Introduction
Secretion is a process by which cells selectively externalize compounds as a part of numerous metabolic exchanges, and is a basic feature of every eukaryotic cell. Besides exocytosis which involves the fusion of secreted vesicles to the plasma membrane and whose mechanism is well characterized (Deak et al., 2008; Südhof and Rothman, 2009; Anantharam et al., 2010), two additional forms of "unconventional" secretion (Fig. 1), apocrine and holocrine, are known. These two forms of secretion are essentially described in morphological terms and involve the release of cellular components or cell portions without homotypic membrane fusion. During apocrine secretion the cell discharges a portion of its cytoplasm followed by a partial or complete recovery and reconstitution of the cell structure and shape. In the case of holocrine secretion the whole content of the cell is fragmented and released into the luminal space of the gland.
A B C
Figure 1. Three forms of secretion. A = merocrine (or exocytosis). The secreted products are transferred into membranes vesicles that are fused to the plasma membrane and released into the extracellular space. B = apocrine. Intra-cellular components are freed in the lumen of the gland by a yet unknown mechanism, presumably by shedding whole portions of the cytoplasm. After release the cytoplasm is partially or totally reconstituted and a new cycle of secretion may take place. C = holocrine. The entire cell is fragmented and “dissolved” into the lumen of the gland.
The apocrine secretion was first described in 1833 by Purkinje and independently in 1839 by Velpeau and later in 1854 by Verneuil as a chronic acneiform infection of the cutaneous apocrine glands, designated as hidradenitis suppurativa (for review see Lasko et al., 2008). Current studies in apocrine secretion deal essentially with lactation of mammary glands, discharge in other exocrine glands, particularly those present in the skin of mammals or humans, and concern also numerous human disorders including breast cancers and psoriasis (Satoh et al., 1996; Gesase and Satoh, 2003, Khalbuss, 2005; Crowson et al., 2006; O'Malley and Bane, 2008; Elayat et al., 2010).
Although the scientific and medical literature in Medline PubMed and Web of Science databases contains more than 90 000 original references related to apocrine and holocrine secretion, most of the works refer to morphology and descriptive pathology, yet provide no basic explanation to the process. In addition the morphological description of apocrine secretion is commonly transferred from one textbook to another without referring to any original article.
In contrast to exocytosis, whose numerous studies in yeast to human have revealed a large number of controlling factors and genes, no factor or gene regulating the apocrine (and/or holocrine) process has been so far characterized. In insects, morphological studies have provided indications of apocrine secretion in the gut of mosquitoes, the salivary glands of the bees, and the male accessory glands of hemipterans (Oliveira et al., 2009; Silva-Zacarin et al., 2007; Freitas et al., 2007, 2010). However, despite all the morphological and developmental studies performed in Drosophila melanogaster, no report deals with the occurrence of apocrine secretion.
In the course of our research on programmed cell death we have unexpectedly discovered that several years ago for the first time described extrusion of cellular components from doomed Drosophila salivary glands is a process of apocrine secretion when whole pieces of cellular contents are released into extracellular space. The apocrine secretion as a type of cell contents release has been described morphologically and defined as such some 178 years ago, however, until today no protein components, factors or genes affecting apocrine secretion were identified. In contrast, widespread and well known exocytosis is an example of very intensely studied secretory mechanism with many dozens of identified factors to be involved (Jahn, 2004; Rutter and Tsuboi, 2004; Sudhof, 2004; Chieregatti and Meldolesi, 2005; Barclay et al., 2005; Snyder and Kelly, 2006; Westerink, 2006). In textbooks, the apocrine secretion is frequently described in association with lactation activity of mammary glands, Harderian gland, and of some exocrine glands (Satoh et al., 1996; Gesase a Satoh, 2003) or notably as a differential diagnostic marker for some benign metaplasias and in many dermatogenic cancers (Griffith, 2005; Khalbuss, 2005; Shah, 2005; Crowson et al., 2006). It should be noted that, for example, epithelial tissue of the mammary gland in humans where milk is released by apocrine mechanism is known to undergo widespread involution after lactation is completed, and this resembles strongly many features of apoptosis. However, to use apocrine/holocrine activity as potentially main diagnostic tool or even to find the way how to treat its unwanted and adverse effects, it is necessary to characterize molecular and genetic clues of this process.
Proposed project is based on rich experience of the research team in this field including methodology that have been build during basic research activity. The main goal of the project is to significanlty contribute to selfsustaining development and wide-spread application of new knowledge and novel methods in biomedical research and practical medicine itself. The proposed research is expected to bring essential new information on apocrine and holocrine secretion in health and disease. Project is based on broad application of proteomic and subsequently genomic methods to identify main and crucial components of apocrine and holocrine secretion that as we expect will share common determinants between different cell types and even evolutionary different species.
Similarly as incidence of skin diseases, notably skin tumours, also incidence of breast cancer is steadily increasing not only in Slovakia but world-wide. Several skin diseases are listed among those which are cited to be apocrine-coupled: apocrine hydrocystoma, cylindromatosis, various apocrine gland cysts with hemosiderotic dermatofibromas, lipomatous mixed tumor of the skin, bilateral axillary metachronous apocrine carcinomas, poromatosis, focal axillary hyperhidrosis, hypohidrotic ectodermal dysplasia, and various forms of almost two centuries known hidradenitis suppurativa. There are also several apocrine-associated types of breast cancer: deciduoid-like myofibroblastoma, fibroadenomatous cancer, and mammary-type fibroepithelial neoplasms. Some salivary gland tumours like intraductal carcinoma, micropapillary mucinous carcinoma, pleomorphic adenoma or sclerosing polycystic adenosis are also known to show positive apocrine histology. This short list provide strong evidence how important basic as well as applied research to this field is to facilitate understanding of apocrine and/or holocrine diseases. The role of research, preventive and clinical medicine from this point of view is to gain insights into molecular mechanism of the apocrine and holocrine processes which finally can contribute to highly individualized and specific treatment of every diagnosed patient that is in good accordance with world-leading trends in this field.
One of the main overal goals is to identify molecular determinant (markers) of listed diseases with special emphasis on factors common to apocrine and holocrine secretion. There is no doubt that in spite of different etiology and histogenetic origin of mentioned diagnoses, the unifying or common feature is apo/holocrine secretory activity, and it is assumed that this mechanism of cellular contents release is controlled by identical or highly related components of the proteome. This assumption is based on our findings from initial phases of the study we have been undertaking several months ago in model organisms as a part of basic research. For the proposed project, in collaboration with clinicians, we collect material (exogenous secretions, mucus, sweat, cerumens, milk etc.) from healthy volunteers and from patients diagnosed for any of above mentioned diseases. After collecting and storage of the material (for the shortest necessary time at -80°C), samples are being extracted for their protein and lipid components, followed by separation of the fractions (using appropriate analytical methods including SDS-PAGE, 2D PAGE and nano-HPLC, GC), and subsequently identified, where especially in the case of proteins the mass spectrometry (MALDI-TOF/TOF) is employed.
Among interesting results already obtained under this study is identification of IgGs, interleukins, cytoskeletal proteins including actin, actinin, spectrins etc. in the apocrine secretion fluid from humans. Some of these proteins are either not present in healthy specimens, or they become significantly upregulated e.g. under hidradenitis suppurativa condition.
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