Pharmacological treatment of systemic sclerosis-associated interstitial lung disease: an updated review and current approach to patient care.

Interstitial lung disease (ILD) has a high prevalence among patients with systemic sclerosis (SSc), carrying high mortality and morbidity. During the last decade, the emergence of new pharmacological therapies for SSc-associated ILD (SSc-ILD) and improved tools for its diagnosis and monitoring have changed the prevailing clinical approach, highlighting the need for early recognition and prompt treatment for SSc-ILD. Furthermore, the recent approval of multiple therapies for SSc-ILD poses challenges for the rheumatologist and pulmonologist in choosing the appropriate therapy for individual clinical scenarios. We review the pathophysiology of SSc-ILD, and the mechanisms of action and rationale behind current therapies. We also review the evidence of the efficacy and safety of immunosuppressive drugs, antifibrotic agents, and immunomodulators from cyclophosphamide and mycophenolate to novel agents such as nintedanib and tocilizumab. We also emphasise the importance of early diagnosis and monitoring and describe our approach to pharmacological therapy for SSc-ILD patients.

Ultrasound quantitative assessment of ventral finger microvasculopathy in systemic sclerosis with Raynaud's phenomena: a comparative study.

OBJECTIVE: To assess the finger vascularity of systemic sclerosis patients with Raynaud's phenomenon (RP-SSc) using various ultrasound techniques. METHODS: All fingers (except thumbs) of 18 RP-SSc patients and 18 controls were imaged at room temperature using four ultrasound vascular imaging techniques. The percent vascular area was quantified by counting blood flow pixels in a 25 mm2 square centred at the nail fold for the dorsal side and in 25 mm2 and 100 mm2 square from the fingertip for the ventral side. The mean vascular intensity was calculated from the corresponding areas for dorsal and ventral sides. RESULTS: The percent vascular areas and mean vascular intensities in RP-SSc were significantly lower than those in controls for both dorsal and ventral sides (p<0.01). The mean vascular intensities showed slightly higher area under the curve (AUC) than the percent vascular areas (0.53-0.91 vs 0.53-0.90) regardless of imaging technique and assessment side. For each imaging technique, the ventral side vascularity showed a higher AUC (0.74-0.91) compared with the dorsal side (0.53-0.81). Moreover, ventral side abnormalities were associated with a history of digital ulcers. CONCLUSIONS: Ultrasound demonstrated potential to quantify finger vascularity of RP-SSc. The ventral side of the fingers showed a higher accuracy in detecting RP-SSc than the dorsal side.

Treatment of Severe Swallowing Dysfunction in Systemic Sclerosis with IVIG: Role of Antimuscarinic Antibodies.

Oropharyngeal and esophageal dysmotility can cause serious clinical complications such as aspiration pneumonia, cachexia, and sarcopenia, with a resulting increase in mortality and disability. The current standard of care for the treatment of SSc-associated swallowing dysfunction is mainly supportive, although severe cases are usually refractory to conventional management. Recent studies have shown that the abnormal production of functional autoantibodies such as anti-cholinergic muscarinic receptor III antibodies may participate in the pathogenesis of SSc-associated gastrointestinal dysmotility and may provide a novel target for therapeutic intervention. We describe two patients with severe and rapid onset of SSc-associated severe swallowing dysfunction and esophageal dysmotility who had failed standard of care therapy, requiring complete enteral and parenteral nutrition. Both patients were positive for the presence of circulating antimuscarinic III receptor antibodies. They were treated with IVIG at a dose of 2 g/Kg/month divided in two consecutive days, for six months. Following IVIG therapy, both patients markedly improved their symptoms as shown by a reduction in their UCLA2.0 score, and achieved an improvement of esophageal motility documented radiologically. Both patients resumed oral feeding and had their feeding tubes removed within the treatment period. None of the patients developed severe adverse events attributable to IVIG, except for low-grade fever during IVIG infusion in one of the cases. These results provide support for the role of functional autoantibodies in the development of SSc-associated gastrointestinal dysfunction.

Progressive multifocal fibrosing neuropathy: description of a novel disease.

