epidermal-growth-factor and Fibrosis

Obstructive nephropathy is a major cause of renal failure, particularly in infants and children. Cellular and molecular mechanisms responsible for the progression of the tubular atrophy and interstitial fibrosis-processes that lead to nephron loss-have been elucidated in the past 5 years. Following urinary tract obstruction and tubular dilatation, a cascade of events results in upregulation of the intrarenal renin-angiotensin system, tubular apoptosis and macrophage infiltration of the interstitium. This is followed by accumulation of interstitial fibroblasts through proliferation of resident fibroblasts and epithelial-mesenchymal transformation of renal tubular cells. Under the influence of cytokines, chemokines and other signaling molecules produced by tubular and interstitial cells, fibroblasts undergo transformation to myofibroblasts that induce expansion of the extracellular matrix. The cellular interactions that regulate development of interstitial inflammation, tubular apoptosis and interstitial fibrosis are complex. Changes in renal gene expression and protein production afford many potential biomarkers of disease progression and targets for therapeutic manipulation. These include signaling molecules and receptors involved in macrophage recruitment and proliferation, tubular death signals and survival factors, and modulators of epithelial-mesenchymal transformation. Targeted gene deletion and various forms of gene therapy have been used in experimental obstructive nephropathy, mostly rodent models of unilateral ureteral obstruction or cell culture techniques. Further refinement of these models is needed to develop a matrix of biomarkers with clinical predictive value, as well as molecular therapies that will prevent or reverse the renal structural and functional consequences of obstructive nephropathy.

Topics: Apoptosis; Biomarkers; Chemokine CCL7; Epidermal Growth Factor; Fibrosis; Genetic Therapy; Humans; Inflammation; Kidney; Monocyte Chemoattractant Proteins; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Ureteral Obstruction

Topics: Asthma; Bronchi; Chronic Disease; Epidermal Growth Factor; Epithelium; Fibrosis; Humans; Models, Immunological; Signal Transduction

Renal insufficiency as a result of congenital obstructive nephropathy is a consequence of impaired renal growth: chronic unilateral ureteral obstruction (UUO) results in greater injury to the immature kidney than to the adult kidney. The neonatal kidney responds to UUO by marked activation of the renin-angiotensin system, which contributes to severe vasoconstriction and progressive interstitial fibrosis of the obstructed kidney. The latter results in part because of activation of transforming growth factor-beta 1 by angiotensin II. Chronic UUO in the neonatal rat delays maturation of the obstructed kidney, possibly in part through suppressed expression of epidermal growth factor. In addition to affecting growth factors, UUO stimulates apoptosis in the obstructed kidney, which is quantitatively greater in the neonate than in the adult. In contrast, expression of clusterin, a glycoprotein that may play a protective role in the response to UUO, is greater in the adult than in the neonatal obstructed kidney. The response of the developing kidney to UUO is similar in a number of respects to cystic kidney disease. This includes a reduction in epidermal growth factor, and increased apoptosis that may result from suppression of bcl-2, an oncoprotein that inhibits apoptosis. Improved knowledge of the cellular and molecular basis for cystic renal disorders should lead to specific intervention in fetuses and infants with congenital obstructive nephropathy, thereby improving renal growth and development.

Topics: Adult; Age Factors; Angiotensin II; Animals; Animals, Newborn; Apoptosis; Clusterin; Epidermal Growth Factor; Fibrosis; Gene Expression Regulation; Glycoproteins; Growth Substances; Hemodynamics; Humans; Infant; Infant, Newborn; Kidney; Kidney Diseases, Cystic; Mice; Mice, Knockout; Models, Biological; Molecular Chaperones; Proto-Oncogene Proteins c-bcl-2; Rats; Renin-Angiotensin System; Transforming Growth Factor beta; Ureteral Obstruction

Noninvasive biomarkers that reflect tubular health and allow early recognition of accelerated graft fibrosis development are warranted. Serum uromodulin (sUmod) and urinary epidermal growth factor (uEGF) originate from kidney tubules and may reflect functional nephron mass. The aim of this study was to investigate the associations between sUmod and uEGF with measured glomerular filtration rate (mGFR) and kidney allograft interstitial fibrosis percentage (IF%) score.. sUmod and uEGF measurements, mGFR by iohexol-clearance and kidney allograft biopsies were obtained from kidney transplant recipients (KTRs) included in the Omega-3 fatty acids in Renal Transplantation (ORENTRA) trial at 8 weeks (baseline) and at 1 year after transplantation (end of study). Associations were analyzed with univariable and multivariable linear regression.. Ninety patients at baseline and 48 patients at end of study had complete study variable assessments. uEGF normalized to urinary creatinine (uEGF/Cr) was associated with mGFR both at baseline (standardized β-coefficient [Std. β-coeff] = 0.457 [p = <0.001]) and at end of study (Std. β-coeff = 0.637 [p = <0.001]). sUmod was only associated with mGFR at end of study (Std. β-coeff = 0.443 [p = 0.002]). uEGF/Cr, sUmod, and mGFR were associated with graft IF% score both at baseline (Std. β-coeff = -0.349 [p = 0.001], -0.274 [p = 0.009] and -0.289 [p = 0.006], respectively) and at end of study (Std. β-coeff = -0.365 [p = 0.011], -0.347 [p = 0.016] and -0.405 [p = 0.004], respectively). The results remained largely unchanged in multivariable analysis.. uEGF/Cr and sUmod were associated with mGFR and graft IF% score. Our results indicate a possible role of uEGF/Cr and sUmod in the follow-up of KTRs.

Topics: Creatinine; Epidermal Growth Factor; Fibrosis; Glomerular Filtration Rate; Humans; Kidney Transplantation; Uromodulin

Activity and chronicity of kidney involvement in ANCA-associated vasculitis (AAV) can be currently reliably evaluated only by kidney biopsy. In this study, we measured a panel of serum and urinary biomarkers collected at the time of kidney biopsy and hypothesized that they could reflect specific histopathological parameters in the biopsy and help to predict prognosis.. We examined a cohort of 45 patients with AAV and 10 healthy controls. Biomarker levels (DKK-3, CD163, EGF, PRO-C6 and C3M) were measured in this study by ELISA. Biopsies were scored with a scoring system for AAV (focal x crescentic x sclerotic x mixed class) and interstitial fibrosis was quantified.. Levels of urinary DKK-3, CD163, EGF, PRO-C6 and C3M significantly differed among biopsy classes in AAV, with urinary DKK-3 and PRO-C6 levels being highest in the sclerotic class and lowest in the focal class, urinary CD163 levels highest in the crescentic class and urinary C3M levels highest in the focal class. Moreover, the urinary biomarkers were able to discriminate focal biopsy class from the other classes. Urinary DKK-3, EGF, PRO-C6 and C3M levels measured at the time of biopsy were also significantly related to the extent of fibrosis and to the final kidney function at the end of follow-up.. This small pilot study suggests that selected urinary biomarkers of fibrosis and inflammation may reflect changes in the kidney biopsy and be prognostic of kidney outcome in patients with AAV.

Topics: Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis; Antibodies, Antineutrophil Cytoplasmic; Biomarkers; Epidermal Growth Factor; Fibrosis; Humans; Inflammation; Kidney; Pilot Projects

The epithelial-mesenchymal transition (EMT) is a multi-step morphogenetic process in which epithelial cells lose their epithelial properties and gain mesenchymal characteristics. The process of EMT has been shown to mediate mammary gland fibrosis. Understanding how mesenchymal cells emerge from an epithelial default state will aid in unravelling the mechanisms that control fibrosis and, ultimately, in identifying therapeutic targets to alleviate fibrosis.. On treatment with EGF and/or HG, qPCR analysis showed a significant increase in the gene expression of EMT markers and downstream signalling genes. The expression of these genes was reduced on treatment with EGF+HG combination in both cell lines. The protein expression of COL1A1 increased as compared to the control in cells treated with EGF or HG alone, but when the cells were treated with EGF and HG together, the protein expression of COL1A1 decreased. ROS levels and cell death increased in cells treated with EGF and HG alone, whereas cells treated with EGF and HG together showed a decrease in ROS production and apoptosis.. This study demonstrates that EGF and HG induce EMT in mammary epithelial cells and may also have a role in fibrosis.

