The implantation of bioengineered scaffolds into lesion-induced gaps of the spinal cord is a promising strategy for promoting functional tissue repair because it can be combined with other intervention strategies. Our previous investigations showed that functional improvement following the implantation of a longitudinally microstructured collagen scaffold into unilateral mid-cervical spinal cord resection injuries of adult Lewis rats was associated with only poor axon regeneration within the scaffold. In an attempt to improve graft-host integration as well as functional recovery, scaffolds were seeded with highly enriched populations of syngeneic, olfactory bulb-derived ensheathing cells (OECs) prior to implantation into the same lesion model. Regenerating neurofilament-positive axons closely followed the trajectory of the donor OECs, as well as that of the migrating host cells within the scaffold. However, there was only a trend for increased numbers of regenerating axons above that supported by non-seeded scaffolds or in the untreated lesions. Nonetheless, significant functional recovery in skilled forelimb motor function was observed following the implantation of both seeded and non-seeded scaffolds which could not be correlated to the extent of axon regeneration within the scaffold. Mechanisms other than simple bridging of axon regeneration across the lesion must be responsible for the improved motor function.
Adult ; Animals ; Axons* ; Collagen ; Forelimb ; Humans ; Rats ; Regeneration* ; Spinal Cord Injuries* ; Spinal Cord* ; Tissue Donors

BACKGROUND: Mesenchymal stem cells (MSCs) and/or biological scaffolds have been used to regenerate articular cartilage with variable success. In the present study we evaluated cartilage regeneration using a combination of bone marrow (BM)-MSCs, Hyalofast™ and/or native cartilage tissue following full thickness surgical cartilage defect in rabbits. METHODS: Full-thickness surgical ablation of the medial-tibial cartilage was performed in New Zealand white (NZW) rabbits. Control rabbits (Group-I) received no treatment; Animals in other groups were treated as follows. Group-II: BMMSCs (1 × 10⁶ cells) + Hyalofast™; Group-III: BMMSCs (1 × 10⁶ cells) + cartilage pellet (CP); and Group-IV: BMMSCs (1 × 10⁶ cells) + Hyalofast™+ CP. Animals were sacrificed at 12 weeks and cartilage regeneration analyzed using histopathology, International Cartilage Repair Society (ICRS-II) score, magnetic resonance observation of cartilage repair tissue (MOCART) score and biomechanical studies. RESULTS: Gross images showed good tissue repair (Groups IV>III>Group II) and histology demonstrated intact superficial layer, normal chondrocyte arrangement, tidemark and cartilage matrix staining (Groups III and IV) compared to the untreated control (Group I) respectively. ICRS-II score was 52.5, 65.0, 66 and 75% (Groups I–IV) and the MOCART score was 50.0, 73.75 and 76.25 (Groups II–IV) respectively. Biomechanical properties of the regenerated cartilage tissue in Group IV closed resembled that of a normal cartilage. CONCLUSION: Hyalofast™ together with BM-MSCs and CP led to efficient cartilage regeneration following full thickness surgical ablation of tibial articular cartilage in vivo in rabbits. Presence of hyaluronic acid in the scaffold and native microenvironment cues probably facilitated differentiation and integration of BM-MSCs.
Animals ; Bone Marrow ; Cartilage* ; Cartilage, Articular ; Chondrocytes ; Cues ; Hyaluronic Acid ; Mesenchymal Stromal Cells* ; New Zealand ; Osteoarthritis ; Rabbits* ; Regeneration*

