Human Vitronectin Research Articles

A Novel Recombinant Vitronectin Variant Supports the Expansion and Differentiation of Pluripotent Stem Cells in Defined Animal-Free Workflows.

An essential aspect of harnessing the potential of pluripotent stem cells (PSCs) and their derivatives for regenerative medicine is the development of animal-free and chemically defined conditions for ex vivo cultivation. PSCs, including embryonic and induced PSCs (iPSCs), are in the early stages of clinical trials for various indications, including degenerative diseases and traumatic injury. A key step in the workflows generating these cells for more widespread clinical use is their safe and robust ex vivo cultivation. This entails optimization of cell culture media and substrates that are safe and consistent while maintaining robust functionality. Here, we describe the design of a human vitronectin (hVTN) variant with improved manufacturability in a bacterial expression system along with improved function in comparison to wild-type VTN and other previously characterized polypeptide fragments. In conjunction with an animal component-free media formulation, our hVTN fragment provides animal-free conditions for the enhanced expansion of iPSCs. This hVTN variant also supports the reprogramming of PBMCs into iPSCs. Furthermore, we show that these iPSCs can be efficiently differentiated into the three major germ layers and cortical neurons, thereby closing the loop on a completely defined animal-free workflow for cell types relevant for regenerative medicine.

Interaction between Yersinia pestis Ail Outer Membrane Protein and the C-Terminal Domain of Human Vitronectin.

Yersinia pestis, the causative agent of plague, is capable of evading the human immune system response by recruiting the plasma circulating vitronectin proteins, which act as a shield and avoid its lysis. Vitronectin recruitment is mediated by its interaction with the bacterial transmembrane protein Ail, protruding from the Y. pestis outer membrane. By using all-atom long-scale molecular dynamic simulations of Ail embedded in a realistic model of the bacterial membrane, we have shown that vitronectin forms a stable complex, mediated by interactions between the disordered moieties of the two proteins. The main amino acids driving the complexation have also been evidenced, thus favoring the possible rational design of specific peptides which, by inhibiting vitronectin recruitment, could act as original antibacterial agents.

Glyceraldehyde 3-Phosphate Dehydrogenase on the Surface of Candida albicans and Nakaseomyces glabratus Cells-A Moonlighting Protein That Binds Human Vitronectin and Plasminogen and Can Adsorb to Pathogenic Fungal Cells via Major Adhesins Als3 and Epa6.

Candida albicans and other closely related pathogenic yeast-like fungi carry on their surface numerous loosely adsorbed "moonlighting proteins"-proteins that play evolutionarily conserved intracellular functions but also appear on the cell surface and exhibit additional functions, e.g., contributing to attachment to host tissues. In the current work, we characterized this "moonlighting" role for glyceraldehyde 3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12) of C. albicans and Nakaseomyces glabratus. GAPDH was directly visualized on the cell surface of both species and shown to play a significant part in the total capacity of fungal cells to bind two selected human host proteins-vitronectin and plasminogen. Using purified proteins, both host proteins were found to tightly interact with GAPDH, with dissociation constants in an order of 10-8 M, as determined by bio-layer interferometry and surface plasmon resonance measurements. It was also shown that exogenous GAPDH tightly adheres to the surface of candidal cells, suggesting that the cell surface location of this moonlighting protein may partly result from the readsorption of its soluble form, which may be present at an infection site (e.g., due to release from dying fungal cells). The major dedicated adhesins, covalently bound to the cell wall-agglutinin-like sequence protein 3 (Als3) and epithelial adhesin 6 (Epa6)-were suggested to serve as the docking platforms for GAPDH in C. albicans and N. glabratus, respectively.

