scholarly journals Immunoreactivity and a new staining method of monocarboxylate transporter 1 located in endothelial cells of cerebral vessels of human brain in distinguishing cerebral venules from arterioles

2021 ◽  
Vol 65 (s1) ◽  
Author(s):  
Yuan Cao ◽  
Dong-Hui Ao ◽  
Chao Ma ◽  
Wen-Ying Qiu ◽  
Yi-Cheng Zhu
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Human Brain ◽  
Smooth Muscle Actin ◽  
Cerebral Vessels ◽  
Monocarboxylate Transporter ◽  
Specific Marker ◽  
Cerebral Veins ◽  
Internal Elastic Lamina ◽  
Monocarboxylate Transporter 1

Distinguishing brain venules from arterioles with arteriolosclerosis is less reliable using traditional staining methods. We aimed to immunohistochemically assess the monocarboxylate transporter 1 (MCT1), a specific marker of venous endothelium found in rodent studies, in different caliber vessels in human brains. Both largeand small-caliber cerebral vessels were dissected from four autopsy donors. Immunoreactivity for MCT1 was examined in all autopsied human brain tissues, and then each vessel was identified by neuropathologists using hematoxylin and eosin stain, the Verhoeff’s Van Gieson stain, immunohistochemical stain with antibodies for α-smooth muscle actin and MCT1 in sequence. A total of 61 cerebral vessels, including 29 arteries and 32 veins were assessed. Immunoreactivity for MCT1 was observed in the endothelial cells of various caliber veins as well as the capillaries, whereas that was immunenegative in the endothelium of arteries. The different labeling patterns for MCT1 could aid in distinguishing various caliber veins from arteries, whereas assessment using the vessel shape, the internal elastic lamina, and the pattern of smooth muscle fibers failed to make the distinction between small-caliber veins and sclerotic arterioles. In conclusion, MCT1 immunohistochemical staining is a sensitive and reliable method to distinguish cerebral veins from arteries.

Abstract 418: Co-Culture of Endothelial Cells, Smooth Muscle Cells and Monocytes in Collagen Scaffold: A Novel i n vitro Model of Atherosclerosis

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Martin Liu ◽  
Angelos Karagiannis ◽  
Matthew Sis ◽  
Srivatsan Kidambi ◽  
Yiannis Chatzizisis
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Type I Collagen ◽  
Smooth Muscle Actin ◽  
Muscle Cells ◽  
Type I ◽  
Collagen Gels ◽  
Human Coronary Artery

Objectives: To develop and validate a 3D in-vitro model of atherosclerosis that enables direct interaction between various cell types and/or extracellular matrix. Methods and Results: Type I collagen (0.75 mg/mL) was mixed with human artery smooth muscle cells (SMCs; 6x10 5 cells/mL), medium, and water. Human coronary artery endothelial cells (HCAECs; 10 5 /cm 2 ) were plated on top of the collagen gels and activated with oxidized low density lipoprotein cholesterol (LDL-C). Monocytes (THP-1 cells; 10 5 /cm 2 ) were then added on top of the HCAECs. Immunofluorescence showed the expression of VE-cadherin by HCAECs (A, B) and α-smooth muscle actin by SMCs (A). Green-labelled LDL-C particles were accumulated in the subendothelial space, as well as in the cytoplasm of HCAECs and SMCs (C). Activated monocytes were attached to HCAECs and found in the subendothelial area (G-I). Both HCAECs and SMCs released IL-1β, IL-6, IL-8, PDGF-BB, TGF-ß1, and VEGF. Scanning and transmission electron microscopy showed the HCAECs monolayer forming gap junctions and the SMCs (D-F) and transmigrating monocytes within the collagen matrix (G-I). Conclusions: In this work, we presented a novel, easily reproducible and functional in-vitro experimental model of atherosclerosis that has the potential to enable in-vitro sophisticated molecular and drug development studies.

