OBJECTIVETo explore an effective method to culture oral mucosa epithelial cells (OMECs) of canine in vitro, and to observe the biological characteristics of OMECs growing on small intestinal submucosa (SIS) in order to provide the experimental basis for epithelium tissue engineering.

METHODSThe primary OMECs were cultivated with DKSFM (defined keratinocyte serum free medium) containing 6% fetal bovine serum (FBS). The morphological characteristics and the growth curve of OMECs were observed. The expressions of OMECs marker (CK19) were examined by immunocytochemistry. The 2nd passage of OMECs were seeded on SIS, OMECs co-cultured with SIS were observed by hematoxylin-eosin staining, immunohistochemical staining, and scanning electron microscope (SEM).

RESULTSOMECs were grown well in DKSFM. Immunohistochemical staining of the 2nd passage cultured canine OMECs with broadly reacting anti-cytokeratin anyibodies (CK19) was positive. OMECs formed a single layer on the surface of SIS, and eight days later the cells were polygong and arranged like slabstone.

CONCLUSIONCulture of canine OMECs in DKSFM containing 6% FBS is a simple and feasible method. SIS has good biocompatibility, it is a kind of good bioscafold in the tissue-engineered epithelium.

Animals ; Cattle ; Cell Culture Techniques ; Cells, Cultured ; Coculture Techniques ; Epithelial Cells ; In Vitro Techniques ; Intestine, Small ; Mouth Mucosa ; Tissue Engineering

Advancement in cell culture protocols, multidisciplinary research approach, and the need of clinical implication to reconstruct damaged or diseased tissues has led to the establishment of three-dimensional (3D) test systems for regeneration and repair. Regenerative therapies, including dental tissue engineering, have been pursued as a new prospect to repair and rebuild the diseased/lost oral tissues. Interactions between the different cell types, growth factors, and extracellular matrix components involved in angiogenesis are vital in the mechanisms of new vessel formation for tissue regeneration. In vitro pre-vascularization is one of the leading scopes in the tissue-engineering field. Vascularization strategies that are associated with co-culture systems have proved that there is communication between different cell types with mutual beneficial effects in vascularization and tissue regeneration in two-dimensional or 3D cultures. Endothelial cells with different cell populations, including osteoblasts, smooth muscle cells, and fibroblasts in a co-culture have shown their ability to advocate pre-vascularization. In this review, a co-culture perspective of human gingival fibroblasts and vascular endothelial cells is discussed with the main focus on vascularization and future perspective of this model in regeneration and repair.
Cell Culture Techniques ; Coculture Techniques* ; Endothelial Cells* ; Extracellular Matrix ; Fibroblasts* ; Humans* ; In Vitro Techniques ; Intercellular Signaling Peptides and Proteins ; Myocytes, Smooth Muscle ; Osteoblasts ; Regeneration ; Tissue Engineering

OBJECTIVETo culture human dental papilla cells (HDPCs)and to study its cytobiological characters in vitro.

METHODSHDPCs were isolated and cultured with explant culture technique in vitro; Type I collagen, fibronection and laminin were detected in HDPCs and its secreted matrix with the immunocyto-chemical stain; HDPCs were incubated in mineralized promoting solution containing 10 mmol/L beta-glycerophosphate, 100 mg/L of ascorbic acid and 10 nmol/L dexamethasone supplemented with 10% FBS and the form of mineralized nodules was tested with Alizarin Red S stainning.

RESULTSCultured HDPCs in vitro were well growing in DMEM/F12. Type I collagen, fibronection and laminin staining were all positive in both HDPCs and its secreted matrix, and laminin was stained with bunchiness in matrix. Mineralized nodules formed after cultured 27 days by Alizarin Red S stainning.

CONCLUSIONHDPCs isolated and cultured are well growing in vitro, have a capability of synthesizing and secreting matrix and in mineralized promoting solution, are able to form mineralizer, so, HDPCs have a capacity of seed cell of tissue engineering regeneration tooth.