Solid-state NMR MAS CryoProbe enables structural studies of human blood protein vitronectin bound to hydroxyapatite

The low sensitivity of nuclear magnetic resonance (NMR) is a major bottleneck for studying biomolecular structures of complex biomolecular assemblies. Cryogenically cooled probe technology overcomes the sensitivity limitations enabling NMR applications to challenging biomolecular systems. Here we describe solid-state NMR studies of the human blood protein vitronectin (Vn) bound to hydroxyapatite (HAP), the mineralized form of calcium phosphate, using a CryoProbe designed for magic angle spinning (MAS) experiments. Vn is a major blood protein that regulates many different physiological and pathological processes. The high sensitivity of the CryoProbe enabled us to acquire three-dimensional solid-state NMR spectra for sequential assignment and characterization of site-specific water-protein interactions that provide initial insights into the organization of the Vn-HAP complex. Vn associates with HAP in various pathological settings, including macular degeneration eyes and Alzheimer's disease brains. The ability to probe these assemblies at atomic detail paves the way for understanding their formation.

Xeno-free culture and proliferation of hPSCs on 2D biomaterials.

Human pluripotent stem cells (human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs)) have unlimited proliferative potential, whereas adult stem cells such as bone marrow-derived stem cells and adipose-derived stem cells have problems with aging. When hPSCs are intended to be cultured on feeder-free or xeno-free conditions without utilizing mouse embryonic fibroblasts or human fibroblasts, they cannot be cultured on conventional tissue culture polystyrene dishes, as adult stem cells can be cultured but should be cultivated on material surfaces grafted or coated with (a) natural or recombinant extracellular matrix (ECM) proteins, (b) ECM protein-derived peptides and specific synthetic polymer surfaces in xeno-free and/or chemically defined conditions. This review describes current developing cell culture biomaterials for the proliferation of hPSCs while maintaining the pluripotency and differentiation potential of the cells into 3 germ layers. Biomaterials for the cultivation of hPSCs without utilizing a feeder layer are essential to decrease the risk of xenogenic molecules, which contributes to the potential clinical usage of hPSCs. ECM proteins such as human recombinant vitronectin, laminin-511 and laminin-521 have been utilized instead of Matrigel for the feeder-free cultivation of hPSCs. The following biomaterials are also discussed for hPSC cultivation: (a) decellularized ECM, (b) peptide-grafted biomaterials derived from ECM proteins, (c) recombinant E-cadherin-coated surface, (d) polysaccharide-immobilized surface, (e) synthetic polymer surfaces with and without bioactive sites, (f) thermoresponsive polymer surfaces with and without bioactive sites, and (g) synthetic microfibrous scaffolds.

Selective Grafting of Protease-Resistant Adhesive Peptides on Titanium Surfaces.

In orthopedic, dental, and maxillofacial fields, joint prostheses, plates, and screws are widely used in the treatment of problems related to bone tissue. However, the use of these prosthetic systems is not free from complications: the fibrotic encapsulation of endosseous implants often prevents optimal integration of the prostheses with the surrounding bone. To overcome these issues, biomimetic titanium implants have been developed where synthetic peptides have been selectively grafted on titanium surfaces via Schiff base formation. We used the retro-inverted sequence (DHVPX) from [351-359] human Vitronectin and its dimer (D2HVP). Both protease-resistant peptides showed increased human osteoblast adhesion and proliferation, an augmented number of focal adhesions, and cellular spreading with respect to the control. D2HVP-grafted samples significantly enhance Secreted Phosphoprotein 1, Integrin Binding Sialoprotein, and Vitronectin gene expression vs. control. An estimation of peptide surface density was determined by Two-photon microscopy analysis on a silanized glass model surface labeled with a fluorescent analog.

A Vitronectin-Derived Peptide Restores Ovariectomy-Induced Bone Loss by Dual Regulation of Bone Remodeling.

