Multiprotein collagen/keratin hydrogel promoted myogenesis and angiogenesis of injured skeletal muscles in a mouse model.

Volumetric loss is one of the challenging issues in muscle tissue structure that causes functio laesa. Tissue engineering of muscle tissue using suitable hydrogels is an alternative to restoring the physiological properties of the injured area. Here, myogenic properties of type I collagen (0.5%) and keratin (0.5%) were investigated in a mouse model of biceps femoris injury. Using FTIR, gelation time, and rheological analysis, the physicochemical properties of the collagen (Col)/Keratin scaffold were analyzed. Mouse C2C12 myoblast-laden Col/Keratin hydrogels were injected into the injury site and histological examination plus western blotting were performed to measure myogenic potential after 15 days. FTIR indicated an appropriate interaction between keratin and collagen. The blend of Col/Keratin delayed gelation time when compared to the collagen alone group. Rheological analysis revealed decreased stiffening in blended Col/Keratin hydrogel which is favorable for the extrudability of the hydrogel. Transplantation of C2C12 myoblast-laden Col/Keratin hydrogel to injured muscle tissues led to the formation of newly generated myofibers compared to cell-free hydrogel and collagen groups (p < 0.05). In the C2C12 myoblast-laden Col/Keratin group, a low number of CD31+ cells with minimum inflammatory cells was evident. Western blotting indicated the promotion of MyoD in mice that received cell-laden Col/Keratin hydrogel compared to the other groups (p < 0.05). Despite the increase of the myosin cell-laden Col/Keratin hydrogel group, no significant differences were obtained related to other groups (p > 0.05). The blend of Col/Keratin loaded with myoblasts provides a suitable myogenic platform for the alleviation of injured muscle tissue.

Interaction of alginate with nano-hydroxyapatite-collagen using strontium provides suitable osteogenic platform.

BACKGROUND: Hydrogels based on organic/inorganic composites have been at the center of attention for the fabrication of engineered bone constructs. The establishment of a straightforward 3D microenvironment is critical to maintaining cell-to-cell interaction and cellular function, leading to appropriate regeneration. Ionic cross-linkers, Ca2+, Ba2+, and Sr2+, were used for the fabrication of Alginate-Nanohydroxyapatite-Collagen (Alg-nHA-Col) microspheres, and osteogenic properties of human osteoblasts were examined in in vitro and in vivo conditions after 21 days. RESULTS: Physicochemical properties of hydrogels illustrated that microspheres cross-linked with Sr2+ had reduced swelling, enhanced stability, and mechanical strength, as compared to the other groups. Human MG-63 osteoblasts inside Sr2+ cross-linked microspheres exhibited enhanced viability and osteogenic capacity indicated by mineralization and the increase of relevant proteins related to bone formation. PCR (Polymerase Chain Reaction) array analysis of the Wnt (Wingless-related integration site) signaling pathway revealed that Sr2+ cross-linked microspheres appropriately induced various signaling transduction pathways in human osteoblasts leading to osteogenic activity and dynamic growth. Transplantation of Sr2+ cross-linked microspheres with rat osteoblasts into cranium with critical size defect in the rat model accelerated bone formation analyzed with micro-CT and histological examination. CONCLUSION: Sr2+ cross-linked Alg-nHA-Col hydrogel can promote functionality and dynamic growth of osteoblasts.

An Insight into Eudragit S100 Preserving Mechanism of Cinnarizine Supersaturation.

