COL1A2 Transcription Research Articles

The role of fibromodulin in myocardial fibrosis in a diabetic cardiomyopathy rat model.

Diabetic cardiomyopathy (DCM) is pathologically characterized by excessive deposition of extracellular matrix proteins, leading to myocardial fibrosis. Fibromodulin (Fmod) plays a crucial role in the pathogenesis of fibrotic diseases. However, the role and mechanism of Fmod in DCM-related myocardial fibrosis remain unclear. In the present study, we established a DCM rat model and an in vitro model of rat primary cardiac fibroblasts (RPCFs) exposed to high glucose. We assessed mRNA and protein expression levels of Col1a1, Col3a1, α-SMA and Fmod in both models. Fmod-overexpressing (ov-Fmod) and Fmod-knockdown (si-Fmod) rat cardiac fibroblasts (RCFs) were generated. Subsequently, whole RNA sequencing was conducted on ov-Fmod RCFs. The gene Col15a1 was evaluated in the DCM rat and all cell models. The correlation between plasma levels of Fmod and Col15a1 in DCM rat models was assessed. Transcription and protein levels of Fmod, Col1a1, Col3a1 and α-SMA were significantly elevated in DCM rat hearts and RPCFs. In ov-Fmod RCFs, fibrosis markers were similarly increased, except for Col3a1, which decreased. The Col1a1/Col3a1 ratio was elevated. Conversely, knocking down Fmod yielded opposite results. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses indicated that Fmod participates in multiple fibrosis-related pathways, affecting Col15a1. Expression of Col15a1 was significantly decreased in all models, compared to controls, except in si-Fmod RCFs. Importantly, Col15a1 and Fmod in plasma exhibited an inverse relationship in DCM. In summary, Fmod is implicated in DCM, with Fmod overexpression downregulating Col15a1 and increasing the Col1a1/Col3a1 ratio. This mechanism may influence diastolic heart failure in DCM by modulating myocardial stiffness and elasticity.

Effect of nHA/CS/PLGA delivering adipose stem cell-derived exosomes and bone marrow stem cells on bone healing—in vitro and in vivo studies

Adipose stem cell-derived exosomes (ADSC-EXO) have been demonstrated to promote osteogenic differentiation of bone marrow stem cells (BMSCs) and facilitate bone regeneration. The present study aims to investigate the effect of ADSC-EXO-loaded nano-hydroxyapatite/chitosan/poly-lactide-co-glycolide (nHA/CS/PLGA) scaffolds on maxillofacial bone regeneration using tissue engineering. ADSC-EXO was isolated and co-cultured with BMSCs, and the osteogenic differentiation of BMSCs was assessed through the detection of mineralized nodule formation, alkaline phosphatase (ALP) activity, and mRNA expression of COL1A1 and runt-related transcription factor 2 (RUNX2). The nHA/CS/PLGA scaffolds were fabricated and loaded with ADSC-EXO and BMSCs, and these tissue engineering complexes were applied to the maxillofacial bone defect region of rabbits to elucidate their bone regeneration effect. The osteogenic differentiation of BMSCs was markedly enhanced when they were co-cultured with ADSC-EXO. This was evidenced by an increase in the formation of mineralized nodule formation, ALP activity, and mRNA expression of COL1A1 and runt-related transcription factor 2 (RUNX2). In vivo experiments demonstrated that the application of ADSC-EXO and BMSCs loaded nHA/CS/PLGA scaffolds effectively repaired maxillofacial bone defects in rabbits. ADSC-EXO has been demonstrated to promote the osteogenic differentiation of BMSCs. The ADSC-EXO and BMSCs loaded nHA/CS/PLGA scaffolds have been shown to facilitate the regeneration of maxillofacial bone defects. This may serve as a potential therapeutic strategy for large-scale bone defects.

TLR-2 and TLR-4 mRNA expression in different grades of histopathological lesions of equine endometrium from follicular phase.