Bone remodeling is tightly regulated through bone resorption and bone formation; imbalances in bone remodeling can cause various pathological conditions such as osteoporosis. Antiresorptive agents commonly used for treating osteoporosis do not substantially reverse osteoporotic bone loss. We evaluated the effects of the RVYFFKGKQYWE motif (residues 270-281; VnP-16) of human vitronectin on the osteogenic differentiation of human mesenchymal stem cells (hMSCs) and osteoclastogenesis of bone marrow-derived macrophages. The effects of VnP-16 were also assessed in a mouse model of estrogen deficiency-induced osteoporosis (ovariectomized female C57BL/6mice). To assay whether VnP-16 can reverse ovariectomy-induced bone loss, synthetic peptides or vehicle were subcutaneously injected into ovariectomized mice once a week for 4weeks (n = 10/group). To evaluate the bone restorative effects of VnP-16, in-vivo micro-computed tomography analysis and histological staining were performed. VnP-16 induced osteogenic differentiation of hMSCs and inhibited the RANKL-RANK-TRAF6 axis in the osteoclastogenesis signaling pathway. Furthermore, systemic administration of VnP-16 reversed ovariectomy-induced bone loss in the femoral neck, distal femur and lumbar spine by increasing osteoblast differentiation and promoting bone formation, and concomitantly decreasing osteoclastogenesis and inhibiting bone resorption. The bone restorative effect of VnP-16 was observed one week after subcutaneous administration, and although the timing of the effect differed according to bone location, it persisted for at least 3weeks. Our findings suggest that VnP-16 is a potential therapeutic agent for treating osteoporosis that mediates its effects through dual regulation of bone remodeling.

The relationship of hs-CRP, vitronectin and NT-proBNP serum levels with the extent and severity of cardiac complications in patients with organophosphate pesticide poisoning.

Acute organophosphate poisoning kills tens of thousands of people annually around the world. These substances are widely used as insecticides in homes, industry, and agricultural environments. Due to the ease of access, they can cause accidental or intentional risks of exposure through the skin or respiratory contact. This study aimed to evaluate the serum levels of hs-CRP, Vitronectin, and NT-proBNP and their relationship with the extent and severity of cardiac complications in patients with organophosphate pesticide poisoning. In this descriptive-comparative study, 160 patients were studied with acute organophosphate poisoning. Also, for better comparison, 40 healthy individuals participated in this study. Diagnosis of organophosphate poisoning was based on clinical findings of serum butyrylcholinesterase levels. The hs-CRP measurement was performed by an autoanalyzer (Abbott, model Alcyon 300, USA) with the ELISA hs-CRP kit (The apDia Company, Belgium). Vitronectin (VN) measurements were performed by ELISA method and Glory science human VN kit with Catalog No: 11668. NT-ProBNP serum levels were analyzed by ProBNP assay kit (Roche, Germany) by ECLIA method using Elecsys 2010 Analyzer. The most important variables studied in this study were the electrical activity and conduction system of the heart, PR distance, QTC interval, and T-wave changes. In this study, most of the patients were women and girls (60.78%). The highest percentage of organophosphate poisoning was in the age group of 15-24 years (37.25%). In most cases (78.43%), poisoning was intentional or suicidal. Evaluation of electrophysiological abnormalities of the heart showed that 89 patients (55.62%) had long QTC interval (>450 msec), 43 cases (26.87%) had possible long QTC (431-450 msec), and 28 cases (17.5%) had normal QTC (<430 msec). Only 9.37% of cases (n = 15) showed an increase in P-R distance, which is characteristic of the first-degree ventricular atrial block. Sinus bradycardia occurred in 57 cases (35.62%) and sinus tachycardia in 43 cases (26.87%); in 60 cases (37.5%), the pulse rate was normal. Smooth T-wave changes were observed in 9.8% of patients and reverse T-wave was observed in 17.6%. A long T-wave was not reported in any case. In only two cases (1.25%) was grade 1 ventricular atrial block and grade 2 and 3 blocks were not observed. In general, there was a significant difference in the hs-CRP, vitronectin, and NT-proBNP serum levels between the patient and control groups in all studied variables. These parameters were also related to the extent and severity of the disease.

