Tropoelastin Gene Expression Research Articles

Contribution of the Elastic Component and Venous Wall Arterialization in Patients with Venous Reflux

Chronic venous disease (CVeD) is defined as a set of disorders affecting the venous system mainly manifested in the form of varicose veins. CVeD is characterized by a sustained venous hypertension, leading to a plethora of functional and structural changes in the vein that may cause valve incompetence and pathologic reflux. In turn, venous reflux aggravates the venous hypertension and enhances the progression of CVeD into the most advanced stages. Previous studies have proposed that there are several alterations in the venous wall preceding the valve dysfunction and venous reflux. Besides, it has also been identified that young patients with CVeD present premature aging and changes in the venous wall composition that may be related to the presence of venous reflux. In this context, the aim of the present study is to examine the possible pathophysiological role of elastic fibers and their precursors in the venous wall of patients with reflux in comparison to those without reflux, considering the variable age in both groups (<50 years and ≥50 years). We performed immunohistochemical and quantitative polymerase chain reaction (PCR) in order to assess the protein and gene expression of tropoelastin, fibrillin-1, fibulins 4 and 5, lysyl oxidase and lysyl oxidase like 1, respectively. In parallel, we assessed the elastin content through histological techniques (orcein stain) in this group of patients. Our results show significant changes in elastic fibers and their precursors in young patients with pathologic reflux when compared with elder patients with reflux and young patients without reflux. These variations suggest that the venous system of young patients with venous reflux appears to present an enhanced dynamism and arterialization of the venous wall, which may be associated with a premature aging and pathological environment of the tissue.

Marked disruption and aberrant regulation of elastic fibres in early striae gravidarum.

Striae gravidarum (SG), or 'stretch marks' of pregnancy, begin as erythematous streaks, and mature over months to years to become permanent scar-like bands that may be hypopigmented, atrophic and lax. To investigate early molecular alterations that may promote laxity of mature SG, we investigated the dermal elastic fibre network, which provides human skin with elastic properties. We obtained skin samples of newly developed, erythematous abdominal SG in healthy pregnant women. The elastic fibre network was examined by Verhoeff elastic staining and immunofluorescence staining of skin sections. Gene expression was measured by real-time polymerase chain reaction. The normal elastic fibre network appeared markedly disrupted in SG, compared with perilesional abdominal skin or control (normal-appearing hip skin). This disruption was accompanied by the emergence of short, disorganized, thin, thread-like 'fibrils', which were observed prominently in the mid-to-deep dermis. These fibrils were rich in tropoelastin (the main component of normal elastic fibres), and persisted into the postpartum period without forming normal-appearing elastic fibres. The emergence of these fibrils was accompanied by increased gene expression of tropoelastin and fibrillin-1, but not other elastic fibre components, including fibrillin-2 and fibulin-1, -2 or -5. In early SG, the elastic fibre network appears markedly disrupted, and newly synthesized tropoelastin-rich fibrils emerge, likely as a result of uncoordinated synthesis of elastic fibre components. Because they are thin and disorganized, tropoelastin-rich fibrils likely do not function as normal elastic fibres do. These observations provide the foundations for elucidating pathogenic mechanisms by which laxity may develop in SG.

Aberrant elastin remodeling in the lungs of O₂-exposed newborn mice; primarily results from perturbed interaction between integrins and elastin.

Excessive localization of elastin from septal tips to alveolar walls is a key feature of bronchopulmonary dysplasia (BPD). The abnormal accumulation of lung elastin, involving the structural and functional interaction of a series of proteins, remains poorly understood. To further investigate the mechanisms accounting for the abnormal accumulation of elastin in the lungs of newborn mice with BPD, we evaluate elastin distribution and its interaction with proteins involved in its aberrant localization, such as integrin αv, fibulin-5 and transforming growth factor β1 (TGF-β1), in lungs of newborn mice exposed to 60% O2 for 21 days. Lung histology revealed aberrant elastin production and impaired lung septation in O2-exposed lungs, while tropoelastin, integrin αv, fibulin-1, fibulin-2 and fibulin-4 gene expression were elevated. Dual staining image analysis of lung sections revealed that co-localization of integrin αv and elastin increased following O2 exposure with elastin distributed throughout the walls of air spaces rather than at septal tips. Furthermore, integrin αv appeared to be induced initially. Concurrently, increased fibulin-5 and TGF-β1 (which may regulate elastic fiber assembly) expression was detected, which may explain the altered lung elastin deposition and defective septation that are observed during BPD. These data support the hypothesis that excessive and aberrant αv integrin expression was initially induced by hyperoxia; αv integrin then interacted with and recruited elastin. These alterations were accompanied by fibulin-5 deposition and TGF-β1 activation, which may impede normal matrix remodeling, thereby contributing to the pathological pulmonary features of BPD.

