The relationship between von willebrand factor antigen and fibronectin in human plasma, endothelial cells and fibroblasts in culture

Giddings, J. C.; Hogg, S. D.; Legg, I.R.; Hughes, I.A.

doi:10.1016/0049-3848(86)90004-6

Cited by 4 publications

(1 citation statement)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The minimal vWF staining in fibroblasts likely represents an artifact. However, expression of vWF in fibroblasts has been reported if cocultured with ECs or medium has been preconditioned (13). Furthermore, the morphological differences between the cell types as described above are visible.…”

Section: Resultsmentioning

confidence: 97%

Isolation and Culture of Fibroblasts, Vascular Smooth Muscle, and Endothelial Cells From the Fetal Rat Ductus Arteriosus

et al. 2011

View full text Add to dashboard Cite

ABSTRACT:The ductus arteriosus (DA), a fetal arterial shunt vessel between the proximal descending aorta and the pulmonary artery, closes shortly after birth. Initial functional closure as a result of the DA's smooth muscle contraction is followed by definite anatomical closure. The latter involves several complex mechanisms like endothelial cushion formation and smooth muscle cell migration resulting in fibrosis and sealing of the vessel. These complex steps indicate highly specialized functions of the DA vascular smooth muscle cells (VSMCs), endothelial cells, and fibroblasts. Herein, we describe a new reproducible method for isolating VSMCs, endothelial cells, and fibroblasts of high viability from fetal rat DA using immunomagnetic cell sorting. Purity of the different cell cultures was assessed by immunohistochemistry and flow cytometry and ranged between 85 and 94%. The capability of the VSMCs to react to hypoxic stimuli was assessed by intracellular calcium and ATP measurements and by VEGF mRNA expression analysis. VSMCs respond to hypoxia with decreases in intracellular calcium concentrations and ATP levels, whereas VEGF mRNA expression increased 3.2-fold. The purified vessel-specific different cell types are suitable for subsequent gene expression profiling and functional studies and provide important tools for improving our understanding of the complex processes involved in the closure of the DA. T he ductus arteriosus (DA) is a shunt which connects the proximal descending aorta and the main pulmonary artery (PA). During fetal life, it serves to bypass the pulmonary circulation. The DA closes shortly after birth, and pulmonary blood flow increases during late gestation (1). After birth, an abrupt increase in oxygen tension results in ductal constriction, which is further promoted by falling levels of prostaglandin E2 because of an increased pulmonary metabolism and the elimination of placental prostaglandin. DA medial smooth muscle contraction leads to wall thickening, luminal obliteration, and shortening of the vessel. Permanent sealing of the DA is a process generated by the infolding of the endothelium, neointima formation, subintimal disruption, and vasa vasorum ingrowth resulting in subsequent vessel fibrosis. Several major molecular pathways are known to be involved in this programmed proliferation during permanent closure of the DA, e.g. NO signaling, the system of VEGF-promoted angiogenesis, and the cyclooxygenase/prostaglandin system (2-5).However, many questions concerning the exact function and cross-talk of these signaling pathways and the molecular basis for failed DA closure still remain unanswered (6). Previous studies investigating molecular events in the duct were in part hampered by the lack of an in vitro model that allows to study the processes in question separately in each of the different cell types of the DA instead of using the whole vessel (7). Herein, we describe an improved and reproducible method to culture organotypic vascular smooth muscle cells (VSMCs), fibroblasts, and e...

show abstract

Section: Resultsmentioning

confidence: 97%

Isolation and Culture of Fibroblasts, Vascular Smooth Muscle, and Endothelial Cells From the Fetal Rat Ductus Arteriosus