Entrapment peripheral neuropathies are clinically characterized by sensory impairment and motor deficits. They are usually caused by mechanical injuries, but they are also a frequent manifestation of metabolic diseases, toxic agent exposure, or systemic fibrotic disorders. Here we describe the clinical, radiological, and histopathological features of a novel progressive fibrotic disorder characterized by progressive multifocal fibrosing neuropathy. We identified two patients who presented with severe and progressive peripheral neuropathic symptoms sequentially affecting multiple sites. These patients presented with severe and progressive multifocal, sequentially additive peripheral neuropathic symptoms. Extensive nerve conduction and radiological studies showed the sequential development of multifocal motor and sensory peripheral neuropathy in the absence of any exposure to known infectious, inflammatory, or fibrotic triggers and the lack of family history of compression neuropathies. Extensive clinical and laboratory test evaluation failed to support the diagnosis of any primary inflammatory or genetic peripheral neuropathy and there was no evidence of any systemic fibrosing disorder including Systemic Sclerosis, lacking cutaneous fibrotic changes and cardiopulmonary abnormalities. The clinical course was progressive with sequential development of motor and sensory deficits of upper and lower extremities displaying proximal predominance. Histopathological study of tissues obtained during nerve release surgeries showed severe perineural fibrosis with marked accumulation of thick collagen bundles encroaching the peripheral nerves. There was no evidence of vasculitic, inflammatory, or vascular fibroproliferative lesions. We suggest that the clinical findings described here represent a previously undescribed fibrotic disorder affecting peripheral nerves, and we propose the descriptive term "Progressive Multifocal Fibrosing Neuropathy" to refer to this novel disorder. Despite the inherent limitations of this early description, we hope this is would contribute to the identification of additional cases.

Serine/threonine kinase inhibition as antifibrotic therapy: transforming growth factor-ß and Rho kinase inhibitors.

Serine/threonine kinases mediate the phosphorylation of intracellular protein targets, transferring a phosphorus group from an adenosine triphosphate molecule to the specific amino acid residues within the target proteins. Serine/threonine kinases regulate multiple key cellular functions. From this large group of kinases, TGF-ß through serine/threonine activity of its receptors and Rho kinase (ROCK) play an important role in the development and maintenance of fibrosis in various human diseases, including SSc. In recent years, multiple drugs targeting and inhibiting these kinases have been developed, opening the possibility of becoming potential antifibrotic agents of clinical value for treating fibrotic diseases. This review analyses the contribution of TGF-ß and ROCK-mediated serine/threonine kinase molecular pathways to the development and maintenance of pathological fibrosis and the potential clinical use of their inhibition.

Stratification in systemic sclerosis according to autoantibody status versus skin involvement: a study of the prospective EUSTAR cohort.

BACKGROUND: The current subclassification of systemic sclerosis into cutaneous subtypes does not fully capture the heterogeneity of the disease. We aimed to compare the performances of stratification into LeRoy's cutaneous subtypes versus stratification by autoantibody status in systemic sclerosis. METHODS: For this cohort study, we assessed people with systemic sclerosis in the multicentre international European Scleroderma Trials and Research (EUSTAR) database. Individuals positive for systemic-sclerosis autoantibodies of two specificities were excluded, and remaining individuals were classified by cutaneous subtype, according to their systemic sclerosis-specific autoantibodies, or both. We assessed the performance of each model to predict overall survival, progression-free survival, disease progression, and different organ involvement. The three models were compared by use of the area under the curve (AUC) of the receiver operating characteristic and the net reclassification improvement (NRI). Missing data were imputed. FINDINGS: We assessed the database on July 26, 2019. Of 16 939 patients assessed for eligibility, 10 711 patients were included: 1647 (15·4%) of 10 709 were male, 9062 (84·6%) were female, mean age was 54·4 (SD 13·8) years, and mean disease duration was 7·9 (SD 8·2) years. Information regarding cutaneous subtype was available for 10 176 participants and antibody data were available for 9643 participants. In the prognostic analysis, there was no difference in AUC for overall survival (0·82, 95% CI 0·81-0·84 for cutaneous only vs 0·84, 0·82-0·85 for antibody only vs 0·84, 0·83-0·86 for combined) or for progression-free survival (0·70, 0·69-0·71 vs 0·71, 0·70-0·72 vs 0·71, 0·70-0·72). However, at 4 years the NRI showed substantial improvement for the antibody-only model compared with the cutaneous-only model in prediction of overall survival (0·57, 0·46-0·71 for antibody only vs 0·29, 0·19-0·39 for cutaneous only) and disease progression (0·36, 0·29-0·46 vs 0·21, 0·14-0·28). The antibody-only model did better than the cutaneous-only model in predicting renal crisis (AUC 0·72, 0·70-0·74 for antibody only vs 0·66, 0·64-0·69 for cutaneous only) and lung fibrosis leading to restrictive lung function (AUC 0·76, 0·75-0·77 vs 0·71, 0·70-0·72). The combined model improved the prediction of digital ulcers and elevated systolic pulmonary artery pressure, but did poorly for cardiac involvement. INTERPRETATION: The autoantibody-only model outperforms cutaneous-only subsetting for risk stratifying people with systemic sclerosis in the EUSTAR cohort. Physicians should be aware of these findings at the time of decision making for patient management. FUNDING: World Scleroderma Foundation.

