Neutrophil predominance in induced sputum from asthmatic patients. Therapeutic implications and role of clara cell 16-KD protein.

UNLABELLED: Eosinophil is considered to be a main protagonist in asthma; however, often discordances between clinical manifestations and response to treatment are observed. We aimed to determine the occurrence of neutrophil predominance in asthma and to identify its characteristics on the basis of clinical-functional features, induced sputum cellular pattern and soluble molecules, to guide the appropriated anti-inflammatory therapy. A total of 41 patients were included in randomized groups: 21-40 year-old, with stable mild-to-severe asthma, steroid-naïve and non-smokers. An induced sputum sample was obtained under basal conditions, a second one after treatment with budesonide (400 ug b.i.d.) or montelukast (10 mg/d) for six weeks, and a final one after a 4-week washout period. By cytospin we evaluated eosinophil (EP) or neutrophil predominance (NP), and in supernatant we determined LTE4, and CC16. Peak expiratory flow variability (PEFV) was measured. A total of 23/41 patients corresponded to EP and 18/41 patients to NP. The PEFV was higher in EP than in NP. LTE4 was higher with NP than with EP. No difference was found for CC16. Montelukast reduced the predominant cell in both subsets, whereas budesonide only reduced eosinophils in EP. Budesonide and montelukast reduced PEFV in EP but not in NP. Considering the total treated-samples in each subset, CC16 level increased significantly in EP. IN CONCLUSION: a NP subset of asthmatic patients was identified. These patients show a lower bronchial lability; the leukotriene pathway is involved which responds to anti-leukotriene treatment. This phenotype shows a poor recovery of CC16 level after treatment.

Neutrophil predominance in induced sputum from asthmatic patients: Therapeutic implications and role of clara cell 16-KD protein / Predominio de neutrófilos en esputo inducido de asmáticos: Implicancias terapéuticas y rol de la proteína 16-kD de la célula clara

Eosinophil is considered to be a main protagonist in asthma; however, often discordances between clinical manifestations and response to treatment are observed. We aimed to determine the occurrence of neutrophil predominance in asthma and to identify its characteristics on the basis of clinical-functional features, induced sputum cellular pattern and soluble molecules, to guide the appropriated anti-inflammatory therapy. A total of 41 patients were included in randomized groups: 21-40 year-old, with stable mild-to-severe asthma, steroid-naïve and non-smokers. An induced sputum sample was obtained under basal conditions, a second one after treatment with budesonide (400 µg b.i.d.) or montelukast (10 mg/d) for six weeks, and a final one after a 4-week washout period. By cytospin we evaluated eosinophil (EP) or neutrophil predominance (NP), and in supernatant we determined LTE4, and CC16. Peak expiratory flow variability (PEFV) was measured. A total of 23/41 patients corresponded to EP and 18/41 patients to NP. The PEFV was higher in EP than in NP. LTE4 was higher with NP than with EP. No difference was found for CC16. Montelukast reduced the predominant cell in both subsets, whereas budesonide only reduced eosinophils in EP. Budesonide and montelukast reduced PEFV in EP but not in NP. Considering the total treated-samples in each subset, CC16 level increased significantly in EP. In conclusion: a NP subset of asthmatic patients was identified. These patients show a lower bronchial lability; the leukotriene pathway is involved which responds to anti-leukotriene treatment. This phenotype shows a poor recovery of CC16 level after treatment.