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.

scholarly journals Dysfunction of brain pericytes in chronic neuroinflammation

2015 ◽  
Vol 36 (4) ◽  
pp. 794-807 ◽  
Author(s):  
Yuri Persidsky ◽  
Jeremy Hill ◽  
Ming Zhang ◽  
Holly Dykstra ◽  
Malika Winfield ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Human Brain ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Neurovascular Unit ◽  
Brain Endothelial Cells ◽  
Monocyte Migration ◽  
Brain Pericytes ◽  
Hiv 1

Brain pericytes are uniquely positioned within the neurovascular unit to provide support to blood brain barrier (BBB) maintenance. Neurologic conditions, such as HIV-1-associated neurocognitive disorder, are associated with BBB compromise due to chronic inflammation. Little is known about pericyte dysfunction during HIV-1 infection. We found decreased expression of pericyte markers in human brains from HIV-1-infected patients (even those on antiretroviral therapy). Using primary human brain pericytes, we assessed expression of pericyte markers (α1-integrin, α-smooth muscle actin, platelet-derived growth factor-B receptor β, CX-43) and found their downregulation after treatment with tumor necrosis factor-α (TNFα) or interleukin-1 β (IL-1β). Pericyte exposure to virus or cytokines resulted in decreased secretion of factors promoting BBB formation (angiopoietin-1, transforming growth factor-β1) and mRNA for basement membrane components. TNFα and IL-1β enhanced expression of adhesion molecules in pericytes paralleling increased monocyte adhesion to pericytes. Monocyte migration across BBB models composed of human brain endothelial cells and pericytes demonstrated a diminished rate in baseline migration compared to constructs composed only of brain endothelial cells. However, exposure to the relevant chemokine, CCL2, enhanced the magnitude of monocyte migration when compared to BBB models composed of brain endothelial cells only. These data suggest an important role of pericytes in BBB regulation in neuroinflammation.

scholarly journals The alpha 6 and beta 4 integrin subunits are expressed by smooth muscle cells of human small vessels: a new localization in mesenchymal cells.

1994 ◽  
Vol 42 (9) ◽  
pp. 1221-1228 ◽  
Author(s):  
O Cremona ◽  
P Savoia ◽  
P C Marchisio ◽  
G Gabbiani ◽  
C Chaponnier
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Smooth Muscle Actin ◽  
Muscle Cells ◽  
Confocal Laser ◽  
Alpha Smooth Muscle Actin ◽  
Excitation Spectra ◽  
Small Vessels ◽  
Von Willebrand

The alpha 6 beta 4 integrin complex is generally thought to be expressed by epithelial cells, where it is localized in specific adhesion structures, the hemidesmosomes. Recent observations have suggested a new localization of the beta 4 integrin chain in small vessels, possibly in endothelial cells, i.e., in cells of mesenchymal origin. In the present study we show that (a) the alpha 6 and beta 4 integrin chains are not expressed by endothelial cells, since they are not localized in von Willebrand factor-producing cells; (b) instead, smooth muscle cells of small vessels are intensely positive to antibodies to both alpha 6 and beta 4 intergrin chains; and (c) in some restricted regions of these smooth muscle cells there is a clear colocalization between alpha-smooth muscle actin and alpha 6 and beta 4 integrin chains, suggesting that a new type of cytoskeletal linkage for the alpha 6 beta 4 integrin complex may occur in mesenchyme-derived cells. Our observations are supported by confocal laser microscopy (CLSM) images of specimens labeled by double immunofluorescence. This technical choice was made to take advantage of the higher resolution offered by CLSM in comparison with conventional immunofluorescence. A careful selection of barrier filters was necessary to separate accurately emission and excitation spectra of the fluorochromes used in this study, resulting in an efficient colocalization analysis.

scholarly journals ELTD1 Activation Induces an Endothelial-EMT Transition to a Myofibroblast Phenotype

