We have previously observed that near-infrared (IR) pre-irradiation protects normal human dermal fibroblasts from ultraviolet (UV) cytotoxicity in vitro. Here, we show that IR pre-irradiation of ...human fibroblasts inhibited UVB activation of caspase-9 and -3, leading us to study early events in the mitochondrial apoptotic pathway after IR irradiation. IR irradiation led to a partial release of cytochrome c and Smac/Diablo but not apoptosis-inducing factor (AIF). This was accompanied by a slight but transient decrease in the mitochondrial membrane potential (Δψm) and by the insertion of Bax into mitochondrial membrane. Early apoptotic events in the mitochondrial pathway thus occurred after IR irradiation despite a lack of caspase-9 and -3 activation. This could be explained by the induction by IR of the expression of heat shock protein Hsp27, which is known to prevent apoptosome assembly. Furthermore, the balance between pro-apoptotic (i.e., Bax) and anti-apoptotic (i.e., Bcl-2 or Bcl-xL) proteins, which was rather pro-apoptotic after IR exposure, became anti-apoptotic 24 h later, suggesting a protective effect. Together, these actions could also contribute to prepare the cell to resist UVB-triggered apoptosis. Finally, isolated rat liver mitochondria-released cytochrome c in response to IR, demonstrating that mitochondria were a primary target of IR radiation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Dextran derivatives can protect heparin binding growth factor implied in wound healing, such as transforming growth factor-β1 (TGF-β1) and fibroblast growth factor-2 (FGF-2). The first aim of this ...study was to investigate the effect of these compounds on human
dermal fibroblasts in culture with or without TGF-β1. Several dextran derivatives obtained by substitution of methylcarboxylate (MC), benzylamide (B) and sulphate (Su) groups were used to determine the effects of each compound on fibroblast growth in vitro. The data indicate
that sulphate groups are essential to act on the fibroblast proliferation. The dextran derivative LS21 DMCBSu has been chosen to investigate its effect on dermal wound healing process. Fibroblasts cultured in collagenous matrices named dermal equivalent were treated with the bioactive polymer
alone or associated to TGF-β1 or FGF-2. Cross-sections of dermal equivalent observed by histology or immunohistochemistry, demonstrated that the bioactive polymer accelerates the collagen matrices organization and stimulates the human type-III collagen expression. This bioactive
polymer induces apoptosis of myofibroblast, property which may be beneficial in treatment of hypertrophic scar. Culture media analyzed by zymography and Western blot showed that this polymer significantly increases the secretion of zymogen and active form of matrix metalloproteinase-2 (MMP-2),
involved in granulation tissue formation. These data suggest that this bioactive polymer has properties which may be beneficial in the treatment of wound healing.
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BFBNIB, DOBA, GIS, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Protein kinase C (PKC) is known to be involved in cellular proliferation and differentiation. In this work, we have investigated
the effects of a novel PKC inhibitor, GF 109203X, on normal human ...fibroblast and keratinocyte growth. GF 109203X selectively
inhibited PKC activity extracted from either fibroblasts (IC50 = 0.01 microM) or keratinocytes (IC50 = 0.4 microM). The inhibitory
effects of GF 109203X on total PKC activity and Ca(2+)-independent PKC activity were similar. Nevertheless, in keratinocytes
Ca(2+)-independent PKC activity represented 95% of total PKC activity, whereas in fibroblasts it corresponded to only 32%
of total PKC activity. GF 109203X also inhibited a cellular function related to PKC activity in living fibroblasts and keratinocytes;
it blocked the inhibitory effect of 12-O-tetradecanoylphorbol-13-acetate on 125I-epidermal growth factor binding. GF 109203X
inhibited fibroblast growth, in terms of tritiated thymidine incorporation and cell counts, in a dose-dependent manner. We
also observed that GF 109203X at 1 microM inhibited serum stimulation of expression of mRNA for c-fos and c-jun, which are
usually involved in cellular proliferation. These results suggest that PKC stimulates fibroblast growth. In contrast, GF 109203X
stimulated keratinocyte growth. We also observed that GF 109203X inhibited c-fos and c-jun mRNA expression in these cells.
In fact, in keratinocytes these proto-oncogenes would be involved in the cellular differentiation process rather than in cellular
proliferation. This suggests that the inhibition of PKC favors keratinocyte proliferation probably by inhibiting their differentiation.
Thus, using GF 109203X, we show that PKC is involved differently in human fibroblast and keratinocyte growth.
Dextran derivatives that mimic the action of heparin have been shown to protect heparinbinding growth factors, such as Transforming Growth Factor-β1 (TGF-β1) and Fibroblast Growth Factor-2 (FGF-2). ...The aim of this study was to investigate the effect of LS21 DMCBSu,
a dextran derivative which contains methylcarboxylate, benzylamide and sulfate groups, both by itself and when combined with TGF-β1 and FGF-2, on the behaviour of fibroblasts. Two systems were assessed: a monolayer culture and three-dimensional collagenous matrices (dermal equivalent).
