Identification and characterization of telocytes in rat testis

Abstract

In this study, we investigated the localization, morphological features and cellular interactions of telocytes in the rat testicular interstitium. Transmission electron microscopy (TEM) and immunohistochemical and immunofluorescence analyses of the rat testicular interstitium showed a distinct layer of telocytes surround the seminiferous tubules along with inner layer of peritubular myoid cells. The majority of the telocytes were made up of a small cell body and moniliform prolongations that contained mitochondria and secretory vesicles. Some other telocytes were observed possessing large cell bodies. Within the testicular interstitium, the telocytes formed a network connecting peritubular myoid cells, Leydig cells as well as blood vessels. Immunohistochemical and double immunofluorescence analyses showed that rat testicular telocytes express CD34 and PDGFRα, but are negative for vimentin and α-SMA. Our findings demonstrate the presence of telocytes in the rat testicular interstitium. These cells interact with peritubular myoid cells, seminiferous tubules, Leydig cells and blood vessels via long telopode extensions, which suggests their vital role in the intercellular communication between different cell types within the rat testis.

Keywords: CD34; PDGFRα; telocytes; telopodes; ultrastructure.

Figure 1

TEM photograph of a peritubular myoid cell in the rat testis. (A) Peritubular myoid cells are seen in the lamina propria of the seminiferous tubule. (B–C) A single peritubular myoid cell (PMC) is shown with actin filaments (indicated by the arrow), mitochondria, Golgi apparatus and secretory vesicles. Note: PMC: peritubular myoid cell; ST: seminiferous tubule; TC: telocyte; BM: basement membrane; Nu: nucleus; M: Mitochondria; G: Golgi apparatus;Vc: Vesicles. Scale Bar = A: 2μm; B: 1μm and C: 500nm.

Figure 2

TEM photograph of a telocyte in rat testis. (A) The telocyte is located surround the seminiferous tubule (indicated by the arrow) and shows a distinct cytoplasmic process with a (C) podom containing mitochondria and the vesicles and (D) podomer.(B) The cell body contains small amount of cytoplasm with mitochondria around the nucleus. Higher magnification illustrates the rectangular area. Nu: nucleus; M: Mitochondria;Vc: vesicles; Pd: Podom; P: podomer. Scale Bar = A: 10μm; B: 2μm; C: 1μm; D: 2μm.

Figure 3

TEM photograph of a telocyte in rat testis. (A) Telocyte overlays the peritubular myoid cell in the lamina propria of seminiferous tubule. (B, D) Cytoplasmic process shows long extensions with (B) mitochondria and (D) vesicles. (C) The telopode seems more developed, with winding, thickened and dichotomous feature. TC: Telocyte; PMC: Peritubular myoid cell; Bm: Basement membrane; Nu: Nucleus; M: Mitochondria; Sv: Secretory vesicle; Vc: Vesicles. Scale Bar = A: 5μm, B–D:1μm.

Figure 4

TEM photograph of another telocyte in the rat testis. (A) The telocyte surrounds the seminiferous tubule and the peritubular myoid cell.(B) The telocyte shows distinctly elongated nucleus (indicated by the arrow) with a small amount cytoplasm. (C) The cytoplasmic process shows few attachment plaques (indicated by the arrows). (D) The cytoplasmic prolongation shows many vesicles. TC: Telocyte; PMC: Peritubular myoid cell; Nu: Nucleus; Bm: Basement membrane; Vc: Vesicles. Scale Bar = A: 4μm; B:2μm; C: 1μm; D: 2μm.

Figure 5

TEM photograph showing two neighboring telocytes in the rat testes. (A) Telopodes (Tps; indicated by arrow) and (B) small secretory vesicles (indicated by the arrows) of the two telocytes are shown.(C) Higher magnified image shows the area between the two telocytes. Note: TC: Telocyte; PMC: Peritubular myoid cells; Nu: Nucleus; Tps: Telopode; Sv: Secretory vesicle. Scale Bar = A–B: 2μm.

Figure 6

TEM photograph shows different types of junctions formed by Tps with other cells in the rat testis. Physical contacts made by Tps with adjacent cells through (A) Mortise and tenon joint, (B) long overlapped structure, (C) closeness (indicated by the arrow), (D, E) short overlapped structure and punctate contact (indicated by the circles). Scale Bar = A–D: 1μm; E: 10μm.

Figure 7

Immunohistochemical analyses of peritubular tissue in the rat testis with anti-αSMA and anti-CD34 antibodies. (A–C) CD34 positive staining (brown) surrounds the seminiferous tubules. Higher magnification (C) shows segmental pattern of CD34 staining. (E–G) αSMA positive staining (brown) also surrounds the seminiferous tubules. Higher magnification (G) shows smooth uniform staining pattern. Higher magnification illustrates the rectangular area. (D, H) shows negative contral group of rat testis. Scale Bar = A, E: 100μm; B, F: 20μm; C, G: 5μm; D: 50μm; H: 20μm.

Figure 8

Double immunofluorescence of telocytes in the rat testis with anti-αSMA and anti-CD34 antibodies. (A–F) Immunofluorescence staining shows positive αSMA (A; green) and CD34 (B; red) staining of the inner and the outer layers surrounding the seminiferous tubules. (G–I) Immunofluorescence staining shows positive CD34 staining (red) around the blood vessel and in endothelial cells, and positive αSMA staining (green) on the vessel wall. Scale Bar = A–C: 20μm; D–I: 10μm.

Figure 9

Double immunofluorescence staining of telocytes in the rat testis with anti-αSMA and anti-PDGFRα antibodies. (A–C) Immunofluorescence images show positive αSMA (green) and PDGFRα (red) staining inside/outside as continuous layers around the seminiferous tubule. (D–F) Immunofluorescence images show higher magnification illustrates the rectangular area in (C). Scale Bar = A–C: 20μm; D–F:10μm.

Figure 10

Double immunofluorescence staining of telocytes in rat testis with anti-CD34 and anti-vimentin antibodies. (A–C) Immunofluorscence staining shows that CD34 (green) and Vimentin (red) do not co-localize around the seminiferous tubule. Vimentin is expressed on Sertoli cells (pointed by arrows) and Leydig cells (pointed by triangles). Scale Bar = A–C: 20μm.

Figure 11

Interactive network of cells including telocytes in the inter-tubular space of rat testes. (A–D) Transmission electron micrographs show that telocytes form interactive networks by extending their telopodes to connect with Leydig cells and surrounding blood vessels (A, C) through various secretory vesicles (D, indicated by the arrows). The circles highlight the direct contact between telopodes and Leydig cells (A–B). (E) Immunofluorescence staining of CD34 (green) and αSMA (red) shows telocytes surrounding the PMC and the blood vessel. Arrows show the telocytes. Higher magnification illustrates the rectangular area. Tp: telopode; Bv: Blood vessel; LC: Leydig cells; PMC: peritubular myoid cells; TC: telocyte. Scale Bar = A: 2μm; B: 1μm; C: 5μm;D: 1μm; E: 10μm.