Gradient nanofibrous chitosan/poly ɛ-caprolactone scaffolds as extracellular microenvironments for vascular tissue engineering

One of the major challenges of tissue-engineered small-diameter blood vessels is restenosis caused by thrombopoiesis. The goal of this study was to develop a 3D gradient heparinized nanofibrous scaffold, aiding endothelial cells lined on the lumen of blood vessel to prevent thrombosis. The vertical graded chitosan/poly ?-caprolactone (CS/PCL) nanofibrous vessel scaffolds were fabricated with chitosan and PCL by sequential quantity grading co-electrospinning. To mimic the natural blood vessel microenvironment, we used heparinization and immobilization of vascular endothelial growth factor (VEGF) in the gradient CS/PCL. The quantity of heparinized chitosan nanofibers increased gradually from the tunica adventitia to the lumen surfaces in the gradient CS/PCL wall of tissue engineered vessel. More heparin reacted to chitosan nanofiber in gradient CS/PCL than in uniform CS/PCL nanofibrous scaffolds. Antithrombogenic properties of the scaffolds were enhanced by the heparinization of these scaffolds, as shown by activated partial thromboplastin time and platelet adhesion assay. Compared to the uniform CS/PCL scaffold, the release of VEGF from the gradient CS/PCL was more stable and sustained, and the burst release of VEGF was reduced approximately 42.5% within the initial 12 h. The adhesion and proliferation of human umbilical vein endothelial cells (HUVEC) were enhanced on the gradient CS/PCL scaffold. Furthermore, HUVEC grew and formed an entire monolayer on the top side of the gradient CS/PCL scaffold. Therefore, use of vertical gradient heparinized CS/PCL nanofibrous scaffolds could provide an approach to create small-diameter blood vessel grafts with innate properties of mammalian vessels of anticoagulation and rapid induction of re-endothelialization.     

Endothelialization and patency of RGD-functionalized vascular grafts in a rabbit carotid artery model

To address the growing demand of small-diameter vascular grafts for cardiovascular disease, it is necessary to develop substitutes with bio-functionalities, such as anticoagulation, rapid endothelialization, and smooth muscle regeneration. In this study, the small-diameter tubular grafts (2.2 mm) were fabricated by electrospinning of biodegradable polymer polycaprolactone (PCL) followed by functional surface coating with an arginine-glycine-aspartic acid (RGD)-containing molecule. The healing characteristics of the grafts were evaluated by implanting them in rabbit carotid arteries for 2 and 4 weeks. Results showed that at both time points, all 10 of the RGD-modified PCL grafts (PCL-RGD) were patent, whereas 4 of the 10 non-modified PCL grafts were occluded due to thrombus formation. Scanning electron microscopy (SEM) data showed abundant platelets adhering on the surface of the midportion of the PCL grafts. In contrast, only few platelets were observed on the PCL-RGD surface, suggesting that RGD modification significantly improved the hemocompatibility of the PCL grafts. Histological analysis demonstrated enhanced cell infiltration and homogeneous distribution within the PCL-RGD grafts in comparison with the PCL grafts. Furthermore, immunofluorescence staining also showed a 3-fold increase of endothelial coverage of the PCL-RGD grafts than that of PCL grafts at those two time points. After 4-week implantation, 65.3 ± 7.6% of the surface area of the PCL-RGD grafts was covered by smooth muscle cell layer, which is almost 23% more than that on the PCL grafts. The present study indicates that RGD-modified PCL grafts exhibit an improved remodeling and integration capability in revascularization.     

Heparin-Conjugated PCL Scaffolds Fabricated by Electrospinning and Loaded with Fibroblast Growth Factor 2

A biodegradable poly(ε-caprolactone) (PCL) was synthesized by ring-opening polymerization of ε-caprolactone catalyzed by Sn(Oct)₂/BDO, followed by the heparin conjugation using EDC/NHS chemistry. The structure of the heparin–PCL conjugate was characterized by ¹H-NMR and GPC. The results of static contact angle and water uptake ratio measurements also confirmed the conjugation of heparin with the polyester. Its in vitro anticoagulation time was substantially extended, as evidenced by activated partial thromboplastin time (APTT) testing. Afterwards the conjugate was electrospun into small-diameter tubular scaffolds and loaded with Fibroblast Growth Factor 2 (FGF2) in aqueous solution. The loading efficiency was assayed by enzyme-linked immunosorbent assay (ELISA); the results indicated that the conjugate holds a higher loading efficiency than the blank polyester. The viability of released FGF2 was evaluated by MTT and cell adhesion tests. The amount and morphology of cells were significantly improved after FGF2 loading onto the electrospun heparin–PCL vascular scaffolds.     