Topics: Epidermal Growth Factor; Epithelial-Mesenchymal Transition; Fibrosis; Humans; Mammary Glands, Human; Reactive Oxygen Species

Alcoholic liver disease (ALD) and other forms of chronic hepatotoxic injury can lead to transforming growth factor β1 (TGFβ1)-induced hepatic fibrosis and compromised liver function, underscoring the need to develop novel treatments for these conditions. Herein, our analyses of liver tissue samples from severe alcoholic hepatitis (SAH) patients and two murine models of ALD reveals that the ALD phenotype was associated with upregulation of the transcription factor ETS domain-containing protein (ELK-3) and ELK-3 signaling activity coupled with downregulation of α/β hydrolase domain containing 10 (ABHD10) and upregulation of deactivating S-palmitoylation of the antioxidant protein Peroxiredoxin 5 (PRDX5). In vitro, we further demonstrate that ELK-3 can directly bind to the ABHD10 promoter to inhibit its transactivation. TGFβ1 and epidermal growth factor (EGF) signaling induce ABHD10 downregulation and PRDX5 S-palmitoylation via ELK-3. This ELK-3-mediated ABHD10 downregulation drives oxidative stress and disrupts mature hepatocyte function via enhancing S-palmitoylation of PRDX5's Cys100 residue. In vivo, ectopic Abhd10 overexpression ameliorates liver damage in ALD model mice. Overall, these data suggest that the therapeutic targeting of the ABHD10-PRDX5 axis may represent a viable approach to treating ALD and other forms of hepatotoxicity.

Topics: Animals; Epidermal Growth Factor; Esterases; Fibrosis; Humans; Liver Cirrhosis; Liver Diseases, Alcoholic; Mice; Proto-Oncogene Proteins c-ets; Transcription Factors

Contrast-induced nephropathy (CIN) is a major clinical problem associated with acute kidney injury during hospitalization. However, effective treatments for CIN are currently lacking. Mesenchymal stem cells (MSCs) have protective effects against kidney injury by suppressing inflammation and fibrosis. We previously showed that MSCs cultured in serum-free medium (SF-MSCs) enhance their anti-inflammatory and anti-fibrotic effects. However, whether SF-MSCs potentiate their anti-apoptotic effects is unknown. Here, we investigated the effects of SF-MSCs on a CIN mouse model.. To create CIN model mice, we removed right kidney at first. One week later, the left renal artery was clamped for 30 min to cause ischemia-reperfusion injury, and mice were injected with iohexol. Then the kidney received 10 Gy of irradiation, and MSCs or SF-MSCs were injected immediately. At 24 h post-injection, mice were sacrificed, and their blood and kidneys were collected to evaluate renal function, DNA damage, and apoptosis. In addition, apoptosis was induced in HEK-293 cells by irradiation and cells were treated with conditioned medium from MSCs or SF-MSCs.. Treatment of CIN model mice with SF-MSCs markedly improved renal function compared with MSCs treatment. Cleaved caspase-3 levels and TUNEL-positive cell numbers were strongly suppressed in CIN model mice treated with SF-MSCs compared with the findings in those treated with MSCs. γH2AX levels, a chromosome damage marker, were reduced by MSCs and further reduced by SF-MSCs. In addition, cleaved caspase-3 in irradiated HEK-293 cells was more strongly suppressed by conditioned medium from SF-MSCs than by that from MSCs. Secretion of epidermal growth factor (EGF) was enhanced by culturing MSCs in serum-free medium. Knockdown of EGF by siRNA attenuated the inhibitory effects of SF-MSCs on CIN-induced renal dysfunction and tubular apoptosis.. These findings strongly suggest that SF-MSCs improve CIN in model mice by exerting anti-apoptotic effects in a paracrine manner. Thus, SF-MSCs represent a potential novel therapy for CIN.

Topics: Acute Kidney Injury; Animals; Caspase 3; Cells, Cultured; Culture Media, Conditioned; Epidermal Growth Factor; Fibrosis; HEK293 Cells; Humans; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice

Kidney biopsies of patients with diabetic nephropathy (DN) and normal kidney function may exhibit interstitial fibrosis (IF) without reduction of glomerular filtration rate (GFR) because of hyperfiltration. The aim of our study was to analyse the performance of a set of biomarkers of tubular injury to estimate the extent of IF in patients with DN and normal kidney function.. This cross-sectional study included 118 adults with DN diagnosed by kidney biopsy and GFR ≥90 mL/min/1.73 m2 and a control group of healthy subjects. We measured the urinary excretion of monocyte chemoattractant protein-1 (MCP-1) neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), liver-type fatty acid-binding protein (L-FABP), β2-microglobulin and dickkopf-3 protein (DKK-3) at the time of kidney biopsy. GFR was measured by chromium-51 labeled ethylenediamine tetraacetic acid (Cr-EDTA) (measured GFR). IF was quantified using a quantitative morphometric procedure. Predictive multivariate models were developed to estimate the IF surface.. Patients with DN showed significantly higher levels of DKK-3, MCP-1 and L-FABP and significantly lower levels of epidermal growth factor (EGF) than healthy controls. There were no significant between-group differences in the levels of β2-microglobulin, KIM-1 or NGAL. IF was negatively associated with EGF and positively with age, proteinuria, MCP-1, DKK-3 and L-FABP, but not with β2-microglobulin, KIM-1, NGAL or GFR. The best model to predict IF surface accounted for 59% of its variability and included age, proteinuria, EGF, DKK-3 and MCP-1.. Our study provides a model to estimate the IF in DN that can be useful to assess the progression of IF in patients with normal kidney function.

Topics: Adult; Biomarkers; Chemokine CCL2; Cross-Sectional Studies; Diabetes Mellitus; Diabetic Nephropathies; Edetic Acid; Epidermal Growth Factor; Fatty Acid-Binding Proteins; Fibrosis; Glomerular Filtration Rate; Humans; Kidney; Lipocalin-2; Proteinuria

Oncofetal protein, CRIPTO, is silenced during homeostatic postnatal life and often re-expressed in different neoplastic processes, such as hepatocellular carcinoma. Given the reactivation of CRIPTO in pathological conditions reported in various adult tissues, the aim of this study was to explore whether CRIPTO is expressed during liver fibrogenesis and whether this is related to the disease severity and pathogenesis of fibrogenesis. Furthermore, we aimed to identify the impact of CRIPTO expression on fibrogenesis in organs with high versus low regenerative capacity, represented by murine liver fibrogenesis and adult murine heart fibrogenesis. Circulating CRIPTO levels were measured in plasma samples of patients with cirrhosis registered at the waitlist for liver transplantation (LT) and 1 year after LT. The expression of CRIPTO and fibrotic markers (αSMA, collagen type I) was determined in human liver tissues of patients with cirrhosis (on a basis of viral hepatitis or alcoholic disease), in cardiac tissue samples of patients with end-stage heart failure, and in mice with experimental liver and heart fibrosis using immuno-histochemical stainings and qPCR. Mouse models with experimental chronic liver fibrosis, induced with multiple shots of carbon tetrachloride (CCl

Topics: Adenoviridae; Animals; Cell Proliferation; Collagen; Disease Models, Animal; End Stage Liver Disease; Epidermal Growth Factor; Fibrosis; GPI-Linked Proteins; Hepatocytes; Intercellular Signaling Peptides and Proteins; Ligands; Liver Cirrhosis; Liver Regeneration; Male; Membrane Glycoproteins; Mice, Inbred C57BL; Myocardium; Neoplasm Proteins; Up-Regulation; Wound Healing

Keloid and hypertrophic scars are skin fibrosis-associated disorders that exhibit an uncontrollable proliferation of fibroblasts and their subsequent contribution to the excessive accumulation of extracellular matrix (ECM) in the dermis. In this study, to elucidate the underlying mechanisms, we investigated the pivotal roles of epidermal growth factor (EGF) in modulating fibrotic phenotypes of keloid and hypertrophic dermal fibroblasts. Our initial findings revealed the molecular signatures of keloid dermal fibroblasts and showed the highest degree of skin fibrosis markers, ECM remodeling, anabolic collagen-cross-linking enzymes, such as lysyl oxidase (LOX) and four LOX-like family enzymes, migration ability, and cell-matrix traction force, at cell-matrix interfaces. Furthermore, we observed significant EGF-mediated downregulation of anabolic collagen-cross-linking enzymes, resulting in amelioration of fibrotic phenotypes and a decrease in cell motility measured according to the cell-matrix traction force. These findings offer insight into the important roles of EGF-mediated cell-matrix interactions at the cell-matrix interface, as well as ECM remodeling. Furthermore, the results suggest their contribution to the reduction of fibrotic phenotypes in keloid dermal fibroblasts, which could lead to the development of therapeutic modalities to prevent or reduce scar tissue formation.