El eosinófilo es considerado la célula protagonista principal en el asma; sin embargo, a menudo se observan discordancias entre las manifestaciones clínicas y la respuesta de los pacientes al tratamiento. Nos propusimos determinar la ocurrencia de predominio de neutrófilos en el asma e identificar las características clínico-funcionales, el patrón celular y las moléculas solubles del esputo inducido, para guiar el tratamiento apropiado anti-inflamatorio. Se incluyeron 41 pacientes: 21 a 40 años de edad, con asma estable leve a grave, no tratados con esteroides tópicos ni sistémicos y no fumadores. Se obtuvo una muestra de esputo inducido en condiciones basales, una segunda muestra después del tratamiento al azar con budesonida (400 µg dos veces al día) o el montelukast (10 mg/d) durante seis semanas, y una final después de un período de lavado de 4 semanas. En el frotis por citocentrifugado se evaluó el predominio de eosinófilos (EP) o neutrófilos (NP), y en el sobrenadante se determinó LTE4, y CC16. Se midió la variabilidad del flujo espiratorio máximo (PEFV). Un total de 23/41 pacientes correspondieron al EP y 18/41 pacientes con NP. El PEFV fue mayor en el EP que en NP. LTE4 fue mayor en NP que en EP. No se encontraron diferencias de los niveles de CC16 en ambos grupos. Montelukast redujo la célula predominante en ambos subgrupos, mientras que budesonida sólo redujo los eosinófilos en EP. Tanto budesonida como montelukast redujeron PEFV en EP, pero no en NP. El nivel de CC16 aumentó significativamente en el EP luego del tratamiento antiinflamatorio. En conclusión: se identificó un subgrupo de asmáticos NP que presentan una menor labilidad bronquial, la vía de los leucotrienos parece estar involucrada y responde al tratamiento anti-leucotrienos. Este fenotipo muestra una escasa recuperación del nivel de CC16 posterior al tratamiento.

The initial segment of the rat epididymis is able to uptake immature germ cells shed by testicular damage.

The initial segment of the caput epididymidis, the most proximal part of the rat epididymis, has specific functional characteristics. In the present study, the behavior of the epididymal epithelium from this region was evaluated after the exposure to a massive number of immature germ cells in the luminal fluid. The experimental release of immature germ cells from the seminiferous tubules was performed by injecting anti-microtubule compounds into the rete testis and the lumen of seminiferous tubules. Twenty-four hours after nocodazole or colchicine administration, a massive phagocytosis of immature spermatogenic cells, recognized as acrosin-positive structures, was easily observed in the epithelium of the initial segment of the epididymis assessed by light and electron microscopy. Immature germ cells were engulfed by epithelial cells, where most of them were found as cell debris at different stages of degradation. No signs of inflammation were observed either in the lumen or in the interstitium. The phagocytosis of immature germ cells was restricted to the epithelium of the initial segment of the epididymis, suggesting a role for this segment as the first selective barrier for the exclusion of abnormal gametes along the male genital tract.

Increased expression of uteroglobin associated with tubal inflammation and ectopic pregnancy.

OBJECTIVE: Evaluation of uteroglobin (UG) expression in the fallopian tube in different tubal diseases. DESIGN: The UG was screened and quantified in samples of fallopian tubes from patients with salpingitis, hydrosalpinx, and ectopic pregnancy by exposing the UG with immunohistochemical techniques. SETTING: University hospital and electron microscopy center. PATIENT(S): Women with pelvic inflammatory disease (PID) and complicated tubal ectopic pregnancy consulting for medical care. INTERVENTION(S): Salpingectomy. MAIN OUTCOME MEASURE(S): Tubal tissues were collected and examined using regular pathologic techniques. The UG immunoreactivity in the tubal epithelium was also assessed. RESULT(S): Fallopian tube epithelium displayed an increased UG expression in patients with PID and complicated tubal pregnancy compared with control patients. CONCLUSION(S): Uteroglobin is present in the human fallopian tube as a secretory protein and appears to be involved in immunosuppressive responses in the fallopian tube.

Budesonide effects on Clara cell under normal and allergic inflammatory condition.