Hollow fibers of poly(lactide-co-glycolide) and poly(ε-caprolactone) blends for vascular tissue engineering applications

At present the manufacture of small-diameter blood vessels is one of the main challenges in the field of vascular tissue engineering. Currently available vascular grafts rapidly fail due to development of intimal hyperplasia and thrombus formation. Poly(lactic-co-glycolic acid) (PLGA) hollow fiber (HF) membranes have previously been proposed for this application, but as we show in the present work, they have an inhibiting effect on cell proliferation and rather poor mechanical properties. To overcome this we prepared HF membranes via phase inversion using blends of PLGA with poly(ε-caprolactone) (PCL). The influence of polymer composition on the HF physicochemical properties (topography, water transport and mechanical properties) and cell attachment and proliferation were studied. Our results show that only the ratio PCL/PLGA of 85/15 (PCL/PLGA85/15) yielded a miscible blend after processing. A higher PLGA concentration in the blend led to immiscible PCL/PLGA phase-separated HFs with an inhomogeneous morphology and variation in the cell culture results. In fact, the PCL/PLGA85/15 blend, which had the most homogeneous morphology and suitable pore structure, showed better human adipose stem cell (hASC) attachment and proliferation compared with the homopolymers. This, combined with the good mechanical and transport properties, makes them potentially useful for the development of small-caliber vascular grafts.     

PEG对肝素化PCL/PEG人工血管膜材料中肝素释放的影响

目的 研究聚乙二醇(PEG)的加入对肝素化聚己内酯/聚乙二醇(PCL/PEG)人工血管膜材料中肝素体外释放的影响.方法 通过共混法和冷冻干燥技术制备不同PEG质量分数(0、5％、10％、15％)的肝素化PCL/PEG膜材料,并通过体外释放实验考察PEG的加入对肝素释放性能的影响.同时,通过X射线衍射、傅里叶红外光谱和差示扫描热分析仪探究PEG的加入对基体结构性能的影响.结果 PEG的加入降低了肝素从基体释放的难度,提高了肝素第1天的平均释放速率和34 d内的累积释放率,且两者在一定程度上随着PEG质量分数的增加而增大.X射线衍射、傅里叶红外光谱和差示扫描热分析结果均表明,肝素的加入会使PCL膜的结晶度在一定程度上有所增大,但整体影响并不显著,且肝素的加入会促进PEG晶粒的生长,肝素和PEG在基体中呈现共域化分布.结论 利用共混法和冷冻干燥技术制备了肝素化PCL/PEG膜材料,并可通过调控PEG的质量分数在一定程度上实现对肝素释放行为的控制,进而预测试样在一定程度上具有抗凝作用,该材料有望用作小口径人工血管膜材料.     

Electrospun tubular scaffolds: On the effectiveness of blending poly(ε-caprolactone) with poly(3-hydroxybutyrate-co-3-hydroxyvalerate)

Tissue engineering can effectively contribute to the development of novel vascular prostheses aimed to overcome the well-known drawbacks of small-diameter grafts. To date, poly(ε-caprolactone) (PCL), a bioresorbable synthetic poly(α-hydroxyester), is considered one of the most promising materials for vascular tissue engineering. In this work, the potential advantage of intimate blending soft PCL and hard poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), a polymer of microbial origin, has been evaluated. Nonwoven mats and small-diameter tubular scaffolds of PCL, PHBV, and PCL/PHBV were fabricated by means of electrospinning technique. Mechanical properties and suture retention strength were investigated according to the international standard for cardiovascular implants. Biological tests demonstrated that both PCL-based scaffolds supported survival and growth of rat cerebral endothelial cells in a short time. The fiber alignment of the electrospun tubular scaffolds contributed to a more rapid and homogeneous cell colonization of the luminal surface. ? 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2012.     