Topics: Adult; Cell Movement; Cells, Cultured; Cicatrix, Hypertrophic; Elastic Modulus; Enzymes; Epidermal Growth Factor; Extracellular Matrix; Female; Fibroblasts; Fibrosis; Humans; Hydrogels; Keloid; Male; Middle Aged; Skin

Urinary levels of EGF may be a noninvasive biomarker of the degree of interstitial fibrosis. However, all the available data are based on studies that examined the EGF/creatinine ratio in spot urine samples. The agreement between EGF/creatinine ratio and 24-hours EGF excretion has not been analyzed, neither has it been established which of these two measurements is a better predictor of the degree of interstitial fibrosis. To investigate whether the EGF/creatinine ratio can predict 24-hours EGF, and which of these two measures is a better predictor of interstitial fibrosis in patients with IgA nephropathy (IgAN).. This is a cross-sectional study including 80 patients with IgAN. EGF levels were measured by ELISA in spot second-morning and 24-hours urine samples. We analyzed the concordance between these two measures and their respective ability to predict interstitial kidney fibrosis.. The intraclass correlation coefficient between 24-hours and spot EGF/creatinine was 0.63 (95% CI: 0.54 - 0.70), bias was 2.7 µg/mL (95% CI: 2.1 - 7.5). Passing-Bablok regression did not show a significant deviation from linearity (p = 0.72). Bland-Altman showed a systematic and proportional error between both EGF measures. Spot EGF/creatinine ratios overestimated the 24-hours EGF at low excretion values and underestimated it at high excretion values. In univariate analyses, 24-hours excretion of EGF was a better predictor of interstitial fibrosis than spot EGF/creatinine ratio (R2: 0.43 vs. 0.30, p = 0.000). In multivariate analyses, the 24-hours excretion of EGF plus GFR, significantly improved the prediction of interstitial fibrosis when compared with GFR alone (R2: 0.52 vs. 0.39, p = 0.000). When spot-urine EGF was introduced instead of the 24-hours excretion, the model was statistically significant but had a lower predictive capacity (R2: 0.46 spot EGF/creatinine vs. R2: 0.52 24-hours EGF excretion, p = 0.000).. The 24-hours excretion of EGF should be considered as the first-choice measure to estimate the interstitial fibrosis. The EGF/creatinine ratio cannot accurately estimate the total EGF excretion of but it also improves the estimation of the fibrosis surface, and, consequently, could be an alternative whenever 24-hours urine samples cannot be obtained.

Topics: Adult; Aged; Biomarkers; Creatinine; Cross-Sectional Studies; Epidermal Growth Factor; Female; Fibrosis; Glomerular Filtration Rate; Glomerulonephritis, IGA; Humans; Kidney Diseases; Male; Middle Aged; Predictive Value of Tests; Sensitivity and Specificity

Besides their importance for the vascular tone, vascular smooth muscle cells (VSMC) also contribute to pathophysiological vessel alterations. Various G-protein coupled receptor ligands involved in vascular dysfunction and remodeling can transactivate the epidermal growth factor receptor (EGFR) of VSMC, yet the importance of EGFR transactivation for the VSMC phenotype is incompletely understood. The aims of this study were (i) to characterize further the importance of the VSMC-EGFR for proliferation, migration and marker gene expression for inflammation, fibrosis and reactive oxygen species (ROS) homeostasis and (ii) to test the hypothesis that vasoactive substances (endothelin-1, phenylephrine, thrombin, vasopressin and ATP) rely differentially on the EGFR with respect to the abovementioned phenotypic alterations. In primary, aortic VSMC from mice without conditional deletion of the EGFR, proliferation, migration, marker gene expression (inflammation, fibrosis and ROS homeostasis) and cell signaling (ERK 1/2, intracellular calcium) were analyzed. VSMC-EGFR loss reduced collective cell migration and single cell migration probability, while no difference between the genotypes in single cell velocity, chemotaxis or marker gene expression could be observed under control conditions. EGF promoted proliferation, collective cell migration, chemokinesis and chemotaxis and leads to a proinflammatory gene expression profile in wildtype but not in knockout VSMC. Comparing the impact of five vasoactive substances (all reported to transactivate EGFR and all leading to an EGFR dependent increase in ERK1/2 phosphorylation), we demonstrate that the importance of EGFR for their action is substance-dependent and most apparent for crowd migration but plays a minor role for gene expression regulation.

Topics: Adenosine Triphosphate; Animals; Cell Movement; Cell Proliferation; Cells, Cultured; Endothelin-1; Enzyme Activation; Epidermal Growth Factor; ErbB Receptors; Extracellular Signal-Regulated MAP Kinases; Fibrosis; Gene Expression Regulation; Genotype; Inflammation; Ligands; Mice, Knockout; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Oxidative Stress; Phenotype; Phenylephrine; Primary Cell Culture; Signal Transduction; Thrombin; Time Factors; Vasopressins

The degree of tubular atrophy and interstitial fibrosis (IFTA) is an important prognostic factor in glomerulonephritis. Imbalance between pro-inflammatory cytokines such as monocyte chemoattractant protein- 1 (MCP-1) and protective cytokines such as epidermal growth factor (EGF) likely determine IFTA severity. In separate studies, elevated MCP-1 and decreased EGF have been shown to be associated with IFTA severity. In this study, we aim to evaluate the predictive value of urinary EGF/MCP-1 ratio compared to each biomarker individually for moderate to severe IFTA in primary glomerulonephritis (GN).. Urine samples were collected at biopsy from primary GN (IgA nephropathy, focal and segmental glomerulosclerosis, minimal change disease, membranous nephropathy). MCP-1 and EGF were analyzed by enzyme-linked immunosorbent assay.. EGF, MCP-1 and EGF/MCP-1 ratio from primary GN, all correlated with IFTA (n=58). By univariate analysis, glomerular filtration rate, EGF, and EGF/MCP-1 ratio were associated with IFTA. By multivariate analysis, only EGF/MCP-1 ratio was independently associated with IFTA. EGF/MCP-1 ratio had a sensitivity of 88% and specificity of 74 % for IFTA. EGF/MCP-1 had good discrimination for IFTA (AUC=0.85), but the improvement over EGF alone was not significant.. EGF/MCP-1 ratio is independently associated IFTA severity in primary glomerulonephritis, but the ability of EGF/MCP-1 ratio to discriminate moderate to severe IFTA may not be much better than EGF alone.

