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Jaime Gutiérrez Cristian A. Droppelmann Osvaldo Contreras Chiaki Takahashi Enrique Brandan 《PloS one》2015,10(8)
Fibroblasts are critical for wound contraction; a pivotal step in wound healing. They produce and modify the extracellular matrix (ECM) required for the proper tissue remodeling. Reversion-inducing cysteine-rich protein with Kazal motifs (RECK) is a key regulator of ECM homeostasis and turnover. However, its role in wound contraction is presently unknown. Here we describe that Transforming growth factor type β1 (TGF-β1), one of the main pro-fibrotic wound-healing promoting factors, decreases RECK expression in fibroblasts through the Smad and JNK dependent pathways. This TGF-β1 dependent downregulation of RECK occurs with the concomitant increase of β1-integrin, which is required for fibroblasts adhesion and wound contraction through the activation of focal adhesion kinase (FAK). Loss and gain RECK expression experiments performed in different types of fibroblasts indicate that RECK downregulation mediates TGF-β1 dependent β1-integrin expression. Also, reduced levels of RECK potentiate TGF-β1 effects over fibroblasts FAK-dependent contraction, without affecting its cognate signaling. The above results were confirmed on fibroblasts derived from the Reck
+/- mice compared to wild type-derived fibroblasts. We observed that Reck
+/- mice heal dermal wounds more efficiently than wild type mice. Our results reveal a critical role for RECK in skin wound contraction as a key mediator in the axis: TGF-β1—RECK- β1-integrin. 相似文献
2.
Authors index 总被引:1,自引:0,他引:1
Lehmann Johannes Weigl Doris Peter Inka Droppelmann Klaus Gebauer Gerhard Goldbach Heiner Zech Wolfgang 《Plant and Soil》1999,210(2):249-262
In a runoff irrigation system in Northern Kenya, we studied the nutrient interactions of sole cropped and alley cropped Sorghum
bicolor (L.) Moench and Acacia saligna (Labill.) H.L. Wendl. The trees were pruned once before the cropping season and the
biomass was used as fodder for animals. The nutrient contents in leaf tissue, soil and soil solution were monitored and the
uptake of applied tracers (15N, Sr) was followed. The grain yield of alley cropped sorghum was similar to or slightly higher than in monoculture and did
not decrease near the tree-crop interface. Foliar N and Ca contents of the crop were higher in the agroforestry combination
than in monoculture, corresponding to higher soil N and Ca contents. Soil solution and soil mineral N dynamics indicate an
increase of N under the tree row and unused soil N at the topsoil in the alley of the sole cropped trees as well as below
60 cm depth in the crop monoculture. The N use efficiency of the tree+crop combination was higher than the sole cropped trees
or crops. Competition was observed for Zn and Mn of both tree and crop whereas for Ca only the tree contents decreased. P,
K, Mg and Fe dynamics were not affected by alley cropping at our site. The lower uptake of applied Sr by trees in alley cropping
compared to those of the monoculture stand suggested a lower competitiveness of the acacia than sorghum, which did not show
lower Sr contents when intercropped. The study showed the usefulness of combining soil and plant analyses together with tracer
techniques identifying nutrient competition, nutrient transfer processes and the complementary use of soil nutrients, as the
main features of the tree-crop combination.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
3.
Yáñez AJ Garcia-Rocha M Bertinat R Droppelmann C Concha II Guinovart JJ Slebe JC 《FEBS letters》2004,577(1-2):154-158
In primary cultured hepatocytes, fructose-1,6-bisphosphatase (FBPase) localization is modulated by glucose, dihydroxyacetone (DHA) and insulin. In the absence of these substrates, FBPase was present in the cytoplasm, but the addition of glucose or DHA induced its translocation to the nucleus. As expected, we observed the opposite effect of glucose on glucokinase localization. The addition of insulin in the absence of glucose largely increased the amount of nuclear FBPase. Moreover, at high concentrations of glucose or DHA, FBPase shifted from the cytosol to the cell periphery and co-localized with GS. Interestingly, the synthesis of Glu-6-P and glycogen induced by DHA was not inhibited by insulin. These results indicate that FBPase is involved in glycogen synthesis from gluconeogenic precursors. Overall, these findings show that translocation may be a new integrative mechanism for gluconeogenesis and glyconeogenesis. 相似文献
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Cristian A. Droppelmann Jaime Guti��rrez Cecilia Vial Enrique Brandan 《The Journal of biological chemistry》2009,284(20):13551-13561
Matrix metalloproteinase-2 (MMP-2) is an important extracellular matrix
remodeling enzyme, and it has been involved in different fibrotic disorders.