Bilayered vascular grafts based on silk proteins

A major block in the development of small diameter vascular grafts is achieving suitable blood vessel regeneration while minimizing the risk of thrombosis, intimal hyperplasia, suture retention, and mechanical failure. Silk-based tubular vessels for tissue engineering have been prepared by molding, dipping, electrospinning, or gel spinning, however, further studies are needed to improve the mechanical and blood compatibility properties. In the present study a bilayered vascular graft based on silk fibroin (SF) was developed. The graft was composed of an inner silk fiber-reinforced SF tube containing heparin and a highly porous SF external layer. Compared with previously fabricated SF tubes the fiber-reinforcement provided a comparable or higher mechanical strength, burst pressure, and suture retention strength, as well as mechanical compliance, to saphenous veins for vascular grafts. Heparin release was sustained for at least 1 month, affording blood compatibility to the grafts. The outer layer of the grafts prepared through lyophilization had a highly porous structure in which the macropore walls were composed of nanofibers similar to extracellular matrix, which offered an excellent environment for cell growth. In vitro studies showed good cytocompatibility and hemocompatibility.     

Controlled release of heparin from poly(epsilon-caprolactone) electrospun fibers

Sustained delivery of heparin to the localized adventitial surface of grafted blood vessels has been shown to prevent the vascular smooth muscle cell (VSMC) proliferation that can lead to graft occlusion and failure. In this study heparin was incorporated into electrospun poly(epsilon-caprolactone) (PCL) fiber mats for assessment as a controlled delivery device. Fibers with smooth surfaces and no bead defects could be spun from polymer solutions with 8%w/v PCL in 7:3 dichloromethane:methanol. A significant decrease in fiber diameter was observed with increasing heparin concentration. Assessment of drug loading, and imaging of fluorescently labeled heparin showed homogenous distribution of heparin throughout the fiber mats. A total of approximately half of the encapsulated heparin was released by diffusional control from the heparin/PCL fibers after 14 days. The fibers did not induce an inflammatory response in macrophage cells in vitro and the released heparin was effective in preventing the proliferation of VSMCs in culture. These results suggest that electrospun PCL fibers are a promising candidate for delivery of heparin to the site of vascular injury.     

Polysaccharide-based biomaterials with on-demand nitric oxide releasing property regulated by enzyme catalysis

The regulatory role of nitric oxide (NO) in cell signaling has been well recognized. Clinically, NO deficiency is known to be associated with severe vascular disorders, especially in patients with long-term diabetes. Exogenous compensation of NO is a promising therapeutic strategy, although the lack of stable NO compounds often lead to unsatisfactory clinical outcomes. In the present study, we report a stable comb-shaped polymer (CS–NO) using glycosylated NO compound as pendent chains and chitosan (CS) as backbone for controlled NO release. The on-demand release of NO is achieved by controlling the decomposition process of the CS–NO polymer, which is blocked by galactose and only occurs in the presence of glycosidase, making the NO releasing kinetic closely correlate with the glycosidase concentration. In addition, due to its high stability, the CS–NO polymers can also be processed into supportive membrane or injectable hydrogel, further demonstrating its clinical potential. Indeed, we report that the NO-releasing membrane inhibited platelet adhesion, prolonged activated partial thromboplastin time (APTT) as shown in the platelet-rich-plasma (PRP) assay. We also observe enhanced human umbilical vein endothelial cell growth yet suppressed vascular smooth muscle cell proliferation on the NO-contained membrane in vitro. Furthermore, in vivo administration of CS–NO solution significantly enhanced angiogenesis in diabetic mice with hind-limb ischemia. Protective effect of CS–NO was also observed against limb necrosis. Given the physiological importance of NO, the CS–NO polymer may be considered a promising option in therapeutic development against vascular disorders and diabetic feet.     

PCL-PU composite vascular scaffold production for vascular tissue engineering: attachment, proliferation and bioactivity of human vascular endothelial cells

A new compliant scaffold suitable for small-diameter vascular grafts has been developed that promotes strong attachment of endothelial cells. Composite scaffolds were produced by wet spinning polycaprolactone (PCL) fibres which form the luminal surface, then electrospinning porous polyurethane (PU) onto the back of the PCL fibres to form the vessel wall substitute. Human endothelial cells demonstrated strong attachment to the composite PCL-PU scaffold, and proliferated to form a monolayer with strong PECAM-1 expression and cobblestone morphology. Attached cells demonstrated abundant release of von Willebrand factor, nitric oxide and ICAM-1 under physiological stimuli, and exhibited an immune response to lipopolysaccharide. The composite scaffold may also deliver bioactive molecules. Active trypsin, used as a test molecule, had a defined 48 h pattern of release from luminal PCL fibres. These data confirm the potential of this novel composite scaffold in vascular tissue engineering.     