Genome Sequences of Mycobacterium smegmatis Phages Purgamenstris and PhancyPhin.

Two novel mycobacteriophages, PhancyPhin and Purgamenstris, were isolated from the Houston, Texas, area. They were isolated in the same year with the soil enrichment method using the host Mycobacterium smegmatis mc2 155. They exhibit a 99.55% nucleotide identity with each other.

Tyrosine kinases in the pathogenesis of tissue fibrosis in systemic sclerosis and potential therapeutic role of their inhibition.

Systemic sclerosis (SSc) is an idiopathic autoimmune disease with a heterogeneous clinical phenotype ranging from limited cutaneous involvement to rapidly progressive diffuse SSc. The most severe SSc clinical and pathologic manifestations result from an uncontrolled fibrotic process involving the skin and various internal organs. The molecular mechanisms responsible for the initiation and progression of the SSc fibrotic process have not been fully elucidated. Recently it has been suggested that tyrosine protein kinases play a role. The implicated kinases include receptor-activated tyrosine kinases and nonreceptor tyrosine kinases. The receptor kinases are activated following specific binding of growth factors (platelet-derived growth factor, fibroblast growth factor, or vascular endothelial growth factor). Other receptor kinases are the discoidin domain receptors activated by binding of various collagens, the ephrin receptors that are activated by ephrins and the angiopoetin-Tie-2s receptors. The nonreceptor tyrosine kinases c-Abl, Src, Janus, and STATs have also been shown to participate in SSc-associated tissue fibrosis. Currently, there are no effective disease-modifying therapies for SSc-associated tissue fibrosis. Therefore, extensive investigation has been conducted to examine whether tyrosine kinase inhibitors (TKIs) may exert antifibrotic effects. Here, we review the role of receptor and nonreceptor tyrosine kinases in the pathogenesis of the frequently progressive cutaneous and systemic fibrotic alterations in SSc, and the potential of TKIs as SSc disease-modifying antifibrotic therapeutic agents.

Increased expression of interferon regulated and antiviral response genes in CD31+/CD102+ lung microvascular endothelial cells from systemic sclerosis patients with end-stage interstitial lung disease.

OBJECTIVES: Systemic sclerosis (SSc) is characterised by severe fibroproliferative vasculopathy, fibrosis in skin and multiple internal organs, and humoral, cellular and innate immunity abnormalities. Vascular alterations are the earliest and most severe SSc manifestations, however, the mechanisms responsible have remained elusive. To investigate the molecular abnormalities involved in SSc-vasculopathy we examined global gene expression differences between highly purified lung microvascular endothelial cells (MVECs) from patients with SSc-interstitial lung disease (SSc-ILD) and normal lung MVECs. METHODS: MVECs were isolated from fresh transplanted lungs from patients with SSc-ILD. Sequential CD31 and CD102 immunopurification was performed to obtain highly purified CD31+/CD102+ lung MVECs. Global gene expression analysis was successfully performed in CD31+/CD102+ MVEC from two SSc-ILD patients and from two normal lungs. RT-PCR, Western blots, and indirect immunofluorescence validated the gene expression results. RESULTS: Numerous interferon-regulated genes (IRGs) including IFI44, IFI44L, IFI6, IFIH1, IFIT1, ISG-15, BST-2/Tetherin, and RSAD2/Viperin, genes encoding innate immunity antiviral responses (OAS1, OAS2, OAS3, OASL) and antiviral MX1 and MX2 proteins, and mesenchymal cell-specific genes were significantly overexpressed in CD31+/CD102+ SSc-ILD lung MVECs. CONCLUSIONS: Highly purified CD31+/CD102+ MVECs from lungs from SSc patients with end stage SSc-ILD displayed remarkable overexpression of numerous IRGs and of genes encoding antiviral innate immune response and antiviral proteins. These observations suggest that interferon-induced and antiviral response proteins may participate in the pathogenesis of SSc vasculopathy and SSc-ILD. The CD31+/CD102+ lung MVECs from SSc-ILD also showed elevated expression of mesenchymal cell-specific genes confirming the presence of endothelial to mesenchymal transition in SSc-ILD.