Topics: Adult; Atrophy; Biomarkers; Chemokine CCL2; Epidermal Growth Factor; Female; Fibrosis; Glomerulonephritis; Humans; Kidney Tubules; Male; Middle Aged; Sensitivity and Specificity; Severity of Illness Index

The aim of this study was to elucidate functional and molecular effects of mycophenolic acid (MPA) on non-lymphatic, kidney epithelial cells treated with transforming growth factor (TGF). MPA effects were studied using HK2 cells incubated with EGF and TGF. The reversibility of these effects was verified using guanosine and 8-aminoguanosine. The following assays were applied: cell proliferation, viability, collagen matrix contraction, scratch wound closure, spindle index, FACS with anti-CD29 and anti-CD326, promoter demethylation of RAS protein activator like 1 (RASAL1), as well as gene expression of RASAL1, integrin 1ß (ITGB1) (CD29) and epithelial cell adhesion molecule (EpCam) (CD326). Cell proliferation was inhibited by increasing concentrations of MPA, whereas neither apoptosis nor cytotoxicity was detected. Stimulation with EGF and/or TGF led to a significant collagen matrix contraction that was successfully inhibited by MPA. In addition, scratch wound closure was inhibited by incubation with TGF alone or with EGF. Under the same conditions, cell morphology (spindle shape) and molecular phenotype (ITGB1(High)EpCam(Low)/ITGB1(Low)EpCam(High)) were both significantly changed, suggesting an epithelial to mesenchymal transformation. Cell morphology and motility, as well as molecular phenotype, were reversible after MPA treatment with TGF transformation in both presence/absence of EGF, thereby suggesting a correlation with the previously described antifibrotic effects of MPA. Dysregulation of TGF signal transduction appears to be related to progression of fibrosis. A TGF-transformed kidney epithelial cell line derived from human proximal tubules was used to study whether the immunosuppressive drug: MPA possesses any functional or molecular antifibrotic effects. Functional and morphological in vitro changes induced by both the TGF and epithelial-growth-factor were reversible by treatment with MPA. An inhibitory effect of MPA on the TGF pathway appears to be responsible for the previously described antifibrotic effects of the MPA in the COL4A3-deficient mouse model of renal fibrosis.

Topics: Animals; Autoantigens; Cell Line; Cell Movement; Cell Proliferation; Cell Survival; Collagen Type IV; Disease Models, Animal; Epidermal Growth Factor; Fibrosis; Humans; Kidney; Mice; Mycophenolic Acid; Signal Transduction; Transforming Growth Factor beta

Mesenchymal stem cells (MSCs) have been shown to improve the outcome of acute renal injury models; but whether MSCs can delay renal failure in chronic kidney disease (CKD) remains unclear. In the present study, the were cultured in media containing various concentrations of basic fibroblast growth factor, epidermal growth factor and ascorbic acid 2-phosphate to investigate whether hepatocyte growth factor (HGF) secretion could be increased by the stimulation of these growth factors. Then, TGF-β1-treated renal interstitial fibroblast (NRK-49F), renal proximal tubular cells (NRK-52E) and podocytes were co-cultured with conditioned MSCs in the absence or presence of ascorbic acid 2-phosphate to quantify the protective effects of conditioned MSCs on renal cells. Moreover, male Sprague-Dawley rats were treated with 1 × 10(6) conditioned MSCs immediately after 5/6 nephrectomy and every other week through the tail vein for 14 weeks. It was found that basic fibroblast growth factor, epidermal growth factor and ascorbic acid 2-phosphate promoted HGF secretion in MSCs. Besides, conditioned MSCs were found to be protective against TGF-β1 induced epithelial-to-mesenchymal transition of NRK-52E and activation of NRK-49F cells. Furthermore, conditioned MSCs protected podocytes from TGF-β1-induced loss of synaptopodin, fibronectin induction, cell death and apoptosis. Rats transplanted with conditioned human MSCs had a significantly increase in creatinine clearance rate, decrease in glomerulosclerosis, interstitial fibrosis and increase in CD4(+)CD25(+)Foxp3(+) regulatory T cells counts in splenocytes. Together, our studies indicated that conditioned MSCs preserve renal function by their anti-fibrotic and anti-inflammatory effects. Transplantation of conditioned MSCs may be useful in treating CKD.

Topics: Animals; Apoptosis; Ascorbic Acid; CD4-Positive T-Lymphocytes; Cells, Cultured; Coculture Techniques; Creatinine; Disease Progression; Epidermal Growth Factor; Epithelial-Mesenchymal Transition; Female; Fibroblast Growth Factor 2; Fibronectins; Fibrosis; Glomerulosclerosis, Focal Segmental; Hepatocyte Growth Factor; Humans; Kidney; Kidney Tubules, Proximal; Lymphocyte Count; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Microfilament Proteins; Middle Aged; Nephrectomy; Podocytes; Rats; Rats, Sprague-Dawley; Renal Insufficiency, Chronic; Transforming Growth Factor beta1; Young Adult

It has been shown that phosphodiesterase type 5 (PDE5) inhibitors preserve smooth muscle (SM) content and ameliorate the fibrotic degeneration normally seen in the corpora cavernosa after bilateral cavernosal nerve resection (BCNR). However, the downstream mechanisms by which these drugs protect the corpora cavernosa remain poorly understood.. To provide insight into the mechanism, we aimed to determine the gene expression profile of angiogenesis-related pathways within the penile tissue after BCNR with or without continuous sildenafil (SIL) treatment.. Five-month-old Fisher rats were subjected to BCNR or sham operation and treated with or without SIL (20 mg/kg/BW drinking water) for 3 days or 45 days (N = 8 rats per group). Total RNAs isolated from the denuded penile shaft and prostate were subjected to reverse transcription and to angiogenesis real-time-polymerase chain reaction arrays (84 genes). Changes in protein expression of selected genes such as epiregulin (EREG) and connective tissue growth factor (CTGF) were corroborated by Western blot and immunohistochemistry.. Genes modulated by BCNR and SIL treatment.. A decreased expression of genes related to SM growth factors such as EREG, platelet-derived growth factor (PDGF), extracellular matrix regulators such as metalloproteinases 3 and 9, endothelial growth factors, together with an upregulation of pro-fibrotic genes such as CTGF and transforming growth factor beta 2 were found at both time points after BCNR. SIL treatment reversed this process by upregulating endothelial and SM growth factors and downregulating pro-fibrotic factors. SIL did not affect the expression of EREG, VEGF, and PDGF in the ventral prostate of BCNR animals.. SIL treatment after BCNR activates genes related to SM preservation and downregulates genes related to fibrosis in the corpora cavernosa. These results provide a mechanistic justification for the use of SIL and other PDE5 inhibitors as protective therapy against corporal SM loss and fibrosis after radical prostatectomy.

Topics: Animals; Disease Models, Animal; Endothelium, Vascular; Epidermal Growth Factor; Epiregulin; Extracellular Matrix; Fibrosis; Gene Expression; Intercellular Signaling Peptides and Proteins; Male; Matrix Metalloproteinase 1; Matrix Metalloproteinase 3; Nerve Tissue; Penis; Phosphodiesterase 5 Inhibitors; Piperazines; Purines; Rats; Sildenafil Citrate; Sulfones; Transforming Growth Factor beta2

To compare tear epidermal growth factor (EGF) concentration in dry eye (DE) conditions and determine correlations between EGF levels and severity of symptoms and ocular surface signs.. In this prospective case-control study, 35 patients with DE, including subgroups with meibomian gland disease (MGD), Sjögren's syndrome (SS) aqueous tear deficiency, or neurotrophic keratopathy (NK), and 17 asymptomatic control subjects were evaluated. Symptoms, Schirmer test, fluorescein clearance test (FCT), EGF concentration, dye staining, and the presence of corneal subepithelial fibrosis and meibomian gland (MG) orifice metaplasia were recorded. Tear EGF and the severity of irritation and ocular surface signs were correlated.. Tear EGF was higher in MGD than in the control (P = 0.03) and was lower in SS than in the control (P < 0.0001; MGD (P < 0.05) and NK (P < 0.01) groups. The DE subgroup with results in the FCT > 3 and Schirmer 1 >or= 8 had higher EGF levels than the group with FCT > 3 and Schirmer 1 < 8 and both groups with good tear clearance (P < 0.01). Tear EGF levels correlated inversely with conjunctival (r = -0.49, P = 0.0032) and corneal (r = -0.39, P = 0.022) dye staining and positively with MG orifice metaplasia (r = 0.36, P = 0.03) and corneal subepithelial fibrosis (r = 0.5, P = 0.0006).. Tear EGF concentration was increased in eyes with MGD, corneal subepithelial fibrosis, and MG orifice metaplasia. Elevated tear EGF may promote development of corneal subepithelial fibrosis and lid margin changes.

