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The cytodifferentiation of peritubular myoid cells was studied in developing rats from fetal day 18 through approachment of puberty. The parameters taken into consideration were 1) the presence of desmin, a component of intermediate filaments in contractile cells; 2) the expression of alkaline phosphatase, a cell surface enzyme present in no other cell type of the seminiferous tubule; 3) the expression of the smooth muscle specific isoform of alpha-actin, a marker of terminal differentiation in smooth muscle cells; 4) cell proliferation rate, evaluated in radioautography as labeling index after incorporation of 3H-thymidine in short-term organ culture; and 5) cytoarchitectural changes detected with scanning electron microscopy. By means of immunofluorescence and cytochemistry it was observed that the three markers are expressed early during life, long before the onset of the first spermatogenic wave; in particular desmin is already present in fetal samples and alkaline phosphatase activity appears a few days after birth, whereas alpha-smooth muscle isoactin is first detected around birth. As for myoid cell replication, the high prenatal labeling index was found to drop soon after birth and to further slow down during the first month of postnatal life, suggesting that myoid cell proliferation is not a major factor in peritubular expansion. SEM examination of developing peritubulum has shown that, when approaching puberty, the myoid cell undergoes a dramatic change in cytoarchitecture, consisting in extreme flattening and cytoplasmic expansion resulting in an apparent increase in peritubular surface.
Anat Rec 1992 May
PMID:Development and cytodifferentiation of peritubular myoid cells in the rat testis. 160 76

Morphological studies have hypothesized different origins for the precursors of the vascular smooth muscle cells (SMCs). The intriguing possibility that intimal SMCs may arise from the endothelium has newly emerged. As a first step towards understanding of the possible mechanisms involved in the transdifferentiation of endothelium into smooth muscle cells, we characterized the in vivo phenotype of the cells located in the aortic wall (distal to the aortic arches). This was accomplished using advanced stages of chicken embryo development. Furthermore, we investigated whether the cells present at the intimal thickening derive from the endothelial cell transdifferentiation. Immunolabeling of serial cryosections suggested that mesenchymal cells observed in the intimal thickening may arise from the endothelium. These cells may persist either as non-muscle throughout the development or possibly convert to cells expressing smooth muscle alpha-actin (SM alpha-actin). To determine whether endothelial cells may actually transdifferentiate into mesenchymal cells, aortic explants from 14-day-old chicken embryos (stage 40) were used. We found that explanted endothelial cells lose their cobblestone-appearance and migrate toward cell-free area. Some of these cells maintain the vWf immunoreactivity, whereas other cells coordinately lose vWf and gain SM alpha-actin expression (transitional cells). Taken together these findings strongly support the possibility that embryonic aortic endothelial transdifferentiate into mesenchymal cells, some of which express SM alpha-actin. Since TGFbeta-3 is considered an essential factor during epithelial to mesenchymal transitions in earlier chicken heart development, we also investigated the distribution of this growth factor at day 14. Our observations indicated that the immunoreactivity for TGFbeta-3 in this stage may be associated with migrating mesenchymal cells and that this immunoreactivity appears to decrease as cell differentiation advances. Therefore, the present study provides evidence that could help to explain 1) the presence of cells displaying a phenotype reminiscent of fetal-like cells in the normal chicken aorta and in the intimal region of the human aorta; 2) the SM lineage diversity in the chicken embryo reported by others; 3) a subpopulation of immature cells in the subendothelial region of the main pulmonary arteries of fetal, neonatal and adult bovines; and 4) the presence of intimal cushions, intimal pads, eccentric and diffuse intimal thickening that are observed in mammalian and avian vessels at birth.
Anat Rec 2000 01 01
PMID:Intimal thickening involves transdifferentiation of embryonic endothelial cells. 1060 48