Generally, supersaturation of weakly basic drug solution in the gastrointestinal tract can be followed by precipitation, and this can compromise the bioavailability of drugs. The purpose of this study was to evaluate the effect of Eudragit® S100 on the pH-induced supersaturation of cinnarizine and to examine the preserving mechanism of cinnarizine supersaturation by Eudragit®. Variables, including pH of media, ionic strength, and degree of supersaturation, were studied to investigate the effects of these parameters on cinnarizine supersaturation in the presence and absence of Eudragit®. The size of the Eudragit® aggregate in solution using dynamic light scattering was determined. The effect of Eudragit® on the transport of cinnarizine through the Caco-2 membrane was also investigated. The particle size study of Eudragit® aggregates showed that the size of these aggregates become large when the pH was lowered. Supersaturation experiments also demonstrated that Eudragit® preserved higher cinnarizine supersaturation with increasing ionic strength of the solution. The phase separation behavior of cinnarizine solution as a function of the degree of the supersaturation could be readily explained by considering the drug amorphous solubility. In vitro permeation studies revealed that the rate of cinnarizine permeation across Caco-2 cells increased in the presence of Eudragit®. According to the obtained results, the aggregation status of Eudragit® and nonspecific hydrophobic cinnarizine-Eudragit® interactions seemed to be essential in determining the effect of Eudragit® on cinnarizine supersaturation.

The osteogenic differentiation of human dental pulp stem cells in alginate-gelatin/Nano-hydroxyapatite microcapsules.

BACKGROUND: Microcapsule is considered as a promising 3D microenvironment for Bone Tissue Engineering (BTE) applications. Microencapsulation of cells in an appropriate scaffold not only protected the cells against excess stress but also promoted cell proliferation and differentiation. Through the current study, we aimed to microcapsulate the human Dental Pulp Stem Cells (hDPSCs) and evaluated the proliferation and osteogenic differentiation of those cells by using MTT assay, qRT-PCR, Alkaline phosphatase, and Alizarine Red S. RESULTS: The SEM results revealed that Alg/Gel microcapsules containing nHA showed a rough and more compact surface morphology in comparison with the Alg/Gel microcapsules. Moreover, the microencapsulation by Alg/Gel/nHA could improve cell proliferation and induce osteogenic differentiation. The cells cultured in the Alg/Gel and Alg/Gel/nHA microcapsules showed 1.4-fold and 1.7-fold activity of BMP-2 gene expression more in comparison with the control group after 21 days. The mentioned amounts for the BMP-2 gene were 2.5-fold and 4-fold more expression for the Alg/Gel and Alg/Gel/nHA microcapsules after 28 days. The nHA, addition to hDPSCs-laden Alg/Gel microcapsule, could up-regulate the bone-related gene expressions of osteocalcin, osteonectin, and RUNX-2 during the 21 and 28 days through the culturing period, too. Calcium deposition and ALP activities of the cells were observed in accordance with the proliferation results as well as the gene expression analysis. CONCLUSION: The present study demonstrated that microencapsulation of the hDPSCs inside the Alg/Gel/nHA hydrogel could be a potential approach for regenerative dentistry in the near future.

Electrospun polyurethane/poly (ɛ-caprolactone) nanofibers promoted the attachment and growth of human endothelial cells in static and dynamic culture conditions.

In this study, the angiogenic capacity of human endothelial cells was studied after being plated on the surface of polyurethane-poly caprolactone (PU/PCL) scaffolds for 72 h. In this study, cells were designated into five different groups, including PU, PU/PCL (2:1), PU/PCL (1:1); PU/PCL (1:2); and PCL. Data revealed that the PU/PCL (2:1) composition had a higher modulus and breakpoint in comparison with the other groups (p < 0.05). Compared to the other groups, the PU/PCL scaffold with a molar ratio of 2:1 had lower the contact angle θ and higher tensile stress (p < 0.05). The mean size of the PU nanofibers was reduced after the addition of PCL (p < 0.05). Based on our data, the culture of endothelial cells on the surface of PU/PCL (2:1) did not cause nitrosative stress and cytotoxic effects under static conditions compared to cells plated on a conventional plastic surface (p > 0.05). Based on data from the static condition, we fabricated a tubular PU/PCL (2:1) construct for six-day dynamic cell culture inside loop air-lift bioreactors. Scanning electron microscopy showed the attachment of endothelial cells to the luminal surface of the PU/PCL scaffold. Cells were flattened and aligned under the culture medium flow. Immunofluorescence imaging showed the attachment of cells to the luminal surface indicated by blue nuclei on the luminal surface. These data demonstrated that the application of PU/PCL substrate could stimulate endothelial cells activity under static and dynamic conditions.

