scholarly journals Identification and isolation of human prostate epithelial stem cells based on α2β1-integrin expression

2001 ◽  
Vol 114 (21) ◽  
pp. 3865-3872 ◽  
Author(s):  
Anne T. Collins ◽  
Fouad K. Habib ◽  
Norman J. Maitland ◽  
David E. Neal
Keyword(s):  
Stem Cells ◽  
Type I Collagen ◽  
Specific Antigen ◽  
Basal Layer ◽  
Basement Membranes ◽  
Type I ◽  
Integrin Subunit ◽  
Basal Cells ◽  
Epithelial Stem Cells

A major impediment to our understanding of the biology of stem cells is the inability to distinguish them from their differentiating progeny. We made use of the known association of stem cells with basement membranes to isolate prostate epithelial stem cells. We show that, in vivo, putative stem cells express higher levels of the α2-integrin subunit than other cells within the basal layer. Approximately 1% of basal cells examined by confocal microscopy were integrin ‘bright’, and these cells can be selected directly from the tissue on the basis of rapid adhesion to type I collagen. This selected population has a basal phenotype, as determined by expression of CK5 and CK14 and lack of expression of the differentiation-specific markers prostate specific antigen (PSA) and prostatic acid phosphatase (PAP), and has a fourfold greater ability to form colonies in vitro than the total basal population. These putative stem cells are distinguished from other basal cells by their ability to generate prostate-like glands in vivo with morphologic and immuno-histochemical evidence of prostate-specific differentiation. These properties are consistent with a stem cell origin. Furthermore, the presence of surface integrins on prostate stem cells suggests that these cells share common pathways with stem cells in other tissues.

APPLICATION OF DENDRIMER/PLASMID hBMP-2 COMPLEXES LOADED INTO β-TCP/COLLAGEN SCAFFOLD IN THE TREATMENT OF FEMORAL DEFECTS IN RATS

2014 ◽  
Vol 26 (01) ◽  
pp. 1450005 ◽  
Author(s):  
Tingwei Bao ◽  
Huiming Wang ◽  
Wentao Zhang ◽  
Xuefeng Xia ◽  
Jiabei Zhou ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Type I Collagen ◽  
Cell Attachment ◽  
Collagen Scaffold ◽  
Bone Defects ◽  
Bone Morphogenetic Protein 2 ◽  
Porous Scaffolds ◽  
Type I

Purpose: Plasmid loading into scaffolds to enhance sustained release of growth factors is an important focus of regenerative medicine. The aim of this study was to build gene-activated matrices (GAMs) and examine the bone augmentation properties. Methods: Generation 5 polyamidoamine dendrimers (G5 dPAMAM)/plasmid recombinant human bone morphogenetic protein-2 (rhBMP-2) complexes were immobilized into beta-tricalcium phosphate (β-TCP)/type I collagen porous scaffolds. After cultured with rat mesenchymal stem cells (rMSCs), transfection efficiencies were examined. The secretion of rhBMP-2 and alkaline phosphatase (ALP) were detected to evaluate the osteogenic properties. Scanning electron microscopy (SEM) was used to observe attachment and proliferation. Moreover, we applied these GAMs directly into freshly created segmental bone defects in rat femurs, and their osteogenic efficiencies were evaluated. Results: Released plasmid complexes were transfected into stem cells and were expressed, which caused osteogenic differentiations of rat mesenchymal stem cells (rMSCs). SEM analysis showed excellent cell attachment. Bioactivity of plasmid rhBMP-2 was maintained in vivo, and the X-ray observation, histological analysis and immunohistochemistry (IHC) of bone tissue demonstrated that the bone healing in segmental femoral defects was enhanced by implantation of GAMs. Conclusions: Such biomaterials offer therapeutic opportunities in critical-sized bone defects.