Clara cells are nonciliated secretory cells implicated in lung homeostasis by the synthesis of immunomodulatory and host defense products, being one of the most important the CC16 protein. In this study, we compared the effects of budesonide (BUD), an inhaled corticoid, on Clara cell biology and its ability to reverse morphofunctional changes induced in an allergic airway hyper-responsiveness mouse model. In normal mice, exposure to BUD induced morphological changes compatible with a state of maximal differentiation on CC16 positive cells which developed a prominent cupola filled up with numerous mitochondria rich in CYP2E1, a member of the cytochrome P450 family. Consequently, CYP2E1 expression raised significantly. Exposure to OVA provoked hypertrophy of Clara cells and an increment in their number per millimeter of basal membrane. These cells acquired a mucous cell phenotype characterized by a notorious expansion of the secretory granular content. Synthesis of CC16 was greatly up-regulated concurrent to the finding of MUC5AC expression and the increment of epidermal growth factor receptor (EGFR). Mitochondrial content decreased significantly with a consequent reduction in CYP2E1 expression. After BUD treatment of OVA-challenged animals, the majority of Clara cells regained their normal morphology and functional characteristics; CYP2E1 levels raised when compared to the OVA exposed group. The BUD potential to differentiate Clara cells appeared to be important for the regression of the profound changes generated by the allergic injury. These results demonstrated the wide range of stimuli that can modify different aspects of Clara cell biology, and highlighted the effects of budesonide as a modulator of P450 enzymes, which probably contributes to a complementary antiinflamatory activity.

Ultrastructural immunolocalization of IGF-1 and insulin receptors in rat pituitary culture: evidence of a functional interaction between gonadotroph and lactotroph cells.

We have investigated the expression of receptors for insulin and insulin-like growth factor 1 (IGF-1) in rat pituitary cells in vitro and examined the morphological and proliferative changes induced in adenohypophyseal cells by insulin and IGF-1. The proliferation of lactotrophs was determined by double-immunostaining for bromodeoxyuridine and prolactin. Incubation with insulin (10, 100 or 1000 ng/ml) or IGF-1 (5, 30 or 100 ng/ml) for 48 or 72 h significantly increased the number of lactotrophs undergoing mitosis. Co-incubation of insulin or IGF-1 with genistein (25 microM), an inhibitor of the tyrosine kinase receptor, reduced the proliferation of lactotrophs elicited by the hormone and the growth factor. The receptors for insulin and IGF-1 were localized in intact pituitary cells by ultrastructural immunocytochemistry with the colloidal gold-protein A technique. Gonadotrophs expressed both receptors, specific labelling being restricted to this cell type. Electron-microscopical observations of pituitary cell cultures incubated with insulin or IGF-1 revealed gonadotroph cells exhibiting the fine-structural features of enhanced protein synthetic activity. These findings suggest that both insulin and IGF-1 are able to induce the proliferation of lactotrophs through an indirect mechanism mediated by a factor synthesized by gonadotroph cells, in addition to stimulating the biosynthetic activity of the gonadotroph in a direct manner.

Pattern of FGF-2 isoform expression correlated with its biological action in experimental prolactinomas.

Fibroblast growth factor-2 (FGF-2) synthesized in the pituitary is involved in the formation and progression of pituitary tumors. The aim of this study was to analyze the pattern expression of two FGF-2 isoforms at different subcellular levels and to determine its correlation with prolactinoma development. Estrogen administration to male rats for 7, 20, and 60 days generated pituitary tumors, with lactotrophs being the prevalent cell type. Ultrastructural immunolabeling showed FGF-2 in the cytosolic and nuclear compartments of somatotrophs, lactotrophs and gonadotrophs, as well as in folliculo-stellate cells of normal rats. Estrogen stimulation increased FGF-2 immunoreactivity in various tumors and enhanced the expression of two FGF-2 isoforms, 18 and 22 kDa, as quantified by western blot. The 18 kDa isoform observed in cytosol extracts reached the highest levels after 60 days of hormonal stimulation and this was related to lactotroph proliferation. However, the 22 kDa FGF-2 isoform was only detected in the nuclear compartment and achieved the maximum expression at 7 days of estrogen treatment, without any correlation with lactotroph proliferation. These results suggest that the 18 kDa FGF-2 may play a role in the modulation of lactotroph proliferation in prolactinomas induced by estrogen. The overproduction of both FGF-2 isoforms appears to be implicated in autocrine-paracrine-intracrine mitogenic loops; this FGF-2 activity could lead to uncontrolled cell growth, angiogenesis, and tumor formation.