Increased synthesis and deposition of collagen(COL) in the extracellular matrix (ECM) of equine endometrium contributes to endometrosis. Toll-like receptors (TLRs) are transmembrane receptors involved in the innate immune response, recognized for their role in antigen recognition and previously associated with equine endometritis. The TLRs not only recognize pathogen-associated molecular patterns but also regulate inflammations, fibrosis and cancer. The aim of this study was to explore the relationship between TLR expression at different stages of Kenney and Doig's (K-D) grading and COL1 expression during the follicular phase of the oestrous cycle. Forty samples of endometrial tissues were collected post-mortem from mares on the follicular phase of the oestrous cycle (10 samples of each K-D category). Relative mRNA transcription of TLR-2, TLR-4 and COL1A2 genes was assessed using qPCR, and COL1 protein expression by Western blot analysis. The COL1A2 transcription increased in category IIB when compared to categories I, IIA and III endometria (p < .01). The relative protein abundance of COL1 showed no significant differences between endometrial categories (p > .05). As for the TLRs mRNA transcription, TLR-2/-4 transcripts increased in IIA when compared to the other K-D endometria categories (p < .05). Our findings suggest that TLRs may be involved in the initiation of the endometrial inflammatory response. Additional studies are needed to explore TLRs' potential role as diagnostic markers for monitoring inflammation progression and fibrosis development, as well as their involvement in the mechanisms underlying fibrotic pathways.

Six months of physical inactivity is insufficient to cause chronic kidney disease in C57BL/6J mice.

Chronic kidney disease (CKD) is a progressive disorder marked by a decline in kidney function. Obesity and sedentary behavior contribute to the development of CKD, though mechanisms by which this occurs are poorly understood. This knowledge gap is worsened by the lack of a reliable murine CKD model that does not rely on injury, toxin, or gene deletion to induce a reduction in kidney function. High-fat diet (HFD) feeding alone is insufficient to cause reduced kidney function until later in life. Here, we employed a small mouse cage (SMC), a recently developed mouse model of sedentariness, to study its effect on kidney function. Wildtype C57BL/6J male mice were housed in sham or SMC housing for six months with HFD in room (22°C) or thermoneutral (30°C) conditions. Despite hyperinsulinemia induced by the SMC+HFD intervention, kidneys from these mice displayed normal glomerular filtration rate (GFR). However, the kidneys showed early signs of kidney injury, including increases in Col1a1 and NGAL transcripts, as well as fibrosis by histology, primarily in the inner medullary/papilla region. High-resolution respirometry and fluorometry experiments showed no statistically significant changes in the capacities for respiration, ATP synthesis, or electron leak. These data confirm the technical challenge in modeling human CKD. They further support the notion that obesity and a sedentary lifestyle make the kidneys more vulnerable, but additional insults are likely required for the pathogenesis of CKD.

Myeloid AMPK signaling restricts fibrosis but is not required for metformin improvements during CDAHFD-induced NASH in mice

Metabolic programming underpins inflammation and liver macrophage activation in the setting of chronic liver disease. Here, we sought to identify the role of an important metabolic regulator, AMP-activated protein kinase (AMPK), specifically within myeloid cells during the progression of non-alcoholic steatohepatitis (NASH) and whether treatment with metformin, a firstline therapy for diabetes and activator of AMPK could stem disease progression. Male and female Prkaa1fl/fl/Prkaa2fl/fl (Flox) control and Flox-LysM-Cre+ (MacKO) mice were fed a low-fat control or a choline-deficient, amino acid defined 45% Kcal high-fat diet (CDAHFD) for 8 weeks, where metformin was introduced in the drinking water (50 or 250 mg/kg/day) for the last 4 weeks. Hepatic steatosis and fibrosis were dramatically increased in response to CDAHFD-feeding compared to low-fat control. While myeloid AMPK signaling had no effect on markers of hepatic steatosis or circulating markers, fibrosis as measured by total liver collagen was significantly elevated in livers from MacKO mice, independent of sex. Although treatment with 50 mg/kg/day metformin had no effect on any parameter, intervention with 250 mg/kg/day metformin completely ameliorated hepatic steatosis and fibrosis in both male and female mice. While the protective effect of metformin was associated with lower final body weight, and decreased expression of lipogenic and Col1a1 transcripts, it was independent of myeloid AMPK signaling. These results suggest that endogenous AMPK signaling in myeloid cells, both liver-resident and infiltrating, acts to restrict fibrogenesis during CDAHFD-induced NASH progression but is not the mechanism by which metformin improves markers of NASH.