Towards understanding the novel adhesin function of Candida albicans phosphoglycerate mutase at the pathogen cell surface: some structural analysis of the interactions with human host extracellular matrix proteins

Although many atypical proteinaceous cell wall components that belong to a group of multitasking, "moonlighting" proteins, have been repeatedly identified in numerous pathogenic microorganisms, their novel extracellular functions and secretion mechanisms remain largely unrecognized. In Candida albicans, one of the most common fungal pathogens in humans, phosphoglycerate mutase (Gpm1) - a cytoplasmic enzyme involved in the glycolysis pathway - has been shown to occur on the cell surface and has been identified as a potentially important virulence factor. In this study, we demonstrated tight binding of C. albicans Gpm1 to the candidal cell surface, thus suggesting that the readsorption of soluble Gpm1 from the external environment could be a likely mechanism leading to the presence of this moonlighting protein on the pathogen surface. Several putative Gpm1-binding receptors on the yeast surface were identified. The affinities of Gpm1 to human vitronectin (VTR) and fibronectin (FN) were characterized with surface plasmon resonance measurements, and the dissociation constants of the complexes formed were determined to be in the order of 10-8 M. The internal Gpm1 sequence motifs, directly interacting with VTR (aa 116-158) and FN (aa 138-175) were mapped using chemical crosslinking and mass spectrometry. Synthetic peptides with matching sequences significantly inhibited formation of the Gpm1-VTR and Gpm1-FN complexes. A molecular model of the Gpm1-VTR complex was developed. These results provide the first structural insights into the adhesin function of candidal surface-exposed Gpm1.

Thymosin‐β4, and Human Vitronectin peptides Grafted to Collagen Tune Adhesion or VEGF Gene Expression in Human Cell Lines**

AbstractIn order to improve collagen bioactivity for regenerative medicine approaches, thymosin‐β4 (Tβ4P) and Human Vitronectin (HVP) derived peptides are grafted to collagen by thiol‐ene Michael addition. Tβ4P and HVP are known to exert a pro‐angiogenic and a pro‐adhesive activity respectively and HVP is involved in osteogenesis promotion. The ability of these peptides to increase collagen cell adhesion and angiogenesis properties is assessed on human cell lines. In particular, HVP‐grafted collagen increased human osteoblast adhesion and cell proliferation: after 24 h, both adhesion and proliferation roughly showed a 4‐fold increase, if compared to pristine collagen. Tβ4P‐grafted collagen promotes Vascular Endothelial Growth Factor (VEGF) gene expression in human vascular cell lines by more than 7 times. These results suggest that HVP‐grafted collagen may be an interesting biomaterial for bone tissue regeneration, while Tβ4P‐grafted collagen is useful for angiogenesis promotion.

Improvement of human embryonic stem cell-derived retinal pigment epithelium cell adhesion, maturation, and function through coating with truncated recombinant human vitronectin.

To explore an xeno-free and defined coating substrate suitable for the culture of H9 human embryonic stem cell-derived retinal pigment epithelial (hES-RPE) cells in vitro, and compare the behaviors and functions of hES-RPE cells on two culture substrates, laminin521 (LN-521) and truncated recombinant human vitronectin (VTN-N). hES-RPE cells were used in the experiment. The abilities of LN-521 and VTN-N at different concentrations to adhere to hES-RPE cells were compared with a high-content imaging system. Quantitative real-time polymerase chain reaction was used to evaluate RPE-specific gene expression levels midway (day 10) and at the end (day 20) of the time course. Cell polarity was observed by immunofluorescent staining for apical and basal markers of the RPE. The phagocytic ability of hES-RPE cells was identified by flow cytometry and immunofluorescence. The cell adhesion assay showed that the ability of LN-521 to adhere to hES-RPE cells was dose-dependent. With increasing coating concentration, an increasing number of cells attached to the surface of LN-521-coated wells. In contrast, VTN-N presented a strong adhesive ability even at a low concentration. The optimal concentration of LN-521 and VTN-N required to coat and adhesion to hES-RPE cells were 2 and 0.25 µg/cm2, respectively. Furthermore, both LN-521 and VTN-N could facilitate adoption of the desired cobblestone cellular morphology with tight junction and showed polarity by the hES-RPE cells. However, hES-RPE cells cultivated in VTN-N had a greater phagocytic ability, and it took less time for these hES-RPE cells to mature. VTN-N is a more suitable coating substrate for cultivating hES-RPE cells.