Toll-like receptor 4 in rat prostate: modulation by testosterone and acute bacterial infection in epithelial and stromal cells.

The prostate gland is the most inflammation-prone organ in the male reproductive tract. However, little information is available regarding the immunobiology of this gland. Toll-like receptor 4 (TLR4) is considered to be a major sensor of danger signals and a key trigger of the innate immune responses. TLRs have also been implicated in the development of different inflammatory diseases in organs in which epithelial-stromal interactions are critical for homeostasis. The purpose of this work was to evaluate the presence and regulation of TLR4 in the rat prostate. Western blot and immunocytochemical studies revealed that constitutive expression of TLR4 in the rat ventral prostate was localized in the epithelial cells, mainly associated with the rough endoplasmic reticulum, as well as in smooth muscle cells in the stroma. In addition, increased concentrations of TLR4 were found in castrated rats, predominantly in hypertrophied smooth muscle cells. On the other hand, using a bacterial prostatitis model, we observed an increment in the TLR4 cytoplasmic content and migration of this receptor to the apical plasmatic membranes of epithelial cells at 24 h and 48 h post-infection. These findings suggest that the prostate gland is able to recognize pathogens and to initiate immune responses. In addition, TLR4 appears to be implicated in the vital stromal-epithelial interactions that maintain prostate homeostasis during prostatitis, as well as following androgen deprivation.

Antagonic effects of oestradiol in interaction with IGF-1 on proliferation of lactotroph cells in vitro.

The effects of IGF-1, 17 beta oestradiol and its functional interaction on lactotrophs cell proliferation were evaluated. In addition we investigated the involvement of PKC alpha, epsilon and phosphorilated ERK, in the mitogenic process. Primary cell cultures of adenohypophysis from female Wistar rats were studied in serum free conditions. The proliferation of lactotrophs was determined by double immunostaining for BrdU and PRL. The incubation with IGF-1 5, 30 or 100 ng/ml during 48 or 72 h increased lactotrophs proliferation two-threefold depending on IGF-1 concentration. Co-incubation of IGF-1 (30 ng/ml) with genistein (25 microM) or BIM (0.5 or 2 microM), lowered of tyrosine kinase receptor or of PKC respectively, inhibited the induced IGF-1 lactotrophs proliferation. 17 beta oestradiol (1, 10 or 100 nM) had not mitogenic effect, whereas in the presence of serum PRL cells proliferation was stimulated. Co-incubation with 1 nM oestradiol and IGF-1 significantly decreased the lactotroph BrdU-labelling achieved with IGF-1. PKC alpha, epsilon and ERK1/2 levels measured by western blot augmented in the presence of IGF-1 and were inhibited with the addition of genistein, supporting a participation of these enzymes in the proliferate process. Co-incubation of IGF-1 with 1 nM oestradiol decreased both PKC isoforms and activated ERK1/2 levels, suggesting that oestradiol would exert its antiproliferative effect by acting on the signalling pathway of IGF-1. The results revealed antagonic effects of oestradiol on lactotroph proliferation depending on its concentration and the presence of IGF-1.

Subcellular localisation of VEGF in different pituitary cells. Changes of its expression in oestrogen induced prolactinomas.

