European Journal of Histochemistry 2010; volume 54:e1 Folliculostellate cells in roectodermic origin of these cells; 3) FSC spa- tial distribution, association with endocrine Correspondence: Fabian Mohamed, Cátedra de pituitary pars distalis of male cells, and the generation of an intercellular Histología y Embriología, Facultad de Química, viscacha: immunohistochemical, communication network suggest that FSC are Bioquímica y Farmacia, Universidad Nacional de morphometric and involved in the pituitary pars distalis San Luis, Avenida Ejército de los Andes 950 - 2º paracrine regulation of the viscacha. piso. CP 5700, San Luis, Argentina. ultrastructural study E-mail: [email protected] M. Acosta, V. Filippa, Key words: Lagostomus, pituitary, folliculostellate F. Mohamed cells, immunohistochemistry, distribution, ultra- Introduction structure. Cátedra de Histología y Embriología, Facultad de Química, Bioquímica y Pituitary folliculostellate cells (FSC) have Acknowledgments: we wish to thanks Dr. R. Farmacia, Universidad Nacional de San been reported in several mammalian species.1 Piezzi, Mrs. T. Fogal, Mrs. A. Bernardi and Mr. J. Luis, Consejo Nacional de Investigaciones At present, however, there is no agreement in Arroyuelo for their technical collaboration. The Científicas y Técnicas (CONICET), the literature in relation with the embryologi- financial support for this study was received from cal origin and exact function of these cells. In CONICET and Universidad Nacional de San Luis San Luis, Argentina (grant number 22/Q603). fact, FSC have been reported to play a support- ing role, to have phagocytotic activity, and to Received for publication: 28 July 2009. act as regulators of hormone secretion and as Accepted for publication: 24 November 2009. Abstract stem cells.2-5 FSC have been found lining the lumen of follicles scattered throughout the This work is licensed under a Creative Commons Attribution 3.0 License (by-nc 3.0). Folliculostellate cells (FSC) have been pituitary pars distalis.5-9 The FSC have been studied using the antibody against S-100 pro- reported in pituitary of several mammalian ©Copyright M. Acosta et al., 2010 6-12 species. FSC morphology and secreted sub- tein, in both mammalian and non-mam- Licensee PAGEPress, Italy stances have been instrumental to the under- malian species.3,4,13 In addition, some authors European Journal of Histochemistry 2010; 54:e1 standing of their function. The purpose of this have reported in human,11 cat, rat and rabbit14 doi:10.4081/ejh.2010.e1 work was to perform an immunohistochemi- and mink7 that FSC were immunostained with cal, morphometric and ultrastructural study of anti-glial fibrillary acidic protein (GFAP) and the pituitary pars distalis FSC in adult male anti-vimentin. However, in pituitary of dog member of the Chinchillidae family. This rodent viscacha and to analyze their relation with and horse these cells did not express GFAP inhabits the southern hemisphere from hormone secreting cells. Immunohistoche- and vimentin, but they expressed the S-100 Paraguay through central Argentina.21,22 In its mistry and image analysis were carried out in protein.15 Soji et al.6 have shown an increase natural habitat, the viscacha is a seasonal different sectors of the gland, from the middle in the number of these cells in relation with breeder.23-27 The pituitary pars intermedia (PI), (sector 1) to the glandular periphery (sector age, suggesting that the expression of the S- pineal gland, and blood-testis barrier of this 5). Transmission electron microscopy with 100 protein indicates the cell morphologic rodent have been studied using lanthanum.28-30 lanthanum as electrodense tracer was used. maturation stage in rats. Moreover, several Piezzi et al.28 have demonstrated the permeabil- FSC formed follicles with PAS-positive colloid authors have demonstrated the existence of ity to lanthanum and the communication of the inside. They expressed S-100 protein mainly different physiological stages of FSC based on PI parenchyma with Rathke’s pouch. In this in both nucleus and cytoplasm. FSC were stel- their morphological characteristics and pituitary zone, the FSC originated follicles with late-like in shape and exhibited short cyto- immunophenotype.7,11,16 PAS-positive colloid inside, expressed S-100, plasmic processes that contacted with blood On the other hand, there are few reports in GFAP and vimentin, and were closely associated vessels and endocrine cells. In addition, some the literature on the distribution of FSC in the with melanotrophs.31 Moreover, an ultrastruc- follicular colloids were immunostained with pituitary parenchyma. Sato et al.12 have report- tural study of the pituitary PD has demonstrat- anti-S-100 protein. A few FSC were immunos- ed that FSC are