BACKGROUND: Stem cell therapy requires a serum-free and/or chemically-defined medium for commercialization, but it is difficult to find one that supports long-term expansion of cells without compromising their stemness, particularly for novel stem cells. METHODS: In this study, we tested the efficiency of StemPro® MSC SFM Xeno Free (SFM-XF), a serum-free medium, for the long-term expansion of human fetal cartilage-derived progenitor cells (hFCPCs) from three donors in comparison to that of the conventional α-Modified Eagle's Medium (α-MEM) supplemented with 10% fetal bovine serum (FBS). RESULTS: We found that SFM-XF supported the expansion of hFCPCs for up to 28–30 passages without significant changes in the doubling time, while α-MEM with 10% FBS showed a rapid increase in doubling time at 10–18 passages depending on the donor. Senescence of hFCPCs was not observed until passage 10 in both media but was induced in approximately 15 and 25% of cells at passage 20 in SFM-XF and α-MEM with 10% FBS, respectively. The colony forming ability of hFCPCs in SFX-XF was also comparable to that in α-MEM with 10% FBS. hFCPCs expressed pluripotency genes like Oct-4, Sox-2, Nanog, SCF, and SSEA4 at passage 20 and 31 in SFM-XF; however, this was not observed when cells were cultured in α-MEM with 10% FBS. The ability of hFCPCs to differentiate into three mesodermal lineages decreased gradually in both media but it was less significant in SFM-XF. Finally we found no chromosomal abnormality after long-term culture of hFCPCs until passage 17 by karyotype analysis. CONCLUSION: These results suggest that SFM-XF supports the long-term expansion of hFCPCs without significant phenotypic and chromosomal changes. This study have also shown that hFCPCs can be mass-produced in vitro, proving their commercial value as a novel source for developing cell therapies.
Aging ; Cartilage* ; Cell- and Tissue-Based Therapy ; Chromosome Aberrations ; Humans* ; In Vitro Techniques ; Karyotype ; Mesoderm ; Stem Cells* ; Tissue Donors

BACKGROUND: The preservation of stem cell viability and characteristics during cell transport from the bench to patients can significantly affect the success of cell therapy. Factors such as suspending medium, time, temperature, cell density, and container type could be considered for transport conditions. METHODS: To establish optimal conditions, human amniotic fluid stem cells' (AFSCs) viabilities were analyzed under different media {DMEM(H), DMEM/F-12, K-SFM, RPMI 1640, α-MEM, DMEM(L), PBS or saline}, temperature (4, 22 or 37 ℃), cell density (1 × 10⁷ cells were suspended in 0.1, 0.5, 1.0 or 2.0 mL of medium) and container type (plastic syringe or glass bottle). After establishing the transport conditions, stem cell characteristics of AFSCs were compared to freshly prepared cells. RESULTS: Cells transported in DMEM(H) showed relatively higher viability than other media. The optimized transport temperature was 4 ℃, and available transport time was within 12 h. A lower cell density was associated with a better survival rate, and a syringe was selected as a transport container because of its clinical convenience. In compare of stem cell characteristics, the transported cells with established conditions showed similar potency as the freshly prepared cells. CONCLUSION: Our findings can provide a foundation to optimization of conditions for stem cell transport.
Amniotic Fluid ; Cell Count ; Cell Survival* ; Cell- and Tissue-Based Therapy ; Female ; Glass ; Humans ; Stem Cells* ; Survival Rate ; Syringes

BACKGROUND: Skin grafts are required in numerous clinical procedures, such as reconstruction after skin removal and correction of contracture or scarring after severe skin loss caused by burns, accidents, and trauma. The current standard for skin defect replacement procedures is the use of autologous skin grafts. However, donor-site tissue availability remains a major obstacle for the successful replacement of skin defects and often limits this option. The aim of this study is to effectively expand full thickness skin to clinically useful size using an automated skin reactor and evaluate auto grafting efficiency of the expanded skin using Yucatan female pigs. METHODS: We developed an automated bioreactor system with the functions of real-time monitoring and remote-control, optimization of grip, and induction of skin porosity for effective tissue expansion. We evaluated the morphological, ultra-structural, and mechanical properties of the expanded skin before and after expansion using histology, immunohistochemistry, and tensile testing. We further carried out in vivo grafting study using Yucatan pigs to investigate the feasibility of this method in clinical application. RESULTS: The results showed an average expansion rate of 180%. The histological findings indicated that external expansion stimulated cellular activity in the isolated skin and resulted in successful grafting to the transplanted site. Specifically, hyperplasia did not appear at the auto-grafted site, and grafted skin appeared similar to normal skin. Furthermore, mechanical stimuli resulted in an increase in COL1A2 expression in a suitable environment. CONCLUSION: These findings provided insight on the potential of this expansion system in promoting dermal extracellular matrix synthesis in vitro. Conclusively, this newly developed smart skin bioreactor enabled effective skin expansion ex vivo and successful grafting in vivo in a pig model.
Bioreactors ; Burns ; Cicatrix ; Contracture ; Extracellular Matrix ; Female ; Hand Strength ; Humans ; Hyperplasia ; Immunohistochemistry ; In Vitro Techniques ; Methods ; Models, Animal ; Porosity ; Skin Transplantation ; Skin* ; Swine ; Tissue Expansion ; Tissue Expansion Devices ; Transplants