Cell Culture Techniques ; Cells, Cultured ; Collagen Type I ; Dental Papilla ; Dexamethasone ; Glycerophosphates ; Humans ; In Vitro Techniques ; Tissue Engineering ; Tooth

In tissue engineering, alginate has been an attractive material due to its biocompatibility and ability to form hydrogels, unless its uncontrollable degradation could be an undesirable feature. Here, we developed a simple and easy method to tune the degradation profile of the fibrous alginate scaffolds by the microfluidic wet spinning techniques, according with the use of isopropyl alcohol for dense packing of alginate chains in the microfiber production and the increase of crosslinking with Ca²⁺ ion. The degradation profiling was analyzed by mass losses, swelling ratios, and also observation of the morphologic changes. The results demonstrated that high packing density might be provided by self-aggregation of polymer chains through high dipole interactions between sheath and core fluids and that the increase of crosslinking rates could make degradation of alginate scaffold controllable. We suggest that the tunable degradation of the alginate fibrous scaffolds may expand its utilities for biomedical applications such as drug delivery, in vitro cell culture, wound healing, tissue engineering and regenerative medicine.
2-Propanol ; Cell Culture Techniques ; Hydrogel ; Hydrogels ; In Vitro Techniques ; Methods ; Microfluidics* ; Polymers ; Regenerative Medicine ; Tissue Engineering ; Wound Healing

BACKGROUND: Tissue engineering is a multidisciplinary field which attracted much attention in recent years. One of the most important issue in tissue engineering is how to obtain high cell numbers and tissue regeneration while maintaining appropriate cellular characteristics in vitro for restoring damaged or dysfunctional body tissues and organs. These demands can be achieved by the use of three dimensional (3D) dynamic cultures of cells combined with cell-adhesive micro-carriers. METHODS: In this study, human mesenchymal stem cells (hMSCs) were cultured in a wave-bioreactor system for up to 100 days, after seeding on Cultisphere-S porous gelatin micro-carriers. Cell counting was performed at the time points of 7, 12, 17, 31 days and compared to those of hMSCs cultured under static condition. Higher growth and proliferation rates was achieved in wave-type dynamic culture, when cell culture continued to day 31. A scanning electron microscope (SEM) photographs, both live and dead and MTT assays were taken to confirm the survival and distribution of cells on porous gelatin micro-carrier surfaces. The results of histological stains such as hematoxylin and eosin, Masson’s trichrome, Alcian blue and Alizarin red S also showed improved proliferation and tissue regeneration of hMSCs on porous gelatin micro-carriers. CONCLUSION: The experimental results demonstrated the effect and importance of both micro-carriers and bioreactor in hMSC expansion on cell proliferation and migration as well as extracellular matrix formation on the superficial and pore surfaces of the porous gelatin micro-carriers, and then their inter-connections, leading to tissue regeneration.
Alcian Blue ; Bioreactors ; Cell Count ; Cell Culture Techniques ; Cell Proliferation ; Coloring Agents ; Eosine Yellowish-(YS) ; Extracellular Matrix ; Gelatin ; Hematoxylin ; Humans ; In Vitro Techniques ; Mesenchymal Stromal Cells ; Regeneration ; Tissue Engineering

The strategy used to generate tissue-engineered bone construct, in view of future clinical application is presented here. Osteoprogenitor cells from periosteum of consenting scoliosis patients were isolated. Growth factors viz TGF-B2, bFGF and IGF-1 were used in concert to increase cell proliferation during in vitro cell expansion. Porous tricalcium phosphate (TCP)-hydroxyapatite (HA) scaffold was used as the scaffold to form 3D bone construct. We found that the addition of growth factors, greatly increased cell growth by 2 to 7 fold. TCP/HA proved to be the ideal scaffold for cell attachment and proliferation. Hence, this model will be further carried out on animal trial.
Bone Regeneration/*physiology ; *Bone Transplantation ; Cell Division/physiology ; Collagen/metabolism ; *Mesenchymal Stem Cell Transplantation ; Organ Culture Techniques ; Periosteum/*cytology ; Tissue Engineering/*methods