Performance evaluation of a novel conceptual bioprocess for clinically-required mass production of hematopoietic cells.

OBJECTIVE: The novel engineered bioprocess, which was designed and modeled to provide the clinically relevant cell numbers for different therapies in our previous work (Kaleybar et al. Food Bioprod Process 122:254-268, https://doi.org/10.1016/j.fbp.2020.04.012 , 2020), was evaluated by using U937 as hematopoietic model cells. RESULTS: The culture system showed a 30-fold expansion of U937 cells in one-step during a 10-day culture period. The cell growth profile, the substrate and oxygen consumptions, and byproduct formations were all in agreement with the model predications during 7 days. The cell proliferation decrease after 7 days was attributed to optional oxygen limiting condition in the last days of culture. The bioreactor culture system revealed also a slight enhancement of lactate dehydrogenase (LDH) production as compared to the 2D conventional culture system, indicating the low impact of shear stress on cellular damage in the dynamic system. CONCLUSIONS: The results demonstrated that the conceptual bioprocess for suspended stem cell production has a great potential in practice although additional experiments are required to improve the system.

Collagen-alginate-nano-silica microspheres improved the osteogenic potential of human osteoblast-like MG-63 cells.

Modular bone tissue engineering is touted as an alternative approach to replace the damaged bone tissue. Hydrogel microcapsules could promote therapeutic properties by providing 3D condition and an increased cell-to-cell interaction. We investigated the osteogenic properties of alginate-nano-silica hydrogels enriched with collagen and gelatin on human osteoblast-like MG-63 cells. For encapsulation, cells were divided into three groups; control (alginate+ nano-silica), collagen (alginate + collagen + nano-silica), and gelatin (alginate + gelatin + nano-silica) and expanded for 28 days. Cell survival was determined by trypan blue staining and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. To confirm the osteogenic potential, we measured the alkaline phosphatase activity. Alizarin red S staining was used to reveal the existence of hydroxyapatite and transcription BMP-2, osteocalcin and osteonectin evaluated by the real-time polymerase chain reaction. Collagen substrate caused a reduced swelling ratio compared with the control and gelatin groups (P < 0.05). Compared with other groups, collagen had potential to improve mechanical strength and generate porous membrane structure. The addition of collagen (4-fold) and gelatin (1.5-fold) increased cell proliferation rate compared with the control (P < 0.05). Biochemical analysis and Alizarin red S staining showed that collagen-induced osteogenesis by induction of alkaline phosphatase and matrix mineralization. The expression of osteocalcin and BMP-2 was increased in cells from the collagen group. As a result, the combination of natural polymers collagen and gelatin with alginate + nano-silica can increase the osteogenic potential of human osteoblasts.

Comparative study of collagen and gelatin in chitosan-based hydrogels for effective wound dressing: Physical properties and fibroblastic cell behavior.

The influence of collagen as an effective substitute for gelatin was investigated on properties of chitosan/gelatin hydrogels for fibroblasts growth and attachment for wound dressing applications. We synthesized hydrogels based on chitosan associated with collagen and gelatin biopolymers (in the ratio of 1:5 and 1:1, respectively). The hydrogels properties such as morphology, swelling ratio, mechanical characteristics, water vapor loss, water vapor transmission rate (WVTR), and biodegradation were analyzed. 1 × 105 human fibroblasts were seeded per ml of hydrogels and maintained for 7 days. Cell viability was assessed by using MTT. The presence of collagen caused reduced swelling ratio, and biodegradation rate compared to chitosan/gelatin hydrogels (p < 0.05). The introduction of collagen into chitosan hydrogels improved the mechanical strength compared to gelatin. Hydrogels with collagen possessed an optimum WVTR compared to the chitosan group and hydrogels with gelatin (p < 0.05). Analyzing the morphology of hydrogels revealed that the addition of collagen leads to a homogenous and interconnected structure. Collagen impregnation promoted cell survival and attachment compared with chitosan hydrogels after 7 days (p < 0.05). Collectively, these results demonstrated the potential of the chitosan/collagen hydrogels for wound dressing applications.

