• Journal of Dental Rehabilitation and Applied Science
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• v.19 no.4
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• pp.281-290
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• 2003

Platelet-rich plasma which is made with the newly developed technique concentrating platelets 3-folds or more is also proven to be very effective method to stimulate and accelerate the healing of bone and soft tissue. This study is aimed to investigate the effect of platelet-rich plasma on the attachment of osteoblast. To evaluate the effect on human, human osteoblast cell line was cultured. Platelet-rich plasma was extracted from the blood of a healthy volunteer. The effect on the attachment was evaluated by MTT assay. To evaluate autocrine and paracrine effect on osteoblast, conditioned medium was made and compared with platelet-rich plasma. By western blot analysis, the expression of fibronectin and vitronectin in experimental groups was examined. The results were as following: The cellular attachment of osteoblast cell line increased depending on the concentration of platelet-rich plasma and conditioned medium. The amount of increasing was similar between two groups. The expression of fibronectin and vitronectin in platelet-rich plasma and conditioned medium is more than control group in western blot analysis. These findings imply that platelet-rich plasma enhance the cellular attachment by inducing fibronectin, vitronectin from osteoblast and maximize the cellular attachment by using the autocrine and paracrine effect of platelet-rich plasma.

• Reproductive and Developmental Biology
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• v.29 no.4
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• pp.259-267
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• 2005

The present study was conducted to investigate gelatinolytic activities in HAM and to determine whether there are any changes in gelatinolytic activity profiles when the cells are cultured in hepatogenic medium. Placenta was obtained during caesarean section of the volunteers, with informed consent. HAM were isolated from amniotic membrane using collagenase type A HAM were cultured in hepatogenic medium for 3 weeks and the conditioned media were obtained at day 7, 14 and 21. The zymographic pattern of gelatinolytic activity of the HAM did not undergo a change during passages. When the HAM were cultured in a fibronectin-coated dishes in a hepatogenic medium, there was no significant difference of the gelatinase pattern between before and after culture. However, when bFGF was added to the culture, a dramatic increase of 62kDa and 59kDa gelatinases was observed. Interestingly, when ITS instead of FN was present, HAM-conditioned medium also showed a similar increase of both gelatinases. Immunoblotting analysis demonstrated that both 62kDa and 59kDa gelatinases were the active form of MMP-2 resulting from the turnover of MMP-2 proform. Futher study will be necessary to determine the relationship between bFGF and active MMP-2 during hepatogenesis of HAM.

• Journal of Dental Rehabilitation and Applied Science
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• v.20 no.1
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• pp.31-41
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• 2004

The osseointegration in implant therapy is achieved following general wound healing mechanism. Platelet play a major role in wound healing process. In addition to blood clot formation, they secrete many growth factors which regulate the attachment, proliferation and differentiation of nearly all cell types. The use of these growth factors is now known to be very effective methods to improve the cellular activity. Platelet-rich plasma which is made with the newly developed technique concentrating platelets 3-folds or more is also proven to be very effective method to stimulate and accelerate the healing of bone and soft tissue. Previous study proved that platelet-rich plasma enhanced the cellular attachment by inducing fibronectin, vitronectin from osteoblast. So, this study was aimed to investigate the effect of platelet-rich plasma on the cellular proliferation and differentiation in vitro. The effect on the proliferation was evaluated by MTT assay. To evaluate autocrine and paracrine effect, conditioned medium was made and compared. By measuring alkaline phosphatase activity, the effect on the cellular differentiation was evaluated. The results were as following: The cellular proliferation of osteoblast cell line increased depending on the concentration of platelet-rich plasma and conditioned medium. The alkaline phosphatase activity increased depending on the concentration of platelet-rich plasma and conditioned medium. These findings imply that platelet-rich plasma enhance the cellular proliferation and differentiation and maximize the cellular activity by using the autocrine and paracrine effect.

• Development and Reproduction
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• v.10 no.1
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• pp.63-73
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• 2006

This study aimed to find out suitable culture conditions for the differentiation of human amniotic membrane-derived stem cells(HAM) into hepatocyte-like cells. Almost homogenous population of fibroblast-like cells was successfully isolated from the amniotic membrane. In comparison to the non-coated plates and in the absence of insulin/transferrin/selenium(ITS), HAM cultured on the fibronectin-coated plates and in the presence of ITS showed the more intense immunocytochemical staining against the albumin. Addition of both fibroblast growth factor(FGF)-1 and -2 to the differentiation medium gave stronger staining compared to the treatment with FGF-1 or -2 alone. Periodic acid Schiff's base staining of glycogen and morphological turnover of fibroblast-like appearance of HAM into round shape matched the results of immunocytochemical studies. When the efficiency of two-step culture method was examined on the differentiation of HAM into hepatocyte-like cells, all of the results of immunocytochemical staining, periodic acid Schiff's staining and morphological change exhibited effective hepatic differentiation of HAM compared to the continuous culture method. Immunoblot analyses of HAM- conditioned media against the albumin showed that the culture of HAM in the presence of both ITS and fibronectin always gave a stronger staining intensity than those in the absence of them, and that the addition of ether mixture of FGF-4 and either FGF-2 or transforming growth $factor(TGF)-{\alpha}$ to the culture medium significantly enhanced the albumin secretion by HAM. Based on these observations, it is suggested that HAM could differentiate into hepatocyte-like cells under a culture condition consisting of fibronectin and ITS, and addition of FGF-4 with either one of FGF-2 or $TGF-{\alpha}$ could enhance the hepatic differentiation of HAM.

• International Journal of Stem Cells
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• v.16 no.1
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• pp.52-65
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• 2023

Background and Objectives: Epithelial-Mesenchymal transition (EMT) is one of the origins of myofibroblasts in renal interstitial fibrosis. Mesenchymal stem cells (MSCs) alleviating EMT has been proved, but the concrete mechanism is unclear. To explore the mechanism, serum-free MSCs conditioned medium (SF-MSCs-CM) was used to treat rat renal tubular epithelial cells (NRK-52E) fibrosis induced by transforming growth factor-β1 (TGF-β1) which ameliorated EMT. Methods and Results: Galectin-3 knockdown (Gal-3 KD) and overexpression (Gal-3 OE) lentiviral vectors were established and transfected into NRK-52E. NRK-52E fibrosis model was induced by TGF-β1 and treated with the SF-MSCs-CM for 24 h after modelling. Fibrosis and autophagy related indexes were detected by western blot and immunocytochemistry. In model group, the expressions of α-smooth muscle actin (α-SMA), fibronectin (FN), Galectin-3, Snail, Kim-1, and the ratios of P-Akt/Akt, P-GSK3β/GSK3β, P-PI3K/PI3K, P-mTOR/mTOR, TIMP1/MMP9, and LC3B-II/I were obviously increased, and E-Cadherin (E-cad) and P62 decreased significantly compared with control group. SF-MSCs-CM showed an opposite trend after treatment compared with model group. Whether in Gal-3 KD or Gal-3 OE NRK-52E cells, SF-MSCs-CM also showed similar trends. However, the effects of anti-fibrosis and enhanced autophagy in Gal-3 KD cells were more obvious than those in Gal-3 OE cells. Conclusions: SF-MSCs-CM probably alleviated the EMT via inhibiting Galectin-3/Akt/GSK3β/Snail pathway. Meanwhile, Gal-3 KD possibly enhanced autophagy via inhibiting Galectin-3/Akt/mTOR pathway, which synergistically ameliorated renal fibrosis. Targeting galectin-3 may be a potential target for the treatment of renal fibrosis.