The connective tissue growth factor (CTGF/CCN2), which is increased in these
pathologies, induces the production of extracellular matrix proteins. To
understand the fibrotic process observed in diverse pathologies, we analyzed
the fibroblast response to CTGF when MMP-2 activity is inhibited. CTGF
increased fibronectin (FN) amount, MMP-2 mRNA expression, and gelatinase
activity in 3T3 cells. When MMP-2 activity was inhibited either by the
metalloproteinase inhibitor GM-6001 or in MMP-2-deficient fibroblasts, an
increase in the basal amount of FN together with a decrease of its levels in
response to CTGF was observed. This paradoxical effect could be explained by
the fact that the excess of FN could block the access to other ligands, such
as CTGF, to integrins. This effect was emulated in fibroblasts by adding
exogenous FN or RGDS peptides or using anti-integrin αV
subunit-blocking antibodies. Additionally, in MMP-2-deficient cells CTGF did
not induce the formation of stress fibers, focal adhesion sites, and ERK
phosphorylation. Anti-integrin αV subunit-blocking antibodies
inhibited ERK phosphorylation in control cells. Finally, in MMP-2-deficient
cells, FN mRNA expression was not affected by CTGF, but degradation of
125I-FN was increased. These results suggest that expression,
regulation, and activity of MMP-2 can play an important role in the initial
steps of fibrosis and shows that FN levels can regulate the cellular response
to CTGF.Extracellular proteolysis is an essential physiological process that
controls the immediate cellular environment and thus plays a key role in
cellular behavior and survival
(1). The members of the matrix
metalloproteinase
(MMP)2 family of
zinc-dependent endopeptidases are major mediators of extracellular proteolysis
by promoting the degradation of extracellular matrix (ECM) components and cell
surface-associated proteins (2,
3). Each one of these enzymes
is negatively regulated by tissue inhibitors of metalloproteinases (TIMPs)
(4) and is secreted as a
zymogen (pro-MMPs) that is activated in the extracellular space
(5–7).
This mechanism is an important form of regulation of gelatinase activity and
in consequence, highly significant for ECM homeostasis. Among the members of
the MMP family, the metalloproteinase type 2 (MMP-2 or gelatinase A) is known
to be a key player in many physiological and pathological processes, such as
cell migration, inflammation, angiogenesis, and fibrosis
(8–11).Fibrotic disorders are typified by excessive connective tissue and ECM
deposition that precludes normal healing of different tissues. ECM
accumulation can be explained in two ways: increasing expression and
deposition of connective tissue proteins and/or decreasing degradation of ECM
proteins (12). Transforming
growth factor type β, a multifunctional cytokine, is strongly
overexpressed, and it is associated to the pathogenesis of these diseases
(13,
14). It stimulates the
expression of connective tissue growth factor (CTGF/CCN2)
(15), a cytokine that is
responsible for transforming growth factor type β fibrotic activity
(16,
17). The role of CTGF in
fibrosis has gained attention in recent years
(16,
18–22).
CTGF overexpression is known to occur in a variety of fibrotic skin disorders
(23,
24), renal
(25), hepatic
(26), and pulmonary fibrosis
(27) and in muscles from
patients with Duchenne muscular dystrophy
(28).On the other hand, several pathologies involving fibrosis show an increase
in MMP expression, including gelatinase A. Augmented expression of MMP-2 was
found in submucous (29), skin
(30), liver
(31), and lung fibrosis
(32,
33) and dystrophic myotubes
from fibrotic muscles of Duchenne muscular dystrophy
(34). It has been shown that
transforming growth factor type β induces an increase in the amount of
MMP-2 in fibroblasts (35) and
that CTGF induces MMP-2 expression in cultured renal interstitial fibroblasts
(36). The putative role
assigned to MMP-2 in fibrotic disorders is related to tissue regeneration
because of the capacity of this enzyme to degrade basal lamina
(37–39).