Regulatory properties of vitronectin and its glycosylation in collagen fibril formation and collagen-degrading enzyme cathepsin K activity

Vitronectin (VN) is a glycoprotein found in extracellular matrix and blood. Collagen, a major extracellular matrix component in mammals, is degraded by cathepsin K (CatK), which is essential for bone resorption under acidic conditions. The relationship between VN and cathepsins has been unclear. We discovered that VN promoted collagen fibril formation and inhibited CatK activity, and observed its activation in vitro. VN accelerated collagen fibril formation at neutral pH. Collagen fibers formed with VN were in close contact with each other and appeared as scattered flat masses in scanning electron microscopy images. VN formed collagen fibers with high acid solubility and significantly inhibited CatK; the IC50 was 8.1–16.6 nM and competitive, almost the same as those of human and porcine VNs. VN inhibited the autoprocessing of inactive pro-CatK from active CatK. DeN-glycosylation of VN attenuated the inhibitory effects of CatK and its autoprocessing by VN, but had little effect on acid solubilization of collagen and VN degradation via CatK. CatK inhibition is an attractive treatment approach for osteoporosis and osteoarthritis. These findings suggest that glycosylated VN is a potential biological candidate for CatK inhibition and may help to understand the molecular mechanisms of tissue re-modeling.

Correlating the Structure and Activity of Y. pestis Ail in a Bacterial Cell Envelope

Understanding microbe-host interactions at the molecular level is a major goal of fundamental biology and therapeutic drug development. Structural biology strives to capture biomolecular structures in action, but the samples are often highly simplified versions of the complex native environment. Here, we present an Escherichia coli model system that allows us to probe the structure and function of Ail, the major surface protein of the deadly pathogen Yersinia pestis. We show that cell surface expression of Ail produces Y. pestis virulence phenotypes in E. coli, including resistance to human serum, cosedimentation of human vitronectin, and pellicle formation. Moreover, isolated bacterial cell envelopes, encompassing inner and outer membranes, yield high-resolution solid-state NMR spectra that reflect the structure of Ail and reveal Ail sites that are sensitive to the bacterial membrane environment and involved in the interactions with human serum components. The data capture the structure and function of Ail in a bacterial outer membrane and set the stage for probing its interactions with the complex milieu of immune response proteins present in human serum.

Structural Insights into the Interactions of Candidal Enolase with Human Vitronectin, Fibronectin and Plasminogen.

Significant amounts of enolase—a cytosolic enzyme involved in the glycolysis pathway—are exposed on the cell surface of Candida yeast. It has been hypothesized that this exposed enolase form contributes to infection-related phenomena such as fungal adhesion to human tissues, and the activation of fibrinolysis and extracellular matrix degradation. The aim of the present study was to characterize, in structural terms, the protein-protein interactions underlying these moonlighting functions of enolase. The tight binding of human vitronectin, fibronectin and plasminogen by purified C. albicans and C. tropicalis enolases was quantitatively analyzed by surface plasmon resonance measurements, and the dissociation constants of the formed complexes were determined to be in the 10−7–10−8 M range. In contrast, the binding of human proteins by the S. cerevisiae enzyme was much weaker. The chemical cross-linking method was used to map the sites on enolase molecules that come into direct contact with human proteins. An internal motif 235DKAGYKGKVGIAMDVASSEFYKDGK259 in C. albicans enolase was suggested to contribute to the binding of all three human proteins tested. Models for these interactions were developed and revealed the sites on the enolase molecule that bind human proteins, extensively overlap for these ligands, and are well-separated from the catalytic activity center.