Meconium Fatty Acid Ethyl Esters as Biomarkers of Late Gestational Ethanol Exposure and Indicator of Ethanol-Induced Multi-Organ Injury in Fetal Sheep

BackgroundMeconium fatty acid ethyl esters (FAEE) constitute a biomarker of heavy fetal ethanol exposure. Our objective was to measure meconium FAEE in fetal sheep following daily, relatively moderate-dose ethanol exposure in late gestation, and to evaluate their utility in identifying fetal organ-system injury.MethodsPregnant ewes received ethanol (0.75 g/kg; n = 14) or saline (n = 8) via 1-h IV infusion daily during the third trimester equivalent, while additional pregnant sheep served as untreated controls (n = 6). The daily ethanol regimen produced similar maximal maternal and fetal plasma ethanol concentrations of 0.11–0.12 g/dL. Ewes and fetuses were euthanized shortly before term, and meconium was collected and analyzed for FAEE (ethyl palmitate, stearate, linoleate, and oleate).ResultsMeconium total FAEE concentration was significantly higher in ethanol-exposed fetuses compared with controls, and a positive cut-off of 0.0285 nmol total FAEE/g meconium had 93.3% sensitivity and specificity for detecting fetal ethanol exposure. When the studied animals (ethanol-exposed and controls) were classified according to meconium FAEE concentration, FAEE-positive and FAEE-negative groups frequently differed with respect to previously examined pathological endpoints, including nephron endowment, lung collagen deposition, cardiomyocyte maturation, and tropoelastin gene expression in cerebral vessels. Furthermore, in all studied animals as a group (ethanol-exposed and controls combined), meconium FAEE concentration was correlated with many of these pathological endpoints in fetal organs.ConclusionsWe conclude that, in fetal sheep, meconium FAEE could serve as a biomarker of daily ethanol exposure in late gestation and could identify fetuses with subtle ethanol-induced toxic effects in various organs. This study illustrates the potential for using meconium FAEE to identify neonates at risk for dysfunction of major organs following in-utero ethanol exposure that does not result in overt physical signs of ethanol teratogenicity.

Transdermal administration of lactoferrin with sophorolipid1This article is part of a Special Issue entitled Lactoferrin and has undergone the Journal's usual peer review process.

Lactoferrin (Lf), a multifunctional glycoprotein, is known to activate dermal fibroblasts. Enhancing percutaneous absorption without decreasing the activity of Lf is critical in making the dermal administration of Lf beneficial. Sophorolipid (SL), a glycolipid-type biosurfactant, is known to form assemblies that may elevate the efficiency of the transdermal delivery of active ingredients. Here, we investigated the role of SL in the transdermal absorption of bovine Lf (bLf) and the effect of SL on the bLf activity on dermal fibroblasts. Transdermal absorption of bLf through a model skin was enhanced by 1.3-fold to 1.7-fold when SL was added. The effects of SL on the bLf activities on dermal fibroblasts were examined by cell proliferation activities and by gene expression levels of elastic fiber components, collagen IV, and hyaluronan synthases, revealing that SL did not depress the effect of bLf to any extent. Instead, the tropoelastin gene expression was upregulated ∼60-fold by bLf alone, which was further increased to ∼160-fold by bLf and SL together, suggesting a significant synergism between bLf and SL. Protein levels of elastin, assessed by immunohistochemistry, correlated well with the results of gene expressions. These results indicate the feasibility of the transdermal administration of bLf with SL.