BACKGROUND: We aimed to determine the effect of fibronectin (FN)-immobilized microgrooved titanium (Ti) on human gingival fibroblast proliferation, gene expression and protein expression. METHODS: Photolithography was used to fabricate the microgrooved Ti, and amine funtionalization (silanization) was used for FN immobilization on titanium surfaces. Cell proliferation, gene expression and protein expression were analyzed, followed by multiple regression analysis for determining the influential factors on cell proliferation. RESULTS: FN-immobilized microgrooved Ti significantly enhanced the fibroblast proliferation in various timelines of culture, among which a burst of fivefold increase is induced at 96 h of culture compared to that on the control smooth Ti. We suggest a presence of the synergistic promotion effect of microgrooves and FN immobilization on fibroblast proliferation. Through a series of analyses on the expression of various genes and proteins involved in cell adhesion and proliferation, cyclin-dependent kinase 6, cyclin D1, integrin α5, oncogene c-Src, osteonectin, paxillin and talin-2 were determined as influential factors on promoting fibroblast proliferation induced by FN-immobilized microgrooved Ti. CONCLUSION: FN-immobilized microgrooved Ti can act as an effective surface for enhancing fibroblast proliferation, and can be used for promoting soft tissue response on the connective tissue attachment zone of biomaterial surfaces.
Cell Adhesion ; Cell Proliferation* ; Connective Tissue ; Cyclin D1 ; Cyclin-Dependent Kinase 6 ; Fibroblasts* ; Fibronectins ; Gene Expression ; Humans* ; Immobilization ; Oncogenes ; Osteonectin ; Paxillin ; Titanium*

BACKGROUND: Sand blasted titanium (Ti) is commonly used in designing endosseous dental implants due to its biocompatibility and ability to form bonds with bone tissues. However, titanium implants do not induce strong interactions with teeth bones. To increase strong interactions between Ti disk implants and teeth bones, the L-glutamic acid grafted hydroxyapatite nanorods (nHA) were immobilized on albumin modified Ti disk implants (Ti-Alb). METHODS: For modification of Ti disk implants by nHA, the L-glutamic acid grafted nHA was synthesized and then immobilized on albumin modified Ti disk implants. Fourier transformed infrared spectroscopy, X-ray photoelectron spectroscopy, scanning electron microscope; energy dispersive spectroscopy and confocal laser scanning microscopy were used to confirm the modification of Ti disk implants. The bioactivity of nHA-modified Ti disk implants was evaluated by seeding MC3T3-E1 cells on Ti-nHA implants. RESULTS: Characterization techniques have confirmed the successful modification of Ti disk implants by L-glutamic acid grafted nHA. The nHA-modified Ti disk implants have shown enhanced adhesion, proliferation and cytotoxicity of MC3T3-E1 cells in comparison to pristine Ti implants. CONCLUSION: The modification of Ti implants by L-glutamic acid grafted nHA has produced highly osteogenic Ti disk plants in comparison to pristine Ti disk implants due to the formation of bioactive surfaces by hydroxyapatite nano rods on Ti disk implants. Ti-nHA disk implants showed enhanced adhesion, proliferation, and MC3T3-E1 cells viability in comparison to pristine Ti disk implants. Thus nHA might be to be useful to enhance the osseointegration of Ti implants with teeth bones.
Bone and Bones ; Dental Implants* ; Durapatite* ; Fourier Analysis ; Glutamic Acid ; Microscopy, Confocal ; Nanotubes ; Osseointegration ; Photoelectron Spectroscopy ; Spectrum Analysis ; Titanium* ; Tooth ; Transplants