Alginate-gelatin encapsulation of human endothelial cells promoted angiogenesis in in vivo and in vitro milieu.

Up to present, many advantages have been achieved in the field of cell-based therapies by applying sophisticated methodologies and delivery approaches. Microcapsules are capable to provide safe microenvironment for cells during transplantation in a simulated physiological 3D milieu. Here, we aimed to investigate the effect of alginate-gelatin encapsulation on angiogenic behavior of human endothelial cells over a period of 5 days. Human umbilical vein endothelial cells were encapsulated by alginate-gelatin substrate and incubated for 5 days. MTT and autophagy PCR array analysis were used to monitor cell survival rate. For in vitro angiogenesis analysis, cell distribution of Tie-1, Tie-2, VEGFR-1, and VEGFR-2 were detected by ELISA. In addition to in vitro tubulogenesis assay, we monitored the expression of VE-cadherin by Western blotting. The migration capacity of encapsulated HUVECs was studied by measuring MMP-2 and MMP-9 via gelatin zymography. The in vivo angiogenic potential of encapsulated HUVECs was analyzed in immune-compromised mouse implant model during 7 days post-transplantation. We demonstrated that encapsulation promoted HUVECs cell survival and proliferation. Compared to control, no significant differences were observed in autophagic status of encapsulated cells (p > 0.05). The level of Tie-1, Tie-2, VEGFR-1, and VEGFR-2 were increased, but did not reach to significant levels. Encapsulation decreased MMP-2, -9 activity and increased the VE-cadherin level in enclosed cells (p < 0.05). Moreover, an enhanced in vivo angiogenic response of encapsulated HUVECs was evident as compared to non-capsulated cells (p < 0.05). These observations suggest that alginate-gelatin encapsulation can induce angiogenic response in in vivo and in vitro conditions.

Partially aerated submerged fixed-film bioreactor for simultaneous removal of carbon and nutrients from high-strength nitrogen wastewaters: effect of aeration rate and C:N:P ratio.

Influence of aeration rate and COD:N:P (C:N:P) ratio on the performance of an upflow partially aerated submerged fixed film (UP/ASFF) bioreactor for simultaneous carbon and nutrient removal from high-strength nitrogen wastewater was investigated during 6 months. Airflow rates at three levels of 1.5, 3, and 4.5 L/min and C:N:P ratios at four levels of 450:300:10, 450:150:10, 450:100:10, and 450:75:10 were selected as the two main input factors. All experiments were performed at constant chemical oxygen demand (COD), phosphorus (P) and hydraulic residence time of 450 mg COD/L, 10 mg PO43- -P/L and 7.3 h, respectively. The results showed when the airflow rate increased from 1.5 to 4.5 L/min, complete COD removal was achieved. At an airflow rate of 4.5 L/min, total nitrogen removal reached a maximum value of 75% for the C:N:P ratio of 450:75:10. A maximum value of 54% for total phosphorus removal, however, was obtained at an airflow rate of 3 L/min for the C:N:P ratio of 450:75:10. Analysis of variance for the obtained data revealed that aeration rate and nitrogen concentration had more impact on phosphorus removal than COD and nitrogen removal. The study demonstrated that the UP/ASFF system has considerable potential for use in simultaneous removal of carbon and nutrients for high-strength nitrogen wastewater.