mainly localized in the basal ed the existence of two types of colloid-contain- tained with anti-glial fibrillary acidic protein region and transition zone of rat pituitary and ing structures: 1) follicles, originated by the (GFAP) and anti-vimentin. The morphometric also showed an association between FSC dis- FSC, and 2) colloid extracellular accumulations, parameters analyzed (percentages of S-100- tribution and vascularization. limited by different types of granulated cells.32 positive total, cellular and colloidal areas) Furthermore, FSC have been reported to The follicular lumen was filled with abundant increased from sector 1 to sector 3 and then synthesize part of the follicular colloid and a electrodense colloid and several cellular debris decreased to sector 5. Hormone secreting close relationship has been described between comprising vacuoles, altered membranes, mito- cells, mainly lactotrophs, gonadotrophs and these cells, the follicular colloid and hormone chondria and secretory granules, which are corticotrophs were associated with FSC and secreting cells.3,4,16-18 probably a product of FSC phagocytotic activity.32 follicles. The ultrastructural study demonstrat- Ultrastructurally, FSC share distinct charac- In addition, the hormone secreting cells have ed that FSC developed junctional complexes teristics in all studied species. They exhibit exhibited regionalization within the PD and desmosomes between their lateral mem- scanty cytoplasm with poorly developed endo- parenchyma.27,33-36 branes. Lanthanum freely penetrated the plasmic reticulum, inconspicuous Golgi com- Based on the above data and our previous spaces between granulated cells and FSC, but plex, moderate amount of mitochondria and knowledge about the viscacha pituitary, the did not penetrate into the follicular lumen. In absence of the secretory granules.1 Some stud- aim of this work was to perform an immuno- conclusion: 1) the differential expression of S- ies in pituitary have also been performed with histochemical, morphometric and ultrastruc- 100 protein, GFAP and vimentin may indicate lanthanum as electrodense tracer.19,20 tural study of the pituitary pars distalis FSC in different physiological stages of FSC; 2) the Our experimental model is the viscacha adult male viscacha and to analyze their rela- expression of these proteins suggests a neu- (Lagostomus maximus maximus), the largest tion with hormone secreting cells. [European Journal of Histochemistry 2010; 54:e1] [page 1] Original Paper Table 1. Features of the antibodies used for immunohistochemical study. Materials and Methods Antibody Immunostained Clone, catalog Time, Temperature cellular type and source Incubation Fourteen adult male viscachas weighing 6.0- 6.5 kg were captured in their habitat near San Anti-S-100 protein Folliculostellate cell (FSC) Polyclonal. AR058-5R. 12 h, 4ºC in a moist chamber Luis, Argentina (33° 20’ south latitude, 760 m Ready-to-Use. BioGenex altitude) during February-March 2008 (sum- Anti-GFAP FSC Polyclonal. AR020-5R. 12 h, 4ºC in a moist chamber mer), using traps placed in their burrows. (glial fibrillary Ready-to-Use. BioGenex According to the results published by Llanos acidic protein) and Crespo37 and Branch et al.38 the body Anti-Vimentin FSC V9. AM074-5M. 12 h, 4ºC in a moist chamber weight of this rodent is in relation with its age. Ready-to-Use. BioGenex Then, the viscachas used in this study were in Anti-Prl (prolactin) Lactotroph Polyclonal. N1549. 60 min, 4ºC in a moist chamber the same growth stage. After being captured, Ready-to-Use. animals were immediately taken to the labora- DakoCytomation tory, anaesthetized with Nembutal (pentobar- Anti-FSHβ FSH-gonadotroph 83/12/ 2A8 2C7. 12 h, 4ºC in a moist chamber bital; 25 mg/Kg ip.) and killed by decapitation. (follicle stimulating MU026-UC. The brains were rapidly exposed and the pitu- hormone) 1:100. BioGenex itaries (weighing 28-30 mg) were excised. The Anti-LHβ LH-gonadotroph 3LH 5B6 YH4. AM030-5M. 12 h, 4ºC in a moist chamber experimental design was approved by the Local (luteinizing 1:200. BioGenex Ethics Committee and was in agreement with hormone) the Guidelines of the National Institutes of Anti-ACTH (1-24) Corticotroph Polyclonal. AR035-5R. 60 min, 4ºC in a moist chamber Health (Bethesda, Md., USA) for the use of (adrenocorticotropin Ready-to-Use. BioGenex experimental animals. hormone) For light microscopy, the pituitaries (n=6) Anti-GH Somatotroph Polyclonal. N1561. 12 h, 4ºC in a moist chamber were sectioned in the middle plane (Figure 1, (growth hormone) Ready-to-Use. line M). Both halves of each pituitary were DakoCytomation fixed in Bouin’s fluid, dehydrated in increasing Anti-TSH Thyrotroph 5404. AM033-5M. 12 h, 4ºC in a moist chamber graded ethanol, cleared in xylene,