OBJECTIVE :To investigale effect of subretinal fluld(SRF)on proliferalion of the cellular elements of PVR. METHOD:The effect of SRF of 28 patients with rhegmatogenous retinal detachment proliferation of the cultured human retinal pigment epithelial cells(RPE),retinal glial cells (RG),and fibroblast (FB)was observed and detected by the methods of cell-counting and 3H-TdR in DNA synthesis. RESULTS:The range of proliferatinn-stimulating activity was 52.5%~233.3%, 36.4% ~ 177.8%，55.4% ~277.8% above the baseline in 1:10 dilution of these 3 kinds ,of cellular elements,and there was no significant difference among them. CONCLUSION ;The stimulating effect of SRF on the cellular proliferation was thougt to be due to the actions from certain growth factors. （Chin J Ocul Fundus Dis,1996,12: 233-235）
OBJECTIVE: To investigate the changes of fibroblast growth factor (FGF) in burn wounds. METHODS: The FGF expression in the center of wound granulation, the edge of wound, the healed part of wound, the normal skin of patients, and the heal course of second degree burn wounds were detected by immunohistochemical methods. RESULTS: The expression intensity of FGF was different in the different sites of third degree burn wounds. The highest contents of FGF was in the center granulation of burn wounds, the less was in the borderline of wound and healed skin, and the least was in the healed skin. FGF expression mainly concentrated in the middle layer of wound, and almost no FGF expression in normal skin. The most FGF expression was occurred at 14 days after injury in second degree of burn wound. CONCLUSION: The changes of FGF in wounds are closely related to the wound healing, and rational use of FGF can promote wound healing.
OBJECTIVE: To build the trestle of tissue engineering for skin with the collagen. METHODS: The collagen was obtained from the baby cattle hide pretreated by Na2S and elastinase and Protease M, then the collagen was dissolved in 0.5 mol/L acetic acid solution. The collagen was treated with Protease N to minimize its immunogenicity. The resulting collagen could be used to build the trestle of tissue engineering for skin because of good biocompatibility. The collagen molecular weight and structure were analyzed by SDS-PAGE. The bioactivity of trestle was tested in the experiment of the mice wound healing and the cell implantation. RESULTS: The SDS-PAGE result of the collagen treated by Protease M showed the typical spectrum of type I collagen. The built trestle was a collagen sponge matrix in which micropore size was 50-200 microns. It could accelerate wound healing and the implanted fibroblasts could proliferate well. CONCLUSION: The collagen treated by Protease N can get good biocompatibilily and is suitable for building the trestles of tissue engineering for skin with good bioactivity.
To investigate the best way of using fibroblast growth factor (FGF) in wound healing, the following experiments were performed. Twelve Wistar rats were chosen. Four 1.5 cm x 1.5 cm middle-thick skin wounds were made in the back of each rat, 2 in each side, and labelled as number 1 to 4. Number 1 wound of each rat was used as control, only PBS was applied to the wound, 50 microliters per time, twice a day from the first day to 11th day. Number 2 wound was sustained medication group, 50 microliters 4 micrograms/ml FGF was applied twice a day from the first day to 11th day; Number 3 wound was early medication group, 50 microliters 8 micrograms/ml FGF was applied twice a day from the first day to 5th day; Number 4 wound was late medication group, 50 microliters 8 micrograms/ml FGF was added twice a day from the 5th day to 11th day. By day 4, 8, 12 and 16, the area of wounds were measured, and the healing time of each wound was recorded. The elastic fiber, collagen fiber and DNA content were measured by immunohistological method. The result showed that the elastic fiber, collagen fiber and DNA content in the groups of FGF used were more than those in the control group. The healing time of the control group was 14.4 days while that of the early meduation group was 13.4 days, late medation group was 13.5 days and sustained medication group was 12.2 days. It was suggested that FGF could accelerate the wound healing, and sustained use of FGF was the best way of giving the drug.
Taking Wistar rats and pigs as models, the basic fibroblast growth factor (bFGF) was studied on wound healing. Five rats and five pigs were used in the experiment. Each rat had 2 roundshapedwound (1.8cm in diameter) and each pig had 4 wounds of the same size as that ofthe rat. One wound of each rat and 2 wound of each pig were dressed with bFGF saline (60U/cm2). While the other wounds of the rats and pigs were dressed with normal saline as control. The wounds treated with bFGF were completely covered with granulation tissue on the 7th day after injury, and in 14 days the wounds were almost completely covered by epithelium. The bFGF would enhance the growth of theepithelial cells, reepithelization of the wound and the tensile strength of thetissue. It was concluded that the bFGF could promote skin regeneration, whichmight be the direct action of bFGF on the bFGF receptors in the basic cells of skin.
Objective To investigate the phenotypic change and proliferation of fibroblasts in human inflammatory strictured bile duct wall. Methods We observed the density and ultrastructure of fibroblasts, and the histologic structure in human normal bile duct wall and inflammatory strictured bile duct wall by light and electron microscope.Results The results showed that fibroblasts were the main source of extracellular matrix production in bile duct wall. The phenotype of fibroblasts in inflammatory strictured bile duct wall changed obviously, quiescent fibroblasts were activated and transformed to myofibroblasts, with massive proliferation. Conclusion These data suggest that massive proliferation of activated fibroblasts and myofibroblasts is the main source of extracellular matrix overproduction which results in inflammatory bile duct stricture.
