scholarly journals Blood platelet formation in vitro. The role of the cytoskeleton in megakaryocyte fragmentation

1990 ◽  
Vol 97 (1) ◽  
pp. 59-70 ◽  
Author(s):  
F. Tablin ◽  
M. Castro ◽  
R.M. Leven
Keyword(s):  
Membrane Vesicles ◽  
Peripheral Zone ◽  
Membrane System ◽  
Blood Platelet ◽  
Collagen Gels ◽  
Process Formation ◽  
Vitro Model ◽  
Platelet Formation

We have developed a unique in vitro model that promotes differentiation of megakaryocytes into platelets. When megakaryocytes isolated from guinea pig bone marrow were cultured on hydrated rat tail collagen gels, cells spontaneously formed elongated, beaded processes that fragmented to yield cytoplasmic pieces with the same size and internal composition as individual platelets. Addition of nocodazole at the initiation of cultures blocked process formation, while addition of nocodazole to cells with previously established processes resulted in their retraction. The addition of taxol to cultures resulted in abnormally thick processes that were tightly adherent to the underlying substratum, and did not bead or fragment. Cytochalasin D accelerated process formation and fragmentation of megakaryocytes cultured on collagen gels by twofold. On the basis of these results, we propose a model for platelet formation in culture that involves the following steps: adherence of megakaryocytes to the underlying extracellular matrix; dilation of the demarcation membrane system and breakdown of the actin-rich peripheral zone; microtubule-based extension of pseudopodia, which are no longer adherent to the substratum; and fragmentation into platelets by the coalescence and fusion of demarcation membrane vesicles with the plasma membrane. We feel that this distinctive culture system closely approximates thrombocytopoiesis in vivo, thus allowing detailed elucidation of this important process.

scholarly journals In vitro rapid organization of endothelial cells into capillary-like networks is promoted by collagen matrices.

1983 ◽  
Vol 97 (5) ◽  
pp. 1648-1652 ◽  
Author(s):  
R Montesano ◽  
L Orci ◽  
P Vassalli
Keyword(s):  
Endothelial Cells ◽  
Three Dimensional ◽  
Collagen Matrix ◽  
Collagen Gels ◽  
Collagen Matrices ◽  
Narrow Lumen ◽  
Capillary Endothelial Cells ◽  
Vitro Model

We have studied the behavior of cloned capillary endothelial cells grown inside a three dimensional collagen matrix. Cell monolayers established on the surface of collagen gels were covered with a second layer of collagen. This induced the monolayers of endothelial cells to reorganize into a network of branching and anastomosing capillary-like tubes. As seen by electron microscopy, the tubes were formed by at least two cells (in transverse sections) delimiting a narrow lumen. In addition, distinct basal lamina material was present between the abluminal face of the endothelial cells and the collagen matrix. These results showed that capillary endothelial cells have the capacity to form vessel-like structures with well-oriented cell polarity in vitro. They also suggest that an appropriate topological relationship of endothelial cells with collagen matrices, similar to that occurring in vivo, has an inducive role on the expression of this potential. This culture system provides a simple in vitro model for studying the factors involved in the formation of new blood vessels (angiogenesis).

scholarly journals The effects of dietary ligands on zinc uptake at the porcine intestinal brush-border membrane

1990 ◽  
Vol 64 (3) ◽  
pp. 733-741 ◽  
Author(s):  
A. J. Turnbull ◽  
P. Blakeborough ◽  
R. P. H. Thompson
Keyword(s):  
Brush Border ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Intestinal Brush Border Membrane ◽  
Intestinal Brush Border ◽  
Zn Uptake ◽  
Zn Concentration ◽  
Vitro Model