Health Assessment Questionnaire-Disability Index (HAQ-DI) use in modelling disease progression in diffuse cutaneous systemic sclerosis: an analysis from the EUSTAR database.

BACKGROUND: Patients with diffuse cutaneous systemic sclerosis (dcSSc) have a poor prognosis. The importance of monitoring subjective measures of functioning and disability, such as the Health Assessment Questionnaire-Disability Index (HAQ-DI), is important as dcSSc is rated by patients as worse than diabetes or hemodialysis for quality of life impairment. This European Scleroderma Trials and Research (EUSTAR) database analysis was undertaken to examine the importance of impaired functionality in dcSSc prognosis. The primary objectives were to identify predictors of death and HAQ-DI score progression over 1 year. HAQ-DI score, major advanced organ involvement, and death rate were also used to develop a comprehensive model to predict lifetime dcSSc progression. METHODS: This was an observational, longitudinal study in patients with dcSSc registered in EUSTAR. Death and HAQ-DI scores were, respectively, analyzed by Cox regression and linear regression analyses in relation to baseline covariates. A microsimulation Markov model was developed to estimate/predict natural progression of dcSSc over a patient's lifetime. RESULTS: The analysis included dcSSc patients with (N = 690) and without (N = 4132) HAQ-DI score assessments from the EUSTAR database. Baseline HAQ-DI score, corticosteroid treatment, and major advanced organ involvement were predictive of death on multivariable analysis; a 1-point increase in baseline HAQ-DI score multiplied the risk of death by 2.7 (p <  0.001) and multiple advanced major organ involvement multiplied the risk of death by 2.8 (p <  0.05). Multivariable analysis showed that baseline modified Rodnan Skin Score (mRSS) and baseline HAQ-DI score were associated with HAQ-DI score progression at 1 year (p <  0.05), but there was no association between baseline organ involvement and HAQ-DI score progression at 1 year. HAQ-DI score, major advanced organ involvement, and death were successfully used to model long-term disease progression in dcSSc. CONCLUSIONS: HAQ-DI score and major advanced organ involvement were comparable predictors of mortality risk in dcSSc. Baseline mRSS and baseline HAQ-DI score were predictive of HAQ-DI score progression at 1 year, indicating a correlation between these endpoints in monitoring disease progression. It is hoped that this EUSTAR analysis may change physician perception about the importance of the HAQ-DI score in dcSSc.

Recurrence of progressive skin involvement following discontinuation or dose reduction of Mycophenolate Mofetil treatment in patients with diffuse Systemic Sclerosis.

BACKGROUND: Rapidly progressive diffuse cutaneous Systemic Sclerosis (rp-dcSSc) is associated with severe internal organ involvement and high mortality. Mycophenolate Mofetil (MMF) has been shown to halt the progression of rp-dcSSc cutaneous and pulmonary involvement in observational and randomized controlled trials, respectively. However, optimal MMF therapy duration has not been established. Here, we describe the clinical evolution of rp-dcSSc patients successfully treated with MMF following MMF therapy discontinuation or dose reduction. METHODS: Twenty-five patients with recent-onset (< 24 mo) rp-dcSSc received MMF as the only SSc disease-modifying therapy. Following MMF discontinuation or dose reduction to or below 1000 mg/day after an average of two years, the Modified Rodnan skin score (mRSS) and Pulmonary function tests (PFT) were serially evaluated for additional 5 years. MMF therapy was re-instituted if the mRSS increased by greater than 20% or if restrictive lung disease developed. RESULTS: From nineteen patients serially evaluated following MMF discontinuation or dose reduction, five patients (26.3%) developed recurrence of rapid skin involvement with an average of 35.9% increase in mRSS from 7.8 to 10.6 points requiring MMF re-institution. Two of these patients also presented worsening respiratory symptoms and reduction of lung volumes in PFTs. Following MMF resumption, mRSS returned to baseline or stabilized and PFTs improved or stabilized. All these patients were maintained on high dose long term MMF treatment. CONCLUSION: Recurrence of severe skin involvement occurred in 26.3% of patients with rp-dcSSc following MMF discontinuation or dose reduction, requiring prompt MMF therapy resumption. These findings confirm the therapeutic benefit of MMF in rp-dcSSc and suggest that MMF treatment should be maintained for longer than 2 years.