Pedunculoside promotes oral ulcer healing in mice by upregulating STAT3 and Smad3

BackgroundPedunculoside (PE) is a pentacyclic triterpenoid saponin found in the dried barks of the Ilex rotunda Thunb. In traditional Chinese medicine, the juice extracted from various Ilex species is commonly used for treating inflammatory diseases. ObjectiveThis study aims to investigate the potential of PE in promoting the healing of oral ulcers in mice and to explore its possible mechanism. Materials and MethodsA mouse model of oral ulcer was established to evaluate the healing promotion effects of PE, and an in-vitro inflammatory cell model was established by using primary fibroblasts isolated from mouse oral mucosal tissue. Gene transcription and protein expression related to cell proliferation, migration, and collagen synthesis were measured. ResultsPE showed the highest cure rate of 85.7% for oral ulcers in mice within 7-days treatment. It significantly promoted cell proliferation and collagen secretion in fibroblast in-vitro model. PE also upregulated the gene transcription of Smad3, Col1a1 and Col3a1. Furthermore, PE antagonized the inhibitory effect of WP1066 on STAT3activation, but did not affect cell migration. ConclusionThis study demonstrates that PE has potential therapeutic effects in promoting oral ulcer healing in mice. The improvement was achieved by the increasing collagen synthesis through the upregulation of STAT3 and Smad3 in mucosa fibroblasts. These results provide a preliminary pharmacological work for developing PE as a potential treatment agent for oral ulcers.

COL1A1 proximal promoter topology regulates its transcriptional response to transforming growth factor β

ABSTRACT Objective The COL1A1 proximal promoter contains two GC-rich regions and two inverted CCAAT boxes. The transcription factors Sp1 and CBF bind to the GC sequence at −122 to −115 bp and the inverted CCAAT box at −101 to −96 bp, respectively, and stimulate COL1A1 transcriptional activity. Methods To further define the regulatory mechanisms controlling COL1A1 expression by Sp1 and CBF, we introduced 2, 4, 6, or 8 thymidine nucleotides (T-tracts) at position −111 bp of the COL1A1 gene promoter to increase the physical distance between these two binding sites and examined in vitro the transcriptional activities of the resulting constructs and their response to TGF-β1.` Results Insertion of 2 or 4 nucleotides decreased COL1A1 promoter activity by up to 70%. Furthermore, the expected increase in COL1A1 transcription in response to TGF-β1 was abolished. Computer modeling of the modified DNA structure indicated that increasing the physical distance between the Sp1 and CBF binding sites introduces a rotational change in the DNA topology that disrupts the alignment of Sp1 and CBF binding sites and likely alters protein–protein interactions among these transcription factors or their associated co-activators. Conclusion The topology of the COL1A1 proximal promoter is crucial in determining the transcriptional activity of the gene and its response to the stimulatory effects of TGF-β1.

Toll-like receptor 5 tunes hepatic and pancreatic stellate cells activation

ObjectiveStellate cells are responsible for liver and pancreas fibrosis and strictly correlate with tumourigenesis. Although their activation is reversible, an exacerbated signalling triggers chronic fibrosis. Toll-like receptors (TLRs) modulate stellate...

Assessment of the viability and quality of equine endometrial explants as a potential ex vivo model for fibrosis research