Objective To compare the efficiency of epidermis cell culture between big graft method and small strip method. Methods The big graft method was to cut the skin tissue reticularly from dermis layer while the epidermis were not cut off. After it was digested fully in trypsin, theepidermis was separated from skin and was used to culture epidermal cells. The small strip method was routine. The time to cut the skin and to separate the epidermis was recorded, and the number and quality of cells were compared between two methods. Results It took 8-10 minutes to cut an area of 5 cm2 skin into small strips and 1-2 minutes into big grafts. It took 10-15 minutes to separate the epidermis from the same area skin by small strip method and 2 minutes by big graft method. The cells showed better vigor and its number was more by big grafts than by small strips.The chance of fibroblast contamination was reduced obviously. Conclusion The big graft method is simpler than the small strip method and can culture more epidermis cells with less chance of fibroblast contamination.
Objective To investigate the effects of bursopentin ( BP5) on expression of extracellular matrix in human lung fibroblasts ( HLFs) and its mechanism.Methods HLFs were cultured in vitro and divided into five groups. The cells in the control group were cultured in DMEMwithout TGF-β1 or BP5. The cells in TGB-β1 treatment group were cultured in DMEMcontaining 5 μg/L TGF-β1 . While in three TGF-β1 + BP5 treatment groups, the cells were cultured in DMEM containing 5 μg/L TGF-β1 and simultaneously intervened with BP5 at three different concentrations ( 2. 5 μg/mL, 5 μg/mL, and 10 μg/mL respectively) . The expression of α-SMA was detected using a fluorescent-labeling strategy. The expressions of Collagen-Ⅰ, p-Smad2/3, p-Smad3, and Smad7 proteins were measured by Western blot. Results The cells in the TGF-β1 treatment group showed positive expression of α-SMA, implying TGF-β1 had induced fibroblasts to differentiate into myofibroblasts. In the TGF-β1 treatment group, the expressions of collagen-Ⅰ( 1. 402 ±0. 158 vs. 0. 605 ±0. 367) , p-Smad2/3 ( 1. 457 ±0. 111 vs. 0. 815 ±0. 039) , and p-Smad3 ( 1. 320 ±0. 147 vs. 0. 623 ±0. 128) increased with statistical significance ( P lt; 0. 01) . Meanwhile the expression of Smad7 reduced ( 0. 614 ±0. 107 vs. 0. 865 ±0. 063, P lt;0. 05) . But in the TGF-β1 + BP5 treatment groups, over-expressions of collagen-Ⅰ, α-SMA, p-Smad2 and p-Smad3 induced by TGF-β1 were obviously inhibited by BP5, especially at the BP5 concentration of 10 μg/mL ( collagen-Ⅰ: 0. 718 ±0. 049 vs. 1. 402 ±0. 158; p-Smad2 /3: 0. 696 ±0. 031 vs. 1. 457 ±0. 111; p-Smad3: 0. 766 ±0. 006 vs. 1. 320 ±0. 147; all P lt; 0. 01) . Otherwise, the up-regulation of Smad7 ( 1. 237 ±0. 173 vs. 0. 614 ±0. 107) was found.Conclusions Bursopentin can reduce the expressions of collagen-Ⅰ and α-SMA protein of fibroblast stimulated by TGF-β1 , maybe through inhibiting TGF-β1 /Smads transduction pathway. It is suggested that bursopentin may have intervention effect on pulmonary fibrosis.
Objective To establish a method of constructing skin-equivalents (SE) by the hair follicle stem cells (HFSC) and the fibroblasts. Methods The K19 immunostainning was employed to localize the HFSC in the human scalp from the cosmetic surgery. The isolated HFSC through the enzyme digestion were seeded on the dermal equivalent (DE) formed by polymerization of the fibroblasts and collagen. After being cultured between the air-liquid interface for 14 days, SE were harvested and used for an evaluation. Results HFSC were located mainly in the outer root sheath in the hair follicle. Based on DE, the growing HFSC could build a fullydeveloped and multilayered epidermis with the basal membrane formedb etween the epidermis and the dermis. The fibroblasts were active and spread evenly in the collagen matrix. Conclusion The hair follicle stem cells located in the outer root sheath can be successfully used to construct skin-equivalents in vitro and have a promising clinical use in the treatment.
Porpose To investigate the optimal concentration of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) on DNA synthesis and their synergism indensity arrested human retinal pigment epithelial (RPE) cells. Methods Growth factor effects in cultured human RPE of the 6th generation were assessed by [3 H]-thymidine incorporation and radioautography. Results EGF and bFGF were potent stimulators when used alone,and their optimal concentrations were 10ng/ml in DMEM and 1ng/ml in 2% serum DMEM.When used in combination (10ng/ml EGF and 10ng/ml bFGF),they caused a significant enhancement of [3 H]-thymidine incorporation about 2.96 times. Conclusion EGF and bFGF were potent stimulators in RPE cells,and demonstrated synergism in their action. (Chin J Ocul Fundus Dis,1998,14:98-100)