Matrix metalloproteinase-1 up-regulation by hepatocyte growth factor in human dermal fibroblasts via ERK signaling pathway involves Ets1 and Fli1

In this study, we clarified the molecular mechanism(s) underlying the regulation of matrix metalloproteinase (MMP)-1 gene by hepatocyte growth factor (HGF) in cultured human dermal fibroblasts. HGF induced MMP-1 protein as well as mRNA at a transcriptional level via extracellular signal-regulated kinase (ERK) signaling pathway. The region in the MMP-1 promoter mediating the inducible responsiveness to HGF, defined by the transient transfection analysis of the serial 5′ deletion constructs, contained an Ets binding site. Mutation of this Ets binding site abrogated the HGF-inducible promoter activity. Ets1 up-regulated the expression of MMP-1 promoter activity, whereas Fli1 had antagonistic effects on them. After HGF treatment, the protein level and the binding activity of Ets1 was increased and those of Fli1 was decreased, which were canceled by PD98059. These results suggest that HGF up-regulates MMP-1 expression via ERK signaling pathway through the balance of Ets1 and Fli1, which may be a novel mechanism of regulating MMP-1 gene expression

Recurrent, Tumor Mutation Burden-High, Cutaneous Angiosarcoma of the Scalp Treated with Pembrolizumab

Introduction: Chemoradiotherapy with taxanes is well-recognized as a first-line therapy for cutaneous angiosarcoma (CAS), but second-line therapy for CAS is still controversial. Case Presentation: In this report, we described a 75-year-old Japanese case of recurrent, tumor mutation burden-high CAS on the scalp treated with pembrolizumab. Our present case survived for 1 year despite of taxane refractory CAS with mediastinal lymph node metastasis, though the administration of anti-PD-1 Abs alone could not fully suppress the tumor progression of CAS. Conclusion: Since various factors such as pro-angiogenic molecules are correlated with the tumor progression in CAS, the administration of anti-PD-1 Abs alone could not fully suppress the tumor progression of CAS. Further novel anticancer drugs are needed in the future for the treatment of CAS

A Possible Contribution of Altered Cathepsin B Expression to the Development of Skin Sclerosis and Vasculopathy in Systemic Sclerosis

Cathepsin B (CTSB) is a proteolytic enzyme potentially modulating angiogenic processes and extracellular matrix remodeling. While matrix metalloproteinases are shown to be implicated in tissue fibrosis and vasculopathy associated with systemic sclerosis (SSc), the role of cathepsins in this disease has not been well studied. The aim of this study is to evaluate the roles of CTSB in SSc. Serum pro-CTSB levels were determined by enzyme-linked immunosorbent assay in 55 SSc patients and 19 normal controls. Since the deficiency of transcription factor Fli1 in endothelial cells is potentially associated with the development of SSc vasculopathy, cutaneous CTSB expression was evaluated by immunostaining in Fli1+/− and wild type mice as well as in SSc and control subjects. The effects of Fli1 gene silencing and transforming growth factor-β (TGF-β) on CTSB expression were determined by real-time PCR in human dermal microvascular endothelial cells (HDMECs) and dermal fibroblasts, respectively. Serum pro-CTSB levels were significantly higher in limited cutaneous SSc (lcSSc) and late-stage diffuse cutaneous SSc (dcSSc) patients than in healthy controls. In dcSSc, patients with increased serum pro-CTSB levels showed a significantly higher frequency of digital ulcers than those with normal levels. CTSB expression in dermal blood vessels was increased in Fli1+/− mice compared with wild type mice and in SSc patients compared with healthy controls. Consistently, Fli1 gene silencing increased CTSB expression in HDMECs. In cultured dermal fibroblasts from early dcSSc, CTSB expression was decreased compared with normal fibroblasts and significantly reversed by TGF-β1 antisense oligonucleotide. In conclusion, up-regulation of endothelial CTSB due to Fli1 deficiency may contribute to the development of SSc vasculopathy, especially digital ulcers, while reduced expression of CTSB in lesional dermal fibroblasts is likely to be associated with skin sclerosis in early dcSSc

Proteasome Inhibitor Bortezomib Ameliorates Intestinal Injury in Mice

Background: Bortezomib is a proteasome inhibitor that has shown impressive efficacy in the treatment of multiple myeloma. In mice, the addition of dextran sulfate sodium (DSS) to drinking water leads to acute colitis that can serve as an experimental animal model for human ulcerative colitis. Methodology/Principal Findings: Bortezomib treatment was shown to potently inhibit murine DSS-induced colitis. The attenuation of DSS-induced colitis was associated with decreased inflammatory cell infiltration in the colon. Specifically, bortezomib-treated mice showed significantly decreased numbers of CD4 + and CD8 + T cells in the colon and mesenteric lymph nodes. Bortezomib treatment significantly diminished interferon (IFN)-c expression in the colon and mesenteric lymph nodes. Furthermore, cytoplasmic IFN-c production by CD4 + and CD8 + T cells in mesenteric lymph nodes was substantially decreased by bortezomib treatment. Notably, bortezomib enhanced T cell apoptosis by inhibiting nuclear factor-kB activation during DSS-induced colitis. Conclusions/Significance: Bortezomib treatment is likely to induce T cell death, thereby suppressing DSS-induced colitis by reducing IFN-c production