There is a lack of experimental models to study endometrosis ex vivo. We evaluated viability, RNA integrity, gene and protein expression of genes relevant to fibrosis in an ex vivo model with mare endometrial explants. Uteri from healthy mares were collected during the reproductive season at an abattoir, and transported on ice to the laboratory. Uteri without signs of inflammation or contamination and an active ovary were processed: endometrial biopsies (∼50mg) were collected, washed (Hanks balanced salt solution + 0.1% BSA and 3xAAM), placed in 24-well plates (1 mL) and pre-incubated in DMEM with the same additives at 37°C, 5%CO2 for 1h. Thereafter, medium was replaced and explants cultured for 0, 24, 48, 72 or 96h. At these time points, viability was assessed using Alamar blue staining, RNA and proteins were extracted using Trizol. RNA integrity and quantity were assessed using RIN measurement (Tape Station Bioanalyzer; Agilent, USA). Proteins were quantified by Kit Pierce™ BCA Protein Assay (Thermo, USA) and visualized in SDS-PAGE (10% polyacrylamide). Statistical differences for viability assays were performed by ANOVA (P<0.05 cutoff). Culturing explants for 24 or 48h did not affect their viability compared to T0, but viability dropped after 72 and 96h (p˂ 0.05). Non-viable explants were used as reference, and time zero was set aspositive control. RNA integrity declined with prolonged culture of explants, RIN values at 96h were 4 to 6, indicative of RNA degradation. Relative expression of candidate genes involved in fibrosis and apoptosis by RT-PCR demonstrated a steady decrease in the expression of αSMA, PTGES, COL1A1 and COL3A1 transcripts at all time points compared to T0 (p<0.05). Conversely, expression of pro-apoptotic gene BAX increased (p<0.05) when explants were culture for 48h and further. We did not study any particular protein, but the overall yield (Pierce assay) and quality of proteins (SDS-PAGE) in the explants also dropped alongside with increased culture, being more evident after 48h. In conclusion, endometrial explants can be cultured in vitro for up to 48h without noticeable decrease of viability, RNA and protein yield and integrity, and with measurable expression of candidate genes. Prolonging culture for longer than 48h negatively affected viability and quality of the explants. The window of time of 48h of culture of endometrial explants could allow for the evaluation of potential anti-fibrotic drugs or treatments for mare endometrosis. Funded by FONDECYT REGULAR 1210349, ANID, Government of Chile.

5-Aza-2′-Deoxycytidine (5-Aza-dC, Decitabine) Inhibits Collagen Type I and III Expression in TGF-β1-Treated Equine Endometrial Fibroblasts

Endometrosis negatively affects endometrial function and fertility in mares, due to excessive deposition of type I (COL1) and type III (COL3) collagens. The pro-fibrotic transforming growth factor (TGF-β1) induces myofibroblast differentiation, characterized by α-smooth muscle actin (α-SMA) expression, and collagen synthesis. In humans, fibrosis has been linked to epigenetic mechanisms. To the best of our knowledge, this has not been described in mare endometrium. Therefore, this study aimed to investigate the in vitro epigenetic regulation in TGF-β1-treated mare endometrial fibroblasts and the use of 5-aza-2'-deoxycytidine (5-aza-dC), an epigenetic modifier, as a putative treatment option for endometrial fibrosis. Methods and Results: The in vitro effects of TGF-β1 and of 5-aza-dC on DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B), COL1A1, COL3A1, and α-SMA transcripts were analyzed in endometrial fibroblasts, and COL1 and COL3 secretion in a co-culture medium. TGF-β1 upregulated DNMT3A transcripts and collagen secretion. In TGF-β1-treated endometrial fibroblasts, DNA methylation inhibitor 5-aza-dC decreased collagen transcripts and secretion, but not α-SMA transcripts. Conclusion: These findings suggest a possible role of epigenetic mechanisms during equine endometrial fibrogenesis. The in vitro effect of 5-aza-dC on collagen reduction in TGF-β1-treated fibroblasts highlights this epigenetic involvement. This may pave the way to different therapeutic approaches for endometrosis.

Minoxidil weakens newly synthesized collagen in fibrotic synoviocytes from osteoarthritis patients

PurposeSynovial fibrosis (SFb) formation and turnover attributable to knee osteoarthritis (KOA) can impart painful stiffness and persist following arthroplasty. To supplement joint conditioning aimed at maximizing peri-operative function, we evaluated the antifibrotic effect of Minoxidil (MXD) on formation of pyridinoline (Pyd) cross-links catalyzed by Plod2-encoded lysyl hydroxylase (LH)2b that strengthen newly synthesized type-I collagen (COL1) in fibroblastic synovial cells (FSCs) from KOA patients. MXD was predicted to decrease Pyd without significant alterations to Col1a1 transcription by FSCs stimulated with transforming growth factor (TGF)β1.MethodsSynovium from 10 KOA patients grouped by SFb severity was preserved for picrosirius and LH2b histology or culture. Protein and RNA were purified from fibrotic FSCs after 8 days with or without 0.5 µM MXD and/or 4 ng/mL of TGFβ1. COL1 and Pyd protein concentrations from ELISA and expression of Col1a1, Acta2, and Plod2 genes by qPCR were compared by parametric tests with α = 0.05.ResultsHistological LH2b expression corresponded to SFb severity. MXD attenuated COL1 output in KOA FSCs but only in the absence of TGFβ1 and consistently decreased Pyd under all conditions with significant downregulation of Plod2 but minimal alterations to Col1a1 and Acta2 transcripts.ConclusionsMXD is an attractive candidate for local antifibrotic pharmacotherapy for SFb by compromising the integrity of newly formed fibrous deposits by FSCs during KOA and following arthroplasty. Targeted antifibrotic supplementation could improve physical therapy and arthroscopic lysis strategies aimed at breaking down joint scarring. However, the effect of MXD on other joint-specific TGFβ1-mediated processes or non-fibrotic components requires further investigation.