2021 ◽  
Vol 22 (20) ◽  
pp. 11293
Author(s):  
Helen Sheldon ◽  
John Alexander ◽  
Esther Bridges ◽  
Lucia Moreira ◽  
Svetlana Reilly ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Wound Repair ◽  
Network Formation ◽  
Smooth Muscle Actin ◽  
Focal Adhesions ◽  
Enrichment Analysis ◽  
Gene Set Enrichment Analysis ◽  
Immune Response Gene ◽  
Cell Contact

ELTD1 is expressed in endothelial and vascular smooth muscle cells and has a role in angiogenesis. It has been classified as an adhesion GPCR, but as yet, no ligand has been identified and its function remains unknown. To establish its role, ELTD1 was overexpressed in endothelial cells. Expression and consequently ligand independent activation of ELTD1 results in endothelial-mesenchymal transistion (EndMT) with a loss of cell-cell contact, formation of stress fibres and mature focal adhesions and an increased expression of smooth muscle actin. The effect was pro-angiogenic, increasing Matrigel network formation and endothelial sprouting. RNA-Seq analysis after the cells had undergone EndMT revealed large increases in chemokines and cytokines involved in regulating immune response. Gene set enrichment analysis of the data identified a number of pathways involved in myofibroblast biology suggesting that the endothelial cells had undergone a type II EMT. This type of EMT is involved in wound repair and is closely associated with inflammation implicating ELTD1 in these processes.

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.

Abstract 724: Synectin is Required for Normal PDGF Secretion and Smooth Muscle Cell Basic Science Recruitment

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Julie M Paye ◽  
Thomas W Chittenden ◽  
Li-Kun Phng ◽  
Holger Gerhardt ◽  
Michael Simons
Keyword(s):  
Endothelial Cells ◽  
Transcriptional Regulation ◽  
Smooth Muscle ◽  
Smooth Muscle Cell ◽  
Muscle Cell ◽  
Smooth Muscle Actin ◽  
Alpha Smooth Muscle Actin ◽  
Arterial Tree ◽  
Mural Cell ◽  
Post Transcriptional Regulation

Background: Synectin, a ubiquitously expressed scaffold and PDZ protein, has been shown to be a key regulator in the formation of arterial vasculature. Primary arterial endothelial cells isolated from synectin−/− mice exhibited defects in PDGF signaling that were not present in arterial synectin+/+ or either venous synectin+/+ or synectin−/− endothelial cells. Methods: To address the role of synectin in mural cell recruitment, retinas from synectin−/− (KO) and synectin+/+ (WT) mice were stained for isolectin and alpha-smooth muscle actin. To determine the mechanisms responsible for defective smooth muscle cells recruitment, primary arterial and venous endothelial (EC) and smooth muscle (SMC) cells were isolated from synectin+/+ and synectin−/− mice. Results: Examination of the retinal vasculature in synectin−/− mice demonstrated poor smooth muscle cell coverage of the forming arterial tree with a number of SMC cells sitting some distance away from the vessels, a defect reminiscent of retinal abnormalities observed in PDGF-B−/− mice. Western blot analysis of lysates from primary KO SMC demonstrated levels of PDGFR-β analogous to WT, although expression of PDGF-B was markedly reduced in lysates from arterial, but not venous, primary KO endothelial cells. Transduction of KO EC with an adenoviral synectin construct was sufficient to restore PDGF-B protein levels. Further, qPCR was used to determine whether defects in PDGF-B protein production were at the transcription or translation level. Interestingly, there was only a minor decrease in the amount of PDGF-B mRNA in KO compared to WT EC, suggesting that synectin deficiency has selectively affected post-transcriptional regulation of PDGF-B expression. Migration of WT and KO SMC in response to PDGF-B or conditioned media from WT and KO endothelial cells was also assessed to determine the cell type specificity of PDGF defects in KO mice. Conclusions: Synectin is required for post-transcriptional regulation of PDGF-B in endothelial cells. Deficits in PDGF-B secretion by synectin−/− endothelial cells result in inadequate smooth muscle recruitment and coverage of arterial vessels.

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