Abstract 233: Quantification of Histological and Biomechanical Properties of Tissue-Engineered Vascular Graft in Inferior Vena Cava

Background: We developed the first tissue engineered vascular graft (TEVG) for use in congenital heart surgery and confirmed its significant potential via an initial clinical trial (Shinoka et al., JTCVS 2005). The primary complication of TEVG at medium term was stenosis attributed to suboptimal neovessel remodeling, which necessitated an investigation of the mechanics and mechanobiology to predict diverse aspects of the neovessel formation. Here we present the novel tools to investigate the evolving biomechanical properties of TEVG in a mouse inferior vena cava (IVC) replacement model. Hypothesis: The biomechanical properties of a TEVG evolve nonlinearly in time from a construct-dominated stiffness to a neovessel dominated stiffness. Novel biaxial biomechanical tests are required to quantify such changes rigorously. Method: Thirty-six CB17 SCID/beige mice were implanted with TEVG (PGA + P[CL/LA]) as IVC interposition graft. Twelve tissue engineered neovessels were harvested at 2, 6, and 12 weeks after implantation. Neovessels were characterized biomechanically using a custom, computer-controlled biaxial testing device, and compared with native veins. Extracellular matrix (ECM) remodeling of the neovessel was characterized histologically, biochemically, and molecular biologically. Result: The biaxial data revealed an improved compliance of the neovessel over time. Similarly, the axial stretch response of the TEVG became more like the native vein at 12 weeks. This response was quantified as a ratio of neo-vessel to native vein and improved to reach 52% at 12 weeks from 11% at 2 weeks. Scaffold mass in vivo, estimated by remaining scaffold on histology, showed a significant decline at 6 and 12 weeks. Gene expression of both type I and III collagen peaked at 2 weeks, and total collagen mass quantified by a Sircol™ assay revealed that total collagen peaked at 2 weeks but decreased gradually to the level of native vein. Gene expression of both tropoelastin and fibrillin-1 peaked at 2 weeks, whereas elastin mass quantified by Fastin™ assay showed a delayed peak level of elastin at 6 weeks. Gene expression of MMP-2 and 9 peaked at 6 and 2 weeks, respectively, implicating robust ECM remodeling at early time points. Conclusion: Our novel histo-mechanical approach is the first to show that neovessel formation is a dynamic process characterized by progressive degradation of the scaffold and increased ECM remodeling, which yields biomechanical properties of the TEVG similar to native vessel within 12 weeks in vivo.

Restoring Elastin With MicroRNA-29

Elastin (Eln) is a highly distensible protein that enables tissues such as lung, skin, and large arteries to expand and return to their original shape.1 Tropoelastin is encoded by a single copy gene that is highly expressed in utero and then strongly downregulated after birth.2,3 Only low maintenance levels of tropoelastin mRNA are found in most elastic tissues in adults.3,4 Repair of severe injuries, such as third-degree burns, produces tissue that is often heavily scarred and severely limited in flexibility, range of motion, and tissue function.5 Similar scarring is seen after prolonged cancer radiation therapy, in sun-damaged skin,6 and in systemic sclerosis.7,8 In the vasculature, Eln haploinsufficient phenotypes include narrowing (stenosis) of the ascending aorta in the perinatal period and hypertension later in life. Overdistention injury of arteries after balloon catheter-based therapies activates repair responses in the adventitia that closely resemble skin wound contractures and can drive lumen narrowing in restenosis.9 These aberrant conditions have in common the production of a collagen-rich extracellular matrix (ECM) that is deficient in elastic fibers. A major cause of the deficiency in elastic fiber production is the failure to upregulate tropoelastin gene expression in postnatal tissues subject to the injuries described above. An article by Zhang et al in this issue of Arteriosclerosis, Thrombosis, and Vascular Biology describes a novel approach to increase Eln production that may change clinical outcomes in conditions such as those described above.10 See accompanying article on page 756 The elastic fiber is built up by a stepwise addition of tropoelastin monomers on a microfibrillar scaffold (Figure). The core of a microfibril is composed of the glycoprotein fibrillin.11 Fibrillin is deposited in the extracellular space and once there interacts with a group of microfibril-associated proteins, …