In this study, the presence of cells in the intimal region of normal adult bovine pulmonary artery (BPA) was examined by analysis of longitudinal sections at the level of light and transmission electron microscopy. In addition, the morphological and immunohistochemical phenotype of these cells as well as the presence of particular extracellular matrix (ECM) components in this region were also determined. Since ECM production and cell proliferation have been demonstrated to be regulated by locally released growth factors such as transforming growth factor beta (TGFbeta), the presence of TGFbeta-1 in this region was also investigated. Our findings reveal the presence of immature or "nonmuscle" cells into the subendothelial space of normal adult BPA. These cells were characterized by the presence of abundant cytoplasmic organelles and scanty microfilaments. Such cells were negative to antibodies against smooth muscle alpha actin (SM alpha-actin), 1E12, and vWf, but not to vimentin. Similar cells have recently been detected in normal adult BPA and canine carotid arteries, but in the medial region. Because of their location in these elastic arteries, the nonmuscle cells are involved not only in the remodeling of the medial region, but also in the neointima or intimal thickening formation by migration from the media to the subendothelial space, where they proliferate and secrete ECM components. However, a limited number of morphological studies and the current investigation describe the presence of scattered nonmuscle cells within the intima of some normal elastic arteries. This would suggest an important role for these resident cells within the intima in normal and pathological processes as well. In addition, our results show the presence, in this region, of TGFbeta-1 and of ECM components that include collagen, elastin, fibronectin, and laminin which are present in normal conditions and during the intima formation in vivo.
Anat Rec 2000 03 01
PMID:Characterization of nonmuscle cells present in the intima of normal adult bovine pulmonary artery. 1070 46

Very little is known regarding structural and functional responses of the vascular bed of skeletal muscle to denervation and about the role of microcirculatory changes in the pathogenesis of post-denervation muscle atrophy. The purpose of the present study was to investigate the changes of the anatomical pattern of vascularization of the extensor digitorum longus muscle in WI/HicksCar rats 1, 2, 4, 7, 12, and 18 months following denervation of the limb. We found that the number of capillaries related to the number of muscle fibers, i.e. the capillary-to-fiber ratio (CFR), decreased by 88%, from 1.55 +/- 0.35 to 0.19 +/- 0.04, during the first 7 months after denervation and then slightly declined at a much lower rate during the next 11 months of observation to 10% of the CFR in normal muscle. Between months 2 and 4 after denervation, the CRF decreased by 2.4 times, from 58% to 24% of the control value. The loss of capillaries during the first 4 months following nerve transection was nearly linear and progressed with an average decrement of 4.16% per week. Electron microscopy demonstrated progressive degeneration of capillaries following nerve transection. In muscle cells close to degenerating capillaries, the loss of subsarcolemmal and intermyofibrillar mitochondria, local disassembly of myofibrils and other manifestations of progressive atrophy were frequently observed. The levels of devascularization and the degree of degenerative changes varied greatly within different topographical areas, resulting in significant heterogeneity of intercapillary distances and local capillary densities within each sample of denervated muscle. Perivascular and interstitial fibrosis that rapidly developed after denervation resulted in the spatial separation of blood vessels from muscle cells and their embedment in a dense lattice of collagen. As a result of this process, diffusion distances between capillaries and the surfaces of muscle fibers increased 10-400 times. Eighteen months after denervation most of the capillaries were heavily cushioned with collagen, and on the average 40% of the muscle cells were completely avascular. Devascularization of the tissue was accompanied by degeneration and death of muscle cells that had become embedded in a dense lattice of collagen. Immunofluorescent staining for the vascular isoform of alpha-actin revealed preservation of major blood vessels and a greater variability in thickness of their medial layer. Hyperplastic growth of the medial layer in some blood vessels resulted in narrowing of their lumens. By the end of month 7 after denervation, large deposits of collagen around arterioles often exceeded their diameters. Identification of oxidative muscle fibers after immunostaining for slow-twitch myosin, as well as using ultrastructural criteria, has shown that after 2 months of denervation oxidative muscle fibers were less susceptible to atrophy than glycolytic fibers. The lower rate of atrophy of type I muscle fibers at early stages of denervation may be explained by their initially better vascularization in normal muscle and their higher capacity to retain capillaries shortly after denervation. Thus, degeneration and loss of capillaries after denervation occurs more rapidly than the loss of muscle fibers, which results in progressive decrease of the CFR in denervated muscle. The change of capillary number in denervated muscle is biphasic: the phase of a rapid decrease of the CFR during the first 7 months after nerve transection is followed by the phase of stabilization. The presence of areas completely devoid of capillaries in denervated muscle and the virtual absence of such areas in normal muscle indicate the development of foci of regional hypoxia during long-term denervation. The anatomical pattern of muscle microvascularization changes dramatically after nerve transection. Each muscle fiber in normal muscle directly contacts on average 3-5 capillaries. (ABSTRACT TRUNCATED)
Anat Rec 2000 03 01
PMID:Remodeling of the vascular bed and progressive loss of capillaries in denervated skeletal muscle. 1070 50

