Distinction between smooth muscle, fibroblasts and endothelial cells in culture by the use of fluoresceinated antibodies against smooth muscle actin

1977 ◽  
Vol 177 (4) ◽  
Author(s):  
JulieH. Chamley ◽  
Ute Gr�schel-Stewart ◽  
GordonR. Campbell ◽  
G. Burnstock
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Muscle Actin ◽  
Cells In Culture

scholarly journals Host-Derived Pericytes and Sca-1+ Cells Predominate in the MART-1− Stroma Fraction of Experimentally Induced Melanoma

2011 ◽  
Vol 59 (12) ◽  
pp. 1060-1075 ◽  
Author(s):  
J. Humberto Treviño-Villarreal ◽  
Douglas A. Cotanche ◽  
Rosalinda Sepúlveda ◽  
Magda E. Bortoni ◽  
Otto Manneberg ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Smooth Muscle ◽  
Blood Vessel ◽  
Smooth Muscle Actin ◽  
Cell Types ◽  
Muscle Actin ◽  
Cellular Composition ◽  
Tumor Capsule ◽  
Form Blood Vessel

Identification of cell types in tumor-associated stroma that are involved in the development of melanoma is hampered by their heterogeneity. The authors used flow cytometry and immunohistochemistry to demonstrate that anti–MART-1 antibodies can discriminate between melanoma and stroma cells. They investigated the cellular composition of the MART-1−, non-hematopoietic melanoma-associated stroma, finding it consisted mainly of Sca-1+ and CD146+ cells. These cell types were also observed in the skin and muscle adjacent to developing melanomas. The Sca-1+ cell population was observed distributed in the epidermis, hair follicle bulges, and tumor capsule. The CD146+ population was found distributed within the tumor, mainly associated with blood vessels in a perivascular location. In addition to a perivascular distribution, CD146+ cells expressed α-smooth muscle actin, lacked expression of endothelial markers CD31 and CD34, and were therefore identified as pericytes. Pericytes were found to be associated with CD31+ endothelial cells; however, some pericytes were also observed associated with CD31−, MART-1+ B16 melanoma cells that appeared to form blood vessel structures. Furthermore, the authors observed extensive nuclear expression of HIF-1α in melanoma and stroma cells, suggesting hypoxia is an important factor associated with the melanoma microenvironment and vascularization. The results suggest that pericytes and Sca-1+ stroma cells are important contributors to melanoma development.

Endothelial cells genetically selected from differentiating mouse embryonic stem cells incorporate at sites of neovascularization in vivo

2002 ◽  
Vol 115 (10) ◽  
pp. 2075-2085 ◽  
Author(s):  
Sandrine Marchetti ◽  
Clotilde Gimond ◽  
Kristiina Iljin ◽  
Christine Bourcier ◽  
Kari Alitalo ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Endothelial Cell ◽  
Smooth Muscle Cell ◽  
Muscle Cell ◽  
Es Cells ◽  
Smooth Muscle Actin ◽  
Embryonic Stem ◽  
Muscle Actin ◽  
Puromycin Selection

Large scale purification of endothelial cells is of great interest as it could improve tissue transplantation, reperfusion of ischemic tissues and treatment of pathologies in which an endothelial cell dysfunction exists. In this study, we describe a novel genetic approach that selects for endothelial cells from differentiating embryonic stem (ES) cells. Our strategy is based on the establishment of ES-cell clones that carry an integrated puromycin resistance gene under the control of a vascular endothelium-specific promoter, tie-1. Using EGFP as a reporter gene, we first confirmed the endothelial specificity of the tie-1 promoter in the embryoid body model and in cells differentiated in 2D cultures. Subsequently, tie-1-EGFP ES cells were used as recipients for the tie-1-driven puror transgene. The resulting stable clones were expanded and differentiated for seven days in the presence of VEGF before puromycin selection. As expected, puromycin-resistant cells were positive for EGFP and also expressed several endothelial markers, including CD31, CD34,VEGFR-1, VEGFR-2, Tie-1, VE-cadherin and ICAM-2. Release from the puromycin selection resulted in the appearance of α-smooth muscle actin-positive cells. Such cells became more numerous when the population was cultured on laminin-1 or in the presence of TGF-β1, two known inducers of smooth muscle cell differentiation. The hypothesis that endothelial cells or their progenitors may differentiate towards a smooth muscle cell phenotype was further supported by the presence of cells expressing both CD31 andα-smooth muscle actin markers. Finally, we show that purified endothelial cells can incorporate into the neovasculature of transplanted tumors in nude mice. Taken together, these results suggest that application of endothelial lineage selection to differentiating ES cells may become a useful approach for future pro-angiogenic and endothelial cell replacement therapies.

scholarly journals Cardiac microvascular endothelial cells express α-smooth muscle actin and show low NOS III activity

1999 ◽  
Vol 276 (5) ◽  
pp. H1755-H1768 ◽  
Author(s):  
Hiroshi Ando ◽  
Thomas Kubin ◽  
Wolfgang Schaper ◽  
Jutta Schaper
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Microvascular Endothelial Cells ◽  
Low Grade ◽  
Muscle Actin ◽  
Microvascular Endothelial