Significant Decrease in Tropoelastin Gene Expression in Fibroblasts from a Japanese Costello Syndrome Patient with Impaired Elastogenesis and Enhanced Proliferation

Costello syndrome is a connective tissue disorder associated with sparse, thin, and fragmented elastic fibers in tissues. In this study we demonstrated a significant decrease in the expression of tropoelastin mRNA in fibroblasts derived from a Japanese Costello syndrome patient with impaired elastogenesis and enhanced proliferation. In contrast, there were no changes in expression of the Harvey ras (HRAS), fibrillin-1, fibulin-5, microfibril-associated glycoprotein-1 (MAGP-1), lysyl oxidase (LOX), or 67-kDa non-integrin elastin-binding protein (EBP) gene. The proliferative activity of the Costello fibroblasts was about 4-fold higher than that of the normal and pathological control ones. However, no mutations were detected in the coding region of HRAS mRNA. Transduction of the bovine tropoelastin (bTE) gene with the lentiviral vector restored the elastic fiber formation and decreased the growth rate in the Costello fibroblasts. These results strongly suggest that the defect of human tropoelastin (hTE) gene expression should induce the impaired elastogenesis and enhanced proliferation of Costello fibroblasts, and that a primary cause other than the HRAS gene mutation should contribute to the pathogenesis in the present Costello case.

Expression of tropoelastin in human periodontal ligament fibroblasts after simulation of orthodontic force

The ability of elastic fibers to respond to mechanical stimuli suggests that they play a central role in physiological adaptation to external stimuli including application of orthodontic force. The purpose of this study was to examine the effect of external pressure simulating orthodontic force on tropoelastin gene expression in cultured human periodontal ligament fibroblasts (hPDLF). External pressure simulation was achieved by centrifugation for 10, 30, 60, 90 and 120 min of hPDLF in a horizontal microplate rotor. Semi-quantitative RT-PCR analysis of tropoelastin mRNA was performed and β-actin was used as an internal invariant control. While centrifugal force on mRNA levels of β-actin showed almost no change, the mRNA levels of tropoelastin increased significantly to a peak level of more than four-fold after 30 min. Thereafter, at 60 min, the mRNA levels remained at more than three-fold. After 90 min, mRNA levels decreased to control levels. The finding that no changes in mRNA levels of β-actin occurred during the first 90 min of centrifugation validates its use as an invariant control gene in such an experimental model. This study demonstrated that tropoelastin is expressed in hPDLF and that the pressure caused significant time-dependent upregulation of the tropoelastin gene. The responsiveness of the tropoelastin gene to force shows its possible clinical importance in orthodontic tooth movement. Further studies, however, are essential in order to learn whether the high expression of the gene in vitro will also be followed by corresponding protein synthesis and deposition in vivo in the extracellular matrix (ECM) of the periodontal ligament (PDL).

Induction of fibulin-5 gene is regulated by tropoelastin gene, and correlated with tropoelastin accumulation in vitro

Fibulin-5 (also known as DANCE) is an elastin-binding protein that is thought to play a role in elastogenesis. We examined the relationship between the gene expression of fibulin-5 and the gene expression and accumulation of tropoelastin by comparing elastin-producing cells (human gingival fibroblasts) with non-elastin-producing cells (human periodontal ligament fibroblasts) by Northern blot analysis. Fibulin-5 gene induction was found only in elastin-producing cells. Induction of the fibulin-5 gene in elastin-producing cells occurred after induction of the tropoelastin gene, and the fibulin-5 level was reduced upon RNA interference-mediated down-regulation of tropoelastin. Fibulin-5 gene induction was also correlated with a rapid increase of tropoelastin accumulation within the cell layer. These results may suggest that the fibulin-5 gene induction is directly or indirectly regulated by tropoelastin gene expression and plays a role in the accumulation of elastic fibers within matrices.

Gene Expression and Accumulation of Fibrillin-1, Fibrillin-2, and Tropoelastin in Cultured Periodontal Fibroblasts

Gene Expression and Accumulation of Fibrillin-1, Fibrillin-2, and Tropoelastin in Cultured Periodontal Fibroblasts

Gene expression and accumulation of fibrillin-1, fibrillin-2, and tropoelastin in cultured periodontal fibroblasts.