The infrared sensory membranes of pit organs of pit vipers have an extremely rich capillary vasculature that forms many vascular loops, each serving a small number of infrared nerve terminals. We clarified the ultrastructure of capillary pericytes in the pit membranes by scanning and transmission electron microscopy, and examined the immunoreactivity in their cytoplasm to two contractile proteins: smooth muscle alpha-actin (SM alpha-actin) and desmin. The capillary pericytes had two major cytoplasmic processes: thickened primary processes that radiate to embrace the endothelial tube and flattened secondary processes that are distributed widely on the endothelium. Coexpression of SM alpha-actin and desmin was observed in the pericytes of entire capillary segments, and SM alpha-actin was characterized by prominent filament bundles directed mainly at right angles to the capillary long axis. This expression pattern was different from that of capillary pericytes of the scales, where SM alpha-actin was expressed diffusely in the cytoplasm. In a series of electron microscopic sections, we often observed the pericyte processes depressing the endothelial wall. We also observed a close relationship of the pericytes with inter-endothelial cell junctions, and pericyte processes connected with the endothelial cells via gap junctions. From these findings, we surmised that capillary pericytes in the pit membrane have a close functional relationship with the endothelium, and through their contractile and relaxing activity regulate capillary bloodflow to stabilize production of infrared nerve impulses.
Anat Rec 2000 11 01
PMID:Ultrastructure of the capillary pericytes and the expression of smooth muscle alpha-actin and desmin in the snake infrared sensory organs. 1106 40

Blood-tissue exchange and homeostasis within the organs depend on various interactions between endothelial and perivascular cells (Buniatian, 2001). Podocytes possess anatomical and cellular features intermediate between those of astrocytes and hepatic stellate cells (HSCs). Podocytes, like HSCs, are associated with fenestrated capillaries and, similar to astrocytes, interact with the capillaries via the basement membrane and participate in permeability-limiting ultrafiltration. The fact that podocytes come in direct contact with xenobiotics prompted us to investigate whether they express metallothionein (MT), an anticytotoxic system characteristic of astrocytes. In comparative studies, cryosections of 1- and 3-month-old rat kidney and adult rat brain, as well as podocytes and astrocytes from early and prolonged primary cultures of glomerular explants and newborn rat brain, respectively, were investigated. The cells were double-labeled with antiserum against glial fibrillary acidic protein (GFAP) and monoclonal antibody (MAb) against the lysine-containing epitope of Cd/Zn-MT-I (MAb MT) or MAb against alpha-actin. In kidney sections, MT immunoreactivity was detected in GFAP-positive glomerular cells and in interstitial fibroblasts. The pattern of staining for MT and GFAP in glomerular cells was similar to that of astrocytes in vivo. In glomerular cell cultures, MT was expressed in cobblestone-like podocytes which contained Wilms' tumor protein and lacked desmin. MT was upregulated at later culture periods, during which podocytes acquired features typical of undifferentiated astrocytes. This study hints at the existence of common regulatory mechanisms of blood-tissue interactions by neural and non-neural perivascular cells. These mechanisms appear to be used in an organ-specific manner.
Anat Rec 2002 Aug 01
PMID:Glial fibrillary acidic protein-positive cells of the kidney are capable of raising a protective biochemical barrier similar to astrocytes: expression of metallothionein in podocytes. 1212 8

Monolayers of retracted endothelial cells exhibiting wounds or zones denuded of cells were obtained from aortic explants from 10- to 12-day-old chicken embryos. Using time-lapse videomicroscopy, we investigated the sequence of events that occurred both during and after closure of the monolayer wounds. Such wound closure (re-endothelialization process) occurred 4-12 hr after removing the explants, depending on wound width and presence of serum. The cells from along the wound edges appeared to move toward one another. We suggest an important role for bFGF and TGFbeta-2 and -3 during this process. Twenty-five hours after removal there were still some areas of retracted cells, and many of the cells displayed a weak von Willebrand's Factor (vWf) immunoreactivity. Surprisingly, after 63-65 hr many of the endothelial cells had become epithelioid in shape and the vWf immunoreactivity appeared increased. This epithelioid phenotype is currently considered typical of cultured vascular non-muscle-like cells and intimal thickening cells. By 5-7 days, the vast majority of cells in the monolayer had acquired an epithelioid morphology, showing a cobblestone appearance. These cells were significantly smaller than polygonal cells. Most importantly, they showed strong vWf immunoreactivity. At the edge of the monolayers we found that the majority of the cells had become epithelioid. Some of them detached from their neighbors and became round in shape and acquired mesenchymal characteristics, some expressing smooth muscle alpha-actin (SM alpha-actin). These findings demonstrate not only that embryonic endothelial cells that are transiently mechanically altered may change their phenotype to an epithelioid phenotype, but also that these cells may eventually transdifferentiate into mesenchymal cells expressing SM alpha-actin. Since some aspects of endothelial cell behavior have been shown to be regulated by locally released growth factors such as TGFbeta and FGF, we also investigated TGFbeta-2 and -3 and bFGF expression. Presence of TGFbeta-2 and -3 and bFGF-immunoreactive epithelioid and mesenchymal cells indicates that these growth factors may be involved in the changes described.
Anat Rec A Discov Mol Cell Evol Biol 2003 Jan
PMID:Mechanically altered embryonic chicken endothelial cells change their phenotype to an epithelioid phenotype. 1249 91