Polymeric biomaterial LS21 DMCBSu and LS21 DMCBSu associated with either TGF-β1 or FGF-2, were added to the monolayer culture on day 3. After 7 days of culture the number of cells was determined. Two treatments were carried out on the dermal equivalents: 9 days of treatment from
day 0 to day 9 of culture and 9 days of treatment from day 21 to day 30 of culture for the premature and the mature dermal equivalents respectively. In the monolayer culture, the bioactive polymer produced a slight increase in fibroblast growth (10% with 10 μg/ml of LS21 DMCBSu)
and promoted the stimulating effect of the growth factors on cell growth. In the premature dermal equivalents growth was stimulated by 20% when 10 μg/ml LS21 DMCBSu was added. The dextran derivative mixed with TGF-β1slightly inhibited the growth effect of the growth factor
in the dermal equivalents. The functionalized dextran with FGF-2 enhanced the stimulating effect of the growth factor in the premature dermal equivalent. A significant increase in cell growth was observed with the fibroblasts treated with the FGF-2 LS21 DMCBSu mixture and FGF-2 (51% and 40%,
respectively). However, none of the described treatments affected the cell growth in the mature dermal equivalent. Furthermore, the dextran derivative had no effect on dermal contraction under these experimental conditions (3D culture).
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BFBNIB, DOBA, GIS, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
In this study, epidermal growth factor (EGF), 20 ng/ml, and keratinocyte growth factor (KGF), 10 ng/ml, were able to stimulate human keratinocyte growth only in presence of GF 109203X 1 microM, a ...selective PKC inhibitor. This suggests that PKC negatively controls keratinocyte growth in response to EGF and KGF. On the other hand, EGF and KGF have no significant effect on PKC activity. In contrast, in human fibroblasts, EGF stimulated fibroblast growth in the presence or not of GF 109203X. Thus, EGF seems to stimulate fibroblast growth in a PKC independent manner. Moreover, EGF didn't modify significantly PKC activity in fibroblasts. KGF had no effect on fibroblasts. These results show differences in the interconnections between PKC and EGF transduction pathway in the modulation of human keratinocyte and fibroblast growth. Moreover, they especially demonstrate that PKC would negatively control human keratinocyte growth in response to EGF and also to KGF.
Dextran derivatives that mimic the action of heparin have been shown to protect heparin-binding growth factors, such as Transforming Growth Factor-beta1 (TGF-beta1) and Fibroblast Growth Factor-2 ...(FGF-2). The aim of this study was to investigate the effect of LS21 DMCBSu, a dextran derivative which contains methylcarboxylate, benzylamide and sulfate groups, both by itself and when combined with TGF-beta1 and FGF-2, on the behaviour of fibroblasts. Two systems were assessed: a monolayer culture and three-dimensional collagenous matrices (dermal equivalent). Polymeric biomaterial LS21 DMCBSu and LS21 DMCBSu associated with either TGF-beta1 or FGF-2, were added to the monolayer culture on day 3. After 7 days of culture the number of cells was determined. Two treatments were carried out on the dermal equivalents: 9 days of treatment from day 0 to day 9 of culture and 9 days of treatment from day 21 to day 30 of culture for the premature and the mature dermal equivalents respectively. In the monolayer culture, the bioactive polymer produced a slight increase in fibroblast growth (10% with 10 microg/ml of LS21 DMCBSu) and promoted the stimulating effect of the growth factors on cell growth. In the premature dermal equivalents growth was stimulated by 20% when 10 microg/ml LS21 DMCBSu was added. The dextran derivative mixed with TGF-beta1 slightly inhibited the growth effect of the growth factor in the dermal equivalents. The functionalized dextran with FGF-2 enhanced the stimulating effect of the growth factor in the premature dermal equivalent. A significant increase in cell growth was observed with the fibroblasts treated with the FGF-2 LS21 DMCBSu mixture and FGF-2 (51% and 40%, respectively). However, none of the described treatments affected the cell growth in the mature dermal equivalent. Furthermore, the dextran derivative had no effect on dermal contraction under these experimental conditions (3D culture).
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BFBNIB, DOBA, GIS, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
We report the pharmacologic effects of retinoids in a human skin-equivalent model. This sophisticated culture system is composed, as in vivo, of a dermis and epidermis, and provides a unique in vitro ...system for studying dermal-epidermal interactions and thus, whether normal dermal fibroblasts influence the effects of retinoids on epidermal growth. Epidermalization was initiated on collagen substrates in which fibroblasts were either viable or lysed by osmotic shock. Retinoic acid, isotretinoin, and acitretin at 10(-6) M or 10(-7) M were added to the cultures just after epidermalization, then every two days. Epidermal growth was determined after 2 weeks in terms of the surface area, DNA content, and tritiated thymidine incorporation during the last 24 h of culture. In the absence of viable fibroblasts, retinoic acid and isotretinoin increased epidermal growth, whereas etretin inhibited it. In contrast, in the presence of viable fibroblasts, retinoic acid and isotretinoin inhibited epidermal growth, whereas etretin had no effect. Thus, retinoic acid and isotretinoin had a similar effect on keratinocyte proliferation that contrasted with that of etretin. Taken as a whole, these results show that fibroblasts, present within a collagen substrate, can modulate the pharmacologic effects of retinoids on epidermal growth.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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