Intestinal brush-border-membrane vesicles were prepared from the porcine small bowel by magnesium precipitation and differential centrifugation, and were functionally intact. The influence of dietary ligands on 65Zn uptake was determined using a 65Zn concentration of 5 μm, an incubation time of 1 min and a reaction temperature of 27°, with a rapid filtration technique. At this low Zn concentration the addition of an excess of folate, histidine or glucose had no effect on Zn uptake. Addition of picolinate, citrate and phytate to the incubation medium significantly reduced Zn uptake at all concentrations of ligand examined. Any inhibitory effects of folic acid in vivo may thuss be due to a mucosal rather than lumen interaction. Those ligands inhibiting absorption may have done so through the formation of Zn-ligand complexes, which are either insoluble, or which reduce the binding of Zn to its mucosal receptor. This in vitro model of Zn absorption is useful for comparing the effects of potential Zn-binding ligands in the diet.

scholarly journals Homologous tumor cell membrane vesicles active preferential self-recognition of tumor cells in vitro

2022 ◽  
Vol 65 (1) ◽  
Author(s):  
Chenghu Wu ◽  
Ailin Yu ◽  
Yue Chen ◽  
Mingbo Fan
Keyword(s):  
Cell Membrane ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Tumor Therapy ◽  
Membrane Vesicles ◽  
Membrane System ◽  
Tumor Cell Membrane ◽  
Self Recognition

AbstractCell membrane vesicles, as delivery carriers of drugs or biological agents in vivo, are an important therapeutic mode in the study of disease treatment. Tumor membrane-derived vesicles have been widely used in tumor therapy because of their good tumor enrichment effect. The most common method is the surface of nanoparticles coated with tumor cell membrane, which can effectively prolong the circulation time of particles in the blood and the enrichment of tumors. In this study, we prepared vesicles of different tumor cell membrane derivate and studied their targeting to tumors detailly. The results showed that homologous vesicles have high targeting to homologous tumor cells. The fluorescence of vesicles in homologous tumor cells was significantly higher than that in other tumor cells. This study will provide a new strategy and guidance for the clinical treatment of cancer based on the tumor cell membrane system. Graphical Abstract

scholarly journals Blood platelet formation at a glance

2020 ◽  
Vol 133 (20) ◽  
pp. jcs244731
Author(s):  
Julie Boscher ◽  
Ines Guinard ◽  
Anita Eckly ◽  
François Lanza ◽  
Catherine Léon
Keyword(s):  
Bone Marrow ◽  
Blood Platelets ◽  
Blood Platelet ◽  
Main Function ◽  
Formation Processes ◽  
Bona Fide ◽  
Platelet Formation

ABSTRACTThe main function of blood platelets is to ensure hemostasis and prevent hemorrhages. The 1011 platelets needed daily are produced in a well-orchestrated process. However, this process is not yet fully understood and in vitro platelet production is still inefficient. Platelets are produced in the bone marrow by megakaryocytes, highly specialized precursor cells that extend cytoplasmic projections called proplatelets (PPTs) through the endothelial barrier of sinusoid vessels. In this Cell Science at a Glance article and the accompanying poster we discuss the mechanisms and pathways involved in megakaryopoiesis and platelet formation processes. We especially address the – still underestimated – role of the microenvironment of the bone marrow, and present recent findings on how PPT extension in vivo differs from that in vitro and entails different mechanisms. Finally, we recapitulate old but recently revisited evidence that – although bone marrow does produce megakaryocytes and PPTs – remodeling and the release of bona fide platelets, mainly occur in the downstream microcirculation.

scholarly journals Culture of isolated bovine megakaryocytes on reconstituted basement membrane matrix leads to proplatelet process formation

Blood ◽  
1990 ◽  
Vol 76 (5) ◽  
pp. 912-924 ◽  
Author(s):  
KS Topp ◽  
F Tablin ◽  
J Levin
Keyword(s):  
Basement Membrane ◽  
Bone Marrow Cells ◽  
Human Platelets ◽  
Velocity Sedimentation ◽  
Thin Filaments ◽  
Membrane Matrix ◽  
Process Formation ◽  
Vitro Model