CGRP induces myofibroblast differentiation and the production of extracellular matrix in MRC5s via autocrine and paracrine signalings.

There are contradictory views on which calcitonin gene-related peptide (CGRP) causes pulmonary fibrosis. Fibrotic potency of CGRP was tested and compared to that of transforming growth factor-β (TGF-β). Myofibroblast differentiation, cell proliferation, and activations of TGF-β and Wnt pathways were examined for 24, 48, and 72 h in A549 and MRC5 cell lines stimulated with CGRP and TGF-β. CGRP-induced cell proliferation in MRC5s early on while cell proliferation in A549 occurred progressively. CGRP promoted fibroblast-myofibroblast differentiation by inducing the transcription of ACTA2, COL1A1, SMAD2/3, and SMAD4 genes, the production of collagen, fibronectin, α-smooth muscle actin, and activation of TGF-β signaling starting from 24 h. Additionally, TGF-β signaling induced by CGRP decreased theDKK1 level and activated the Wnt signaling in MRC5s. After CGRP stimulation, Wnt7a levels were increased from 24to 72 h, while Wnt5a levels were elevated at 72 h in MRC5s. CGRP did not induce epithelial-mesenchymal transition in A549s, unlike TGF-β.A comparison of fibrotic potency of CGRP and TGF-β showed that TGF-β is a powerful profibrotic molecule and induces earlier myofibroblast differentiation. Even so, CGRP promotes myofibroblast differentiation and extracellular matrix production by inducing Smad-dependent-TGF-β and Wnt signalings via autocrine and paracrine signalings in MRC5s.

Metallopeptidades 2 and 9 genes epigenetically modulate equine endometrial fibrosis.

Endometrium type I (COL1) and III (COL3) collagen accumulation, periglandular fibrosis and mare infertility characterize endometrosis. Metalloproteinase-2 (MMP-2), MMP-9 and tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) are involved in collagen turnover. Since epigenetic changes may control fibroproliferative diseases, we hypothesized that epigenetic mechanisms could modulate equine endometrosis. Epigenetic changes can be reversed and therefore extremely promising for therapeutic use. Methylation pattern analysis of a particular gene zone is used to detect epigenetic changes. DNA methylation commonly mediates gene repression. Thus, this study aimed to evaluate if the transcription of some genes involved in equine endometrosis was altered with endometrial fibrosis, and if the observed changes were epigenetically modulated, through DNA methylation analysis. Endometrial biopsies collected from cyclic mares were histologically classified (Kenney and Doig category I, n = 6; category IIA, n = 6; category IIB, n = 6 and category III, n = 6). Transcription of COL1A1, COL1A2, COL3A1, MMP2, MMP9, TIMP1, and TIMP2 genes and DNA methylation pattern by pyrosequencing of COL1A1, MMP2, MMP9, TIMP1 genes were evaluated. Both MMP2 and MMP9 transcripts decreased with fibrosis, when compared with healthy endometrium (category I) (P < 0.05). TIMP1 transcripts were higher in category III, when compared to category I endometrium (P < 0.05). No differences were found for COL1A1, COL1A2, COL3A1 and TIMP2 transcripts between endometrial categories. There were higher methylation levels of (i) COL1A1 in category IIB (P < 0.05) and III (P < 0.01), when compared to category I; (ii) MMP2 in category III, when compared to category I (P < 0.001) and IIA (P < 0.05); and (iii) MMP9 in category III, when compared to category I and IIA (P < 0.05). No differences in TIMP1 methylation levels were observed between endometrial categories. The hypermethylation of MMP2 and MMP9, but not of COL1A1 genes, occurred simultaneously with a decrease in their mRNA levels, with endometrial fibrosis, suggesting that this hypermethylation is responsible for repressing their transcription. Our results show that endometrosis is epigenetically modulated by anti-fibrotic genes (MMP2 and MMP9) inhibition, rather than fibrotic genes activation and therefore, might be promising targets for therapeutic use.

