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

Tissue engineering applies the principle of engineering and life sciences towards the development of biological substitute that restore, maintain or improve tissue or organ function. Scientists grow tissues or organs in vitro and implant them when the body is unable to prompt into healing itself. This presentation aims to highlight the potential clinical application of engineered tissues being researched on at the Tissue Engineering Centre, Universiti Kebangsaan Malaysia Medical Centre.

The emergence of tissue engineering and stem cell research has created a tremendous response amongst scientist in Malaysia. However, despite the enthusiastic to embark on the research we have to carefully divert the research towards our needs. This is due to our responsibility to address the mounting problem of communicable diseases here and a very limited funding. As commercialization is a key objective the combination of products towards treating or diagnosing communicable and non-communicable diseases in the developing country is another important factor. The discussion here is mainly on the evolution of tissue engineering in Malaysia and taking a model of tissue engineering in otolaryngology.

Skin is the largest organ in human system and plays a vital role as a barrier against environment and pathogens. Skin regeneration is important in tissue engineering especially in cases of chronic wounds. With the tissue engineering technology, these skins equivalent have been use clinically to repair burns and wounds. Consented redundant skin samples were obtained from patients aged 9 to 65 years old. Skin samples were digested with dispase, thus separating the epidermis and the dermis layer. The epidermis layer was trypsinized and cultured in DKSFM in 6-well plate at 37 degrees C and 5% CO2. Once confluent, the culture were trypsinized and the cells were pooled. Cells were counted using haemacytometer. Doubling time and viability were calculated and analysed. From the result, we conclude that doubling time and viability of in vitro keratinocytes cultured in DKSFM media is not age dependant.
Age Factors ; Burns/physiopathology ; Burns/*therapy ; Cell Aging/*physiology ; Cell Division/physiology ; Cell Survival/physiology ; Chronic Disease ; Keratinocytes/*cytology ; *Skin Transplantation ; Statistics ; Tissue Engineering/*methods ; Wound Healing/physiology ; Wounds and Injuries/physiopathology ; Wounds and Injuries/*therapy

Human adipose-derived stem cells (HADSC) have demonstrated the capacity of differentiating into bone depending on the specific induction stimuli and growth factors. However, investigation on stem cell characteristic after osteogenic differentiation is still lacking. The goal of this study was to investigate the differential expression of sternness and osteogenic genes in non-induced HADSC compared with HADSC after osteogenic induction using quantitative Real Time RT-PCR. Our results showed that OCT-4, REX-1, FZD9, OSC, RUNX, and ALP were up regulated after osteogenic induction. This may indicated that HADSCs after osteogenic induction still possessed some stemness properties.

The regulation roles of insulin-like growth factor-1 (IGF-1) with basic fibroblast growth factor (bFGF) and transforming growth factor beta 2 (TGFbeta2) in human nasal septum chondrocytes monolayer culture and cartilage engineering was investigated in this study. The role of IGF-1 with bFGF and TGFbeta2 was investigated by measuring chondrocyte growth kinetic and collagen genes expression. IGF-1 together with bFGF and TGFbeta2 promote cartilage tissue engineering, increase type II collagen expression and enhance the histological features of engineered cartilage.
Cartilage/*transplantation ; Cell Division/physiology ; Chondrocytes/*cytology ; Collagen Type II/genetics ; Culture Media, Serum-Free ; Gene Expression/physiology ; Growth Substances/*physiology ; Insulin-Like Growth Factor I/*physiology ; Tissue Engineering/*methods

Bone marrow harvested by aspiration contains connective tissue progenitor cells which can be selectively isolated and induced to express bone phenotype in vitro. The osteoblastic progenitor can be estimated by counting the number of cells attach using the haemacytometer. This study was undertaken to test the hypothesis that human aging is associated with a significant change on the number of osteoblastic progenitors in the bone marrow. Bone marrow aspirates were harvested from 38 patients, 14 men (age 11-70) and 24 women (age 10-70) and cultured in F12: DMEM (1:1). In total 15 bone marrow samples have been isolated from patients above 40 years old (men/women) of age. Fourteen (93.3%) of this samples failed to proliferate. Only one (6.7%) bone marrow sample from a male patient, aged 59 years old was successfully cultured. Seventy percent (16/23) of the samples from patient below than 40 years old were successfully cultured. However, our observation on the survival rate for cells of different gender from patient below 40 years old does not indicate any significant difference. From this study, we conclude that the growth of bone marrow stromal cells possibly for bone engineering is better from bone marrow aspirates of younger patient.
Age Factors ; Bone Marrow Cells/*cytology ; Cell Aging/*physiology ; Cell Division/*physiology ; Cell Survival/physiology ; Mesenchymal Stem Cells/*cytology ; Osteoblasts/*cytology ; Sex Factors ; Stromal Cells/cytology ; *Tissue Engineering

The angiogenic potential of native skin (NS), keratinocytes single skin equivalent (SSE-K), fibroblasts single skin equivalent (SSE-F) and bilayered skin equivalent secreting angiogenic growth factors such as transforming growth factor beta1 (TGF-beta1), vascular endothelial growth factor (VEGF), keratinocyte growth factor (KGF) and basic fibroblast growth factor (bFGF) in the in vitro systems at 24, 48, 72 hours and 7 days was compared using Enzyme-Linked Immunosorbent Assay (ELISA). Bilayered skin equivalent exhibit highest release of growth factors within 24 hours to 7 days of culture compared to NS, SSE-K and SSE-F. This proved the potential of bilayered skin equivalent in producing and sustaining growth factors release to enhance angiogenesis, fibroblasts proliferation, matrix deposition, migration and growth of keratinocytes.

Bone marrow derived Mesenchymal stem cells (MSCs) were evaluated as an alternative source for tissue engineering of peripheral nerves. Human MSCs were subjected to a series of treatment with a reducing agent, retinoic acid and a combination of trophic factors. This treated MSCs differentiated into Schwann cells were characterized in vitro via flow cytometry analysis and immunocytochemically. In contrast to untreated MSCs, differentiated MSCs expressed Schwann cell markers in vitro, as we confirmed by flow cytometry analysis and immunocytochemically. These results suggest that human MSCs can be induced to be a substitute for Schwann cells that may be applied for nerve regeneration since it is difficult to grow Schwann cells in vitro.

This study was conducted to explore the feasibility of culturing conjunctiva epithelial cells in serum-free and feeder layer-free culture system with regard to the cell morphology and immunocytochemistry of the rabbit bulbar, fornix and palpebral conjunctiva epithelia. The results showed that epithelium cells from all the three conjunctiva regions can be cultured in a serum-free and feeder layer-free environment. We obtained highest epithelial growth from fornix region with minimum invasion of fibroblast cells compared to other area. All cultured cells were stained positive for cytokeratin 19 and MUC5AC and negative for cytokeratin 3. These findings suggested that fornix was a better source of cells for the development of tissue engineered conjunctiva for future clinical application.