Abstract We have enriched for bovine megakaryocytes and identified a culture system that may provide an in vitro model for platelet formation. Mature megakaryocytes with an unusually high ploidy distribution were obtained after differential centrifugation and velocity sedimentation of bone marrow cells through gradients of bovine serum albumin (BSA). The cell membranes of isolated megakaryocytes and megakaryocytes in vivo stained with antisera to human platelets and human platelet membrane GPIIIa. The microenvironment of bovine megakaryocytes in vivo was investigated using antibodies to types I and IV collagen and laminin. In an attempt to duplicate the microenvironment in vitro, bovine megakaryocytes were cultured on a reconstituted basement membrane matrix (Matrigel). The cells adhered to the gel, extended radial lamellipodia, and occasionally formed lengthy pseudopodia. Ultrastructural examination of these cells showed widening and coalescence of the megakaryocyte demarcation membranes (DMS), and inclusion of platelet granules, thin filaments, and microtubules in the processes. Very few DMS vesicles were present distally in the processes. The culture of megakaryocytes on a reconstituted basement membrane may closely model the in vivo megakaryocyte microenvironment and allow the study of thrombocytopoiesis in vitro.

scholarly journals Culture of isolated bovine megakaryocytes on reconstituted basement membrane matrix leads to proplatelet process formation

Blood ◽  
1990 ◽  
Vol 76 (5) ◽  
pp. 912-924 ◽  
Author(s):  
KS Topp ◽  
F Tablin ◽  
J Levin
Keyword(s):  
Basement Membrane ◽  
Bone Marrow Cells ◽  
Human Platelets ◽  
Velocity Sedimentation ◽  
Thin Filaments ◽  
Membrane Matrix ◽  
Process Formation ◽  
Vitro Model

We have enriched for bovine megakaryocytes and identified a culture system that may provide an in vitro model for platelet formation. Mature megakaryocytes with an unusually high ploidy distribution were obtained after differential centrifugation and velocity sedimentation of bone marrow cells through gradients of bovine serum albumin (BSA). The cell membranes of isolated megakaryocytes and megakaryocytes in vivo stained with antisera to human platelets and human platelet membrane GPIIIa. The microenvironment of bovine megakaryocytes in vivo was investigated using antibodies to types I and IV collagen and laminin. In an attempt to duplicate the microenvironment in vitro, bovine megakaryocytes were cultured on a reconstituted basement membrane matrix (Matrigel). The cells adhered to the gel, extended radial lamellipodia, and occasionally formed lengthy pseudopodia. Ultrastructural examination of these cells showed widening and coalescence of the megakaryocyte demarcation membranes (DMS), and inclusion of platelet granules, thin filaments, and microtubules in the processes. Very few DMS vesicles were present distally in the processes. The culture of megakaryocytes on a reconstituted basement membrane may closely model the in vivo megakaryocyte microenvironment and allow the study of thrombocytopoiesis in vitro.

The Effects of Brinolase (Fibrinolytic Enzyme from Aspergillus Oryzae) on Platelet Aggregation of Dog and Man

1972 ◽  
Vol 28 (01) ◽  
pp. 031-048 ◽  
Author(s):  
W. H. E Roschlau ◽  
R Gage
Keyword(s):  
Platelet Aggregation ◽  
Aspergillus Oryzae ◽  
Degradation Products ◽  
Blood Platelet ◽  
Human Platelets ◽  
Fibrinolytic Enzyme ◽  
Inhibitory Effects ◽  
Blood Platelet Aggregation

SummaryInhibition of blood platelet aggregation by brinolase (fibrinolytic enzyme from Aspergillus oryzae) has been demonstrated with human platelets in vitro and with dog platelets in vivo and in vitro, using both ADP and collagen as aggregating stimuli. It is suggested that the optimal inhibitory effects of brinolase occur indirectly through the generation of plasma fibrinogen degradation products, without compromising platelet viability, rather than by direct proteolysis of platelet structures.