Influence of shear force on ex vivo expansion of hematopoietic model cells in a stirred tank bioreactor.

To evaluate shear stress influence on ex vivo expansion of hematopoietic cell lineages for clinical application, in this study, human pro-monocytic cell (namely U937 cell line) was selected as a hematopoietic stem cell (HSC) model and cultured in suspension mode at two different agitation rates (50, 100 rpm) in the stirred bioreactor. At the agitation rate of 50 rpm, the cells achieved higher expansion folds (27.4 fold) with minimal morphological changes as well as apoptotic cell death, while at 100 rpm the expansion fold decreased after 5-day of culture in suspension culture in comparison with static culture and reached 24.5 fold at the end of the culture. The results of glucose consumption and lactate production were also in agreement with the data of fold expansion and indicated the preference of culture in the stirred bioreactor when agitated at 50 rpm. This study indicated the stirred bioreactor system with an agitation rate of 50 rpm and surface aeration may be used as a potential dynamic culture system for clinical applications of hematopoietic cell lineage. The current experiments shed data related to the effect of shear stress on human U937 cells, as a hematopoietic cell model, to set a protocol for expansion of HSCs for biomedical applications.

Developing a Multichannel Bioreactor with a Collagen Scaffold, ECM, and Cryoprecipitate to Significantly Produce Platelets from Umbilical Cord Blood Stem Cells.

Background: Platelets play a key role in the treatment of thrombocytopenia. Nowadays, platelets (PLTs) are only obtained through blood donation. However, due to the limitations in their preparation and storage, they are produced in laboratories, especially through bioreactors that convert megakaryocytes from stem cells into large-scale injectable PLTs. Materials and Methods: In this study, the CD34 cells isolated from cord blood were differentiated into megakaryocytes. A 6-chamber bioreactor with a two-layer collagen scaffold, several ECM factors, and human cryoprecipitate were used to simulate the structure of the bone marrow. After the addition of megakaryocytes to the scaffold, PLTs were produced due to the flow pressure and the interaction between the scaffold structure and the ECM factors. Results: CD41 + cells were expanded 100 times as much as CD34 + cord blood stem cells. The mean PLT release from one megakaryocyte in the pure collagen scaffold was 17.42 PLTs. Once fibrin, fibronectin, hyaluronic acid, and cryoprecipitates were added to collagen, the mean PLT release was 21.4, 22.4, 23.9, and 27.37, respectively. With the simultaneous addition of three factors to collagen (CFFH) and then four factors (CFFHC), the number of PLTs reached 30.52 and then 34. Conclusion: Functional PLTs can be produced on a large scale with a multi-chamber bioreactor using a combination of ECM and cryoprecipitate with collagen scaffolding.

Phenolated alginate hydrogel induced osteogenic properties of mesenchymal stem cells via Wnt signaling pathway.

Osteogenic properties of phenolated alginate (1.2 %) hydrogel containing collagen (0.5 %)/nano-hydroxyapatite (1 %) were studied on human mesenchymal stem cells in vitro. The phenolation rate and physical properties of the hydrogel were assessed using nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FTIR), Scanning electron microscope (SEM), swelling ratio, gelation time, mechanical assay, and degradation rate. The viability of encapsulated cells was monitored on days 7, 14, and 21 using an MTT assay. Osteoblast differentiation was studied using western blotting, and real-time PCR. Using PCR array analysis, the role of the Wnt signaling pathway was also investigated. Data showed that the combination of alginate/collagen/nanohydroxyapatite yielded proper mechanical features. The addition of nanohydroxyapatite, and collagen reduced degradation, swelling rate coincided with increased stiffness. Elasticity and pore size were also diminished. NMR and FTIR revealed suitable incorporation of collagen and nanohydroxyapatite in the structure of alginate. Real-time PCR analysis and western blotting indicated the expression of osteoblast-related genes such as Runx2 and osteocalcin. PCR array revealed the induction of numerous genes related to Wnt signaling pathways during the maturation of human stem cells toward osteoblast-like cells. In vivo data indicated that transplantation of phenolated alginate/collagen/nanohydroxyapatite hydrogel led to enhanced de novo bone formation in rats with critical-sized calvarial defects. Phenolated alginate hydrogel can promote the osteogenic capacity of human amniotic membrane mesenchymal stem cells in the presence of nanohydroxyapatite and collagen via engaging the Wnt signaling pathway.