The elastic system fibers consist of three types--oxytalan, elaunin, and elastic fibers--differing in their relative microfibril and elastin contents. All three types are found in human gingiva, but human periodontal ligaments contain only elastin-free fibers. We examined cultured human gingival fibroblasts (HGF) and cultured human periodontal ligament fibroblasts (HPLF) to determine the gene expression of fibrillin-1 and fibrillin-2 (the major components of microfibrils) and of tropoelastin. In addition, we assessed the degree of accumulation of these proteins in the extracellular matrix. Northern blot analysis revealed that the level of expression of fibrillin-1 and fibrillin-2 was higher in HGF than in HPLF. However, examination of matrix samples from HGF and HPLF cell layers showed that there was no difference in fibrillin-1 accumulation, although fibrillin-2 accumulated to a much greater extent in the HGF-derived matrix. Tropoelastin was expressed only in and around HGF. These results show a correlation between gene expression and the accumulation of tropoelastin and fibrillin-2 in HGF.

Biphasic response of tropoelastin at the poststenotic dilation segment of the rabbit aorta

Objective: The purpose of this study was to assess the time course of tropoelastin gene expression in the poststenotic dilatation segment of rabbit aorta with experimental coarctation. Methods: Midthoracic aortic coarctation was created in rabbits to produce a PSD. The time points of the study after coarctation were 1, 3, and 7 days and 2, 4, and 8 weeks (n = 3 each). Additional animals (n = 6) were subjected to hypercholesterolemia for analysis of tropoelastin expression in intimal lesions. Northern and Western blot analyses were used to quantitate tropoelastin messenger RNA (mRNA) and protein, and immunohistochemistry was used to analyze tropoelastin distribution. Results: Thoracic aortic coarctation produced a moderate stenosis, which resulted in PSD. mRNA levels in the PSD segment decreased at days 1 and 3, followed by an increase at 2 and 4 weeks ( P < .05 versus controls). This biphasic change in tropoelastin mRNA was associated with increase in tropoelastin protein levels at 2 and 4 weeks ( P < .05 versus controls). PSD diameter reached a maximum at 4 weeks and did not increase significantly thereafter. The number of medial elastic laminae in PSD was reduced slightly, but media thickness was unchanged. Intimal lesions were much smaller in the PSD segment than in the proximal segment in animals with hypertension superimposed with hypercholesterolemia. Moreover, tropoelastin protein distributed not only in the intima but also in the media of the PSD. Conclusion: Tropoelastin gene expression is regulated in a biphasic pattern and precedes PSD formation. The differential distribution of tropoelastin in the media suggests a role for tropoelastin in the poststenotic adaptation response, which may provide increased elasticity to the PSD wall. (J Vasc Surg 2002;36:605-12.)

Gene Expression of Tropoelastin Is Enhanced in the Aorta Proximal to the Coarctation in Rabbits

To assess elastin biosynthesis in the aortic wall in response to acute elevation of blood pressure, we studied the aortic gene expression of tropoelastin in a rabbit midthoracic aortic coarctation model. The time points of the study were 1, 3, and 7 days and 2, 4, and 8 weeks after coarctation. Additional animals were subjected to hypercholesterolemia for analysis of tropoelastin expression in the intimal lesion. mRNA for tropoelastin was quantitated by Northern blot analysis and its distribution was revealed by in situ hybridization. The 65-kDa tropoelastin was analyzed by Western blotting and immunohistochemistry. Tropoelastin mRNA proximal to the coarctation was increased at 2 weeks and returned to baseline by 8 weeks (P < 0.05 versus control). Changes in 65-kDa tropoelastin corresponded to those of mRNA. Tropoelastin gene was expressed mainly in the intima and in the outer media at the proximal region to the stenoses, which was particularly remarkable in the intimal lesion. The results indicate that tropoelastin gene expression was enhanced in the early remodeling response to elevated blood pressure. The distribution of newly synthesized tropoelastin in the outer media suggests a reenforcement role of tropoelastin, which preserves mechanical resiliency in response to changes in tensile stress.