Activation of human keratinocyte migration on type I collagen and fibronectin

1990 ◽  
Vol 96 (2) ◽  
pp. 197-205
Author(s):  
M. Guo ◽  
K. Toda ◽  
F. Grinnell
Keyword(s):  
Type I Collagen ◽  
Cultured Cells ◽  
Focal Adhesions ◽  
Human Keratinocytes ◽  
Type I ◽  
Integrin Subunit ◽  
Basal Cells ◽  
Keratinocyte Migration ◽  
Beta 1 Integrin ◽  
Cells Cultured

The purpose of our studies was to learn more about the regulation of keratinocyte migration. Human keratinocytes freshly harvested from skin were relatively immotile cells, whereas keratinocytes harvested from cell culture migrated on type I collagen or fibronectin as measured in a phagokinesis assay. Development of migratory competence by keratinocytes varied depending on the culture substratum. Cells cultured on plastic were activated more quickly and to a greater extent than cells cultured on dermis. The effect of the culture substratum on migratory competence was reversible. That is, cells cultured on plastic showed reduced activity after subculture on dermis. Cells cultured on dermis showed increased activity after subculture on plastic. Freshly isolated as well as cultured keratinocytes contained beta 1 integrin subunits, but only cultured cells were able to organize the subunits into focal adhesions. These adhesion sites also contained vinculin. In epidermal explants, beta 1 integrin subunits were mostly in basal cells, often more prominent between lateral cell borders than at the epidermal-dermal interface. In keratinocytes that migrated out of skin explants, there appeared to be an increase in the intensity of beta 1 integrin subunit immunostaining, possibly because of the change in shape of migrating cells. Also, beta 1 integrin subunits were found around and beneath migrating keratinocytes. These results show that changes in the distribution of beta 1 integrin subunits accompany development of migratory competence.

scholarly journals Differentiation-related expression of a major 64K corneal keratin in vivo and in culture suggests limbal location of corneal epithelial stem cells.

1986 ◽  
Vol 103 (1) ◽  
pp. 49-62 ◽  
Author(s):  
A Schermer ◽  
S Galvin ◽  
T T Sun
Keyword(s):  
Stem Cells ◽  
Strong Support ◽  
Transitional Zone ◽  
Basal Cells ◽  
Epithelial Stem Cells ◽  
Corneal Epithelial ◽  
Differentiated Cells ◽  
Limbal Epithelium ◽  
Cell Layers

In this paper we present keratin expression data that lend strong support to a model of corneal epithelial maturation in which the stem cells are located in the limbus, the transitional zone between cornea and conjunctiva. Using a new monoclonal antibody, AE5, which is highly specific for a 64,000-mol-wt corneal keratin, designated RK3, we demonstrate that this keratin is localized in all cell layers of rabbit corneal epithelium, but only in the suprabasal layers of the limbal epithelium. Analysis of cultured corneal keratinocytes showed that they express sequentially three major keratin pairs. Early cultures consisting of a monolayer of "basal" cells express mainly the 50/58K keratins, exponentially growing cells synthesize additional 48/56K keratins, and postconfluent, heavily stratified cultures begin to express the 55/64K corneal keratins. Cell separation experiments showed that basal cells isolated from postconfluent cultures contain predominantly the 50/58K pair, whereas suprabasal cells contain additional 55/64K and 48/56K pairs. Basal cells of the older, postconfluent cultures, however, can become AE5 positive, indicating that suprabasal location is not a prerequisite for the expression of the 64K keratin. Taken together, these results suggest that the acidic 55K and basic 64K keratins represent markers for an advanced stage of corneal epithelial differentiation. The fact that epithelial basal cells of central cornea but not those of the limbus possess the 64K keratin therefore indicates that corneal basal cells are in a more differentiated state than limbal basal cells. These findings, coupled with the known centripetal migration of corneal epithelial cells, strongly suggest that corneal epithelial stem cells are located in the limbus, and that corneal basal cells correspond to "transient amplifying cells" in the scheme of "stem cells----transient amplifying cells----terminally differentiated cells."

scholarly journals Estrogen-induced breast cancer is the result of disruption of asymmetric cell division of the stem cell