Collagen and nano-hydroxyapatite interactions in alginate-based microcapsule provide an appropriate osteogenic microenvironment for modular bone tissue formation.

The addition of nano-hydroxyapatite (nHA) and collagen (Col) to the alginate (Alg) microcapsule hydrogel reduced swelling and degradation ratios while the compressive strength increased compared to Alg, Alg-Col, and Alg-nHA groups. MTT assay and Calcein-AM staining revealed an enhanced MG-63 osteoblasts viability in the Alg-nHA-Col hydrogel compared to the other groups. SEM showed the attachment of MG-63 osteoblasts inside Alg-Col hydrogels. Non-significant differences were found in antioxidant capacity of cells inside the Alg-nHA-Col hydrogel compared to the Alg group. Hematoxylin-Eosin staining showed the distribution of MG-63 osteoblasts inside microspheres. Calcium deposits, alkaline phosphatase (ALP) activity with the increase of intracellular calcium were found in Alg-nHA-Col group. Western blotting showed that levels of osteocalcin, ColA2, Sox-9, and ColA1 also significantly increased compared to the Alg, Alg-Col, Alg-nHA groups. The present study demonstrated that the addition of mineral nHA and protein (Col) into the Alg improves osteogenic potential and provides a 3D platform for modular bone tissue engineering.

PCL/Col I-based magnetic nanocomposite scaffold provides an osteoinductive environment for ADSCs in osteogenic cues-free media conditions.

BACKGROUND: The bone tissue engineering (BTE) approach has been introduced as an alternative to conventional treatments for large non-healing bone defects. Magnetism promotes stem cells' adherence to biocompatible scaffolds toward osteoblast differentiation. Furthermore, osteogenic differentiation media are expensive and any changes in its composition affect stem cells differentiation. Moreover, media growth factors possess a short half-life resulting in the rapid loss of their functions in vivo. With the above in mind, we fabricated a multilayered nanocomposite scaffold containing the wild type of Type I collagen (Col I) with endogenous magnetic property to promote osteogenesis in rat ADSCs with the minimum requirement of osteogenic differentiation medium. METHODS: Fe3O4 NPs were synthesized by co-precipitation method and characterized using SEM, VSM, and FTIR. Then, a PCL/Col I nanocomposite scaffold entrapping Fe3O4 NPs was fabricated by electrospinning and characterized using SEM, TEM, AFM, VSM, Contact Angle, tensile stretching, and FTIR. ADSCs were isolated from rat adipose tissue and identified by flow cytometry. ADSCs were loaded onto PCL/Col I and PCL/Col I/Fe3O4-scaffolds for 1-3 weeks with/without osteogenic media conditions. The cell viability, cell adhesion, and osteogenic differentiation were evaluated using MTT assay, SEM, DAPI staining, ALP/ARS staining, RT-PCR, and western blotting, respectively. RESULTS: SEM, VSM, and FTIR results indicated that Fe3O4 was synthesized in nano-sized (15-30 nm) particles with spherical-shaped morphology and superparamagnetic properties with approved chemical structure as FTIR revealed. According to SEM images, the fabricated magnetic scaffolds consisted of nanofiber (500-700 nm). TEM images have shown the Fe3O4 NPs entrapped in the scaffold's fiber without bead formation. FTIR spectra analysis confirmed the maintenance of the natural structure of Col I, PCL, and Fe3O4 upon electrospinning. AFM data have shown that MNPs incorporation introduced stripe-like topography to nanofibers, while the depth of the grooves has decreased from 800 to 500 nm. Flow cytometry confirmed the phenotype of ADSCs according to their surface markers (i.e., CD29 and CD105). Additionally, Fe3O4 NP improved nanocomposite scaffold strength, wettability, porosity, biocompatibility and also facilitates the ALP activity, calcium-mineralization. Finally, magnetic nanocomposite scaffolds upregulated osteogenic-related genes or proteins' expression (e.g., Col I, Runx2, OCN, ON, BMP2) in seeded ADSCs with/without osteo-differentiation media conditions. CONCLUSIONS: Together, these results indicate that Fe3O4 NPs within the natural structure of Col I increase osteogenic differentiation in osteogenic cues-free media conditions. This effect could be translated in vivo toward bone defects healing. These findings support the use of natural ECM materials alongside magnetic particles as composite scaffolds to achieve their full therapeutic potential in BTE treatments.