Vascular endothelial growth factor (VEGF) is an important angiogenic factor in the pituitary gland. The objective of this study was to unveil the VEGF subcellular localisation in different pituitary cell types and to evaluate changes in its expression at different time intervals after oestrogen stimulation. A relevant feature demonstrated was the identification of this cytokine in the nucleus and cytoplasm of lactotrophs, somatotrophs and gonadotrophs, as well as in follicle-stellate cells of male rats. Oestrogen treatment increased the number of VEGF immunopositive cells and its expression detected differentially by western blot in both nucleus and cytoplasm of pituitary cells when compared to the control. At ultrastructural level VEGF appeared associated with nucleolus and euchromatin involving a possible internal autocrine loop. In lactotrophs, the predominant cell of the tumour, VEGF was immunodetected in RER, Golgi complex, and vesicular organelles, supporting further the association with an auto-paracrine effect exerted by VEGF. The nucleus/cytoplasm ratio of VEGF revealed a prevalent accumulation of VEGF in the cytoplasm. The presence of VEGF in the nucleus may probably be associated with a translocation to this cell compartment. This study demonstrated a cytoplasmic and nuclear immunolocalisation of VEGF in normal and tumoural adenohypophyseal cells. In the course of prolactinoma development, the oestrogen stimulated VEGF expression in tumoural cells, promoting a vascular adaptation which contributes to growth and progression of the tumour.

Soluble p-cadherin found in human semen.

Cadherins constitute a family of calcium-dependent cell-cell adhesion molecules. P (placental)-cadherin is a 118-kd protein expressed by basal cells in epithelial tissues. P-cadherin also has been described as a soluble protein in certain biological fluids, including human serum and breast milk. Here, we report the presence of an 80-kD fragment of P-cadherin in human semen. No significant differences were found in semen samples from fertile and nonfertile patients. Our results add evidence to previous data indicating that soluble fragments of P-cadherin have a widespread distribution in bodily fluids and suggest that soluble P-cadherin might have functions other than basal epithelial cell-cell adhesion.

Immunolocalization of Pit-1 in gonadotroph nuclei is indicative of the transdifferentiation of gonadotroph to lactotroph cells in prolactinomas induced by estrogen.

The pituitary protein transcription factor (Pit-1) regulates the differentiation and proliferation of somatotrophs, lactotrophs, and thyrotrophs and the c-Myc oncoprotein plays a critical role in somatotroph and lactotroph differentiation. Both were involved in the genesis of pituitary tumors. The combined analysis of Pit-1 and c-Myc expression and the morphometric and biochemical parameters of the lactotroph population after treatment with estrogen for 7, 20, and 60 days provided new information on molecular mechanisms implicated in the formation of prolactinomas. Estrogen treatment for 7 days caused a significant proliferation of lactotrophs (70%) and this increase reached an additional 55% at 60 days. The proliferation of lactotrophs was concurrent with higher serum and pituitary prolactin levels. An augmentation of Pit-1 and c-Myc expression in both cytoplasmic and nuclear extracts after estrogen can be associated with lactotroph proliferation. Moreover, the multistep correlation analysis revealed that the expression of nuclear Pit-1 was the strongest predictor of prolactinoma development. Also the Pit-1 immunolocalization in nuclei of gonadotrophs suggests the activation of genes involved in transdifferentiation of gonadotroph to lactotroph. Therefore, the understanding of the Pit-1 function may help in the design of strategies to control the secretion and proliferation of pituitary tumors of the somatomammotrope lineage.

Regulation of uteroglobin/Clara cell protein expression after acute lung exposure to an organophosphoreted insecticide.