2010 ◽  
Vol 1 (2) ◽  
Author(s):  
Jose Russo ◽  
Kara Snider ◽  
Julia S. Pereira ◽  
Irma H. Russo
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Type I Collagen ◽  
Asymmetric Cell Division ◽  
Type I ◽  
In Vivo Models ◽  
Pregnancy And Lactation ◽  
In Vitro Systems

AbstractStem cells have the unique potential to divide asymmetrically to generate daughters with distinct fates, one which remains a stem cell and the other which turns into a cell committed to differentiation. By dividing asymmetrically, stem cells maintain the stem cell pool and simultaneously generate committed cells that reconstitute the organ, for example, to prepare the breast for a new pregnancy after involution from a previous pregnancy and lactation process. In addition to the in vivo models of mammary morphogenesis, there are in vitro systems that make the ductulogenic pattern of breast epithelia growth more amenable to study in critically determined conditions. The human breast epithelial cells MCF-10F formed tubules when grown in type I collagen and we demonstrated that treatment of these cells with 17β-estradiol (E

scholarly journals Extracellular matrix components of the mouse thymus microenvironment: ontogenetic studies and modulation by glucocorticoid hormones.

1991 ◽  
Vol 39 (11) ◽  
pp. 1539-1546 ◽  
Author(s):  
J Lannes-Vieira ◽  
M Dardenne ◽  
W Savino
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Type I Collagen ◽  
Basement Membranes ◽  
Thymic Epithelial Cell ◽  
Epithelial Cell Line ◽  
Type I ◽  
Extracellular Matrix Components

The present investigation was an ontogenetic study on the distribution of extracellular matrix (ECM) components in the thymic microenvironment of C57BL/6 mice (comprising young and old adults and developing embryos) and NZB mice. In addition, we evaluated the in vivo and in vitro influence of hydrocortisone treatment on basement membrane protein production by a thymic epithelial cell line. In young normal animals, Type I collagen was restricted to the interstitial spaces of the capsule and septa, where Type IV collagen, fibronectin, and laminin could be detected in the basement membranes. In addition, fibronectin-containing fibers were seen within the medulla of the thymic lobules. The ECM distribution pattern in the developing embryos was distinct from that observed in adults, since a fine meshwork of basement membrane-containing proteins was clearly seen throughout the parenchyma. Moreover, aging normal and NZB mice exhibited a denser ECM pattern than young adult normal animals. Treatment with hydrocortisone, both in vivo and in vitro, resulted in enhancement of ECM expression, detected in mice as early as 2 hr post injection and lasting for several days. Considering that the fluctuations of ECM expression parallel important events in thymocyte differentiation, we discuss the possibility that the two phenomena may be associated.

scholarly journals Effects of chitosan-collagen dressing on wound healing in vitro and in vivo assays

2021 ◽  
Vol 19 ◽  
pp. 228080002198969
Author(s):  
Min-Xia Zhang ◽  
Wan-Yi Zhao ◽  
Qing-Qing Fang ◽  
Xiao-Feng Wang ◽  
Chun-Ye Chen ◽  
...  
Keyword(s):  
Wound Healing ◽  
Wound Dressing ◽  
Type I Collagen ◽  
Inhibition Zone ◽  
Negative Control ◽  
Type I ◽  
Nih3t3 Cells ◽  
Post Operation

The present study was designed to fabricate a new chitosan-collagen sponge (CCS) for potential wound dressing applications. CCS was fabricated by a 3.0% chitosan mixture with a 1.0% type I collagen (7:3(w/w)) through freeze-drying. Then the dressing was prepared to evaluate its properties through a series of tests. The new-made dressing demonstrated its safety toward NIH3T3 cells. Furthermore, the CCS showed the significant surround inhibition zone than empty controls inoculated by E. coli and S. aureus. Moreover, the moisture rates of CCS were increased more rapidly than the collagen and blank sponge groups. The results revealed that the CCS had the characteristics of nontoxicity, biocompatibility, good antibacterial activity, and water retention. We used a full-thickness excisional wound healing model to evaluate the in vivo efficacy of the new dressing. The results showed remarkable healing at 14th day post-operation compared with injuries treated with collagen only as a negative control in addition to chitosan only. Our results suggest that the chitosan-collagen wound dressing were identified as a new promising candidate for further wound application.