Calcium and hydroxyapatite binding site of human vitronectin provides insights to abnormal deposit formation

The human blood protein vitronectin (Vn) is a major component of the abnormal deposits associated with age-related macular degeneration, Alzheimer's disease, and many other age-related disorders. Its accumulation with lipids and hydroxyapatite (HAP) has been demonstrated, but the precise mechanism for deposit formation remains unknown. Using a combination of solution and solid-state NMR experiments, cosedimentation assays, differential scanning fluorimetry (DSF), and binding energy calculations, we demonstrate that Vn is capable of binding both soluble ionic calcium and crystalline HAP, with high affinity and chemical specificity. Calcium ions bind preferentially at an external site, at the top of the hemopexin-like (HX) domain, with a group of four Asp carboxylate groups. The same external site is also implicated in HAP binding. Moreover, Vn acquires thermal stability upon association with either calcium ions or crystalline HAP. The data point to a mechanism whereby Vn plays an active role in orchestrating calcified deposit formation. They provide a platform for understanding the pathogenesis of macular degeneration and other related degenerative disorders, and the normal functions of Vn, especially those related to bone resorption.

An atmospheric pressure plasma jet to tune the bioactive peptide coupling to polycaprolactone electrospun layers

The surface chemistry of scaffolds for tissue regeneration can guide cells growth. In this study, we present a novel method to functionalize electrospun Polycaprolactone (PCL) scaffolds allowing the tuning of biomolecule superficial concentration by varying only one process parameter. The method is based on the deposition of NH2 functional groups starting form (3-Aminopropyl) triethoxysilane (APTES) as precursor by a novel Atmospheric Pressure Plasma Jet (APPJ) and by a successive selective covalent linking of these amines with a synthetic Human Vitronectin adhesive cue (HVP). The addition, in the peptide C-terminus, of an aldehyde group ensures the selective ligation by alkylimino-de-oxo-bisubstitution between the primary amine and HVP. By this method, we managed to alter the HVP surface concentration just varying the deposition time of the plasma process; this resulted in different surface coverage of the plasma coating, which in turn led to diverse amount of linked HVP. Coating stability, morphology and coverage was assessed by infrared and photo-electron spectroscopies and by electron microscopy. As a function of the coverage a variation on peptide concentration was revealed by Total Nitrogen method and confirmed by biological assays, which demonstrated an increase of human osteoblasts viability as a function of peptide concentration.

Characterization of an Extensive Interface on Vitronectin for Binding to Plasminogen Activator Inhibitor-1: Adoption of Structure in an Intrinsically Disordered Region.

Small-angle neutron scattering (SANS) measurements were pursued to study human vitronectin, a protein found in tissues and the circulation that regulates cell adhesion/migration and proteolytic cascades that govern hemostasis and pericellular proteolysis. Many of these functions occur via interactions with its binding partner, plasminogen activator inhibitor-1 (PAI-1), the chief inhibitor of proteases that lyse and activate plasminogen. We focused on a region of vitronectin that remains uncharacterized from previous X-ray scattering, nuclear magnetic resonance, and computational modeling approaches and which we propose is involved in binding to PAI-1. This region, which bridges the N-terminal somatomedin B (SMB) domain with a large central β-propeller domain of vitronectin, appears unstructured and has characteristics of an intrinsically disordered domain (IDD). The effect of osmolytes was evaluated using circular dichroism and SANS to explore the potential of the IDD to undergo a disorder-to-order transition. The results suggest that the IDD favors a more ordered structure under osmotic pressure; SANS shows a smaller radius of gyration (Rg) and a more compact fold of the IDD upon addition of osmolytes. To test whether PAI-1 binding is also coupled to folding within the IDD structure, a set of SANS experiments with contrast variation were performed on the complex of PAI-1 with a vitronectin fragment corresponding to the N-terminal 130 amino acids (denoted the SMB-IDD because it contains the SMB domain and IDD in linear sequence). Analysis of the SANS data using the Ensemble Optimization Method confirms that the SMB-IDD adopts a more compact configuration when bound to PAI-1. Calculated structures for the PAI-1:SMB-IDD complex suggest that the IDD provides an interaction surface outside of the primary PAI-1-binding site located within the SMB domain; this binding is proposed to lead to the assembly of higher-order structures of vitronectin and PAI-1 commonly found in tissues.