GIRK3 deletion facilitates kappa opioid signaling in chondrocytes, delays vascularization and promotes bone lengthening in mice.

Long bones are formed and repaired through the process of endochondral ossification. Activation of G protein-coupled receptor (GPCR) signaling pathways is crucial for skeletal development and long bone growth. G protein-gated inwardly-rectifying K+ (GIRK) channel genes are key functional components and effectors of GPCR signaling pathways in excitable cells of the heart and brain, but their roles in non-excitable cells that directly contribute to endochondral bone formation have not been studied. In this study, we analyzed skeletal phenotypes of Girk2-/-, Girk3-/- and Girk2/3-/- mice. Bones from 12-week-old Girk2-/- mice were normal in length, but femurs and tibiae from Girk3-/- and Girk2/3-/- mice were longer than age-matched controls at 12-weeks-old. Epiphyseal chondrocytes from 5-day-old Girk3-/- mice expressed higher levels of genes involved in collagen chain trimerization and collagen fibril assembly, lower levels of genes encoding VEGF receptors, and produced larger micromasses than wildtype chondrocytes in vitro. Girk3-/- chondrocytes were also more responsive to the kappa opioid receptor (KOR) ligand dynorphin, as evidenced by greater pCREB expression, greater cAMP and GAG production, and upregulation of Col2a1 and Sox9 transcripts. Imaging studies showed that Kdr (Vegfr2) and endomucin expression was dramatically reduced in bones from young Girk3-/- mice, supporting a role for delayed vasculogenesis and extended postnatal endochondral bone growth. Together these data indicate that GIRK3 controls several processes involved in bone lengthening.

Decreased expression of transcription factor Homeobox A11 and its potential target genes in bladder cancer

Bladder cancer (BC) ranks as the ninth most commonly diagnosed cancer worldwide. The presence of a transcription factor (TF) has been uncovered as a significant contributor to the pathophysiological changes of cancers. In present study, we elucidated the expression and clinical significance of Homeobox A11 (HOXA11) in BC for the first time, and originally investigated HOXA11 as a TF. Employing in-house immunohistochemistry (IHC), we incorporated 137 BC and 34 non-BC cases to detect the expression of HOXA11 protein in BC tissues. HOXA11-related RNA-sequencing (RNA-seq) expression and RNA microarrays were collected from public databases, the “sva” and “limma” R packages were implemented to integrate and normalize the RNA-seq data and microarrays separately. Integration expression was carried out to further evaluate the HOXA11 expression by utilizing the standard mean difference (SMD). The expression level of HOXA11 in various BC cell lines was also evaluated. We further systematically analyzed the downstream target genes of HOXA11 in BC by utilizing Chromatin Immunoprecipitation Sequencing (ChIP-seq) profiles, differentially expressed genes (DEGs), and HOXA11-related genes. Modification of histone marks on the promoter region of target genes were also discovered by histone ChIP-seq data. Results of the IHC and RNA-seq revealed the protein and mRNA expression of HOXA11 was significantly decreased in BC tissues compared to non-BC tissues (2.98 ± 1.48 vs. 8.23 ± 2.64; 6.87 ± 1.54 vs. 8.38 ± 1.42). Five platforms significantly revealed the down-regulation of HOXA11 expression in BC (GPL96, GPL570, GPL6102, GPL6884, and GPL13497). A similar decreased trend was discovered in BC tissues in expression integration with the incorporated SMD reaching −0.843 (−1.362 ~ −0.325, p = 0.001) and −1.051 (−1.674 ~ −0.428, p = 0.001). Expression of HOXA11 was down-regulated in most of the BC cell lines. COL1A1 was considered as a final HOXA11 target gene and positively related to HOXA11 with the correlation coefficient as 0.584 (95% CI: 0.371–0.739, p < 0.001). HOXA11 regulates COL1A1 expression in BC via H3K27ac modification. The expression of COL1A1 was down-regulated with the SMD reached −0.312 (p < 0.001). In conclusion, HOXA11 expression is markedly decreased and might promote the transcription of COL1A1 to inhibit BC.

