Dynamic changes in mitral valve extracellular matrix, tissue mechanics and function in a mouse model of Marfan syndrome.

OBJECTIVE: Mouse models of Marfan syndrome (MFS) with Fibrillin 1 (Fbn1) variant C1041G exhibit cardiovascular abnormalities, including myxomatous valve disease (MVD) and aortic aneurism, with structural extracellular matrix (ECM) dysregulation. In this study, we examine the structure-function-mechanics relations of the mitral valve related to specific transitions in ECM composition and organization in progressive MVD in MFS mice from Postnatal day (P)7 to 1 year-of-age. APPROACH AND RESULTS: Mechanistic links between mechanical forces and biological changes in MVD progression were examined in Fbn1C1041G/+ MFS mice. By echocardiography, mitral valve dysfunction is prevalent at 2 months with a decrease in cardiac function at 6 months, followed by a preserved cardiac function at 12 months. Mitral valve (MV) regurgitation occurs in a subset of mice at 2-6 months, while progressive dilatation of the aorta occurs from 2 to 12 months. Mitral valve tissue mechanical assessments using a uniaxial Permeabilizable Fiber System demonstrate decreased stiffness of MFS MVs at all stages. Histological and microscopic analysis of ECM content, structure, and fiber orientation demonstrate that alterations in ECM mechanics, composition, and organization precede functional abnormalities in Fbn1C1041G/+MFS MVs. At 2 months, ECM abnormalities are detected with an increase in proteoglycans and decreased stiffness of the mitral valve. By 6-12 months, collagen fiber remodeling is increased with abnormal fiber organization in MFS mitral valve leaflets. At the same time, matrifibrocyte gene expression characteristic of collagen-rich connective tissue is increased, as detected by RNA in situ hybridization and qPCR. Together, these studies demonstrate early prevalence of proteoglycans at 2 months followed by upregulation of collagen structure and organization with age in MVs of MFS mice. CONCLUSIONS: Altogether, our data indicate dynamic regulation of mitral valve structure, tissue mechanics, and function that reflect changes in ECM composition, organization, and gene expression in progressive MVD. Notably, increased collagen fiber organization and orientation, potentially dependent on increased matrifibrocyte cell activity, is apparent with altered mitral valve mechanics and function in aging MFS mice.

Curcumin Promotes Diabetic Foot Ulcer Wound Healing by Inhibiting miR-152-3p and Activating the FBN1/TGF-ß Pathway.

The objective of this study was to investigate the mechanism of curcumin in diabetic foot ulcer (DFU) wound healing. A DFU rat model was established, and fibroblasts were cultured in a high-glucose (HG) environment to create a cell model. Various techniques, including Western blot, RT‒qPCR, flow cytometry, Transwell, cell scratch test and H&E staining, were employed to measure the levels of relevant genes and proteins, as well as to assess cell proliferation, apoptosis, migration, and pathological changes. The results showed that miR-152-3p was overexpressed in DFU patients, while FBN1 was underexpressed. Curcumin was found to inhibit fibroblast apoptosis, promote proliferation, migration, and angiogenesis in DFU rats, and accelerate wound healing in DFU rats. In addition, overexpression of miR-152-3p weakened the therapeutic effect of curcumin, while overexpression of FBN1 reversed the effects of the miR-152-3p mimic. Further investigations into the underlying mechanisms revealed that curcumin expedited wound healing in DFU rats by restoring the FBN1/TGF-ß pathway through the inhibition of miR-152-3p. In conclusion, curcumin can suppress the activity of miR-152-3p, which, in turn, leads to the rejuvenation of the FBN1/TGF-ß pathway and accelerates DFU wound healing.

MFAP2 promotes HSCs activation through FBN1/TGF-ß/Smad3 pathway.

Liver fibrosis is a chronic inflammatory process characterized by the accumulation of extracellular matrix (ECM), which contributes to cirrhosis and hepatocellular carcinoma. Increasing evidence suggests that the activation of hepatic stellate cells (HSCs) under an inflammatory state leads to the secretion of collagens, which can cause cirrhosis. In this study, we analysed data from the Gene Expression Omnibus (GEO) databases to identify differentially expressed genes (DEGs) between quiescent and fibrotic HSCs. We found that Microfibril Associated Protein 2 (MFAP2) was elevated in carbon tetrachloride (CCl4)-induced liver fibrosis and Transforming Growth Factor-Beta 1 (TGF-ß1)-activated HSCs. Knockdown of MFAP2 inhibited HSC proliferation and partially attenuated TGF-ß-stimulated fibrogenesis markers. Bioinformatics analysis revealed that Fibrillin-1 (FBN1) was correlated with MFAP2, and the expression of FBN1 was significantly upregulated after MFAP2 overexpression. Silencing MFAP2 partially attenuated the activation of HSCs by inhibiting HSC proliferation and decreasing collagen deposits. In vitro results showed that the inhibition of MFAP2 alleviated hepatic fibrosis by inhibiting the activation and inducing the apoptosis of active HSCs in a CCl4-induced mouse model. In conclusion, our results suggest that MFAP2 is a potential target for the clinical treatment of liver fibrosis.