Vitamin K promotes mineralization, osteoblast-to-osteocyte transition, and an anticatabolic phenotype by γ-carboxylation-dependent and -independent mechanisms

2009 ◽  
Vol 297 (6) ◽  
pp. C1358-C1367 ◽  
Author(s):  
Gerald J. Atkins ◽  
Katie J. Welldon ◽  
Asiri R. Wijenayaka ◽  
Lynda F. Bonewald ◽  
David M. Findlay
Keyword(s):  
Bone Formation ◽  
Vitamin K ◽  
Type I Collagen ◽  
Vitamin K2 ◽  
Type I ◽  
Vitamin K1 ◽  
Vitamin K3 ◽  
Positive Effect

The vitamin K family members phylloquinone (vitamin K1) and the menaquinones (vitamin K2) are under study for their roles in bone metabolism and as potential therapeutic agents for skeletal diseases. We have investigated the effects of two naturally occurring homologs, phytonadione (vitamin K1) and menatetrenone (vitamin K2), and those of the synthetic vitamin K, menadione (vitamin K3), on human primary osteoblasts. All homologs promoted in vitro mineralization by these cells. Vitamin K1-induced mineralization was highly sensitive to warfarin, whereas that induced by vitamins K2 and K3 was less sensitive, implying that γ-carboxylation and other mechanisms, possibly genomic actions through activation of the steroid xenobiotic receptor, are involved in the effect. The positive effect on mineralization was associated with decreased matrix synthesis, evidenced by a decrease from control in expression of type I collagen mRNA, implying a maturational effect. Incubation in the presence of vitamin K2 or K3 in a three-dimensional type I collagen gel culture system resulted in increased numbers of cells with elongated cytoplasmic processes resembling osteocytes. This effect was not warfarin sensitive. Addition of calcein to vitamin K-treated cells revealed vitamin K-dependent deposition of mineral associated with cell processes. These effects are consistent with vitamin K promoting the osteoblast-to-osteocyte transition in humans. To test whether vitamin K may also act on mature osteocytes, we tested the effects of vitamin K on MLO-Y4 cells. Vitamin K reduced receptor activator of NF-κB ligand expression relative to osteoprotegerin by MLO-Y4 cells, an effect also seen in human cultures. Together, our findings suggest that vitamin K promotes the osteoblast-to-osteocyte transition, at the same time decreasing the osteoclastogenic potential of these cells. These may be mechanisms by which vitamin K optimizes bone formation and integrity in vivo and may help explain the net positive effect of vitamin K on bone formation.

Cells in regenerating deer antler cartilage provide a microenvironment that supports osteoclast differentiation

2001 ◽  
Vol 204 (3) ◽  
pp. 443-455
Author(s):  
C. Faucheux ◽  
S. Nesbitt ◽  
M. Horton ◽  
J. Price
Keyword(s):  
Type I Collagen ◽  
Mononuclear Cells ◽  
Osteoclast Differentiation ◽  
Cathepsin K ◽  
Collagen Matrix ◽  
Tartrate Resistant Acid Phosphatase ◽  
Type I ◽  
Deer Antler