scholarly journals Functional and Transcriptional Adaptations of Blood Monocytes Recruited to the Cystic Fibrosis Airway Microenvironment In Vitro

2021 ◽  
Vol 22 (5) ◽  
pp. 2530
Author(s):  
Bijean D. Ford ◽  
Diego Moncada Giraldo ◽  
Camilla Margaroli ◽  
Vincent D. Giacalone ◽  
Milton R. Brown ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Bactericidal Activity ◽  
Scavenger Receptor ◽  
Receptor Expression ◽  
Blood Monocytes ◽  
Bacterial Killing ◽  
Blood Neutrophils ◽  
Vitro Model

Cystic fibrosis (CF) lung disease is dominated by the recruitment of myeloid cells (neutrophils and monocytes) from the blood which fail to clear the lung of colonizing microbes. In prior in vitro studies, we showed that blood neutrophils migrated through the well-differentiated lung epithelium into the CF airway fluid supernatant (ASN) mimic the dysfunction of CF airway neutrophils in vivo, including decreased bactericidal activity despite an increased metabolism. Here, we hypothesized that, in a similar manner to neutrophils, blood monocytes undergo significant adaptations upon recruitment to CFASN. To test this hypothesis, primary human blood monocytes were transmigrated in our in vitro model into the ASN from healthy control (HC) or CF subjects to mimic in vivo recruitment to normal or CF airways, respectively. Surface phenotype, metabolic and bacterial killing activities, and transcriptomic profile by RNA sequencing were quantified post-transmigration. Unlike neutrophils, monocytes were not metabolically activated, nor did they show broad differences in activation and scavenger receptor expression upon recruitment to the CFASN compared to HCASN. However, monocytes recruited to CFASN showed decreased bactericidal activity. RNASeq analysis showed strong effects of transmigration on monocyte RNA profile, with differences between CFASN and HCASN conditions, notably in immune signaling, including lower expression in the former of the antimicrobial factor ISG15, defensin-like chemokine CXCL11, and nitric oxide-producing enzyme NOS3. While monocytes undergo qualitatively different adaptations from those seen in neutrophils upon recruitment to the CF airway microenvironment, their bactericidal activity is also dysregulated, which could explain why they also fail to protect CF airways from infection.

scholarly journals Radiofrequency Irradiation Modulates TRPV1-Related Burning Sensation in Rosacea

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1424
Author(s):  
Seyeon Oh ◽  
Myeongjoo Son ◽  
Joonhong Park ◽  
Donghwan Kang ◽  
Kyunghee Byun
Keyword(s):  
Burning Sensation ◽  
In Vivo Model ◽  
Molecular Evidence ◽  
Exposed Skin ◽  
Heat Treated ◽  
Vitro Model ◽  
Effective Use ◽  
Inflammatory Molecules

Rosacea is a skin inflammatory condition that is accompanied by not only redness and flushing but also unseen symptoms, such as burning, stinging, and itching. TRPV1 expression in UVB-exposed skin can lead to a painful burning sensation. Upregulated TRPV1 expression helps release neuropeptides, including calcitonin gene-related peptide, pituitary adenylate cyclase-activating polypeptide, and vasoactive intestinal peptide, which can activate macrophage and inflammatory molecules. In this study, we found that radiofrequency (RF) irradiation reduced TRPV1 activation and neuropeptide expression in a UVB-exposed in vivo model and UVB- or heat-treated in an in vitro model. RF irradiation attenuated neuropeptide-induced macrophage activation and inflammatory molecule expression. Interestingly, the burning sensation in the skin of UVB-exposed mice and patients with rosacea was significantly decreased by RF irradiation. These results can provide experimental and molecular evidence on the effective use of RF irradiation for the burning sensation in patients with rosacea.

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