Tropoelastin Expression by Periodontal Fibroblasts

Elastic system fibers are load-bearing proteins found in periodontal tissue. There are three types--oxytalan, elaunin, and elastic fibers--which differ in their relative microfibril and elastin contents. Oxytalan fibers are known to be distributed in the periodontal ligaments and gingiva, whereas elaunin and elastic fibers are present only in the gingiva. We examined gene expression and accumulation of tropoelastin in the cell-matrix layers of human gingival fibroblasts (HGF) and periodontal ligament fibroblasts (HPLF) in vitro. HGF and HPLF were cultured in MEM containing 10% newborn calf serum for 8 wks. Northern blotting and RT-PCR analyses showed that only HGF expressed mRNA encoding tropoelastin. Western blotting analysis demonstrated 77-kDa protropoelastin and 68-kDa tropoelastin only in the cell-matrix layer of HGF cultured for 8 wks. These results suggest that the different tropoelastin expression patterns reflect the difference between HGF and HPLF phenotypes.

Tropoelastin Expression by Periodontal Fibroblasts

Elastic system fibers are load-bearing proteins found in periodontal tissue. There are three types—oxytalan, elaunin, and elastic fibers—which differ in their relative microfibril and elastin contents. Oxytalan fibers are known to be distributed in the periodontal ligaments and gingiva, whereas elaunin and elastic fibers are present only in the gingiva. We examined gene expression and accumulation of tropoelastin in the cell-matrix layers of human gingival fibroblasts (HGF) and periodontal ligament fibroblasts (HPLF) in vitro. HGF and HPLF were cultured in MEM containing 10% newborn calf serum for 8 wks. Northern blotting and RT-PCR analyses showed that only HGF expressed mRNA encoding tropoelastin. Western blotting analysis demonstrated 77-kDa protropoelastin and 68-kDa tropoelastin only in the cell-matrix layer of HGF cultured for 8 wks. These results suggest that the different tropoelastin expression patterns reflect the difference between HGF and HPLF phenotypes.

Birth associated changes in pulmonary arterial connective tissue gene expression in the normal and hypertensive lung

To determine the temporal and spatial expression of the connective tissue precursors, procollagen and tropoelastin mRNA in normal and pulmonary hypertensive porcine pulmonary arteries from birth onwards. Using in situ hybridisation, connective tissue gene expression for procollagen alpha1(I) and alpha1(III) and tropoelastin was studied in intrapulmonary arteries from normal piglets, 5 min-16 weeks, and from piglets made pulmonary hypertensive by exposure to hypobaric hypoxia for 3 days, from birth, 3 or 14 days of age. In addition, Type III pN-procollagen, tropoelastin and collagen I and III were studied by immunohistochemistry. Quantitative or semi-quantitative techniques were applied to both in situ and immunohistochemical studies. Procollagen alpha1(I) and alpha1(III) mRNA expression increased rapidly in the media and adventitia between birth and 3 days of age (P<0.05). The increase was transient and the number of cells expressing procollagen mRNA decreased to the low newborn number after 6 days of age. Type III pN-procollagen immunostaining was greatest in newborn elastic and muscular arteries and then decreased. Collagen I and III increased mainly after 6 days of age. In animals exposed to chronic hypobaric hypoxia from birth, the increase in procollagens I and III mRNA was prevented. Exposure to hypoxia from 3 or 14 days led to little change in either gene expression or in procollagen and mature collagen from the normal. Tropoelastin gene expression was high at birth in the endothelium and media for the first 6 days, and then decreased. Normally, tropoelastin decreased in the media and increased in the adventitia after 16 days of age. Hypoxia had no effect on the mRNA but led to increased tropoelastin. We demonstrated marked, rapid changes in temporal and cell specific connective tissue gene expression in normal pulmonary arteries immediately after birth as the vasculature remodels. Each gene appeared to have its own timetable of expression and responded differently to hypoxia-induced hypertension.

Mechanical strain and dexamethasone selectively increase surfactant protein C and tropoelastin gene expression.

