Reaching out: junctions between cardiac telocytes and cardiac stem cells in culture

Abstract

Telocytes (TCs) were previously shown by our group to form a tandem with stem/progenitor cells in cardiac stem cell (CSC) niches, fulfilling various roles in cardiac renewal. Among these, the ability to 'nurse' CSCs in situ, both through direct physical contact (junctions) as well as at a distance, by paracrine signalling or through extracellular vesicles containing mRNA. We employed electron microscopy to identify junctions (such as gap or adherens junctions) in a co-culture of cardiac TCs and CSCs. Gap junctions were observed between TCs, which formed networks, however, not between TCs and CSCs. Instead, we show that TCs and CSCs interact in culture forming heterocellular adherens junctions, as well as non-classical junctions such as puncta adherentia and stromal synapses. The stromal synapse formed between TCs and CSCs (both stromal cells) was frequently associated with the presence of electron-dense nanostructures (on average about 15 nm in length) connecting the two opposing membranes. The average width of the synaptic cleft was 30 nm, whereas the average length of the intercellular contact was 5 μm. Recent studies have shown that stem cells fail to adequately engraft and survive in the hostile environment of the injured myocardium, possibly as a result of the absence of the pro-regenerative components of the secretome (paracrine factors) and/or of neighbouring support cells. Herein, we emphasize the similarities between the junctions described in co-culture and the junctions identified between TCs and CSCs in situ. Reproducing a CSC niche in culture may represent a viable alternative to mono-cellular therapies.

Keywords: adherens junctions; cardiac stem cells; cardiac telocytes; gap junctions; stromal synapse; telopodes.

Figure 1

Light microscopy (1 μm semithin sections of epoxy resin embedded cells, stained with toluidine blue), reveals close contacts between cardiac stem cells (CSC) and cardiac telocytes (TC) after 24 hrs (A) and 48 hrs (B) of culture. Notably, CSCs markedly increased in number after 48 hrs of culture (one imaged cell is undergoing mitosis – *).

Figure 2

(A) Low magnification transmission electron microscopy images of TC–CSC after 24 hrs of culture show adherens and gap junctions in a network of telopodes (Tp1‐Tp3). (B) Higher magnification of AJs (white arrows) between telopodes Tp1 and Tp2 shown in image A (rectangular mark B). (C) Higher magnification of gap junction between Tp1 and Tp3 shown in image A (rectangular mark C), highlighting that telopodes connect through different types of junctions.

Figure 3

Transmission electron microscopy images of TC–CSC culture after 24 hrs. (A, B) Serial sections show close contacts (black arrows) between cardiac stem cells (CSC ₁‐CSC ₃) and telopodes (Tp) of telocytes. (C, D) Higher magnification of rectangular marked areas in images A and B highlight the interface between cardiac stem cell CSC ₁ and a telopode (Tp). An oblique sectioned stromal synapse (arrows) is visible between Tp and CSC ₁. The length of the stromal synapse is about 2 μm (green line). Various types of vesicles may be seen at the interface between cells.

Figure 4

(A) Transmission electron microscopy images of TC–CSC culture after 48 hrs shows a telopode (Tp) in close contact with a cardiac stem cell (CSC). (B, C, D) Marked areas from image A are shown at higher magnification in the corresponding panels. A planar contact (stromal synapse) between TC and CSC can be seen associated with a number of electron‐dense structures (arrows). A puncta adherentia junction (arrowhead) is visible between TC and CSC in image C. Cp – coated pit.

Figure 5

Transmission electron microscopy shows similar intercellular connections (plain stromal synapses) between telocytes (TC) and cardiac stem cells (CSC) in culture (A) and in tissue (B). Telopodes (Tp) connect with a cardiac stem cell (CSC) through small electron‐dense structures (arrows). Endo: endothelial cell; P: pericyte; N: nerve ending; CM: cardiomyocyte.

Figure 6

(A) Transmission electron microscopy of human atrial tissue represents a glimpse into the complex environment of the cardiac stem cell niche, which comprises: telocytes (TC), cardiac stem cells (CSC), capillaries (Endo: endothelial cell; P: pericytes) and nerve endings (N). CM – adult cardiomyocyte. Arrows indicate the close contacts between a telopode (Tp) and a CSC. (B) Higher magnification of the rectangular marked area in image A.

Figure 7

Schematic representation of plain stromal synapses between the cell membranes of telopodes (Tp) and those of cardiac stem cells (CSC) in culture (A) and in tissue (B), the former spanning a distance of approximately 20 nm.

Figure 8

Schematic representation highlighting the different types of intercellular communication in TC‐CSC cultures. (modified with permission from 39).