We established a culture system of porcine coronary microvascular endothelial cells (MVEC) with high cellular yield and purity >98%. Endothelial origin was confirmed by immunostaining, immunoblotting and fluorescence-activated cell sorter (FACS) analysis using low-density lipoprotein uptake, CD31, von Willebrand factor, and the lectin Dolichos biflorus agglutinin. MVEC were positive for α-smooth muscle actin in culture and in myocardium, as confirmed by FACS. Of the primary MVEC, ∼30% expressed nitric oxide synthase (NOS) III in numbers decreasing from the first passage (6 ± 1%) to the second passage (4 ± 1%; P < 0.001 vs. primary isolates), whereas ∼100% of aortic endothelial cells (AEC) expressed NOS III. In AEC, NOS III activity (pmol citrulline ⋅ mg protein−1 ⋅ min−1) was 80 ± 10 and was nearly abolished in the absence of calcium (5 ± 1, P < 0.001). In primary MVEC, however, NOS III activity in the presence and absence of calcium was 20 ± 4 and 25 ± 5, respectively. We conclude that cardiac MVEC, in contrast to AEC, contain α-smooth muscle actin, show low-grade NOS III activity, and provide a suitable in vitro system for the study of endothelial pathophysiology.

scholarly journals Pulmonary endothelial cells exhibit sexual dimorphism in their response to hyperoxia

2018 ◽  
Vol 315 (5) ◽  
pp. H1287-H1292 ◽  
Author(s):  
Yuhao Zhang ◽  
Xiaoyu Dong ◽  
Jasmine Shirazi ◽  
Jason P. Gleghorn ◽  
Krithika Lingappan
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Cell Migration ◽  
Smooth Muscle Actin ◽  
Microvascular Endothelial Cells ◽  
Muscle Actin ◽  
Mesenchymal Transition ◽  
Pulmonary Microvascular Endothelial Cells ◽  
Hyperoxia Exposure ◽  
Microvascular Endothelial

Abnormal pulmonary vascular development is a critical factor in the pathogenesis of bronchopulmonary dysplasia (BPD). Despite the well-established sex-specific differences in the incidence of BPD, the molecular mechanism(s) behind these are not completely understood. Exposure to a high concentration of oxygen (hyperoxia) contributes to BPD and creates a profibrotic environment in the lung. Our objective was to elucidate the sex-specific differences in neonatal human pulmonary microvascular endothelial cells (HPMECs) in normoxic and hyperoxic conditions, including the propensity for endothelial-to-mesenchymal transition. HPMECs (18- to 24-wk gestation donors, 6 male donors and 5 female donors) were subjected to hyperoxia (95% O2 and 5% CO2) or normoxia (air and 5% CO2) up to 72 h. We assessed cell migration and angiogenesis at baseline. Cell proliferation, viability, and expression of endothelial (CD31) and fibroblast markers (α-smooth muscle actin) were measured upon exposure to hyperoxia. Female HPMECs had significantly higher cell migration when assessed by the wound healing assay (40.99 ± 4.4%) compared with male HPMECs (14.76 ± 3.7%) and showed greater sprouting (1710 ± 962 μm in female cells vs. 789 ± 324 in male cells) compared with male endothelial cells in normoxia. Hyperoxia exposure decreased cell viability (by 9.8% at 48 h and 11.7% at 72 h) and proliferation (by 26.7% at 72 h) markedly in male HPMECs, whereas viability was sustained in female endothelial cells. There was greater expression of α-smooth muscle actin (2.5-fold) and decreased expression (5-fold) of CD31 in male HPMECs upon exposure to hyperoxia. The results indicate that cellular sex affects response in HPMECs in normoxia and hyperoxia. NEW & NOTEWORTHY Cellular sex affects response in human neonatal pulmonary microvascular endothelial cells in normoxia and hyperoxia. Under normoxic conditions, female human neonatal pulmonary microvascular endothelial cells display greater migration and angiogenic sprouting compared with male endothelial cells. Compared with female endothelial cells, hyperoxia exposure decreased cell viability and proliferation and increased α-smooth muscle actin and decreased CD31 expression in male endothelial cells, indicating an increased endothelial-mesenchymal transition.

scholarly journals Congenital Cutaneous Kaposiform Hemangioendothelioma in a Newborn Calf

2020 ◽  
Vol 43 (2) ◽  
pp. 193-196
Author(s):  
Erkmen Tuğrul Epikmen ◽  
Ahmet Aydogan ◽  
Hamdi Avci ◽  
Sümbül Serap Birincioğlu
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Tumor Cells ◽  
Histopathological Examination ◽  
Smooth Muscle Actin ◽  
Whole Body ◽  
Muscle Actin ◽  
Alpha Smooth Muscle Actin ◽  
Kaposiform Hemangioendothelioma ◽  
The Masses

AbstractA one-day-old female Holstein calf was presented with subcutaneous masses spread over the whole body. Macroscopically, the masses were firm in touch, greyish-white in colour, 0.5-2 cm in diameter range. Histopathological examination confirmed the cutaneous Kaposiform hemangioendothelioma (KHE). Microscopic examination of the tumor revealed sheets of spindled endothelial cells forming vascular slits. Immunohistochemically, the tumor cells and capillaries gave strongly positive reaction for CD31 while vimentin, alpha smooth muscle actin and cytokeratin AE1/AE3 were negative. In this case, macroscopical, detailed histhopathological and immunohistochemical findings of congenital KHE reported firstly in a newborn calf.