Uteroglobin (UG) or Clara cell protein (CC16), the main secretory product of bronchiolar Clara cells, plays an important protective role in the respiratory tract against inflammatory processes. In the lung, protein secretion is regulated by glucocorticoids, but also proinflammatory cytokines, such as interferon-gamma (IFN-gamma) and TNF-alpha, have been found to modulate the expression of this peptide. We have previously demonstrated that the acute exposure to an organophosphoreted insecticide induces an enhanced production of UG/CC16 by Clara cells. In the present report, we worked with intact and adrenalectomised (ADX) animals to study the mechanism involved in the UG/CC16 increase caused by the insecticide and the role played by a glucocorticoid (dexamethasone; DEX). In intact rats we found that DEX treatment could not reproduce such an increase of UG/CC16 synthesis with pharmacological doses. In ADX rats, even though glucocorticoid deprivation provoked a strong inhibition of UG/CC16 synthesis, the exposure to the organophosphoreted insecticide stimulated the synthesis of the protein, shown by the great accumulation of secretory granules in the cytoplasm of Clara cells and the increase of UG/CC16 detected by immunocytochemistry and western blot. These results imply that glucocorticoids are not essential to trigger the increase of UG/CC16 in response to an injury, and they also suggest an involvement of other molecules associated with inflammation. In coincidence with these observations, we have found that IFN-gamma, a proinflammatory cytokine, increased after insecticide exposition in both groups, intact and ADX, mainly in ADX rats. The stimulation of UG/CC16 synthesis occurring during inflammatory processes of the respiratory tract caused by acute inhalation of a toxicant appears to be functional without the intervention of glucocorticoids and mediated by IFN-gamma as a mechanism for local control of the inflammatory response.

Apoptotic cell death of oestrogen activated lactotrophs induced by tamoxifen.

Stimulation and inhibition of lactotroph cells cause remarkable morphological and functional changes. In keeping with these changes, the size of the lactotroph cell population undergoes striking alterations due to proliferation or cell death. Factors involved in the induction of apoptosis of pituitary cells are not well established. We demonstrated earlier that oestrogens prevent lactotroph cells of female rats to die by apoptosis induced by bromocryptine treatment, a fact that can be reversed in ovariectomised rats. In this study, we developed experimental models for in vivo and in vitro studies to gain further insight on the survival effect of oestrogens on lactotrophs. In rats pretreated with oestrogens, tamoxifen generates a massive cell death by apoptosis as validated by the TUNEL technique and DNA electrophoresis of pituitary gland. On electron microscope observations, numerous lactotrophs exhibited progressive morphological changes in the nuclei compatible with the apoptotic process. The cells remaining intact also exhibit signs of inhibition due to a significant transformation of regular lactotrophs in atypical subtypes. In pituitary cell cultures exposed to tamoxifen and oestrogen simultaneously, most of the lactotrophs displayed features of apoptosis in the nucleus. The present reports gathered new evidences on the apoptogenic potential of tamoxifen on lactotroph cells, and corroborates the contribution of oestrogens to sustain both a balanced population of lactotrophs and a competent secretory activity. The concept that opposed activities, such as inhibition and stimulation, can activate apoptosis is also strengthen by these observations.

Regulated expression and ultrastructural localization of galectin-1, a proapoptotic beta-galactoside-binding lectin, during spermatogenesis in rat testis.

Galectin-1, a highly conserved beta-galactoside-binding protein, induces apoptosis of activated T cells and suppresses the development of autoimmunity and chronic inflammation. To gain insight regarding the potential role of galectin-1 as a novel mechanism of immune privilege, we investigated expression and ultrastructural localization of galectin-1 in rat testis. Galectin-1 expression was assessed by Western blot analysis and immunocytochemical localization in testes obtained from rats aged from 9 to 60 days. Expression of this carbohydrate-binding protein was developmentally regulated, and its immunolabeling exhibited a stage-specific pattern throughout the spermatogenic process. Immunogold staining using the anti-galectin-1 antibody revealed the typical Sertoli cell profile in the seminiferous epithelium, mainly at stages X-II. During spermiation (stages VI-VIII), a strong labeling was observed at the luminal pole of seminiferous epithelium, localized on apical stalks of Sertoli cells, on heads of mature spermatids, and on bodies of residual cytoplasm. Moreover, spermatozoa released into the lumen showed a strong immunostaining. Following spermiation (stage VIII), galectin-1 expression was restored at the basal portion of Sertoli cells and progressively spread out through the whole cells as differentiation of germinal cells proceeded. Immunoelectron microscopy confirmed distribution of galectin-1 in nuclei and cytoplasmic projections of Sertoli cells and on heads and tails of late spermatids and residual bodies. Surface localization of galectin-1 was evidenced in spermatozoa from caput epididymis. Thus, the regulated expression of galectin-1 during the spermatogenic cycle suggests a novel role for this immunosuppressive lectin in reproductive biology.