Serum response factor (SRF), a transcription factor ubiquitously involved in the processes of cellular proliferation and differentiation, has been implicated in cardiac and skeletal muscle development because of its strong expression in embryonic muscle lineages, and its necessity for the transcription of transiently transfected muscle genes that contain SRF binding sites. This study was designed to ascertain whether SRF is required for the expression of an endogenous SRF-dependent gene during differentiation of early embryonic cardiac myocytes by introducing a dominant-negative SRF construct via retroviral delivery. Although no effect on overt cellular differentiation was detected, semi-quantitative RT-PCR revealed that expression of the SRF-dependent gene cardiac alpha-actin was inhibited, whereas expression of the non-SRF-dependent genes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and cardiac troponin-C was unaffected. No effect on myocyte proliferation was detected. Curiously, immunohistochemical localization of SRF protein suggested that whereas endogenous SRF was homogeneously dispersed throughout the cytoplasm and nucleus, the dominant-negative SRF protein was concentrated in the nucleus. These results extend previous findings using transiently transfected genes to the endogenous level, indicating that SRF is required for the full expression of muscle genes that contain SRF binding sites during cardiac myocyte differentiation.
Anat Rec A Discov Mol Cell Evol Biol 2003 Apr
PMID:Inhibition of the cardiac alpha-actin gene in embryonic cardiac myocytes by dominant-negative serum response factor. 1262 74

In a recent paper written with the purpose of shedding light on the question of whether genomic GC levels are related to temperature in vertebrates, Ream et al. [Mol. Biol. Evol. 20 (2003) 105] offered an analysis of two sets of homologous genes: those coding for alpha-actin and lactate dehydrogenase-A (LDH-A). The conclusion was that "there is no consistent relationship between adaptation temperature and the percentage of thermal stability-enhancing G+C base pairs in protein-coding genes". We argue here that the data presented neither prove nor suggest such a conclusion because of conceptual and methodological errors.
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PMID:LDH-A and alpha-actin as tools to assess the effects of temperature on the vertebrate genome: some problems. 1460 4

The electromyographic properties of the cremaster muscle (CM) are quite different from other skeletal muscles. It shows excessive spontaneous discharges, and the motor unit shape and firing frequency of the CM muscle differ from that of limb muscles. In this study, CM of six adult cadavers and six orchiectomy specimens were used to reveal the detailed histology of the muscle and provide an anatomophysiological explanation for these unusual electromyographic properties. Routine histochemical stains revealed the CM was composed of several distinct bundles of smooth and striated muscle fibers within connective tissue. The smooth muscle fibers that were more profuse than previously known and were not arranged in layers, but widely dispersed between striated muscle fibers. Bielschowsky silver staining technique, anti-neurofilament and anti-synaptophysin immunostaining showed the presence of multiple motor end-plates observed as a series of small dots or lines running along the striated muscle fibers and several nerve endings on a single muscle fiber. Myosin immunostaining confirmed the CM is a slow-twitch muscle, and alpha-actin smooth muscle immunostaining confirmed the presence of a large number of smooth muscle fibers. There were also small multipolar neurons forming nerve plexuses between smooth muscle fibers. Anti-GFAP immunostaining confirmed the presence of glial cells similar to astrocytes. In conclusion, the findings of this detailed anatomical study showed the CM, widely known as a striated muscle, contains a large number of smooth muscle fibers, and the spontaneous electromyographic discharges are due to the presence of multiple motor end-plates and dense innervation.
Anat Rec (Hoboken) 2008 Jul
PMID:Morphology and innervation of the human cremaster muscle in relation to its function. 1844 92


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