Selective inhibitors of bromodomain BD1 and BD2 of BET proteins modulate radiation-induced profibrotic fibroblast responses.

Radiotherapy can induce various adverse effects including fibrosis in cancer patients. Radiation-induced aberrant expression of profibrotic genes has been associated with dysregulated epigenetic mechanisms. Pan-BET (bromodomain and extraterminal domain) inhibitors, such as JQ1 and I-BET151, have been reported to attenuate the profibrotic response after irradiation. Despite their profound preclinical efficacy, the clinical utility of pan-inhibitors is limited due to observed cytotoxicicities. Recently, inhibitors were developed that selectively target the first (BD1) and second (BD2) bromodomain of the BET proteins (iBET-BD1 [GSK778] and iBET-BD2 [GSK046]). Here, their potential to attenuate radiation-induced fibroblast activation with low-toxicity was investigated. Our results indicated that cell proliferation and cell cycle progression in fibroblasts from BJ cells and six donors were reduced when treated with I-BET151 and iBET-BD1, but not with iBET-BD2. After irradiation, induction of DGKA and profibrotic markers, especially COL1A1 and ACTA2, was attenuated with all BET inhibitors. H3K27ac enrichment was similar at the DGKA enhancer region after I-BET151 treatment and irradiation, but was reduced at the COL1A1 transcription start site and the ACTA2 enhancer site. iBET-BD2 did not change H3K27ac levels in these regions. BRD4 occupancy at these regions was not altered by any of the compounds. Cell migration activity was measured as a characteristic independent of extracellular matrix production and was unchanged in fibroblasts after irradiation and BET inhibitor-treatment. In conclusion, iBET-BD2 efficiently suppressed radiation-induced expression of DGKA and profibrotic markers without showing cytotoxicity. Thus BD2-selective targeting is a promising new therapeutic avenue for further investigations to prevent or attenuate radiotherapy-induced fibrosis.

Coupling induction of osteogenesis and type H vessels by pulsed electromagnetic fields in ovariectomy-induced osteoporosis in mice.

The growth of blood vessels and osteogenesis are coupled in bone tissue. A specialized subset of CD31hiEndomucinhi (CD31hiEmcnhi) vascular endothelium in bone has been identified to positively regulate bone formation. Pulsed electromagnetic field (PEMF) can promote the facture healing and reverse the loss of bone mass. However, the underlying mechanisms mediating in the positive effects of PEMF on bone mass accrual remain unclear. In the ovariectomized (OVX) osteoporotic mouse model, PEMF with specific parameters was administrated after 12weeks of surgery and continued for 8weeks. μCT analysis, quantitative PCR and Elisa assays were used to assess the PEMF-induced the osteogenesis, while immunostaining and flow cytometry were used to evaluate the abundance of CD31hiEmcnhi endothelium in the metaphysis near the growth plate. Administration of PEMF substantially countered OVX-induced bone loss as shown by greater trabecular bone, higher expression of Osterix, PDGFB and Col-1a1 transcripts, and modulation of bone anabolic and catabolic activity. The PEMF-induced osteogenesis was coupled by the expansion of CD31hiEmcnhi endothelium as demonstrated by CD31 and Endomucin double-positive immunostaining and flow cytometry. Concurrently, the higher level of HIF-α was found in PEMF-treated mice than in vehicle controls. Notably, inhibition of HIF-1α considerably reduced PEMF-induced osteogenesis, and led to a remarkable decrease of CD31hiEmcnhi vessels in the PEMF-treated OVX mice. The present study demonstrated the PEMF-induced coupling promotion of osteogenesis and CD31hiEmcnhi endothelial cells in a mouse model of postmenopausal osteoporosis. This coupling effect might be mediated in HIF-1α signaling in CD31hiEmcnhi endothelium. These findings open up new directions of al that might enable therapeutic improvement of osteogenesis in patients with osteoporosis.

Right predominant electrical remodeling in a pure model of pulmonary hypertension promotes reentrant arrhythmias.