Prolactin-releasing activity of GHRP-5 (Momany peptide) on lactotrophs in vivo and in vitro.

In the present study the in vivo and in vitro effects of GHRP-5 on the PRL-releasing activity in correlation with the morphological changes of lactotroph cells and their transcriptional activity were evaluated. The in vivo treatment (12 micrograms/100 g BW/day for 3 days) of male rats with GHRP-5 does not induce any significant changes in serum PRL levels. In contrast, the addition of GHRP-5 to pituitary cell cultures increased significantly the release of PRL. This effect is enhanced in cell cultures of enriched lactotrophs, increasing significantly the secretion of PRL, the concentrations of which were 50% higher than that of untreated control cells. The administration of GHRP-5 provokes several changes in the fine structure of lactotrophs, compatible with an increased secretory activity. After the GHRP-5 treatment the different lactotroph subtypes persist but the subtype I displaying secretory granules of larger size (500-900 nm) and a significant development of the Golgi apparatus and RER were more frequently observed. These results can be correlated with a significant augmentation in PRL mRNA after the GHRP-5 treatment. In spite of that no variations in serum PRL levels were observed in vivo, following GHRP-5 treatment, the lactotroph population experienced evident fine structure modifications, concordant with an upsurge of PRL synthesis. These observations confirmed a direct action of GHRP-5 on receptors expressed by lactotrophs. The differential actions of GHRP-5 on in vivo and in vitro designs confirm a different effectiveness of this secretagogue to induce PRL secretion.

Prolactin-releasing activity of GHRP-5 (Momany peptide) on lactotrophs in vivo and in vitro

In the present study the in vivo and in vitro effects of GHRP-5 on the PRL-releasing activity in correlation with the morphological changes of lactotroph cells and their transcriptional activity were evaluated. The in vivo treatment (12 micrograms/100 g BW/day for 3 days) of male rats with GHRP-5 does not induce any significant changes in serum PRL levels. In contrast, the addition of GHRP-5 to pituitary cell cultures increased significantly the release of PRL. This effect is enhanced in cell cultures of enriched lactotrophs, increasing significantly the secretion of PRL, the concentrations of which were 50 higher than that of untreated control cells. The administration of GHRP-5 provokes several changes in the fine structure of lactotrophs, compatible with an increased secretory activity. After the GHRP-5 treatment the different lactotroph subtypes persist but the subtype I displaying secretory granules of larger size (500-900 nm) and a significant development of the Golgi apparatus and RER were more frequently observed. These results can be correlated with a significant augmentation in PRL mRNA after the GHRP-5 treatment. In spite of that no variations in serum PRL levels were observed in vivo, following GHRP-5 treatment, the lactotroph population experienced evident fine structure modifications, concordant with an upsurge of PRL synthesis. These observations confirmed a direct action of GHRP-5 on receptors expressed by lactotrophs. The differential actions of GHRP-5 on in vivo and in vitro designs confirm a different effectiveness of this secretagogue to induce PRL secretion.

Prolactin-releasing activity of GHRP-5 (Momany peptide) on lactotrophs in vivo and in vitro

