pulmonary embolism, but in the second period it was only 19.4%. This percentage would be improved with more widely used heparin prevention (early and sufficient dose).

Pathological and experimental investigations in a case of gigantism.

A pituitary adenoma was transsphenoidally removed from a 4.5-year-old girl suffering from gigantism. Prior to the operation both the growth hormone (GH) and the prolactin (PRL) levels in the serum were elevated. By light microscopy the tumor appeared to be an acidophilic adenoma. Two distinct cell types, the densely granulated and the sparsely granulated cells, could be distinguished by electron microscopy. Double immunolabeling revealed the presence of GH alone in some densely granulated cells and PRL alone in some sparsely granulated cells, as well as GH and PRL co-localized in both of the morphologically distinguished cell types. Both cell types were identified in the monolayer and the suspension cultures by electron microscopy. GH and PRL concentrations in the culture media were measured by radioimmunoassay. The basal secretion of growth hormone was almost uniform during the 3-week cell culture period. GH and PRL release was significantly inhibited by bromocriptine. Our studies revealed a bimorphous and bihormonal mixed adenoma in childhood.

Characterization of human gliomas by a monoclonal antibody both on tissue culture and paraffin-embedded sections.

A monoclonal antibody designated OITIC3-11 was produced against GFAP positive human glioblastoma multiforme tumour cells. The specificity of the monoclonal antibody was tested on different types of human brain tumours and on normal adult brain both on tissue cultures and paraffin-embedded sections. The OITIC3-11 monoclonal antibody reacted with 16 of 18 malignant and 1 of 6 benign gliomas but did not react with meningioma, pituitary adenoma, metastatic brain tumours and normal adult brain tissue.

Intrasellar hamartoma associated with pituitary adenoma.

The histological, ultrastructural and immunocytochemical features are reported of an intrasellar neuronal and lipomatous hamartoma associated with pituitary growth hormone (GH) cell adenoma and acromegaly. Electron microscopy demonstrated a close contact between neurons and adenomatous GH cells. By immunohistochemistry the adenoma cells revealed a positive staining for GH and prolactin. The neurons of hamartoma showed neurosecretory activity which might have induced the development of pituitary GH cell adenoma.

[Clinical evaluation of autopsy in the aged]. / Klinische Bewertung der Autopsie im Senium.

On a general medical department with 54 beds, 801 patients (8.6%) expired between 1980 and 1984 in 780 of them (97.3%) autopsy has revealed. 58.4% of expired patients were over seventy years. In this age group autopsy data diverged from the underlying disease in 9.58%, however this ratio of patients under seventy was 8.56%, as well. Mistake of clinical diagnosis in the direct cause of death was 18.9% in the group of under seventy and 21.3% of over seventy. The highest ratio of mistakes in the underlying disease has occurred in diseases of malignancies (progressing with age). Referring to the direct cause of death, undiagnosed pulmonary embolism had in each age group an equal high ratio. Because of the augmented multimorbidity with age the autopsy in the elderly can serve for an improvement of diagnosis.

Proteinase inhibitors while influencing hormone release do not affect cell morphology of hypophyseal cultures.

Cultured cells from adult rat anterior pituitaries and intermediate lobes were treated with proteinase inhibitor substrate analogues (Boc-DPhe-Pro-Arginal [BOC-DPPA], DPhe-Pro-Arginal [DPPA], BOC-DPhe-Leu-Lysinal [BOC-DPLL], BOC-DPhe-Phe-Lysinal [BOC-DPPL]) to elucidate their effect on cell morphology. It was established that BOC-DPPA and DPPA (which in previous studies stimulated alpha-MSH release [6]) caused a slight decrease in the number of immunoreactive secretory granules in melanotrophs. BOC-DPLL, which inhibited growth hormone and prolactin release, did not alter the fine structural features of cultured cells. No difference was observed in the membrane turnover traced by cationic ferritin when cells were treated with BOC-DPPL. We suggest that substrate analogues used are harmless to pituitary cells.

Dissimilar responsiveness of cultured corticotrophs and melanotrophs to tripeptide aldehydes.

Cultured cells from adult rat anterior pituitaries or intermediate lobes were treated with the proteinase inhibitor tripeptide aldehydes BOC-DPhe-Pro-Arg-H (Boc-fPRH) and DPhe-Pro-Arg-H (fPRH), ovine corticotropin-releasing factor (oCRF), and bromocriptine. One millimolar fPRH stimulated basal, and slightly enhanced oCRF-induced ACTH release by melanotrophs in short-term experiments. The basal release of alpha-MSH was also stimulated by the drug. In long-term experiments, fPRH elevated markedly both the release and the intracellular level of ACTH; BOC-fPRH caused an increased alpha-MSH release. Tritiated fPRH had no preference for POMC-producing cells and BOC-fPRH or fPRH were harmless to the cell morphology. In anterior pituitary cell cultures, fPRH diminished slightly basal and oCRF-induced ACTH release. Bromocriptine was ineffective on corticotrophs, however, in melanotrophs it inhibited ACTH release markedly with or without fPRH in the medium. The dissimilar responsiveness of the corticotrophs and melanotrophs to the peptide aldehydes may be interpreted in terms of their differing membrane receptors or intracellular mechanism of stimulus-secretion coupling.