et al. 2011

View full text Add to dashboard Cite

show abstract

Fluorescence Labeling of the Capillary Network in Rat Brains

Theilen

Kuschinsky

1992

J Cereb Blood Flow Metab

View full text Add to dashboard Cite

Summary:To assess the reliability of fluorescence meth ods for a quantitative staining of brain capillaries, three different immunohistochemical fluorescent markers were used in the rat brain. Staining of the basement membrane by antibodies directed against fibronectin was compared, in the same brain section, with simultaneous staining of the vascular endothelium constituents nonmuscle myosin or von Willebrand factor (factor VIII). These stainings all resulted in identical patterns, which demonstrates their suitability for capillary staining in the brain. It has beenIn a previous study (Gobel et aI., 1990), we have postulated a continuous perfusion of all capillaries in the brains of conscious rats. The fluorescent mi croscopical method as used in this study has been criticized since higher capillary counts were mea sured than have been reported by groups that used light microscopical histochemical methods (Weiss, 1989; Kikano et aI., 1989; Anwar et aI., 1990). The discrepancy was ascribed to the fixation procedure employed in our experiments, which "causes changes in the FITC label which may cause clump ing or movement to noncapillary areas" (Weiss, 1988). It was the aim of the present study to validate the use of fluorescent markers of brain capillaries and to exclude the possibility of artifacts caused by the staining procedure. MATERIALS AND METHODSThe experiments were performed on male Sprague Dawley rats weighing 270-350 g.Received April 25, 1991; revised June 27, 1991; accepted July I, 1991.Address correspondence and reprint requests to Prof. Dr. w. Kuschinsky at Physiologisches Institut der Universitat Heidel berg, 1m Neuenheimer Feld 326, W-6900 Heidelberg, F.R.G.Abbreviations used: DTAF , 4.5-[(4,6-dichlorothriazin-2-yl)aminolfluorescein; PBS, phosphate-buffered saline; TRITe, tetramethylrhodamine B isothiocyanate. 347claimed that fixation of the tissue results in the appear ance of spurious capillary spots. Such a fixation artifact could be excluded using nonmuscle myosin staining. These results validate the methods of quantitative fluo rescent microscopical staining of capillary morphology in the brain and therefore support our concept of a contin uous perfusion of all capillaries in the brains of conscious rats. Key Words: Brain capillaries-Capillary perfusion Fibronectin-Indirect immunofluorescence-Nonmuscle myosin-von Wille brand factor. Double staining of capillary morphologyAfter decapitation, the brains of nine rats were pro cessed as described previously (Gobel et aI., 1990, Method 3). Fixation in pure acetone was performed for 30 s. The sections were overlaid with 70 j.Ll of the first of four antibodies (AI or Bl), and then they were incubated in a humid chamber at room temperature for 30 min. There after, the sections were washed three times in phosphate buffered saline (PBS) for 5 min. The margins were wiped dry, and the sections were overlaid with the second an tibody (A2 or B2). This procedure was repeated twice (A3 or B3, A4 or B4). The following sequence (1-4) of the staining steps yi...

show abstract

Nanotransfection-based vasculogenic cell reprogramming drives functional recovery in a mouse model of ischemic stroke

et al. 2021

View full text Add to dashboard Cite

Ischemic stroke causes vascular and neuronal tissue deficiencies that could lead to substantial functional impairment and/or death. Although progenitor-based vasculogenic cell therapies have shown promise as a potential rescue strategy following ischemic stroke, current approaches face major hurdles. Here, we used fibroblasts nanotransfected with Etv2, Foxc2, and Fli1 (EFF) to drive reprogramming-based vasculogenesis, intracranially, as a potential therapy for ischemic stroke. Perfusion analyses suggest that intracranial delivery of EFF-nanotransfected fibroblasts led to a dose-dependent increase in perfusion 14 days after injection. MRI and behavioral tests revealed ~70% infarct resolution and up to ~90% motor recovery for mice treated with EFF-nanotransfected fibroblasts. Immunohistological analysis confirmed increases in vascularity and neuronal cellularity, as well as reduced glial scar formation in response to treatment with EFF-nanotransfected fibroblasts. Together, our results suggest that vasculogenic cell therapies based on nanotransfection-driven (i.e., nonviral) cellular reprogramming represent a promising strategy for the treatment of ischemic stroke.

show abstract

The relationship between von willebrand factor antigen and fibronectin in human plasma, endothelial cells and fibroblasts in culture

Cited by 4 publications

References 23 publications

Isolation and Culture of Fibroblasts, Vascular Smooth Muscle, and Endothelial Cells From the Fetal Rat Ductus Arteriosus

Isolation and Culture of Fibroblasts, Vascular Smooth Muscle, and Endothelial Cells From the Fetal Rat Ductus Arteriosus

Fluorescence Labeling of the Capillary Network in Rat Brains

Nanotransfection-based vasculogenic cell reprogramming drives functional recovery in a mouse model of ischemic stroke

Contact Info

Product

Resources

About