The effects of heparin releasing hydrogels on vascular smooth muscle cell phenotype

Poly(ethylene glycol) diacrylate (PEGDA) hydrogel scaffolds were engineered to promote contractile smooth muscle cell (SMC) phenotype via controlled release of heparin. The scaffold design was evaluated by quantifying the effects of free heparin on SMC phenotype, engineering hydrogels to provide controlled release of heparin, and synthesizing cell-adhesive, heparin releasing hydrogels to promote contractile SMC phenotype. Heparin inhibited SMC proliferation and up-regulated expression of contractile SMC phenotype markers, including smooth muscle α-actin, calponin, and SM-22α, in a dose-dependent fashion (6 μg/ml to 3.2 mg/ml). Heparin release from PEGDA hydrogels was controlled by altering PEGDA molecular weight (MW 1000–6000) and concentration at polymerization (10–30% w/w), yielding release profiles ranging from hours to weeks in duration. Heparin released from PEGDA gels, formulated for optimized heparin loading and release kinetics (30% w/w PEGDA, MW 3000), stimulated SMCs to up-regulate contractile marker mRNA. A cell-instructive scaffold construct was prepared by polymerizing a thin hydrogel film, with pendant RGD peptides for cell attachment, over the optimized hydrogel depots. SMCs seeded on these constructs had elevated levels of contractile marker mRNA after 3 d of culture compared with SMCs on control constructs. These results indicate that RGD-modified, heparin releasing PEGDA gels can act as cell-instructive scaffolds that promote contractile SMC phenotype.     

New polyurethane compositions containing high amounts of covalently bonded heparin

Heparin was bonded covalently to hydrophilic polyurethanes. Polymers were characterized by spectroscopic techniques (IR and ¹H, ¹³C NMR), molecular weight measurements and differential scanning calorimetry. The heparin content of the polymer films (up to more than 500 μg/cm²) was determined by a calorimetric system, and its biological activity was evaluated in vitro by APTT (activated partial thromboplastin time) measurements with a tenfold increase of the clotting time (about 300 s) with respect to the clotting time of human plasma (30 s). When heparin is bonded to a polymer provided with a hydrophilic polyfunctional spacer, the heparin content of the films is remarkably higher than previously described (about twice the reported values), and also the biological activity of the bonded heparin is favourably affected.     

Human endothelial cell growth on mussel-inspired nanofiber scaffold for vascular tissue engineering

The endothelialization of prosthetic scaffolds is considered to be an effective strategy to improve the effectiveness of small-diameter vascular grafts. We report the development of a nanofibrous scaffold that has a polymeric core and a shell mimicking mussel adhesive for enhanced attachment, proliferation, and phenotypic maintenance of human endothelial cells. Polycaprolactone (PCL) was chosen as a core material because of its good biodegradability and mechanical properties suitable for tissue engineering. PCL was electrospun into nanofibers with a diameter of approximately 700 nm and then coated with poly(dopamine) (PDA) to functionalize the surface of PCL nanofibers with numerous catechol moieties similar to mussel adhesives in nature. The formation of a PDA ad-layer was analyzed using multiple techniques, including scanning electron microscopy, Raman spectroscopy, and water contact angle measurements. When PDA-coated PCL nanofibers were compared to unmodified and gelatin-coated nanofibers, human umbilical vein endothelial cells (HUVECs) exhibited highly enhanced adhesion and viability, increased stress fiber formation, and positive expression of endothelial cell markers (e.g., PECAM-1 and vWF).     

Heparin stability: effects of diluent, heparin activity, container, and pH.