Topics: Case-Control Studies; Dry Eye Syndromes; Epidermal Growth Factor; Epithelium, Corneal; Eye Proteins; Eyelid Diseases; Female; Fibrosis; Fluorophotometry; Humans; Immunoassay; Male; Meibomian Glands; Metaplasia; Middle Aged; Prospective Studies; Tears

Epithelial-mesenchymal transition (EMT) is an important process that contributes to renal fibrogenesis. TGF-beta1 and EGF stimulate EMT. Recent studies suggested that parathyroid hormone-related protein (PTHrP) promotes fibrogenesis in the damaged kidney, apparently dependent on its interaction with vascular endothelial growth factor (VEGF), but whether it also interacts with TGF-beta and EGF to modulate EMT is unknown. Here, PTHrP(1-36) increased TGF-beta1 in cultured tubuloepithelial cells and TGF-beta blockade inhibited PTHrP-induced EMT-related changes, including upregulation of alpha-smooth muscle actin and integrin-linked kinase, nuclear translocation of Snail, and downregulation of E-cadherin and zonula occludens-1. PTHrP(1-36) also induced EGF receptor (EGFR) activation; inhibition of protein kinase C and metalloproteases abrogated this activation. Inhibition of EGFR activation abolished these EMT-related changes, the activation of ERK1/2, and upregulation of TGF-beta1 and VEGF by PTHrP(1-36). Moreover, inhibition of ERK1/2 blocked EMT induced by either PTHrP(1-36), TGF-beta1, EGF, or VEGF. In vivo, obstruction of mouse kidneys led to changes consistent with EMT and upregulation of TGF-beta1 mRNA, p-EGFR protein, and PTHrP. Taken together, these data suggest that PTHrP, TGF-beta, EGF, and VEGF might cooperate through activation of ERK1/2 to induce EMT in renal tubuloepithelial cells.

Topics: Animals; Cell Differentiation; Cell Line; Disease Models, Animal; Epidermal Growth Factor; Epithelial Cells; ErbB Receptors; Fibrosis; Humans; Kidney; Kidney Tubules; Mesoderm; Mice; Mice, Transgenic; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Parathyroid Hormone-Related Protein; Peptide Fragments; Transforming Growth Factor beta1; Vascular Endothelial Growth Factor A

Epidermal growth factor-loaded gelatin microspheres were tested for potential to accelerate tracheal allograft reepithelialization.. Epidermal growth factor-loaded gelatin microspheres were prepared by optimal double-phase emulsified condensation polymerization. One hundred age-matched allogeneic mice were randomly allocated to local administration of 1 mg epidermal growth factor-loaded gelatin microspheres (containing 1 microg epidermal growth factor), 1 mg gelatin microspheres, 1 microg epidermal growth factor, or nothing (control, all groups n = 25) during orthotopic transplant of C57BL/6 donor tracheal segments into BALB/c recipients without immunosuppressors. On days 7, 14, 21, 35, and 52 after transplant, 5 mice per group were killed and evaluated by histologic assessment and scanning electronic microscopy for reepithelialization and fibrosis of tracheal grafts.. Mean diameter of gelatin microspheres was 107 microm. Microspheres could not be fully degraded until 35 days after transplant in vivo. On days 7, 14, and 21, epithelium score and ratio of lamina propria to tracheal cartilage were not statistically different between mice with epidermal growth factor-loaded gelatin microspheres and other groups. On days 35 and 52, however, epithelium score was higher and ratio of lamina propria to tracheal cartilage was lower in epidermal growth factor-loaded gelatin microsphere recipients; these mice also had almost complete differentiation of regenerated epithelium into ciliated columnar epithelium on days 35 and 52, earlier than in other groups.. Gelatin microspheres act as a functional vector for epidermal growth factor. Sustained local application of epidermal growth factor could accelerate reepithelialization of tracheal allografts.

Topics: Animals; Cell Differentiation; Cell Proliferation; Cilia; Delayed-Action Preparations; Drug Carriers; Epidermal Growth Factor; Epithelial Cells; Fibrosis; Gelatin; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Microscopy, Electron, Scanning; Microspheres; Regeneration; Respiratory Mucosa; Surface Properties; Time Factors; Trachea; Transplantation, Homologous

Topics: Animals; Cicatrix; Epidermal Growth Factor; Fibrosis; Humans; Inflammation; Interferon-gamma; Interleukin-10; Ligands; Mice; Models, Biological; Receptors, CXCR3; Signal Transduction; Stress, Mechanical; Wound Healing

Tumors are characterized by extracellular matrix (ECM) remodeling and stiffening. The importance of ECM remodeling to cancer is appreciated; the relevance of stiffening is less clear. We found that breast tumorigenesis is accompanied by collagen crosslinking, ECM stiffening, and increased focal adhesions. Induction of collagen crosslinking stiffened the ECM, promoted focal adhesions, enhanced PI3 kinase (PI3K) activity, and induced the invasion of an oncogene-initiated epithelium. Inhibition of integrin signaling repressed the invasion of a premalignant epithelium into a stiffened, crosslinked ECM and forced integrin clustering promoted focal adhesions, enhanced PI3K signaling, and induced the invasion of a premalignant epithelium. Consistently, reduction of lysyl oxidase-mediated collagen crosslinking prevented MMTV-Neu-induced fibrosis, decreased focal adhesions and PI3K activity, impeded malignancy, and lowered tumor incidence. These data show how collagen crosslinking can modulate tissue fibrosis and stiffness to force focal adhesions, growth factor signaling and breast malignancy.

Topics: Aging; Animals; Breast Neoplasms; Collagen; Epidermal Growth Factor; Extracellular Matrix; Female; Fibrosis; Genes, ras; Humans; Integrins; Mammary Glands, Human; Mice; Mice, Inbred BALB C; Protein-Lysine 6-Oxidase; Signal Transduction

Cell-specific targeting to renal tubular cells is an interesting approach to enhance the accumulation of drugs in the kidney. Low molecular weight proteins are rapidly filtered and extensively accumulate in proximal tubular cells. We therefore have used lysozyme (LZM, 14 kDa) as a tubular cell-specific carrier for the delivery of kinase inhibitors. Two different kinase inhibitors (LY364947 and erlotinib, directed to either the TGF-beta receptor kinase or the EGF receptor) were individually conjugated to LZM via a novel platinum-based linker (Universal Linkage System; ULS). The cellular handling and pharmacological efficacy of the conjugates were evaluated in cultured proximal tubular cells (HK-2 cells). Both conjugates were efficiently internalized via endocytosis. TGF-beta or EGF activated HK-2 cells showed a strong activation of the studied kinases and the conjugates inhibited these events, as was demonstrated by Western blotting of phosphorylated downstream mediators and quantitative gene expression analysis. In conclusion, we have developed tubular cell-specific kinase inhibitor-LZM conjugates via a novel linker strategy, which both showed to be effective in vitro. Future in vivo studies should show their potential for the treatment of renal diseases.