Deer antlers are a rare example of mammalian epimorphic regeneration. Each year, the antlers re-grow by a modified endochondral ossification process that involves extensive remodelling of cartilage by osteoclasts. This study identified regenerating antler cartilage as a site of osteoclastogenesis in vivo. An in vitro model was then developed to study antler osteoclast differentiation. Cultured as a high-density micromass, cells from non-mineralised cartilage supported the differentiation of large numbers of osteoclast-like multinucleated cells (MNCs) in the absence of factors normally required for osteoclastogenesis. After 48 h of culture, tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells (osteoclast precursors) were visible, and by day 14 a large number of TRAP-positive MNCs had formed (783+/−200 per well, mean +/− s.e.m., N=4). Reverse transcriptase/polymerase chain reaction (RT-PCR) showed that receptor activator of NF κ B ligand (RANKL) and macrophage colony stimulating factor (M-CSF) mRNAs were expressed in micromass cultures. Antler MNCs have the phenotype of osteoclasts from mammalian bone; they expressed TRAP, vitronectin and calcitonin receptors and, when cultured on dentine, formed F-actin rings and large resorption pits. When cultured on glass, antler MNCs appeared to digest the matrix of the micromass and endocytose type I collagen. Matrix metalloproteinase-9 (MMP-9) may play a role in the resorption of this non-mineralised matrix since it is highly expressed in 100 % of MNCs. In contrast, cathepsin K, another enzyme expressed in osteoclasts from bone, is only highly expressed in resorbing MNCs cultured on dentine. This study identifies the deer antler as a valuable model that can be used to study the differentiation and function of osteoclasts in adult regenerating mineralised tissues.

Tissue specificity of type I collagen gene expression is determined at both transcriptional and post-transcriptional levels

1984 ◽  
Vol 4 (9) ◽  
pp. 1843-1852
Author(s):  
R J Focht ◽  
S L Adams
Keyword(s):  
Gene Expression ◽  
Rna Structure ◽  
Type I Collagen ◽  
Translational Efficiency ◽  
Type I ◽  
Collagen Gene ◽  
Differentiated Cells ◽  
Collagen Gene Expression

We analyzed the control of type I collagen synthesis in four kinds of differentiated cells from chicken embryos which synthesize very different amounts of the protein. Tendon, skin, and smooth muscle cells were found to have identical amounts of type I collagen RNAs; however, the RNAs had inherently different translatabilities, which were observed both in vivo and in vitro. Chondrocytes also had substantial amounts of type I collagen RNAs, even though they directed no detectable synthesis of the protein either in vivo or in vitro. Type I collagen RNAs in chondrocytes display altered electrophoretic mobilities, suggesting that in these cells the reduction in translational efficiency may be mediated in part by changes in the RNA structure. These data indicate that control of type I collagen gene expression is a complex process which is exerted at both transcriptional and post-transcriptional levels.

The influence of the haematocrit on primary haemostasis in vitro

2005 ◽  
Vol 94 (12) ◽  
pp. 1213-1218 ◽  
Author(s):  
Marco Eugster ◽  
Walter H. Reinhart
Keyword(s):  
Platelet Function ◽  
Type I Collagen ◽  
Inverse Correlation ◽  
Type I ◽  
Membrane Pore ◽  
Closure Time ◽  
Function Analyzer ◽  
Platelet Function Analyzer

SummaryPrimary haemostasis consists of platelet adhesion to subendothelial collagen, their activation and aggregation and finally the formation of a platelet plug. Erythrocytes are involved in this process because they flow in the center of the vessel and push platelets towards the site of action on the vessel wall and enhance shear forces, which activate platelets. In the platelet function analyzer PFA-100® (Dade Behring, Düdingen, Switzerland), the in vivo situation is simulated in vitro with blood being aspirated at high shear rates (5000s-1) through a capillary into a membrane pore with a diameter of 150 μm coated with type I collagen and either epinephrine or adenosine diphosphate. Aggregating platelets plug the pore and stop the flow, which is measured as the closure time. We analysed the influence of erythrocytes on platelet function analyzer measurements by systematic variation of the haematocrit (20,30,40,and 50%) at constant platelet counts of 289±61 ×103/μl plasma, or 152±30 ×103/μl blood, 96±9 ×103/μl blood and 54±5 ×103/μl blood, respectively. An inverse correlation was found between haematocrit and closure time under all circumstances. A decrease of the platelet count by 50 ×103 /μl could be compensated for by a 10% increase in haematocrit. The haematocrit must, therefore, be taken into consideration for the correct interpretation of PFA-100® measurements. Our data also provide a pathophysiological rationale to reduce the risk of bleeding in patients with thrombocytopenia and anaemia by normalizing the haematocrit with erythrocyte transfusions.

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