Primary osteoporosis in men: an unmet medical need.

Osteoporosis is a skeletal disease characterized by loss of bone strength and increased risk of fractures. Even though fracture prevalence is higher in women, fractures also constitute a significant public health issue in older men. Men are screened less and more frequently undertreated than female patients. It is the goal of this review, to summarize updated information about the current understanding of pathophysiology and clinical aspects of diagnosis and treatment of osteoporosis in men.

Role of microRNA in the pathogenesis of systemic sclerosis tissue fibrosis and vasculopathy.

Systemic Sclerosis (SSc) pathogenesis involves multiple immunological, vascular and fibroproliferative abnormalities that contribute to a severe and complex clinical picture. Vasculopathy and fibroproliferative alterations are two hallmark pathological processes in SSc that are responsible for the most severe clinical manifestations of the disease and determine its clinical outcome and mortality. However, the pathogenesis of SSc vasculopathy and of the uncontrolled SSc fibrotic process remain incompletely understood. Recent investigations into the molecular pathways involved in these processes have identified an important role for epigenetic processes that contribute to overall disease progression and have emphasized microRNAs (miRNAs) as crucial epigenetic regulators. MiRNAs hold unique potential for elucidating SSc pathogenesis, improving diagnosis and developing effective targeted therapies for the disease. This review examines the important role that miRNAs play in the development and regulation of vascular and fibroproliferative alterations associated with SSc pathogenesis and their possible participation in the establishment of pathogenetic connections between these two processes. This review also emphasizes that further understanding of the involvement of miRNA in SSc fibrosis and vasculopathy will very likely provide novel future research directions and allow for the identification of groundbreaking therapeutic interventions within these processes. MiR-21, miR- 31, and miR-155 are of particular interest owing to their important involvement in both SSc vasculopathy and fibroproliferative alterations.

Abrogation of transforming growth factor-ß-induced tissue fibrosis in mice with a global genetic deletion of Nox4.

Excessive connective tissue deposition in skin and various internal organs is characteristic of systemic sclerosis (SSc). The profibrotic growth factor TGF-ß plays a crucial role in SSc pathogenesis. The expression of NADPH oxidase 4 (NOX4), a critical mediator of oxidative stress, is potently stimulated by TGF-ß. Here, we evaluated the effect of NOX4 on the development of TGF-ß-induced tissue fibrosis. C57BL6/J control mice and Nox4 knockout mice were implanted subcutaneously with osmotic pumps containing either saline or 2.5 µg TGF-ß1. After 28 days, skin and lung samples were isolated for histopathologic analysis, measurement of hydroxyproline content and gene expression analysis. Histopathology of skin and lungs from normal C57BL6/J mice treated with TGF-ß1 showed profound dermal fibrosis and peribronchial and diffuse interstitial lung fibrosis. In contrast, TGF-ß-treated Nox4 knockout mice showed normal skin and lung histology. Hydroxyproline levels in TGF-ß-treated C57BL6/J mice skin and lungs demonstrated significant increases, however, hydroxyproline content of TGF-ß-treated Nox4 knockout mice tissues was not changed. Expression of various profibrotic and fibrosis-associated genes was upregulated in skin and lungs of TGF-ß1-treated C57BL6/J mice but was not significantly changed in TGF-ß1-treated Nox4 knockout mice. The induction of skin and lung tissue fibrosis by TGF-ß1 parenteral administration in mice was abrogated by the genetic deletion of Nox4 confirming that NOX4 is an essential mediator of the profibrotic effects of TGF-ß. These results suggest Nox4 inhibition as a potential therapeutic target for SSc and other fibroproliferative disorders.

