Effect of Serum Media on Fibroblast Proliferation and Collagen Synthesis

Expanding cells ex-vivo is very important in tissue- engineering. Culture medium is usually supplemented with fetal bovine serum(FBS) in most of the experiments. However, cells grown in bovine serum media may posses the possibilities of disseminating bovine diseases and/or stimulating the patient's immune reactions. To overcome these problems, autologous or homologous serum should be used instead of the FBS. The purpose of this study is to compare cell proliferation and collagen synthesis depending on the kind of sera mixed on media and to provide a guideline on applying established experimental data to clinical cases. Human dermal fibroblasts were obtained from four patients. Five thousand cells per well in 96-well plates were incubated DMEM/F- 12 Nutrient with varying serum mixture; 10% autologous serum, 10% homologous serum, and 10% FBS. Five days after incubation fibroblast proliferation and collagen production were determined by MTT assay and CICP enzyme immunoassay. The mean cell number were; 3.95x104/well, 2.97x104/well and 2.30x104/well, respectively. The average amounts of collagen synthesized were; 238.13ng/ml, 204.88ng/ml, and 163.88ng/ml in each. These results show that the use of human serum mixture may contribute to, not only preventing disseminated infection of bovine diseases. but also increase cell proliferation and collagen synthesis without simulating the patient's immune reactions.

Effect of Granulocyte-macrophage Colony-stimulating Factor and Ascorbic Acid Co-supplementation on the Fibroblast Proliferation and Collagen Synthesis

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is naturally generating protein that has the healing effect in normal wounds as well as infected nonhealing wounds and ulcers. Ascorbic acid has also been known to stimulate fibroblast stimulation and collagen synthesis. However, there are no reports about the effect of GM-CSF and ascorbic acid co-supplementation in wound healing. Therefore, in this report, we examined the effect of GM-CSF and ascorbic acid co-supplementation on the proliferation of human dermal fibroblasts and collagen synthesis which play a crucial role in wound healing process in vitro. To determine an optimal GM-CSF and ascorbic acid concentration for human dermal fibroblast proliferation and collagen synthesis, the cells were incubated with various concentrations of GM-CSF and ascorbic acid. The activity of fibroblast proliferation was determined by MTT assay. To measure the amount of collagen production, the collagen type I carboxy-terminal propeptide enzyme immunoassay was used. The best fibroblast proliferation was observed at the co-supplementation of 0.5 microgram/ml GM-CSF with 25 microgram/ml ascorbic acid and 1 microgram/ml GM-CSF with 12.5 microgram/ml ascorbic acid. Maximal stimulation of collagen synthesis was observed at the co-supplementation of 1 microgram/ml GM-CSF with 12.5 microgram/ml ascorbic acid. The collagen synthesis per cell was also maximal at concentration of 1 microgram/ml GM-CSF with 12.5 microgram/ml ascorbic acid. This results demonstrates that GM-CSF and ascorbic acid co-supplementation increases human dermal fibroblast proliferation and collagen synthesis and the concentration is the critical factor in vitro.

Changes in Carpal Tunnel Pressure Following Increments of the Carpal Tunnel Release

Complete division of transverse carpal ligament (TCL) is accepted as a standard procedure for carpal tunnel syndrome (CTS). In some cases however, after complete release of TCL, the loss of grip and pinch power occurs frequently. Previous study revealed that normal intracanal pressure is 25-30mmHg, therefore There were the possibility that complete division of TCL is an over-correction for CTS. The purpose of this experiment was to measure the carpal tunnel pressure during the incremental division of TCL. Twenty patients who were confirmed as CTS were selected from September 2002 to February 2003. By step-wise division of TCL, the pressure was obtained for 3 times serially. Comparative pressure changes between the pre-division and post-division were analyzed by SRM (standard responsiveness mean) and ES (effect size). In Carpal tunnel syndrome, the mean intracanal pressure was 215.5mmHg. After partial release of the portion which was revealed as most severely compressed area on inching test, the carpal pressure decreased to 70.6 mmHg. Releasing the remaining portion of TCL resulted in another 30% decompression. Effect size of partial release was 2.86, and after release of the remaining portion of TCL the effect size was 1.19. Comparing the effect size, as much effect as 70% of the total release (4.15) can be obtained by partial release.