Transforming growth factor beta 1 promotes the differentiation of endothelial cells into smooth muscle-like cells in vitro

1992 ◽  
Vol 103 (2) ◽  
pp. 521-529 ◽  
Author(s):  
E. Arciniegas ◽  
A.B. Sutton ◽  
T.D. Allen ◽  
A.M. Schor
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Muscle Actin ◽  
Tgf Beta ◽  
Alpha Smooth Muscle Actin ◽  
Aortic Endothelial Cells

Alpha-smooth muscle actin is considered a reliable marker for distinguishing between arterial smooth muscle and endothelial cells. Several authors have reported heterogeneity in the expression of this actin isoform in atherosclerotic lesions. Such heterogeneity appears to result from the presence of different smooth muscle cell phenotypes (contractile and synthetic) in these lesions. In the present study, we show that bovine aortic endothelial cells, which are characterised by the presence of Factor VIII-related antigen (FVIII) and by the absence of alpha-smooth muscle actin (alpha-SM actin) may be induced to express the latter when exposed to TGF-beta 1. FVIII was detected by immunofluorescence, alpha-SM actin was detected by immunofluorescence and immunoblotting. The number of cells expressing alpha-SM actin increased with time of incubation with TGF-beta 1, and this increase occurred concomitantly with a decrease in the expression of FVIII. Double immunofluorescence demonstrated the presence of cells that expressed both FVIII and alpha-SM actin after 5 days of incubation with TGF-beta 1. With longer incubation times (10-20 days) the loss of FVIII expression was complete and over 90% of the cells expressed alpha-SM actin. Ultrastructurally, cells in control cultures showed the typical features of endothelial cells. In the TGF-beta 1-treated cultures, cells which appeared indistinguishable from contractile and synthetic smooth muscle cells were observed. Withdrawal of TGF-beta 1 after 10 days incubation resulted in the re-appearance of polygonal cells which were FVIII-positive and alpha-SM actin-negative.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Survival of the posterior lamellar cornea graft keratocytes and endothelial cells cultivated in the modified corneal preservation media

2021 ◽  
pp. 32-39
Author(s):  
S.A. Borzenok ◽  
◽  
B.E. Malyugin ◽  
D.S. Ostrovskiy ◽  
A.K. Ahmedov ◽  
...  
Keyword(s):  
Cell Culture ◽  
Endothelial Cells ◽  
Smooth Muscle ◽  
Cell Cultures ◽  
Corneal Endothelium ◽  
Smooth Muscle Actin ◽  
Tissue Bank ◽  
Muscle Actin ◽  
Human Donor ◽  
Storage Media

Purpose. To study the survival of keratocytes and endothelial cells of a human donor cornea storage in the standard and the new media which was specifically designed for optimized cornea hydration. Material and methods. 2D cell cultures of keratocytes and endothelial cells obtained from the Eye tissue bank were used for culture in improved storage media over a period of 14 and 7 days subsequently. To confirm phenotype characteristics, the cells were stained by the following markers: for keratocytes – Lumikan, Keratocan, and α-smooth muscle actin; for endothelial cells – ZO-1 and Na/K-ATPase. The onset of apoptosis in cell culture of keratocytes were detected with Cytochrome C, BAX, and Caspase 3 and 8. Viability of cell cultures after the cultivation was carried out using a commercial set of "Live and Dead". Morphology of the endothelial cells was assessed using an electron scanning microscope. Results. It was shown that the 2D keratocyte culture cultured in the improved storage media expressed specific markers: Lumican, Keratocan, and did not express α-smooth muscle actin. There were no markers of apoptosis in the cell culture of keratocytes after 14 days of cultivation. Corneal endothelium cultured in the improved storage media expresses ZO-1, Na/K-ATPase and presented hexagonal cell shape morphology according to electron microscopy. Conclusion. The improved storage media allow to preserve the unique phenotype of keratocytes, with a slight decrease in proliferative cells activity during 14 days. The media maintain a viable and functional corneal endothelium for at least seven days of cultivation. Key words: cell culture; corneal endothelium; keratocyte; posterior lamellar graft, corneal storage media.

AB0237 Endothelin-1 induces alpha-smooth muscle actin (alpha-SMA) expression in cultured human endothelial cells

2013 ◽  
Vol 71 (Suppl 3) ◽  
pp. 651.2-651
Author(s):  
S. Soldano ◽  
P. Montagna ◽  
R. Brizzolara ◽  
A. Sulli ◽  
B. Villaggio ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Muscle Actin ◽  
Human Endothelial Cells ◽  
Alpha Smooth Muscle Actin ◽  
Endothelin 1
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