Redox Dysregulation of Vascular Smooth Muscle Sirtuin-1 in Thoracic Aortic Aneurysm in Marfan Syndrome.

BACKGROUND: Thoracic aortic aneurysms (TAAs) are abnormal aortic dilatations and a major cardiovascular complication of Marfan syndrome. We previously demonstrated a critical role for vascular smooth muscle (VSM) SirT1 (sirtuin-1), a lysine deacetylase, against maladaptive aortic remodeling associated with chronic oxidative stress and aberrant activation of MMPs (matrix metalloproteinases). METHODS: In this study, we investigated whether redox dysregulation of SirT1 contributed to the pathogenesis of TAA using fibrillin-1 hypomorphic mice (Fbn1mgR/mgR), an established model of Marfan syndrome prone to aortic dissection/rupture. RESULTS: Oxidative stress markers 3-nitrotyrosine and 4-hydroxynonenal were significantly elevated in aortas of patients with Marfan syndrome. Moreover, reversible oxidative post-translational modifications (rOPTM) of protein cysteines, particularly S-glutathionylation, were dramatically increased in aortas of Fbn1mgR/mgR mice, before induction of severe oxidative stress markers. Fbn1mgR/mgR aortas and VSM cells exhibited an increase in rOPTM of SirT1, coinciding with the upregulation of acetylated proteins, an index of decreased SirT1 activity, and increased MMP2/9 activity. Mechanistically, we demonstrated that TGFß (transforming growth factor beta), which was increased in Fbn1mgR/mgR aortas, stimulated rOPTM of SirT1, decreasing its deacetylase activity in VSM cells. VSM cell-specific deletion of SirT1 in Fbn1mgR/mgR mice (SMKO-Fbn1mgR/mgR) caused a dramatic increase in aortic MMP2 expression and worsened TAA progression, leading to aortic rupture in 50% of SMKO-Fbn1mgR/mgR mice, compared with 25% of Fbn1mgR/mgR mice. rOPTM of SirT1, rOPTM-mediated inhibition of SirT1 activity, and increased MMP2/9 activity were all exacerbated by the deletion of Glrx (glutaredoxin-1), a specific deglutathionylation enzyme, while being corrected by overexpression of Glrx or of an oxidation-resistant SirT1 mutant in VSM cells. CONCLUSIONS: Our novel findings strongly suggest a causal role of S-glutathionylation of SirT1 in the pathogenesis of TAA. Prevention or reversal of SirT1 rOPTM may be a novel therapeutic strategy to prevent TAA and TAA dissection/ruptures in individuals with Marfan syndrome, for which, thus far, no targeted therapy has been developed.

[EBF1 Promotes the Sensitivity of Cervical Cancer Cells to Cisplatin via Activating FBN1 Transcription].

Cisplatin (DDP) is widely used in the chemotherapy of cervical cancer (CC), the fourth most common female malignancy worldwide. However, some patients progress to chemotherapy resistance, which leads to chemotherapy failure, tumor recurrence, and poor prognosis. Therefore, strategies to identify the regulatory mechanisms underlying CC development and increase tumor sensitivity to DDP will help improve patient survival. This research was designed to ascertain the mechanism of EBF1-dependent regulation of FBN1 which promotes chemosensitivity of CC cells. The expression of EBF1 and FBN1 was measured in CC tissues resistant or sensitive to chemotherapy and in DDP-sensitive or -resistant cells (SiHa and SiHa-DDP cells). SiHa-DDP cells were transduced with lentiviruses encoding EBF1 or FBN1 to evaluate the influence of these two proteins on cell viability, expression of MDR1 and MRP1, and cell aggressiveness. Moreover, the interaction between EBF1 and FBN1 was predicted and demonstrated. Finally, to further verify the EBF1/FB1-dependent mechanism of DDP sensitivity regulation in CC cells a xenograft mouse model of CC was established using SiHa-DDP cells transduced with lentiviruses carrying EBF1 gene and shRNA directed to FBN1 EBF1 and FBN1 showed decreased expression in CC tissues and cells, particularly in those resistant to chemotherapy. Transduction of SiHa-DDP cells with lentiviruses encoding EBF1 or FBN1 lead to decreased viability, IC50, proliferation capacity, colony formation ability, aggressiveness, and increased cell apoptosis. We have shown that EBF1 activates FBN1 transcription by binding to FBN1 promoter region. Additionally, it was revealed that FBN1 silencing reversed the promoting effect of EBF1 overexpression on chemosensitivity of CC cells in vivo. EBF1 facilitated chemosensitivity in CC cells by activating FBN1 transcription.

Developmental and hormonal regulation of FBN1 and OR4M1 mRNA in bovine granulosa cells.