Chitosan has similar structure to glycosaminoglycans in the tissue, thus may be a good candidates as tissue engineering scaffold. However, to improve their cell attachment ability, we try to incorporate this natural polymer with collagen by combining it via cross-linking process. In this preliminary study we evaluate the cell attachment ability of chitosan-collagen scaffold versus chitosan scaffold alone. Chitosan and collagen were dissolved in 1% acetic acid and then were frozen for 24 hours before the lyophilizing process. Human skin fibroblasts were seeded into both scaffold and were cultured in F12: DMEM (1:1). Metabolic activity assay were used to evaluate cell attachment ability of scaffold for a period of 1, 3, 7 and 14 days. Scanning electron micrographs shows good cell morphology on chitosan-collagen hybrid scaffold. In conclusion, the incorporation of collagen to chitosan will enhance its cell attachment ability and will be a potential scaffold in tissue engineering.
Cell Adhesion/*physiology ; *Chitosan ; *Collagen ; Energy Metabolism/physiology ; Fibroblasts/cytology ; Microscopy, Electron, Scanning ; Organ Culture Techniques/*methods ; *Polymers ; Tissue Engineering/*methods

Patient own fibrin may act as the safest, cheapest and immediate available biodegradable scaffold material in clinical 1 tissue engineering. This study investigated the feasibility of using patient own fibrin isolated from whole blood to construct a new human cartilage, skin and bone. Constructed in vitro tissues were implanted on the dorsal part of the nude mice for in vivo maturation. After 8 weeks of implantation, the engineered tissues were removed for histological analysis. Our results demonstrated autologous fibrin has great potential as clinical scaffold material to construct various human tissues.
*Biocompatible Materials ; *Bone Transplantation ; Cartilage/*transplantation ; Cell Division/physiology ; Culture Media ; *Fibrin ; Fibroblasts/cytology ; Mesenchymal Stem Cells/cytology ; Mice, Nude ; Organ Culture Techniques ; Periosteum/cytology ; *Skin Transplantation ; *Tissue Engineering

Cellular mechanics is a branch of tissue engineering and cellular engineering. As one of the important method, loading different mechanical stimuli to culturing cells in vitro so as to study the influence that the stress has on the cells is one of the important fields of cellular mechanics. This paper reviews the experimental methods for mechanically stimulating the cells in vitro, according to the different loading modalities, the methods can be categorized into micropipette aspiration, compression loading, substrate distention, fluid shear, etc. And it also points out their advantages and disadvantages.
Cell Culture Techniques ; Cells, Cultured ; Humans ; Stress, Mechanical ; Tissue Engineering

As one of the top 10 breakthrough and emerging technologies in the world in 2018, cultured meat has attracted extensive attention due to its advantages of traceable origin, food safety and green sustainable development. Europe and the United States have invested a lot of resources to focus on research about cultured meat, which will affect our domestic meat and food market in the future. At present, the challenge of cultured meat production is how to efficiently simulate the growth environment of animal muscle tissue and realize large-scale production in bioreactor. Although cell tissue engineering has been deeply studied and achieved varying successful application, it is still difficult to obtain large-scale cultured meat production due to the high cost and technical requirements. Therefore, the development of efficient and safe cell culture technology is an urgent problem for large-scale cultured meat production, which can effectively reduce costs and achieve industrial application. In this review, we summarize the research progress of animal cell tissue culture technology used for cultured meat, and highlighted the current challenges and possible strategies in further applications.
Animals ; Bioreactors ; Cell Culture Techniques ; Meat ; Tissue Engineering ; United States