Because MMP-2 expression is up-regulated in these pathologies but still a high
ECM deposition is observed, we propose that this accumulation could be
explained by a diminution of the MMP-2 enzymatic activity.In this article, we demonstrate that CTGF increases fibronectin (FN)
amount, MMP-2 expression, and gelatinase activity in 3T3 fibroblasts. More
significantly, we show that MMP-2-deficient cells have an increased basal
amount of FN and show a response to CTGF that is opposite to that of control
cells. This paradoxical effect could be explained by the increase in the FN
amount that blocks the integrins (at least integrins with αV
subunit), which can act like CTGF receptors. 相似文献
7.
Joel L. Asenjo Heide C. Ludwig Cristian A. Droppelmann Juan G. Cárcamo Ilona I. Concha Alejandro J. Yáñez María L. Cárdenas Athel Cornish-Bowden Juan C. Slebe 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
Fructose-1,6-bisphosphatase, a major enzyme of gluconeogenesis, is inhibited by AMP, Fru-2,6-P2 and by high concentrations of its substrate Fru-1,6-P2. The mechanism that produces substrate inhibition continues to be obscure.Methods
Four types of experiments were used to shed light on this: (1) kinetic measurements over a very wide range of substrate concentrations, subjected to detailed statistical analysis; (2) fluorescence studies of mutants in which phenylalanine residues were replaced by tryptophan; (3) effect of Fru-2,6-P2 and Fru-1,6-P2 on the exchange of subunits between wild-type and Glu-tagged oligomers; and (4) kinetic studies of hybrid forms of the enzyme containing subunits mutated at the active site residue tyrosine-244.Results
The kinetic experiments with the wild-type enzyme indicate that the binding of Fru-1,6-P2 induces the appearance of catalytic sites with lower affinity for substrate and lower catalytic activity. Binding of substrate to the high-affinity sites, but not to the low-affinity sites, enhances the fluorescence emission of the Phe219Trp mutant; the inhibitor, Fru-2,6-P2, competes with the substrate for the high-affinity sites. Binding of substrate to the low-affinity sites acts as a “stapler” that prevents dissociation of the tetramer and hence exchange of subunits, and results in substrate inhibition.Conclusions
Binding of the first substrate molecule, in one dimer of the enzyme, produces a conformational change at the other dimer, reducing the substrate affinity and catalytic activity of its subunits.General significance
Mimics of the substrate inhibition of fructose-1,6-bisphosphatase may provide a future option for combatting both postprandial and fasting hyperglycemia. 相似文献8.
9.
Yánez AJ Nualart F Droppelmann C Bertinat R Brito M Concha II Slebe JC 《Journal of cellular physiology》2003,197(2):189-197
The importance of renal and hepatic gluconeogenesis in glucose homeostasis is well established, but the cellular localization of the key gluconeogenic enzymes liver fructose-1,6-bisphosphatase (FBPase) and cytosolic phosphoenolpyruvate carboxykinase (PEPCK) in these organs and the potential contribution of other tissues in this process has not been investigated in detail. Therefore, we analyzed the human tissue localization and cellular distribution of FBPase and PEPCK immunohistochemically. The localization analysis demonstrated that FBPase was expressed in many tissues that had not been previously reported to contain FBPase activity (e.g., prostate, ovary, suprarenal cortex, stomach, and heart). In some multicellular tissues, this enzyme was detected in specialized areas such as epithelial cells of the small intestine and prostate or lung pneumocytes II. Interestingly, FBPase was also present in pancreas and cortex cells of the adrenal gland, organs that are involved in the control of carbohydrate and lipid metabolism. Although similar results were obtained for PEPCK localization, different expression of this enzyme was observed in pancreas, adrenal gland, and pneumocytes type I. These results show that co-expression of FBPase and PEPCK occurs not only in kidney and liver, but also in a variety of organs such as the small intestine, stomach, adrenal gland, testis, and prostate which might also contribute to gluconeogenesis. Our results are consistent with published data on the expression of glucose-6-phosphatase in the human small intestine, providing evidence that this organ may play an important role in the human glucose homeostasis. 相似文献
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