Electrophysiological (EP) properties have been studied mainly in the monocrotaline model of pulmonary arterial hypertension (PAH). Findings are confounded by major extrapulmonary toxicities, which preclude the ability to draw definitive conclusions regarding the role of PAH per se in EP remodeling. The purpose of this study was to investigate the EP substrate and arrhythmic vulnerability of a new model of PAH that avoids extracardiopulmonary toxicities. Sprague-Dawley rats underwent left pneumonectomy (Pn) followed by injection of the vascular endothelial growth factor inhibitor Sugen-5416 (Su/Pn). Five weeks later, cardiac magnetic resonance imaging was performed invivo, optical action potential (AP) mapping exvivo, and molecular analyses invitro. Su/Pn rats exhibited right ventricular (RV) hypertrophy and were highly prone to pacing-induced ventricular tachycardia/fibrillation (VT/VF). Underlying this susceptibility was disproportionate RV-sided prolongation of AP duration, which promoted formation of right-sided AP alternans at physiological rates. While propagation was impaired at all rates in Su/Pn rats, the extent of conduction slowing was most severe immediately before the emergence of interventricular lines of block and onset of VT/VF. Measurement of the cardiac wavelength revealed a decrease in Su/Pn relative to control. Nav1.5 and total connexin 43 expression was not altered, while connexin 43 phosphorylation was decreased in PAH. Col1a1 and Col3a1 transcripts were upregulated coinciding with myocardial fibrosis. Once generated, VT/VF was sustained by multiple reentrant circuits with a lower frequency of RV activation due to wavebreak formation. In this pure model of PAH, we document RV-predominant remodeling that promotes multiwavelet reentry underlying VT. The Su/Pn model represents a severe form of PAH that allows the study of EP properties without the confounding influence of extrapulmonary toxicity.

Lycopene nanoparticles promotes osteoblastogenesis and inhibits adipogenesis of rat bone marrow mesenchymal stem cells.

Lycopene is a carotenoid and antioxidant with potent singlet oxygen quenching ability that reduces oxidative stress and promotes bone health. However, the cellular mechanisms by which lycopene influences bone metabolism are not known. The present study investigated the effects of lycopene nanoparticles on the differentiation of rat bone marrow-derived mesenchymal stem cells into osteoblasts or adipocytes. In osteogenic medium, lycopene supplementation dose-dependently enhanced osteoblast differentiation, as evidenced by the transcription of Alpl, Runx2, Col1a1, Sp7, and Bglap, higher alkaline phosphatase activity, osteocalcin secretion and extracellular matrix mineralisation seen with Alizarin red S staining, and increased haem oxygenase levels. By contrast, lycopene in adipogenic medium inhibited adipocyte differentiation evidenced by decreases in the transcription of Tnfsf11, Tnfrsf11b, Pparg, Lpl, and Fabp4 and reduced fat accumulation observed by Oil Red O staining. Lycopene nanoparticles may promote bone health and are considered as a potential candidate for the prevention and/or treatment of bone loss conditions.

The Inhibitory Effect of Noscapine on the In Vitro Cathepsin G-Induced Collagen Expression in Equine Endometrium.

Cathepsin G (CAT) is a protease released by neutrophils when forming neutrophil extracellular traps that was already associated with inducing type I collagen (COL1) in equine endometrium in vitro. Endometrosis is a fibrotic condition mainly characterized by COL1 deposition in the equine endometrium. The objective was to evaluate if noscapine (an alkaloid for cough treatment with anti-neoplastic and anti-fibrotic properties) would reduce COL1A2 transcription (evaluated by qPCR) and COL1 protein relative abundance (evaluated by western blot) induced by CAT in equine endometrial explants from follicular and mid-luteal phases treated for 24 or 48 h. The explants treated with CAT increased COL1 expression. Noscapine decreased COL1A2 transcription at both estrous cycle phases, but COL1 relative protein only at the follicular phase, both induced by CAT. Additionally, the noscapine anti-fibrotic action was found to be more effective in the follicular phase. The CAT treatment caused more fibrosis at the longest period of treatment, while noscapine acted better at the shortest time of treatment. Our results showed that noscapine could act as an anti-fibrotic drug in equine endometrosis by inhibiting CAT in vitro. Noscapine offers a new promising therapeutic tool for treating fibrosis as a single non-selective agent to be considered in the future.