Topics: Cells, Cultured; Chemistry, Pharmaceutical; Drug Carriers; Endocytosis; Epidermal Growth Factor; ErbB Receptors; Erlotinib Hydrochloride; Fibrosis; Gene Expression Regulation; Humans; Intercellular Signaling Peptides and Proteins; Kidney Tubules, Proximal; Kinetics; Muramidase; Phosphorylation; Protein Kinase Inhibitors; Pyrazoles; Pyrroles; Quinazolines; Receptors, Transforming Growth Factor beta; Signal Transduction; Transforming Growth Factor beta

Apoptosis and epithelial-mesenchymal transdifferentiation (EMT) occur in stressed tubular epithelial cells and contribute to renal fibrosis. Transforming growth factor (TGF)-beta(1) promotes these responses and we examined whether the processes were interdependent in vitro. Direct (caspase inhibition) and indirect [epidermal growth factor (EGF) receptor stimulation] strategies were used to block apoptosis during TGF-beta(1) stimulation, and the subsequent effect on EMT was assessed. HK-2 cells were exposed to TGF-beta(1) with or without preincubation with ZVAD-FMK (pan-caspase inhibitor) or concomitant treatment with EGF plus or minus preincubation with LY-294002 (PI3-kinase inhibitor). Cells were then assessed for apoptosis and proliferation by flow cytometry, crystal violet assay, and Western blotting. Markers of EMT were assessed by microscopy, immunofluorescence, real-time RT-PCR, Western blotting, PAI-1 reporter assay, and collagen gel contraction assay. TGF-beta(1) caused apoptosis and priming for staurosporine-induced apoptosis. This was blocked by ZVAD-FMK. However, ZVAD-FMK did not prevent EMT following TGF-beta(1) treatment. EGF inhibited apoptosis and facilitated TGF-beta(1) induction of EMT by increasing proliferation and accentuating E-cadherin loss. Additionally, EGF significantly enhanced TGF-beta(1)-induced collagen I gel contraction. EGF increased Akt phosphorylation during EMT, and the prosurvival effect of this was confirmed using LY-294002, which reduced EGF-induced Akt phosphorylation and reversed its antiapoptotic and proproliferatory effects. TGF-beta(1) induces EMT independently of its proapoptotic effects. TGF-beta(1) and EGF together lead to EMT. EGF increases proliferation and resistance to apoptosis during EMT in a PI3-K Akt-dependent manner. In vivo, EGF receptor activation may assist in the selective survival of a transdifferentiated, profibrotic cell type.

Topics: Apoptosis; Blotting, Western; Caspase Inhibitors; Cell Differentiation; Cell Survival; Epidermal Growth Factor; Epithelium; ErbB Receptors; Fibrosis; Humans; Kidney; Kidney Tubules, Proximal; Mesoderm; Phosphatidylinositol 3-Kinases; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta; Transforming Growth Factor beta1

Hepatic ischemia-reperfusion injury is an inevitable consequence during liver surgery. The outcome is particularly poor in cirrhotic livers, which are more prone to hepatic ischemia-reperfusion injury. We aim to study whether FTY720 could attenuate hepatic ischemia-reperfusion injury both in normal and in cirrhotic livers. We applied a 70% liver-ischemia (60 min) model in rats with normal or cirrhotic livers. FTY720 was given 20 min before ischemia and 10 min before reperfusion (1 mg/kg, i.v.). Liver tissues and blood were sampled at 20 min, 60 min, 90 min, 6 h and 24 h after reperfusion for detection of MAPK-Egr-1, Akt pathways and caspase cascade. Hepatic ultrastructure and apoptosis were also compared. FTY720 significantly improved liver function in the rats with normal and cirrhotic livers. Akt pathway was activated at 6 and 24 h after reperfusion. FTY720 significantly down-regulated Egr-1, ET-1, iNOS and MIP-2 accompanied with up-regulation of A20, IL-10, HO-1 and Hsp70. MAPK (Raf-MEK-Erk) pathway was down-regulated. Hepatic ultrastructure was well maintained and fewer apoptotic liver cells were found in the FTY720 groups. In conclusion, FTY720 attenuates ischemia-reperfusion injury in both normal and cirrhotic livers by activation of cell survival Akt signaling and down-regulation of Egr-1 via Raf-MEK-Erk pathway.

Topics: Animals; Apoptosis; Blotting, Western; Chemokine CXCL2; Chemokines, CXC; DNA Primers; Down-Regulation; Endothelin-1; Epidermal Growth Factor; Extracellular Signal-Regulated MAP Kinases; Fibrosis; Fingolimod Hydrochloride; Gene Expression Regulation; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Hepatocytes; HSP70 Heat-Shock Proteins; Immunosuppressive Agents; In Situ Nick-End Labeling; Inflammation; Intercellular Signaling Peptides and Proteins; Interleukin-10; Liver; Male; MAP Kinase Signaling System; Microscopy, Electron; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; p38 Mitogen-Activated Protein Kinases; Propylene Glycols; Proteins; raf Kinases; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Sphingosine; Time Factors; Up-Regulation

Fibroblast proliferation, differentiation into myofibroblasts, and increased collagen synthesis are key events during both normal wound repair and fibrotic lesion formation. Here we report that these biological responses to TGF-beta by fibroblasts are regulated via a CTGF-dependent pathway in concert with either EGF or IGF-2. Our studies indicate these responses to TGF-beta are mutually exclusive, and cells that are proliferating do not express alpha-SMA or elevated levels of collagen synthesis. Cells expressing alpha-SMA do not exhibit DNA synthesis but do coexpress higher levels of types I and III collagen mRNA. Thus, fibroblast proliferation and differentiation are controlled by combinatorial signaling pathways involving not only components of the TGF-beta/CTGF pathway, but also signaling events induced by EGF and IGF-2-activated receptors. Collectively, our studies indicate TGF-beta functions as a classic embryonic inducer, initiating a cascade that is controlled by other factors in the cellular environment. We propose a model for this process with regard to wound repair and fibrotic lesion formation that is likely applicable to other instances of CTGF action during embryogenesis.

Topics: Actins; Animals; Cell Differentiation; Cell Line; Collagen; Connective Tissue Growth Factor; DNA Replication; Epidermal Growth Factor; Fibroblasts; Fibrosis; Gene Expression Regulation; Humans; Immediate-Early Proteins; Insulin-Like Growth Factor II; Intercellular Signaling Peptides and Proteins; Kidney; Myoblasts; Rats; Recombinant Proteins; RNA, Messenger; Signal Transduction; Transforming Growth Factor beta; Transforming Growth Factor beta1; Wound Healing

Growth factors produced by airway epithelial cells may be important in the pathogenesis of subepithelial fibrosis, a distinctive lesion of chronic human asthma.. To examine the relationship between the development of subepithelial fibrosis and the expression of transforming growth factor-beta 1 (TGF-beta 1) and ligands for the epidermal growth factor receptor.. BALB/c mice sensitized to ovalbumin were chronically challenged by inhalation of low levels of antigen, leading to development of subepithelial fibrosis and other changes of airway wall remodelling. Growth factor expression was assessed by immunohistochemistry and enzyme immunoassay.. Allergic sensitization directly correlated with airway epithelial expression of both the cleaved, potentially biologically active form of TGF-beta 1 and of amphiregulin in response to allergen challenge. Accumulation of TGF-beta 1 was related to remodelling of the airway wall in chronic asthma, whereas expression of amphiregulin did not exhibit a similar relationship. Production of epithelial cell-derived TGF-beta 1 appeared to be regulated by IL-13, while both IL-13 and CD4(+) T cells regulated accumulation of TGF-beta 1. In contrast to results reported in high-level exposure models of airway fibrosis, eosinophils did not appear to be a significant source of TGF-beta 1.. Airway epithelial cell-derived TGF-beta 1 has a potentially crucial role in the development of airway wall remodelling in asthma. Immunological mechanisms may regulate the release and accumulation of TGF-beta 1.