DNA synthesis in lactotrophs of primary rat anterior pituitary cell cultures. Effects of thyroliberin, bromocriptine and somatostatin.

Prolactin immunostaining in combination with thymidine autoradiography was used to characterize changes in the DNA-synthesizing activity of lactotrophs in primary monolayer cultures of the rat anterior pituitary gland treated for 3 days with thyroliberin (TRH), somatostatin (SRIF) and bromocriptine (CB 154). The number of lactotrophs labelled with 3H-thymidine within the total pool of labeled pituitary cells was used to estimate DNA synthesis in prolactin-producing cells. TRH (10 ng/ml) stimulated DNA synthesis in the whole population of cultured cells but not in lactotrophs. TRH only weakly counteracted the noticeable inhibitory effect of CB 154 (0.75 microM/l) on thymidine incorporation into lactotrophs. SRIF (20 ng/ml) inhibited DNA synthesis in lactotrophs to a lesser extent than CB 154. The combination of methods used in this paper may be useful for studying the selective effects of regulators on the proliferative activity of various pituitary cell types in vivo and in culture.

Responsiveness of intermediate lobe cells to neural mediators in culture.

Immunologically and biologically active ACTH, as well as biologically active alpha-MSH were secreted by monolayer cultures of rat pituitary intermediate lobe cells. The release of the immunoreactive ACTH and the bioactive alpha-MSH was inhibited by dopamine and by the dopamine agonist bromocriptine. The inhibition is probably mediated by specific dopaminergic receptors, since simultaneously added haloperidol prevented the dopamine induced hormone release inhibition. Pro-Leu-Gly-NH2, a potent inhibitor of the alpha-MSH secretion, had no effect on the release of ACTH. Synthetic ovine CRF stimulated the release of both immunoreactive ACTH and bioactive alpha-MSH in dose-dependent fashion. Corticosterone was without effect on hormone release in the cultures derived from both adrenalectomized and sham-operated animals.

Calcium modifies the effects of thyroliberin and somatostatin on hormone release from cell cultures of the rat anterior pituitary.

Role of calcium (Ca2+) in the effects of thyroliberin (TRH) and somatostatin (SRIF) on the release of growth hormone (GH), prolactin (PRL) and thyroid stimulating hormone (TSH) from the rat adenohypophyseal cells in primary monolayer cultures has been studied. Decrease of extracellular Ca2+ diminished the stimulatory effects of TRH on TSH and PRL release. Ca2+ is also an important factor in the mechanism of SRIF action. Data obtained in the experiments with high Ca2+ levels in the medium indicate that some antagonistic interrelationship exists between Ca2+ and SRIF. These results suggest that the participation of cAMP alone is not sufficient for stimulus-secretion coupling. Another messenger, namely Ca2+, is necessary for the effects of hypothalamic hormones. On the other hand, the contribution of Ca2+ to the secretory process in mammotrophs, somatotrophs and thyrotrophs is not equal. PRL and TSH secretion is more dependent on the presence of extracellular Ca2+ than the release of GH.

Surface topography of cultured anterior pituitary cells of the rat.

Anterior pituitary cells of the adult rat were studied by scanning electron microscopy after 3, 6 and 8 days in culture. Epithelial (parenchymal) cells and fibroblasts could be differentiated by their characteristic cell surface structures and by their associations with each other. The surface of solitary and grouped epithelial cells is uneven and rich in smaller or larger blebs, wrinkles and microvilli. A few solitary cilia were found on the surface of some cells. The individual cells forming epithelial cell groups adhere to each other with numerous, parallel, slender extensions. The fibroblasts found in monolayers have extensions with various lengths contacting other cells or the inert substratum. Microvilli arise in various frequency from their smooth surface. Transmission electron microscopy confirmed the presence of the surface structures detected in the scanning electron microscope.

Ba2+ affects growth hormone and prolactin secretion as well as cell morphology in rat anterior pituitary cultures.

The effects of barium ions (Ba2+) on the release of growth hormone (GH), and prolactin (PRL) and ultrastructural changes accompanying altered hormone secretion in primary cultures of the rat anterior pituitary gland were studied in the presence or absence of calcium ions (Ca2+) in the medium. In four days old cultures previously labelled with 14C-L-leucine radiolabelled PRL release was stimulated in the presence of both Ba2+ and Ca2+, while GH release remained unchanged. In the absence of Ca2+, Ba2+ caused a more marked PRL and a significant GH release. In seven days old cultures, Ba2+ stimulated immunoreactive GH release in the presence of Ca2+ three-fold and a dramatic GH release occurred when Ca2+ was omitted from the incubation medium. Many parenchymal cells contracted profoundly in the presence of Ba2+ without Ca2+ in the incubation medium and the discharge of secretory material was frequent in a number of granular cells that did not contract. The observation on contraction suggests that Ba2+ may activate the microtubular-microfilamentous system in pituitary parenchymal cells.