In the present study the in vivo and in vitro effects of GHRP-5 on the PRL-releasing activity in correlation with the morphological changes of lactotroph cells and their transcriptional activity were evaluated. The in vivo treatment (12 micrograms/100 g BW/day for 3 days) of male rats with GHRP-5 does not induce any significant changes in serum PRL levels. In contrast, the addition of GHRP-5 to pituitary cell cultures increased significantly the release of PRL. This effect is enhanced in cell cultures of enriched lactotrophs, increasing significantly the secretion of PRL, the concentrations of which were 50 higher than that of untreated control cells. The administration of GHRP-5 provokes several changes in the fine structure of lactotrophs, compatible with an increased secretory activity. After the GHRP-5 treatment the different lactotroph subtypes persist but the subtype I displaying secretory granules of larger size (500-900 nm) and a significant development of the Golgi apparatus and RER were more frequently observed. These results can be correlated with a significant augmentation in PRL mRNA after the GHRP-5 treatment. In spite of that no variations in serum PRL levels were observed in vivo, following GHRP-5 treatment, the lactotroph population experienced evident fine structure modifications, concordant with an upsurge of PRL synthesis. These observations confirmed a direct action of GHRP-5 on receptors expressed by lactotrophs. The differential actions of GHRP-5 on in vivo and in vitro designs confirm a different effectiveness of this secretagogue to induce PRL secretion.(AU)

Prolactin-releasing activity of GHRP-5 (Momany peptide) on lactotrophs in vivo and in vitro.

In the present study the in vivo and in vitro effects of GHRP-5 on the PRL-releasing activity in correlation with the morphological changes of lactotroph cells and their transcriptional activity were evaluated. The in vivo treatment (12 micrograms/100 g BW/day for 3 days) of male rats with GHRP-5 does not induce any significant changes in serum PRL levels. In contrast, the addition of GHRP-5 to pituitary cell cultures increased significantly the release of PRL. This effect is enhanced in cell cultures of enriched lactotrophs, increasing significantly the secretion of PRL, the concentrations of which were 50

higher than that of untreated control cells. The administration of GHRP-5 provokes several changes in the fine structure of lactotrophs, compatible with an increased secretory activity. After the GHRP-5 treatment the different lactotroph subtypes persist but the subtype I displaying secretory granules of larger size (500-900 nm) and a significant development of the Golgi apparatus and RER were more frequently observed. These results can be correlated with a significant augmentation in PRL mRNA after the GHRP-5 treatment. In spite of that no variations in serum PRL levels were observed in vivo, following GHRP-5 treatment, the lactotroph population experienced evident fine structure modifications, concordant with an upsurge of PRL synthesis. These observations confirmed a direct action of GHRP-5 on receptors expressed by lactotrophs. The differential actions of GHRP-5 on in vivo and in vitro designs confirm a different effectiveness of this secretagogue to induce PRL secretion.

Prototipo de fuente lumínica pra terapia fotodinámica disenado en el Centro de Microscopía Electrónica / Prototype of light source for photodynmic therapy in Centre of Electron Mycroscopy

La Terapia Fotodinámica (PDT) es un tratamiento del cáncer basado en la acumulación específica de una droga fotosensible en el tejido maligno. Su posterior radiación con una longitud de onda apropiada, induce la producción de singuletes de oxígeno responsable de la peroxidación de las organelas y la muerte de las células neoplásicas. En el Centro de Microscopía Electrónica de la Universidad Nacional de Córdoba se diseñó y construyó un prototipo de fuente de irradiación no coherente de 630nm el que posibilitó la aplicación de PDT por primera vez en nuestro país. Este prototipo ha sido aplicado satisfactoriamente en el tratamiento de la queratosis actínica. Fueron tratadas 100 lesiones en 27 pacientes utilizando como fotosensibilizador al ácido d amino levulínico (ALA) al 20 por ciento La activación lumínica duró de 5 a 20 minutos dependiendo de la extensión y profundidad de la lesión. Los resultados obtenidos fueron los siguientes: Remisión Completa de las lesiones (RC) 84por ciento, Remisión parcial (RP) 10 por ciento, Sin respuesta (SR) 0 por ciento y Sin datos (SD) 6 por ciento. En el último grupo están incluidos aquellos pacientes que no retornaron para su evaluación. La alta efectividad, sumada a la inmejorable respuesta cosmética y la reducida agresividad, hacen de PDT el método de elección en el tratamiento de esta patología. El prototipo utilizado en este estudio demostró ser además de no invasivo y bien tolerado, altamente efectivo.