Combination of polyglycerol sebacate coated with collagen for vascular engineering.

Introduction: Here, we monitored the cytocompatibility of scaffolds consisting of poly (glycerol sebacate) (PGS) coated with collagen (Col) for endothelial cell activity after 72 hours. Methods: Human endothelial cells were allocated into Control, PGS, and PGS+Col groups. Scaffolds were characterized using FTIR and HNMR spectroscopy. Contact angel analysis and SEM were used to study wettability, surface morphology, and cell attachment. Cell survival was assessed using LDH leakage assay. Levels of Tie-1, Tie-2, VE-Cadherin, and VEGFR-2 were measured using western blotting and real-time PCR. Results: FTIR and HNMR analyses revealed the proper blending in PGS+Col group. SEM imaging exhibited a flat surface in the PGS group while thin Col fibers were detected in PGS+Col surface. The addition of Col to the PGS reduced the contract angle values from 97.3˚ to 81.1˚. Compared to PGS substrate alone, in PGS+Col group, cells appropriately attached to the surface. PGS and PGS+Col did not alter the leakage of LDH to the supernatant compared to control cells, showing the cytocopatiblity of PGS-based scaffolds. SOD and NO levels were increased significantly in PGS (p<0.05) and PGS+Col groups (p<0.001), respectively. We found that PGS+Col decreased Tie-1 content in endothelial cells whereas protein levels of Tie-2 and VE-Cadherin and expression of VEGFR-2 remained unchanged compared to PGS and control groups. Conclusion: Simultaneous application of Col and PGS can stimulate normal endothleial cell morphology without the alteration of tyrosine kinases receptors and cadherin.

Alginate-chitosan core-shell microcapsule cultures of hepatic cells in a small scale stirred bioreactor: impact of shear forces and microcapsule core composition.

A small scale stirred bioreactor was designed and the effect of different agitation rates (30, 60 and 100 rpm) was investigated on HepG2 cells cultured in alginate-chitosan (AC) core-shell microcapsule in terms of the cell proliferation and liver-specific function. The microencapsulated hepatic cells could proliferate well when they were cultured for 10 days at 30 rpm while the cell-laden microcapsules showed no cell proliferation at 100 rpm in the bioreactor system. Albumin production rate, as an important liver function, increased also 1.8- and 1.5- fold under stirring rate of 30 rpm compared to the static culture and 60 rpm of agitation, respectively. Moreover, In comparison with the static culture, about 1.5-fold increment in urea production was observed at 30 rpm. Similarly, the highest expressions of albumin and P450 genes were found at 30 rpm stirring rate, which were 4.9- and 19.2-fold of the static culture. Addition of collagen to the microcapsule core composition (ACol/C) could improve the cell proliferation and functionality at 60 rpm in comparison with the cell-laden microcapsules without collagen. The study demonstrated the hepatic cell-laden ACol/C microcapsule hydrogel cultured in the small scale stirred bioreactor at low mixing rate has a great potential for mass production of the hepatic cells while maintaining liver-specific functions.