Vitronectins produced by human cirrhotic liver and CCl4-treated rats differ in their glycosylation pattern and tissue remodeling activity.

Liver cirrhosis (LC) is a disease characterized by pathological accumulation and alteration of extracellular matrix (ECM) proteins; the interaction between two such proteins, collagen and vitronectin (VN), is considered to be the key to controlling ECM remodeling in liver cirrhosis. If it is possible to control the modification of oligosaccharides on VN, it may be possible to retard progression of liver cirrhosis. In this study, we examined the relationship between changes in VN glycosylation and activity related to the remodeling of hepatic tissue in human LC and a rat model of LC generated using carbon tetrachloride (CCl4). Plasma concentrations of VN in human LC declined to approximately two‐thirds that in normal plasma, but the ratio of active VN, which has collagen‐binding activities, increased 2.8 times in LC plasma. In contrast, purified LC‐VN exhibited similar binding activities toward type I, IV, and V collagens to those of normal VN. Lectin reactivities and carbohydrate analyses of LC‐VN revealed that branching, fucosylation, and sialylation of N‐glycans were higher than those of normal VN. On the other hand, the plasma level of rat CCl4‐VN increased and the ratio of active molecules to collagen in plasma decreased. Increased fucosylation of LC‐VN was not detected in carbohydrates of CCl4‐VN. The changes in rat VN due to CCl4 treatment did not correspond to the changes in plasma levels of human VN caused by LC, the ratio of active molecules, or carbohydrate composition, thereby indicating that CCl4‐treated rats are not an appropriate model for studying VNs in human LC. Glycosidase treatment of VNs supported the hypothesis that the collagen‐binding activity of VN is modulated by alterations of glycosylation during LC, which may contribute to (a) the matrix incorporation of VN and (b) tissue fibrosis.

Structure and Function of Human Vitronectin, a Key Mediator of Host-Pathogen Interactions

Structure and Function of Human Vitronectin, a Key Mediator of Host-Pathogen Interactions

Evaluation of reagents used to coat the hollow-fiber bioreactor membrane of the Quantum® Cell Expansion System for the culture of human mesenchymal stem cells

The addition of a coating reagent to promote cell adherence is necessary to prepare the membrane surface of the Quantum® Cell Expansion System hollow-fiber bioreactor for the culture of mesenchymal stem cells. In this study, the efficacy of 8 potential coating reagents has been compared in terms of the doubling times of their cell populations, cell morphology, characterization via flow cytometry, and capacity for trilineage differentiation. Human fibronectin (FN), pooled human cryoprecipitate (CPPT), and recombinant human vitronectin (VN) were successful as coating reagents, and each product has advantages in different cell culture contexts. Mesenchymal stem cells harvested from Quantum cultured with each of these 3 compounds as coating reagents all met International Society for Cellular Therapy standards for plastic adherence, surface marker expression, and successful trilineage differentiation. No significant differences were observed among the doubling times from Quantum harvests using FN, CPPT, or VN as coating reagents (P = 0.31). Coating with gelatin, human serum albumin, collagen I, poly‑l‑lysine, and poly‑d‑lysine resulted in significantly lower harvest yield; these agents are not recommended for use as coating reagents in the Quantum system.