Topics: Allergens; Amphiregulin; Animals; Asthma; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Chronic Disease; Disease Models, Animal; EGF Family of Proteins; Epidermal Growth Factor; Epithelial Cells; Female; Fibrosis; Glycoproteins; Immunoenzyme Techniques; Intercellular Signaling Peptides and Proteins; Interleukin-13; Ligands; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Mucosa; Trachea; Transforming Growth Factor beta; Transforming Growth Factor beta1

Endothelin-1 (ET-1) has been implicated in pathologic remodelling and tissue repair processes in the heart. We investigated the effects of ET-1 on growth and collagen synthesis responses in cardiac fibroblasts isolated from human hearts. We also studied the receptor subtype(s) mediating such responses and the factors regulating their expression.. Fibroblasts were isolated from cardiac transplant recipient hearts and characterised by immunocytochemistry. Serum-starved cells were exposed to ET-1 and incorporation of [3H]proline and thymidine were measured as indexes of collagen and DNA synthesis respectively. Blocking experiments utilised the selective ETA receptor antagonist BQ123 and the ETB antagonist BQ788.. ET-1 elicited a potent collagen synthesis response in cardiac fibroblasts, with a maximum 29+/-5% increase that was abolished by BQ123. Cardiac fibroblasts responded to ET-1 with a concentration-dependent decrease in DNA synthesis rate. The effects of ET-1 were similar to those of TGF-beta. Radioligand binding studies revealed the presence of high-affinity ET-1 binding sites on these cells, which were upregulated by treatment with the growth factors PDGF and EGF but downregulated by TGF-beta.. These results therefore implicate ET-1 as a trophic agent in the human heart with the ability to influence the development of cardiac fibrosis.

Topics: Cell Proliferation; Cells, Cultured; Collagen; DNA; Endothelin A Receptor Antagonists; Endothelin-1; Epidermal Growth Factor; Fibroblasts; Fibrosis; Humans; Myocardium; Oligopeptides; Peptides, Cyclic; Piperidines; Platelet-Derived Growth Factor; Receptor, Endothelin A; Transforming Growth Factor beta

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is expressed in both normal pancreatic islets and in pancreatic cancers, but its role in pancreatic physiology and disease is not known. This report examines the effects of HB-EGF overexpression in mouse pancreas.. Transgenic mice were established using a tissue-specific promoter to express an HB-EGF complementary DNA in pancreatic beta cells, effectively elevating HB-EGF protein 3-fold over endogenous levels.. Mice overexpressing HB-EGF in pancreatic islets showed both endocrine and exocrine pancreatic defects. Initially, islets from transgenic mice failed to segregate alpha, beta, delta, and PP cells appropriately within islets, and had impaired separation from ducts and acini. Increased stroma was detected within transgenic islets, expanding with age to cause fibrosis of both endocrine and exocrine compartments. In addition to these structural abnormalities, subsets of transgenic mice developed profound hyperglycemia and/or proliferation of metaplastic ductal epithelium. Both conditions were associated with severe stromal expansion, suggesting a role for islet/stromal interaction in the onset of the pancreatic disease initiated by HB-EGF. Supporting this conclusion, primary mouse fibroblasts adhered to transgenic islets when the 2 tissues were cocultured in vitro, but did not interact with nontransgenic islets.. An elevation in HB-EGF protein in pancreatic islets led to altered interactions among islet cells and among islets, stromal tissues, and ductal epithelium. Many of the observed phenotypes appeared to involve altered cell adhesion. These data support a role for islet factors in the development of both endocrine and exocrine disease.

Topics: Animals; Cell Communication; Cell Division; Epidermal Growth Factor; Epithelial Cells; Fibroblasts; Fibrosis; Gene Expression; Heparin-binding EGF-like Growth Factor; Intercellular Signaling Peptides and Proteins; Islets of Langerhans; Metaplasia; Mice; Mice, Transgenic; Pancreatic Diseases; Pancreatic Ducts; Phenotype

Tubular atrophy is a major feature of most renal diseases and is closely associated with loss of renal function. The present study sought to investigate tubular epithelial cell apoptosis in experimental diabetic nephropathy and to explore the role of pro-apoptotic [transforming growth factor-beta (TGF-beta) and anti-apoptotic growth factors [epidermal growth factor (EGF)]. The effects of renoprotective therapy with blockade of the renin-angiotensin system (RAS) also were examined.. Six-week-old female Ren-2 rats were injected with streptozotocin (STZ) and maintained diabetic for 12 weeks. Further groups of diabetic rats were treated with the angiotensin-converting enzyme (ACE) inhibitor, perindopril, or the angiotensin II type 1 (AT1) receptor antagonist, valsartan, for 12 weeks.. Widespread apoptosis, identified immunohistochemically by single stranded DNA and TUNEL, was noted in the tubules of diabetic Ren-2 rats. These changes were associated with a 50% decrease in EGF expression and a twofold increase in TGF-beta1 mRNA. Treatment of diabetic Ren-2 rats with either valsartan (20 mg/kg/day) or perindopril (6 mg/kg/day) reduced apoptosis to control levels in association with supranormal levels of EGF mRNA (P < 0.01) and a reduction in TGF-beta1 gene expression (P < 0.05) to that of control rats.. Tubular apoptosis is a prominent feature of diabetic Ren-2 rats that is attenuated by blockade of the RAS in association with modulation of pro- and anti-apoptotic growth factor expression.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; Animals, Genetically Modified; Antihypertensive Agents; Apoptosis; Atrophy; Autoradiography; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Epidermal Growth Factor; Female; Fibrosis; Gene Expression; In Situ Hybridization; In Situ Nick-End Labeling; Kidney Tubules; Nephritis, Interstitial; Perindopril; Rats; Renin-Angiotensin System; RNA, Messenger; Tetrazoles; Transforming Growth Factor beta; Valine; Valsartan

During progression of chronic renal disease, qualitative and quantitative changes in the composition of tubular basement membranes (TBMs) and interstitial matrix occur. Transforming growth factor (TGF)-beta(1)-mediated activation of tubular epithelial cells (TECs) is speculated to be a key contributor to the progression of tubulointerstitial fibrosis. To further understand the pathogenesis associated with renal fibrosis, we developed an in vitro Boyden chamber system using renal basement membranes that partially mimics in vivo conditions of TECs during health and disease. Direct stimulation of TECs with TGF-beta(1)/epithelial growth factor results in an increased migratory capacity across bovine TBM preparations. This is associated with increased matrix metalloproteinase (MMP) production, namely MMP-2 and MMP-9. Indirect chemotactic stimulation by TGF-beta(1)/EGF or collagen type I was insufficient in inducing migration of untreated TECs across bovine TBM preparation, suggesting that basement membrane integrity and composition play an important role in protecting TECs from interstitial fibrotic stimuli. Additionally, neutralization of MMPs by COL-3 inhibitor dramatically decreases the capacity of TGF-beta(1)-stimulated TECs to migrate through bovine TBM preparation. Collectively, these results demonstrate that basement membrane structure, integrity, and composition play an important role in determining interstitial influences on TECs and subsequent impact on potential aberrant cell-matrix interactions.

Topics: Adult; Animals; Basement Membrane; Cattle; Cell Movement; Cells, Cultured; Chemotaxis; Electrophoresis, Polyacrylamide Gel; Epidermal Growth Factor; Epithelial Cells; Extracellular Matrix; Fibrosis; Humans; Kidney Failure, Chronic; Kidney Tubules; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Rabbits; Transforming Growth Factor beta; Transforming Growth Factor beta1

Epidermal growth factor (EGF) plays an important role in renal tubular regeneration after ischemic injury in kidney. The present study reports the association between the renin-angiotensin system (RAS) and EGF, and the effect of angiotensin II blockade with losartan (LSRT) on EGF expression in an experimental model of chronic cyclosporine (CsA) nephrotoxicity in rats.. Two separate experiments were performed. In the first experiment, rats on the normal-salt diet (NSD; 0.3%) or low-salt diet (LSD; 0.05%) were treated with or without LSRT for four weeks. In the second experiment, rats on the NSD or LSD were given vehicle (VH group, olive oil, 1 mg/kg per day) or CsA (15 mg/kg per day) or CsA (15 mg/kg per day) plus LSRT (100 mg/L per day). Renal function, histopathology, TUNEL staining, plasma renin activity (PRA), and the expression of renin and EGF were studied.. Normal rats on the LSD showed significantly increased EGF expression (cortex, 2.6-fold; medulla, 1.7-fold) and significantly decreased EGF expression with the LSRT treatment compared with the rats treated with the NSD (cortex, 74.8 vs. 10%; medulla, 22.5 vs. 5%). In contrast, the CsA-treated rats on the LSD had a significantly lower EGF expression (cortex, 98 vs. 53%; medulla, 94 vs. 14%); however, concomitant administration of LSRT increased the EGF expression (cortex, 91- vs. 3.8-fold; medulla, 19- vs. 2.4-fold) compared with the rats on the NSD. In the normal and CsA-treated LSD rats, EGF expression was well correlated with PRA. In addition, EGF expression was well correlated with the interstitial fibrosis score (r = 0.664, P < 0.01) or number of TUNEL-positive cells (r = 0.822, P < 0.01) in CsA-treated LSD rats.. These results suggest that angiotensin II blockade with LSRT decreases EGF expression in normal rats on the LSD, but it protects EGF expression in CsA-induced nephrotoxicity. This finding provides a new perspective on the renoprotection of angiotensin II blockade in chronic CsA nephrotoxicity.