Fabrication of alginate-based hydrogel cross-linked via horseradish peroxidase for articular cartilage engineering.

OBJECTIVE: We aimed to detect the effect of a couple of parameters including Alg, H2O2, and HRP concentrations on the gelation time of Alg-based hydrogels using an enzymatic cross-linked procedure. RESULTS: NMR, UV-Vis, and ATR-FTIR analyses confirmed the conjugation of Ph to the Alg backbone. Data showed gelation time was delayed with the increase and reduction of H2O2 and HRP, respectively. We noted that hydrogel consisted of 1.2% (w/v) Alg, 5 U HRP, and 100 mM H2O2 yielded an appropriate gelation time with appropriate mechanical properties. The addition of 0.5% (v/v) Col developed hydrogel increased the gelation time. The data showed that Alg, HRP, and H2O2 with the ratio of 1:0.54:0.54 had proper physicochemical features for cartilage engineering.

Influence of gelatin and collagen incorporation on peroxidase-mediated injectable pectin-based hydrogel and bioactivity of fibroblasts.

Pectin has recently attracted increasing attention for biomedical and pharmaceutical applications. Due to the lack of adhesion molecules in polysaccharides, phenolic hydroxyl conjugated gelatin was added to enzymatically-gellable peroxidase-modified pectin derivative and compared with phenolic hydroxyl -pectin/collagen. Both pectin and gelatin were modified by tyramine hydrochloride in the presence of EDC/NHS. The phenolic hydroxyl -pectin/phenolic hydroxyl -gelatin, phenolic hydroxyl-pectin/collagen, and phenolic hydroxyl -pectin hydrogels were prepared using horseradish peroxidase and hydrogen peroxide. The hydrogels were characterized by gelation time analysis. Morphology, enzymatic biodegradation, mechanical and swelling properties as well as water vapor transmission rate were also evaluated. Fibroblasts were cultured for 7 days, and the survival rate was evaluated using conventional MTT assay. Hydrogels composed of Ph-pectin/Ph-gelatin showed decreased biodegradation rate, and WVTR and further improved mechanical performance in comparison with other groups. Both phenolic hydroxyl -pectin/collagen and phenolic hydroxyl -pectin/phenolic hydroxyl -gelatin hydrogels exhibited porous structures. The hydrogels composed of collagen promoted cell survival rate 1.4 and 3.5 times compared to phenolic hydroxyl -gelatin and phenolic hydroxyl -pectin based hydrogels at the end of 7 days, respectively (p < 0.001). The study demonstrated the potential of enzymatically-gellable pectin-based hydrogels as cost-effective frameworks for use in tissue engineering applications.

Tissue Engineering Strategies to Increase Osteochondral Regeneration of Stem Cells; a Close Look at Different Modalities.

The homeostasis of osteochondral tissue is tightly controlled by articular cartilage chondrocytes and underlying subchondral bone osteoblasts via different internal and external clues. As a correlate, the osteochondral region is frequently exposed to physical forces and mechanical pressure. On this basis, distinct sets of substrates and physicochemical properties of the surrounding matrix affect the regeneration capacity of chondrocytes and osteoblasts. Stem cells are touted as an alternative cell source for the alleviation of osteochondral diseases. These cells appropriately respond to the physicochemical properties of different biomaterials. This review aimed to address some of the essential factors which participate in the chondrogenic and osteogenic capacity of stem cells. Elements consisted of biomechanical forces, electrical fields, and biochemical and physical properties of the extracellular matrix are the major determinant of stem cell differentiation capacity. It is suggested that an additional certain mechanism related to signal-transduction pathways could also mediate the chondro-osteogenic differentiation of stem cells. The discovery of these clues can enable us to modulate the regeneration capacity of stem cells in osteochondral injuries and lead to the improvement of more operative approaches using tissue engineering modalities.