Endothelial cell-specific activation of transforming growth factor-ß signaling in mice induces cutaneous, visceral, and microvascular fibrosis.

In this study, we tested the hypothesis that constitutive endothelial cell-specific activation of TGF-ß signaling induces tissue fibrosis and vasculopathy resembling the characteristic fibrotic and vascular alterations of systemic sclerosis. Transgenic mice with inducible expression of a constitutively active TGF-ß receptor I specifically in endothelial cells were generated by intercrossing mice harboring a constitutively active TGF-ß receptor I with a mouse strain containing the endothelial cell-specific Cdh5 gene promoter directing the tamoxifen-inducible expression of the Cre-ERT2 cassette. Administration of tamoxifen to these mice would result in constitutive TGF-ß activation and signaling confined to endothelial lineage cells. The effects of constitutive TGF-ß endothelial cell activation were assessed by histopathological examination of skin and various internal organs, tissue hydroxyproline analysis, and assessment of expression of myofibroblast differentiation and TGF-ß signaling genes employing real-time PCR and immunohistochemical staining of lung vessels for endothelial- and myofibroblast-specific proteins. Constitutive TGFß-1 signaling in endothelial cells resulted in cutaneous and visceral fibrosis with prominent fibrotic involvement of the lungs and severe perivascular and subendothelial fibrosis of small arterioles. A marked increase in the expression of fibrosis-associated genes and of genes indicative of myofibroblast activation was also found. Confocal microscopy of lung vessels showed evidence consistent with the induction of endothelial-to-mesenchymal transition (EndoMT). Taken together, our data indicate that transgenic mice with constitutive endothelial cell-specific activation of TGF-ß signaling display severe cutaneous, pulmonary, and microvascular fibrosis resembling the fibrotic and microvascular alterations characteristic of systemic sclerosis.

Treatment of Rapidly Progressive Systemic Sclerosis: Current and Futures Perspectives.

INTRODUCTION: Systemic Sclerosis (SSc) is a systemic autoimmune disease characterized by severe and often progressive cutaneous, pulmonary, cardiac and gastrointestinal tract fibrosis, cellular and humoral immunologic alterations, and pronounced fibroproliferative vasculopathy. There is no effective SSc disease modifying therapy. Patients with rapidly progressive SSc have poor prognosis with frequent disability and very high mortality. AREAS COVERED: This paper reviews currently available therapeutic approaches for rapidly progressive SSc and discuss novel drugs under study for SSc disease modification. EXPERT OPINION: The extent, severity, and rate of progression of SSc skin and internal organ involvement determines the optimal therapeutic interventions for SSc. Cyclophosphamide for progressive SSc-associated interstitial lung disease and mycophenolate for rapidly progressive cutaneous involvement have shown effectiveness. Methotrexate has been used for less severe skin progression and for patients unable to tolerate mycophenolate. Rituximab was shown to induce improvement in SSc-cutaneous and lung involvement. Autologous bone marrow transplantation is reserved for selected cases in whom poor survival risk outweighs the high mortality rate of the procedure. Novel agents capable of modulating fibrotic and inflammatory pathways involved in SSc pathogenesis, including tocilizumab, pirfenidone, tyrosine kinase inhibitors, lipid lysophosphatidic acid 1, and NOX4 inhibitors are currently under development for the treatment of rapidly progressive SSc.

Stimulation of Transforming Growth Factor-ß1-Induced Endothelial-To-Mesenchymal Transition and Tissue Fibrosis by Endothelin-1 (ET-1): A Novel Profibrotic Effect of ET-1.