Physical forces derived from fetal breathing movements and hormones such as glucocorticoids are implicated in regulating fetal lung development. To elucidate whether the different signaling pathways activated by physical and hormonal factors are integrated and coordinated at the cellular and transcriptional levels, organotypic cultures of mixed fetal rat lung cells were subjected to static culture or mechanical strain in the presence and absence of dexamethasone. Tropoelastin and collagen type I were used as marker genes for fibroblasts, whereas surfactant protein (SP) A and SP-C were used as marker genes for distal epithelial cells. Mechanical strain, but not dexamethasone, significantly increased SP-C mRNA expression. Tropoelastin mRNA expression was upregulated by both mechanical strain and dexamethasone. No additive or synergistic effect was observed when cells were subjected to mechanical stretch in the presence of dexamethasone. Neither mechanical strain nor dexamethasone alone or in combination had any significant effect on the expression of SP-A mRNA. Dexamethasone decreased collagen type I mRNA expression, whereas mechanical strain had no effect. The increases in tropoelastin and SP-C mRNA levels induced by mechanical strain and/or dexamethasone were accompanied by increases in their heterogeneous nuclear RNA. In addition, the stretch- and glucocorticoid-induced alterations in tropoelastin and SP-C mRNA expression were abrogated with 10 microg/ml actinomycin D. These findings suggest that tropoelastin and SP-C genes are selectively stimulated by physical and/or hormonal factors at the transcriptional level in fetal lung fibroblasts and distal epithelial cells, respectively.

Hypoxia downregulates tropoelastin gene expression in rat lung fibroblasts by pretranslational mechanisms.

Elastolytic lung injury disrupts cell barriers, flooding alveoli and producing regional hypoxia. Abnormal O2 tensions may alter repair of damaged elastin fibers. To determine the effect of hypoxia on extravascular elastin formation, we isolated rat lung fibroblasts and cultured them under a variety of O2 conditions. Hypoxia downregulated tropoelastin mRNA in a dose- and time-related fashion while upregulating glyceraldehyde-3-phosphate dehydrogenase mRNA levels. The changes in tropoelastin gene expression were not due to cell toxicity as measured by chromium release and cell proliferation studies. Neither cycloheximide nor actinomycin D abrogated this effect. Hypoxia induced early decreases in tropoelastin mRNA stability; minor suppression of gene transcription occurred later. When returned to 21% O2, tropoelastin mRNA recovered to control levels in part by upregulating tropoelastin gene transcription. Taken together, these data indicate that hypoxia regulates tropoelastin gene expression and may alter repair of acutely injured lung.

Wavelength-Specific Synergy Between Ultraviolet Radiation and Interleukin-1α in the Regulation of Matrix-Related Genes: Mechanistic Role for Tumor Necrosis Factor-α

Ultraviolet light causes both acute and chronic changes in extracellular matrix. We sought to examine the effects of different ultraviolet wavelengths on expression of matrix-related genes in fibroblasts. We previously reported that tropoelastin gene expression in vivo decreases with acute ultraviolet B exposure, and interleukin-1 alpha-mediated upregulation of tropoelastin is blocked in vitro after ultraviolet B radiation. In this study, we found that only ultraviolet B, but not ultraviolet A or ultraviolet A1, blocked the ability of interleukin-1 alpha to stimulate tropoelastin expression in vitro. Ultraviolet B and interleukin-1 alpha synergistically increased tumor necrosis factor-alpha secretion by fibroblasts, a finding not seen with ultraviolet B alone nor with ultraviolet A or ultraviolet A1 combined with interleukin-1 alpha. Keratinocytes showed a similar ultraviolet B-specific induction of tumor necrosis factor-alpha production. Addition of tumor necrosis factor-alpha to cultured fibroblasts blocked interleukin-1 alpha-induced stimulation of tropoelastin message, and addition of anti-tumor necrosis factor-alpha antibodies restored the responsiveness of tropoelastin and collagen messages to exogenous interleukin-1 alpha after ultraviolet B exposure. We conclude that interleukin-1 alpha in combination specifically with ultraviolet B induces fibroblasts to secrete tumor necrosis factor-alpha, and that this ultraviolet B-specific induction of tumor necrosis factor-alpha secretion is responsible for effects of ultraviolet B on the expression of matrix-related genes in the skin.