The effects have been studied of diluent, heparin activity after dilution, container, and pH on the stability of heparin solutions stored under conditions resembling those present during heparin infusion by intravenous drip or syringe pump. Heparin activity was measured by activated partial thromboplastin time and thrombin clotting time (and, in one set of studies, also by factor Xa inhibitor assay and protamine sulphate neutralisation). Heparin activity was stable for 6 hours regardless of storage conditions. After 24 hours heparin activity was stable when the drug was diluted in 0.9% saline and stored in plastic, but a small loss of activity was observed in several studies after dilution in 5% dextrose or storage in glass. A more extensive comparison confirmed a 3-5% loss in heparin activity over 24 hours after dilution in 5% dextrose. Changing the pH to 3.5 or 10.0 had little effect on storage stability. We conclude that heparin activity in vitro remains stable during short infusions but recommend dilution in 0.9% saline and a plastic container when a heparin solution is infused over 24 hours.     

Heparin monitoring and thrombosis.

The activated parial thromboplastin time is susceptible to changing concentrations of factor VII even in the presence of heparin. Heparin delays, but does not abolisn, thrombin generation, and this delay is obviated by factor VIII. It is not known if activated parial thromboplastin time shortened by increased factor VIII demands an increase in heparin administration.     

Anwendung von niedermolekularem Heparin bei Hämodialysepatienten

Summary Low-molecular-weight (LMW) heparin has been compared to standard unfractionated (UF) heparin in a total of 49 patients on hemodialysis and hemofiltration in order to determine the necessary therapeutic dose and its effect on the coagulation system. A LMW heparin dose corresponding to 50% of the normal UF heparin dose was found to produce similar plasma heparin levels (anti-FXa-U/ml) in particular on minimal heparinization. At higher doses, UF heparin produced a more marked increase in plasma-heparin than did LMW heparin. Highly significant differences were found between UF and LMW heparin in their effects on PTT and thrombin time. Partial thromboplastin time (PTT) increased under UF heparin by an average of 120 s whereas LMW heparin only produced an increase of 5–7 s. Thrombin time was increased by 250–280 s under UF heparin and by 5–8 s under LMW heparin. With this LMW heparin dose of 50% of the UF heparin dose, no thrombosis of the extracorporal system occurred and no macroscopic detectable thrombotic material was found in the dialyzers or filters. No significant differences were observed between the effects of UF and LMW heparin on Factor VIII activity and fibrin monomers, so that a difference in coagulation activation between the two heparins can be excluded. Furthermore, there were no changes in thromboplastin time according to Quick, fibrinogen, antithrombin III, plasminogen, and a₂-antiplasmin. Thus effective Anti-FXa levels and by similar antithrombotic activity, LMW heparin will probably present less of a bleeding risk because of its reduced effect on PTT and thrombin time. LMW heparin therefore appears to be a good alternative to UF heparin for patients with renal insufficiency requiring dialysis. LMW heparin is indicated in particular in patients at bleeding risk, with diabetic retinopathy, on therapy with oral anticoagulants or platelet aggregation inhibitors, and with thrombocytopenia.

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Abkürzungsverzeichnis F Faktor - FFA Free Fatty Acids=freie Fettsäuren - LMW-Heparin Low Molecular Weight Heparin=niedermolekulares Heparin - LPL Lipoprotein-Lipase - PTT Partial Thromboplastin Time=Partielle Thromboplastinzeit - UF-Heparin unfraktioniertes Heparin     

Enhanced osteogenesis and angiogenesis by PCL/chitosan/Sr-doped calcium phosphate electrospun nanocomposite membrane for guided bone regeneration

Abstract

Membranes play pivotal role in guided bone regeneration (GBR) technique for reconstruction alveolar bone. GBR membrane that is able to stimulate both osteogenic and angiogenic differentiation of cells may be more effective in clinic practice. Herein, we fabricated the Sr-doped calcium phosphate/polycaprolactone/chitosan (Sr-CaP/PCL/CS) nanohybrid fibrous membrane by incorporating 20?wt% bioactive Sr-CaP nanoparticles into PCL/CS matrix via one-step electrospinning method, in order to endow the membrane with stimulation of osteogenesis and angiogenesis. The physicochemical properties, mechanical properties, Sr²⁺ release behavior, and the membrane stimulate bone mesenchymal stem cell (BMSCs) differentiation were evaluated in comparison with PCL/CS and CaP/PCL/CS membranes. The SEM images revealed that the nanocomposite membrane mimicked the extracellular matrix structure. The release curve presented a 28-day long continuous release of Sr²⁺ and concentration which was certified in an optimal range for positive biological effects at each timepoint. The in vitro cell culture experiments certified that the Sr-CaP/PCL/CS membrane enjoyed excellent biocompatibility and remarkably promoted rat bone mesenchymal stem cell (BMSCs) adhesion and proliferation. In terms of osteogenic differentiation, BMSCs seeded on the Sr-CaP/PCL/CS membrane showed a higher ALP activity level and a better matrix mineralization. What’s more, the synergism of the Sr²⁺ and CaP from the Sr-CaP/PCL/CS membrane enhanced BMSCs angiogenic differentiation, herein resulting in the largest VEGF secretion amount. Consequently, the Sr-CaP/PCL/CS nanohybrid electrospun membrane has promising applications in GBR.     