TGF-ß-induced endothelial-to-mesenchymal transition (EndoMT) is a newly recognized source of profibrotic activated myofibroblasts and has been suggested to play a role in the pathogenesis of various fibrotic processes. Endothelin-1 (ET-1) has been implicated in the development of tissue fibrosis but its participation in TGF-ß-induced EndoMT has not been studied. Here we evaluated the role of ET-1 on TGF-ß1-induced EndoMT in immunopurified CD31+/CD102+ murine lung microvascular endothelial cells. The expression levels of α-smooth muscle actin (α-SMA), of relevant profibrotic genes, and of various transcription factors involved in the EndoMT process were assessed employing quantitative RT-PCR, immunofluorescence histology and Western blot analysis. TGF-ß1 caused potent induction of EndoMT whereas ET-1 alone had a minimal effect. However, ET-1 potentiated TGF-ß1-induced EndoMT and TGF-ß1-stimulated expression of mesenchymal cell specific and profibrotic genes and proteins. ET-1 also induced expression of the TGF-ß receptor 1 and 2 genes, suggesting a plausible autocrine mechanism to potentiate TGF-ß-mediated EndoMT and fibrosis. Stimulation of TGF-ß1-induced skin and lung fibrosis by ET-1 was confirmed in vivo in an animal model of TGF-ß1-induced tissue fibrosis. These results suggest a novel role for ET-1 in the establishment and progression of tissue fibrosis.

Endothelial to Mesenchymal Transition (EndoMT) in the Pathogenesis of Human Fibrotic Diseases.

Fibrotic diseases encompass a wide spectrum of clinical entities including systemic fibrotic diseases such as systemic sclerosis, sclerodermatous graft versus host disease, nephrogenic systemic fibrosis, and IgG4-associated sclerosing disease, as well as numerous organ-specific disorders including radiation-induced fibrosis, and cardiac, pulmonary, liver, and kidney fibrosis. Although their causative mechanisms are quite diverse, these diseases share the common feature of an uncontrolled and progressive accumulation of fibrous tissue macromolecules in affected organs leading to their dysfunction and ultimate failure. The pathogenesis of fibrotic diseases is complex and despite extensive investigation has remained elusive. Numerous studies have identified myofibroblasts as the cells responsible for the establishment and progression of the fibrotic process. Tissue myofibroblasts in fibrotic diseases originate from several sources including quiescent tissue fibroblasts, circulating CD34+ fibrocytes, and the phenotypic conversion of various cell types including epithelial and endothelial cells into activated myofibroblasts. However, the role of the phenotypic transition of endothelial cells into mesenchymal cells (Endothelial to Mesenchymal Transition or EndoMT) in the pathogenesis of fibrotic disorders has not been fully elucidated. Here, we review the evidence supporting EndoMT's contribution to human fibrotic disease pathogenesis.

Endothelial Cells Expressing Endothelial and Mesenchymal Cell Gene Products in Lung Tissue From Patients With Systemic Sclerosis-Associated Interstitial Lung Disease.

OBJECTIVE: To examine whether lung endothelial cells (ECs) from patients with systemic sclerosis (SSc)-associated interstitial lung disease (ILD) express mesenchymal cell-specific proteins and gene transcripts, indicative of the occurrence of endothelial-to-mesenchymal phenotypic transition (EndoMT). METHODS: Lung tissue from 6 patients with SSc-associated pulmonary fibrosis was examined by histopathology and immunohistochemistry. Confocal laser microscopy was utilized to assess the simultaneous expression of EC and myofibroblast molecular markers. CD31+CD102+ ECs were isolated from the lung tissue of 2 patients with SSc-associated ILD and 2 normal control subjects, and the expression of EC and mesenchymal cell markers and other relevant genes was analyzed by quantitative polymerase chain reaction, immunofluorescence microscopy, and Western blotting. RESULTS: Immunohistochemical staining revealed cells expressing the EC-specific marker CD31 in the subendothelial, perivascular, and parenchymal regions of the lungs from all SSc patients. Confocal microscopy identified cells displaying simultaneous expression of von Willebrand factor and α-smooth muscle actin in small and medium-sized arterioles in the SSc lung tissue but not in normal control lungs. CD31+CD102+ ECs isolated from SSc lungs expressed high levels of mesenchymal cell-specific genes (type I collagen, type III collagen, and fibronectin), EC-specific genes (type IV collagen and VE-cadherin), profibrotic genes (transforming growth factor ß1 and connective tissue growth factor), and genes encoding EndoMT-related transcription factors (TWIST1 and SNAI2). CONCLUSION: Cells coexpressing EC- and mesenchymal cell-specific molecules are present in the lungs of patients with SSc-associated ILD. CD31+CD102+ ECs isolated from SSc lungs simultaneously expressed mesenchymal cell- and EC-specific transcripts and proteins. Collectively, these observations demonstrate the occurrence of EndoMT in the lungs of patients with SSc-associated ILD.