The Morphology And Relative Studies Of Telocytes In Human Esophagus

As a novel type of interstitial cells, telocytes were initially considered as interstitial Cajal-like Cells (ICLC). With the increasing histological studies, telocytes were identified different from ICLC and other interstitial cell, such as fibroblast, fibrocyte, fibroblast-like cells and mesenchymal cells, so they were formally renamed as telcoytes by L.M.Popsecu in2010. Telocytes consist of a small cell body and extremely long processes called telopodes. It has been suggested that telocytes were in closed relationship with immunocytes, nerve endings, capillaries, smooth muscle cell fibers and a variety of other mesenchymal cells. Possibly, telocytes play a role in upper signal integration, transferring, regulation of the blood supply, tissue repairing and regeneration. Up to now, telocytes have been identified in many organs and tissues in mammal. However, there is still not enough data about the distribution and functions of telocytes in human body. The aim of the present study is to identify the existence of telocytes in human esophagus and provide evidence for the widespread of this new type of cell in human body, and to demonstrate its possible biological functions in human esophageal tissue.The study was approved by the Ethic Committee of Fudan University, Zhongshan Hospital. All collections of human tissues had the informed consent from the patients.Normal human esophagus samples were collected from the corresponding normal tissues adjacent to resection margins from patients who had no anticancer treatment before tumor resection. All samples were examined and confirmed as the normal by the veteran pathologist. A total of10primary esophagus tissue samples for study were obtained. Human adjacent normal esophagus tissue samples were obtained and divided into:1) formalin fixing for histology and immunochemical staining;2) fresh tissues in PBS at pH7.4for electronic microscopy and histology;3) fresh tissues for isolation and primary cell culture of telocytes. Transmission electron microscopy (TEM). For ultrastructural analysis, tissue samples from human esophagus were cut into small pieces about1mm3within1minute after being excised from body and immediately immersed and fixed in a solution of4%glutaraldehyde (pH7.3,4℃). After several steps of pretreatments, the samples were cut into sections of50nm for TEM. Digital pictures were obtained using a high resolution digital camera connected to the TEM. Telocytes existed both in mucous membrane and muscle layer in human esophagus, which possessed typical morphology of small cellular body with2-3telopodes-extremely long with bifurcate cellular prolongation. Telopodes consist of an alternation of thin segments (podomers) and thick segments (podoms), and appeared like labyrinthine and network structure.Immunohistochemistry (IHC). Specimens of esophagus were isolated and fixed with10%neutral formalin and embedded in paraffin. HE sections were made to confirm samples normal esophageal tissue. IHC with c-kit demonstrated the presence of typical telocyte morphology:deeply stained cell body with thin and long telopodes, lying among interstitial cells. IHC with vimentin showed formation of telocytes, with plasma stained clearly. IHC with CD34showed CD34was expressed on telocytes body and their prolongations.Primary cell culture and vital stainings. The tissues of human esophagus were cut and harvested under sterile conditions, after several steps of digesting, centrifuging and resuspended prossessing, telocytes were successfully maintained in primary culture and could easily be identified before reaching confluence. The cells were examined under an inverted Olympus microscope equipped with a digital camera, Olympus20×objective. Starting with the3rd day in culture, teloctyes appeared with characteristic long, moniliform processes, telopode. We used several vital stainings to display the morphology of telocytes. Both methylene blue and Giemsa staining reveals that in living cells, telocytes were very long (several tens of μm), and had an uneven caliber, with dilated portions, resembling "beads on a string", and they also had thick emergence from the cell body, followed by gradual thinning. This characteristic feature made telopode clearly different from dendritic cells, fibroblasts and myofibroblasts et al. Janus green B is a well-known vital staining with high affinity for mitochondria and it is used to assess viability and localize mitochondria. The initial dark green-blue color, due to mitochondria stained with Janus green B, became a brownish gray one, and finally decolorized illustrates the staining of telocytes body and dilations of the processesTelocytes are interstitial cells located in connective tissue, among other specific cellular types like epithelial, muscular, nervous cells, or immunocytes. In our study, telocytes were found both in mucus and muscular wall of esophagus. Typical telocytes with long telopodes were observed among capillary and smooth muscle cells, and there were network structures or labyrinthine structure between telopodes. We also observed telopodes were adjacent to macrophage. Our data do not provide direct information about the presumptive function of telocytes. However, besides the conventional role of mechanical support for the tissues, we think that telocytes play a role in the immune system and in the intercellular communication and regulation, since their connection with macrophage and the existence of3D network that is consisted of telopodes. It is accepted that telocytes might also play a role in the repair and regeneration of the tissues, since they are in close vicinity of blood capillaries and muscle cells. It is reasonable to assume that some esophageal diseases, such as achalasia and reflux esophagitis, esophagus cancer, are associated with dysfunction of telocytes. Further researches are urgently required to identify the biological function and the mechanism of telocytes in the repair and regeneration of injured tissues.