Topics: Angiotensin II; Animals; Apoptosis; Blood Pressure; Cyclosporine; Diet, Sodium-Restricted; Epidermal Growth Factor; Fibrosis; Immunohistochemistry; In Situ Nick-End Labeling; Kidney Glomerulus; Losartan; Male; Rats; Rats, Sprague-Dawley; Renin; Renin-Angiotensin System

Obstructive nephropathy is a primary cause of renal failure in infancy. Chronic unilateral ureteral obstruction (UUO) in the neonatal rat results in reduced renal expression of epidermal growth factor (EGF), renal tubular epithelial (RTE) cell apoptosis and interstitial fibrosis. We wished to determine whether these changes could be prevented by exogenous administration of EGF.. Thirty-three Sprague-Dawley rats underwent UUO within the first 48 hours of life, and received daily injections of either EGF (0.1 mg/kg/day) or saline (control) for the following seven days, after which obstructed and intact opposite kidneys were removed for study. These were compared to 11 sham-operated rats that received either no injections, EGF injections, or saline injections. Renal cell proliferation was determined by proliferating cell nuclear antigen, apoptosis was measured by the TUNEL technique, and the distribution of vimentin, clusterin, transforming growth factor-beta 1 (TGF-beta 1), and alpha-smooth muscle actin were determined by immunohistochemistry. Tubular dilation, tubular atrophy, and interstitial collagen deposition were quantitated by histomorphometry.. Compared to controls, EGF treatment increased RTE cell proliferation in the obstructed kidney by 76%, decreased apoptosis by 80%, and reduced vimentin, clusterin and TGF-beta 1 immunostaining (all P < 0.05). EGF treatment reduced tubular dilation by 50%, atrophic tubules by 30%, and interstitial fibrosis by 50% (all P < 0.05). There was no significant effect of EGF on renal alpha smooth muscle actin distribution. There was no effect of saline or EGF injections on kidneys from sham-operated rats for any of the parameters studied.. We conclude that EGF stimulates RTE cell proliferation and maturation and reduces apoptosis in the neonatal rat kidney subjected to chronic UUO. These effects may contribute to the reduction in tubular dilation, tubular atrophy, and interstitial fibrosis. By preserving renal development, administration of EGF attenuates the renal injury resulting from chronic UUO.

Topics: Animals; Animals, Newborn; Apoptosis; Cell Division; Chronic Disease; Clusterin; Complement Inactivator Proteins; Epidermal Growth Factor; Fibrosis; Glycoproteins; Kidney; Kidney Tubules; Ligation; Molecular Chaperones; Proliferating Cell Nuclear Antigen; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta; Ureter; Ureteral Obstruction; Urothelium; Vimentin

It has been assumed that lens epithelial cells (LECs) existing at the capsulotomy edge have been traumatized through anterior capsulotomy in cataract extraction. In this study, the correlation between traumatized LECs remaining at the anterior capsulotomy edge and epidermal growth factor (EGF) found in the aqueous humor, a cell growth factor though to affect cell morphology, was determined. Anterior lens capsules with adhering LECs were obtained following anterior capsulotomy performed during cataract surgery to first confirm the presence of EGF receptors on LECs, which are needed for EGF to be biologically active. Besides, to identify any EGF receptors on traumatized LECs, I next intentionally traumatized the cells by pressing them with a forceps from the anterior capsular side. It has been found that the LECs containing EGF receptors were always those existing at the edge of the anterior capsular opening and LECs containing EGF receptors existed along the pressed region too. The present results indicate that traumatized LECs along the capsulotomy edge have undergone changes to manifest EGF receptors, thus allowing EGF from the aqueous humor to become more active. The physiological effect of EGF upon these LECs may therefore be one of the causative factors of fibrous opacification of the anterior capsulotomy edge after cataract extraction.

Topics: Aged; Aged, 80 and over; Aqueous Humor; Cataract; Cataract Extraction; Coloring Agents; Epidermal Growth Factor; Epithelial Cells; ErbB Receptors; Fibrosis; Humans; Lens Capsule, Crystalline; Middle Aged; Postoperative Complications; Trypan Blue; Wounds, Nonpenetrating

The difference in the lifespan of dy and mdx mice could be due to different muscle regeneration capabilities. In mdx an involvement of bFGF in stimulating regeneration has been postulated. The aim of our work was to detect the presence, and to study the distribution, of muscular and connective tissue growth factors in mdx and dy mice at different stages of muscle pathology. From 7 to 10 weeks of age the difference between the two dystrophic mice becomes evident. At 13 weeks the dy mouse presents a predominance of fibrosis and degenerative muscular phenomena while the main pathological feature in mdx mouse is the muscle regeneration. In both animal models fibrosis proliferation is correlated to the presence of EGF and its receptor and TGF beta 1. bFGF was localized to regenerating and degenerating fibers in both dy and mdx mice. The bFGF presented a normal pattern in mdx mice at 20 weeks when regenerative and degenerative phenomena were no longer present. Our data suggest that growth factors could influence the outcome of muscular regenerative and degenerative processes.

Topics: Animals; Connective Tissue; Epidermal Growth Factor; ErbB Receptors; Fibroblast Growth Factor 2; Fibrosis; Growth Substances; Immunohistochemistry; Lymphotoxin-alpha; Mice; Mice, Inbred C57BL; Muscles; Muscular Dystrophies

The roles of growth factors in the pathogenesis of various forms of acute and chronic renal disease are largely putative. Nevertheless, there is a growing body of information that links specific growth factors to particular forms of renal injury. In all instances, it is supposed that such associations are not necessarily unique and that multiple cytokines probably interact to determine the pattern of injury or the regenerative response to such injury. Regeneration of tubular epithelium after acute tubular necrosis involves upregulation of the epidermal growth factor (EGF) receptor. Early studies of exogenously administered EGF indicate that the severity and duration of renal failure may be attenuated by this growth factor. Thus far, the observed responses have been limited and the role of EGF as a therapeutic agent requires more study. The mechanism of generation of tubulointerstitial injury in most forms of renal disease is difficult to understand. Early in vitro studies of growth factor production by tubular cells (in the absence of any infiltrating cells) indicate that platelet-derived growth factor produced by the medullary collecting duct is mitogenic for renal medullary fibroblasts, suggesting a paracrine growth system in this region of the kidney. Insulin-like growth factor I has also been shown to be produced by collecting duct cells. Its production is increased by EGF, and its association with certain forms of renal hypertrophy, i.e., diabetes and hypersomatotrophic states, implies its participation in the hypertrophic growth response. Platelet-derived growth factor is a potent mitogen for glomerular mesangial cells, and its production is regulated by a variety of cytokines.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Cell Division; Epidermal Growth Factor; Fibrosis; Glomerular Mesangium; Growth Substances; Humans; Hypertrophy; Insulin-Like Growth Factor I; Kidney; Kidney Diseases; Kidney Glomerulus; Nephrectomy; Platelet-Derived Growth Factor; Rats; Regeneration; Transforming Growth Factor beta