Surface Engineering of Poly(ethylene terephthalate) for Durable Hemocompatibility via a Surface Interpenetrating Network Technique

Heparin was covalently bonded on chemically inert PET substrate using a surface modification technique–surface interpenetrating network with the purpose of fabricating long‐lasting biocompatible materials as vascular grafts. FTIR and XPS spectra confirmed the successful heparinization of PET (PET‐Hep). The density of surface‐immobilized heparin as quantified by a colorimetric method could reach 2.4 μg cm^?2 (in the reported optimal range: 1.5–3.0 μg cm^?2). The hemocompatibility of the heparin‐immobilized PET was improved as evidenced by a platelet adhesion test: significantly less platelet adhesion on PET‐Hep (11.60%) than on untreated PET (48.91%). An MTT assay indicated PET‐Hep was nontoxic to human dermal fibroblast cells. After an initial 5.24% loss of heparin from PET‐Hep in the first 14 h immersion in PBS buffer solution, no further leaching of heparin was found.     

The characteristics,biodistribution and bioavailability of a chitosan-based nanoparticulate system for the oral delivery of heparin

Heparin is a potent anticoagulant; however, it is poorly absorbed in the gastrointestinal tract. In this study, we developed a nanoparticle (NP) system shelled with chitosan (CS) for oral delivery of heparin; the NPs were prepared by a simple ionic gelation method without chemically modifying heparin. The drug loading efficiency of NPs was nearly 100% because a significantly excess amount of CS was used for the CS/heparin complex preparation. The internal structure of the prepared NPs was examined by small angle X-ray scattering (SAXS). The obtained SAXS profiles suggest that the NPs are associated with a two-phase system and consist of the CS/heparin complex microdomains surrounded by the CS matrix. The stability of NPs in response to pH had a significant effect on their release of heparin. No significant anticoagulant activity was detected after oral administration of the free form heparin solution in a rat model, while administration of NPs orally was effective in the delivery of heparin into the blood stream; the absolute bioavailability was found to be 20.5%. The biodistribution of the drug carrier, ^99mTc-labeled CS, in rats was studied by the single-photon emission computed tomography after oral administration of the radio-labeled NPs. No significant radioactivity was found in the internal organs, indicating a minimal absorption of CS into the systemic circulation. These results suggest that the NPs developed in the study can be employed as a potential carrier for oral delivery of heparin.     

Embroidered and Surface Modified Polycaprolactone-Co-Lactide Scaffolds as Bone Substitute: <Emphasis Type="Italic">In Vitro</Emphasis> Characterization

The aim of this study was to evaluate an embroidered polycaprolactone-co-lactide (trade name PCL) scaffold for the application in bone tissue engineering. The surface of the PCL scaffolds was hydrolyzed with NaOH and coated with collagen I (coll I) and chondroitin sulfate (CS). It was investigated if a change of the surface properties and the application of coll I and CS could promote cell adhesion, proliferation, and osteogenic differentiation of human mesenchymal stem cells (hMSC). The porosity (80%) and pore size (0.2–1 mm) of the scaffold could be controlled by embroidery technique and should be suitable for bone ingrowth. The treatment with NaOH made the polymer surface more hydrophilic (water contact angle dropped to 25%), enhanced the coll I adsorption (up to 15%) and the cell attachment (two times). The coll I coated scaffold improved cell attachment and proliferation (three times). CS, as part of the artificial matrix, could induce the osteogenic differentiation of hMSC without other differentiation additives. The investigated scaffolds could act not just as temporary matrix for cell migration, proliferation, and differentiation in bone tissue engineering but also have a great potential as bioartificial bone substitute.