Serum Adhesion Molecule Levels as Prognostic Markers in Patients with Early Systemic Sclerosis: A Multicentre, Prospective, Observational Study

Objective: To assess the utility of circulating adhesion molecule levels as a prognostic indicator of disease progression in systemic sclerosis (SSc) patients with early onset disease. Methods: Ninety-two Japanese patients with early onset SSc presenting with diffuse skin sclerosis and/or interstitial lung disease were registered in a multicentre, observational study. Concentrations of intercellular adhesion molecule (ICAM) -1, E-selectin, L-selectin, and P-selectin in serum samples from all patients were measured by enzyme-linked immunosorbent asssay (ELISA). In 39 patients, adhesion molecule levels were measured each year for four years. The ability of baseline adhesion molecule levels to predict subsequent progression and severity in clinical and laboratory features were evaluated statistically. Results: At their first visit, serum levels of ICAM-1, E-selection, P-selectin were significantly elevated and serum L-selectin levels were significantly reduced in patients with SSc compared with healthy controls. Overall, serum ICAM-1 levels at each time point were significantly inversely associated with the %vital capacity (VC) of the same time and subsequent years by univariate analysis. The initial serum ICAM-1 levels were significantly inversely associated with the %VC at the fourth year by multiple regression analysis. The initial serum P-selectin levels were significantly associated with the health assessment questionnaire disability index (HAQ-DI) at the fourth year by multiple regression analysis. Initial adhesion molecule levels were not significantly associated with other clinical features including skin thickness score. Baseline adhesion molecule levels were not significantly associated with subsequent rate of change of clinical parameters. Conclusion: In patients with SSc, serum levels of ICAM-1 and P-selectin may serve as prognostic indicators of respiratory dysfunction and physical disability, respectively. Further longitudinal studies of larger populations are needed to confirm these findings

The Pathogenesis of Systemic Sclerosis: An Understanding Based on a Common Pathologic Cascade across Multiple Organs and Additional Organ-Specific Pathologies

Systemic sclerosis (SSc) is a multisystem autoimmune and vascular disease resulting in fibrosis of various organs with unknown etiology. Accumulating evidence suggests that a common pathologic cascade across multiple organs and additional organ-specific pathologies underpin SSc development. The common pathologic cascade starts with vascular injury due to autoimmune attacks and unknown environmental factors. After that, dysregulated angiogenesis and defective vasculogenesis promote vascular structural abnormalities, such as capillary loss and arteriolar stenosis, while aberrantly activated endothelial cells facilitate the infiltration of circulating immune cells into perivascular areas of various organs. Arteriolar stenosis directly causes pulmonary arterial hypertension, scleroderma renal crisis and digital ulcers. Chronic inflammation persistently activates interstitial fibroblasts, leading to the irreversible fibrosis of multiple organs. The common pathologic cascade interacts with a variety of modifying factors in each organ, such as keratinocytes and adipocytes in the skin, esophageal stratified squamous epithelia and myenteric nerve system in gastrointestinal tract, vasospasm of arterioles in the heart and kidney, and microaspiration of gastric content in the lung. To better understand SSc pathogenesis and develop new disease-modifying therapies, it is quite important to understand the complex pathogenesis of SSc from the two distinct perspectives, namely the common pathologic cascade and additional organ-specific pathologies

Fli1 represses transcription of the human α2(I) collagen gene by recruitment of the HDAC1/p300 complex.

Fli1, a member of the Ets transcription factor family, is a key repressor of the human α2(I) collagen (COL1A2) gene. Although our previous studies have delineated that TGF-β induces displacement of Fli1 from the COL1A2 promoter through sequential post-translational modifications, the detailed mechanism by which Fli1 functions as a potent transcriptional repressor of the COL1A2 gene has not been fully investigated. To address this issue, we carried out a series of experiments especially focusing on protein-protein interaction and epigenetic transcriptional regulation. The combination of tandem affinity purification and mass spectrometry identified HDAC1 as a Fli1 interacting protein. Under quiescent conditions, HDAC1 induced deacetylation of Fli1 resulting in an increase of Fli1 DNA binding ability and p300 enhanced this process by promoting the formation of a Fli1-HDAC1-p300 complex. TGF-β-induced phosphorylation of Fli1 at threonine 312 led to disassembly of this protein complex. In quiescent dermal fibroblasts Fli1, HDAC1, and p300 occupied the -404 to -237 region, including the Fli1 binding site, of the COL1A2 promoter. TGF-β induced Fli1 and HDAC1 dissociation from the COL1A2 promoter, while promoting Ets1 and p300 recruitment. Furthermore, acetylation levels of histone H3 around the Fli1 binding site in the COL1A2 promoter inversely correlated with the DNA occupancy of Fli1 and HDAC1, while positively correlating with that of Ets1 and p300. In the functional studies, HDAC1 overexpression magnified the inhibitory effect of Fli1 on the COL1A2 promoter. Moreover, pharmacological blockade of HDAC1 by entinostat enhanced collagen production in dermal fibroblasts. Collectively, these results indicate that under quiescent conditions Fli1 recruits HDAC1/p300 to the COL1A2 promoter and suppresses the expression of the COL1A2 gene by chromatin remodeling through histone deacetylation. TGF-β-dependent phosphorylation of Fli1 at threonine 312 is a critical step regulating the remodeling of the Fli1 transcription repressor complex, leading to transcriptional activation of the COL1A2 gene