Myometrial interstitial cells and the coordination of myometrial contractility

Abstract

A strict regulation of contractility in the uterus and fallopian tube is essential for various reproductive functions. The uterus contributes, through either increased contractility or periods of relative quiescence, to: (i) expulsion of menstrual debris, (ii) sperm transport, (iii) adequate embryo placement during implantation, (iv) enlarging its capacity during pregnancy and (v) parturition. The dominant cell population of the uterine wall consists of smooth muscle cells that contain the contractile apparatus responsible for the generation of contractile force. Recent interest has focused on a new population of cells located throughout the myometrium on the borders of smooth muscle bundles. These cells are similar to interstitial cells of Cajal (ICC) in the gut that are responsible for the generation of electrical slow waves that control peristalsis. A precise role for myometrial Cajal-like interstitial cells (m-ICLC) has not been identified. m-ICLC express the c-kit receptor, involved in creating and maintaining the ICC phenotype in the gastrointestinal tract. However, both acute and prolonged inhibition of this receptor with the c-kit antagonist imatinib mesylate does not appear to affect the spontaneous contractility of myometrium. Calcium imaging of live tissue slices suggests that contractile signalling starts on the borders of smooth muscle bundles where m-ICLC are located and recently the possible role of extracellular ATP signalling from m-ICLC has been studied. This manuscript reviews the evidence regarding tissue-level signalling in the myometrium with a particular emphasis on the anatomical and possible functional aspects of m-ICLC as new elements of the contractile mechanisms in the uterus.

Fig 1

Human myometrium. (A) Methylene blue vital staining, before cryofixation (cryosectioning). Note the selective affinity of an m-ICLC for the blue dye. (B) Silver impregnation after fixation and paraffin embedding. A pyriform m-ICLC cell with a very long, moniliform process. Original magnification: 1000×. Reprinted from Ref. [78], with permission from Elsevier.

Fig 2

Photographic reconstruction of human m-ICLC in culture establishing contacts with smooth muscle cells. In compliance with our experience, silver impregnation is one of the choice methods for revealing the typical moniliform aspect of m-ICLC in culture. Inset displaying the same interlaced distribution of m-ICLC (*) by methylene blue vital staining. Both methods reveal weaker stained myocytes. Scale bar = 10 μm. Reprinted from Ref. [63], with permission from FCMM.

Fig 3

Vital staining methods (efficient in pointing out the very long processes) performed on m-ICLC in culture. Primary confluent cultures (day 8) from pregnant human myometrium. (A) Representative m-ICLC with high affinity for methylene blue. (B) m-ICLC stained with Janus green B, a marker for mitochondria in living cells. Note the positive mitchondria in m-ICLC dilations (arrows) original magnification 40×. Modified from Ref. [8], with permission from FCMM.

Fig 4

(A) Human myometrium. Primary semi-confluent cell culture (day 4), phase contrast microscopy. Photographic reconstruction of an m-ICLC with a very long, moniliform cytoplasmic processes (arrows) emerging from cell body. Higher magnification (40×). (B) Human pregnant myometrium (39 weeks of gestation). Semi-thin sections (0.5- to –1-μm thick) of uterine muscular layer embedded in Epon resin and stained with toluidine blue. One may observe the very long process of m-ICLC squeezing between obliquely cut smooth muscle cells. Original magnification 100×.

Fig 5

Electron micrograph of human non-pregnant myometrium. Note the presence of an m-ICLC (blue) with a long thin cytoplasmic process that suddenly comes out from the cellular body and runs between smooth myocytes (brown). One may observe, on the length of the cellular process, the presence of cytoplasmic dilations housing mitochondria. Original magnification, ×9,100. TEM (photographic reconstruction) illustrating a thin (wontedly below 0.1-μm thick) cytoplasmic process of m-ICLC expanded from the cell body.

Fig 6

Human myometrium cells in culture (the second passage); immunofluorescence labelling for c-kit; FITC-conjugated secondary antibodies (green) were used to visualize the reaction (green), m-ICLC that display characteristic morphologic feature (long, moniliform processes), express c-kit and contact adjacent cells; Hoechst 33342 (blue) for nuclear counterstaining. Original magnification 60×. Reprinted from Ref. [8], with permission from FCMM.

Fig 7

Artistic view of possible interconnectivity of m-ICLC in the uterine wall. (A) Myometrial background with smooth muscle cells, nerve fibers and some connective tissue cells. (B) Close proximity of m-ICLC processes with smooth muscle cells, unmyelinated nerves, capillaries, collagen fibres and immunoreactive cells. Scale bar – approximately 7 μm (erytrocyte’s diameter).

Fig 8

Human myometrial cell culture, fourth passage. Immunocytochemical staining for the estrogen and progesterone receptors. (A) Immunocytochemical detection of estrogen receptor – dark stained nuclei (*), counterstaining with methyl green for negative nuclei. (B) ICLC stained positive for progesterone receptor. (C) Double staining (*) for CD117/c-kit (red) and estrogen receptor (black). (D) Double staining for CD117/c-kit (red) and progesterone receptor (black). Scale bar = 10 μm.

Fig 9

Representative traces of myometrial human smooth muscle inhibition by imatinib 20 μM. Reprinted from Ref. [78], with permission from Elsevier.

Fig 10

Extracellular single-unit recording performed on m-ICLC cultures (human pregnant myometrium), at the third passage. (A) Notice the spontaneous electrical activity in MIC. (B) Spontaneous field potential pattern. Reprinted from Ref. [8], with permission from FCMM.

Fig 11

Schematic representation of a smooth muscle bundle. (A) Myometrial ICs (green) produce a synchronous signal along the border of the smooth muscle bundles. (B) ATP and possibly other messenger molecules released by myometrial ICs provide the stimulus for the generation of a contraction. (C) The contraction starts from the border of the smooth muscle bundle and passes towards the centre via a calcium wave (arrows) transmitted through gap junctions.