BACKGROUND: The purpose of this study is to examine physical characteristics of and initial biological properties to anodized titanium treated with poly(D,L-lactide-co-glycolide) (PLG) mixed with recombinant human bone morphogenic protein-2 (rhBMP-2). METHODS: Titanium specimens were prepared in groups of four as follows: group NC was anodized under 300 V as control; group PC was anodized then dropped and dried with solution 0.02 ml PLG; group D was anodized then dropped and dried with solution 0.02 ml PLG/rhBMP-2 (3.75 µg per disc); and group E was anodized then coated with 0.02 ml PLG/rhBMP-2 (3.75 µg per disc) by electrospray. Human osteoblastic-like sarcoma cells were cultured. Cell proliferation and alkaline phosphatase (ALP) activity test were carried out. Runx-2 gene was investigated by the reverse transcription-polymerase chain reaction. Immunofluorescence outcome of osteogenic proteins was observed. RESULTS: After 3 days, there were significantly higher proliferations compared rhBMP-2 loaded titanium discs with rhBMP-2 unloaded discs. The ALPase activity on rhBMP-2 loaded titanium discs was significantly higher than in rhBMP-2 unloaded discs. The expression level of Runx2 mRNA presented the highest on the PLG/rhBMP-2-coated surface. CONCLUSION: PLG polymers mixed with rhBMP-2 might improve proliferation, differentiation and osteogenic protein formation of cells on the anodized titanium.
Alkaline Phosphatase ; Cell Differentiation ; Cell Proliferation ; Fluorescent Antibody Technique ; Humans ; Polymers ; RNA, Messenger ; Sarcoma ; Titanium*

BACKGROUND: Biopolymeric in situ hydrogels play a crucial role in the regenerative repair and replacement of infected or injured tissue. They possess excellent biodegradability and biocompatibility in the biological system, however only a few biopolymeric in situ hydrogels have been approved clinically. Researchers have been investigating new advancements and designs to restore tissue functions and structure, and these studies involve a composite of biometrics, cells and a combination of factors that can repair or regenerate damaged tissue. METHODS: Injectable hydrogels, cross-linking mechanisms, bioactive materials for injectable hydrogels, clinically applied injectable biopolymeric hydrogels and the bioimaging applications of hydrogels were reviewed. RESULTS: This article reviews the different types of biopolymeric injectable hydrogels, their gelation mechanisms, tissue engineering, clinical applications and their various in situ imaging techniques. CONCLUSION: The applications of bioactive injectable hydrogels and their bioimaging are a promising area in tissue engineering and regenerative medicine. There is a high demand for injectable hydrogels for in situ imaging.
Biopolymers* ; Hydrogel* ; Hydrogels* ; Regenerative Medicine ; Tissue Engineering*

BACKGROUND: Injectable biomaterials have attracted increasing attention for volume restoration and tissue regeneration. The main aim of this review is to discuss the current status of the injectable biomaterials for correction of tissue defects in plastic and reconstructive surgery. METHODS: Requirements of injectable biomaterials, mechanism of in situ gelation, characteristics, and the combinational usage of adipose-derived stem cells (ADSCs) and growth factors were reviewed. RESULTS: The ideal injectable biomaterials should be biocompatible, non-toxic, easy to use, and cost-effective. Additionally, it should possess adequate mechanical properties and stability. In situ gelation method includes physical, chemical, enzymatic and photo-initiated methods. Natural and synthetic biomaterials carry their pros and cons due to their inherent properties. The combined use of ADSCs and growth factors provides enhanced potential for adipose tissue regeneration. CONCLUSION: The usage of injectable biomaterials has been increasing for the tissue restoration and regeneration. The future of incorporating ADSCs and growth factors into the injectable biomaterials is promising.
Adipose Tissue ; Biocompatible Materials* ; Intercellular Signaling Peptides and Proteins ; Methods ; Plastics* ; Regeneration ; Stem Cells