Recent studies have reported hormonal regulation of expression of fibrillin 1 (FBN1), the gene that encodes asprosin, in bovine theca cells, however, hormonal regulation of gene expression of FBN1 and the asprosin receptor, olfactory receptor 4M1 (OR4M1), has not been evaluated in granulosa cells (GC). This study was designed to characterize FBN1 and OR4M1 gene expression in GC during development of bovine dominant ovarian follicles, and to determine the hormonal regulation of FBN1 and OR4M1 mRNA expression in GC. GC FBN1 mRNA abundance was greater (P < 0.05) in medium (5.1-8 mm) estrogen inactive (EI) follicles than in large (>8.1 mm) or small (1-5 mm) EI follicles. In comparison, GC OR4M1 mRNA abundance was greater (P < 0.05) in small EI follicles than in large or medium EI follicles. Abundance of OR4M1 mRNA in GC of follicles collected on days 3 to 4 (early growth phase) and on days 5 to 6 (late growth phase) was similar, whereas FBN1 mRNA abundance was greater (P < 0.05) on days 5 to 6 vs days 3 to 4. Hormonal regulators for FBN1 mRNA abundance in cultured small-follicle GC were identified: TGFß1 causing a 2.45-fold increase, WNT3A causing a 1.45-fold increase, and IGF1 causing a 65% decrease. Steroids, leptin, insulin, growth hormone, follicle stimulating hormone, fibroblast growth factor 9 and epidermal growth factor had no effect on FBN1 mRNA abundance. Abundance of OR4M1 mRNA in GC was regulated by progesterone with 3.55-fold increase, but other hormones did not affect GC OR4M1 mRNA abundance. Findings indicate that both FBN1 and OR4M1 gene expression are hormonally and developmentally regulated in bovine follicles, and thus may affect asprosin production and its subsequent role in ovarian follicular function in cattle.

Fibrillin-1 regulates endothelial sprouting during angiogenesis.

Fibrillin-1 is an extracellular matrix protein that assembles into microfibrils which provide critical functions in large blood vessels and other tissues. Mutations in the fibrillin-1 gene are associated with cardiovascular, ocular, and skeletal abnormalities in Marfan syndrome. Here, we reveal that fibrillin-1 is critical for angiogenesis which is compromised by a typical Marfan mutation. In the mouse retina vascularization model, fibrillin-1 is present in the extracellular matrix at the angiogenic front where it colocalizes with microfibril-associated glycoprotein-1, MAGP1. In Fbn1C1041G/+ mice, a model of Marfan syndrome, MAGP1 deposition is reduced, endothelial sprouting is decreased, and tip cell identity is impaired. Cell culture experiments confirmed that fibrillin-1 deficiency alters vascular endothelial growth factor-A/Notch and Smad signaling which regulate the acquisition of endothelial tip cell/stalk cell phenotypes, and we showed that modulation of MAGP1 expression impacts these pathways. Supplying the growing vasculature of Fbn1C1041G/+ mice with a recombinant C-terminal fragment of fibrillin-1 corrects all defects. Mass spectrometry analyses showed that the fibrillin-1 fragment alters the expression of various proteins including ADAMTS1, a tip cell metalloprotease and matrix-modifying enzyme. Our data establish that fibrillin-1 is a dynamic signaling platform in the regulation of cell specification and matrix remodeling at the angiogenic front and that mutant fibrillin-1-induced defects can be rescued pharmacologically using a C-terminal fragment of the protein. These findings, identify fibrillin-1, MAGP1, and ADAMTS1 in the regulation of endothelial sprouting, and contribute to our understanding of how angiogenesis is regulated. This knowledge may have critical implications for people with Marfan syndrome.

Lineage-Specific Induced Pluripotent Stem Cell-Derived Smooth Muscle Cell Modeling Predicts Integrin Alpha-V Antagonism Reduces Aortic Root Aneurysm Formation in Marfan Syndrome Mice.

BACKGROUND: The role of increased smooth muscle cell (SMC) integrin αv signaling in Marfan syndrome (MFS) aortic aneurysm remains unclear. Herein, we examine the mechanism and potential efficacy of integrin αv blockade as a therapeutic strategy to reduce aneurysm progression in MFS. METHODS: Induced pluripotent stem cells (iPSCs) were differentiated into aortic SMCs of the second heart field (SHF) and neural crest (NC) lineages, enabling in vitro modeling of MFS thoracic aortic aneurysms. The pathological role of integrin αv during aneurysm formation was confirmed by blockade of integrin αv with GLPG0187 in Fbn1C1039G/+ MFS mice. RESULTS: iPSC-derived MFS SHF SMCs overexpress integrin αv relative to MFS NC and healthy control SHF cells. Furthermore, integrin αv downstream targets (FAK [focal adhesion kinase]/AktThr308/mTORC1 [mechanistic target of rapamycin complex 1]) were activated, especially in MFS SHF. Treatment of MFS SHF SMCs with GLPG0187 reduced p-FAK/p-AktThr308/mTORC1 activity back to control SHF levels. Functionally, MFS SHF SMCs had increased proliferation and migration compared to MFS NC SMCs and control SMCs, which normalized with GLPG0187 treatment. In the Fbn1C1039G/+ MFS mouse model, integrin αv, p-AktThr308, and downstream targets of mTORC1 proteins were elevated in the aortic root/ascending segment compared to littermate wild-type control. Mice treated with GLPG0187 (age 6-14 weeks) had reduced aneurysm growth, elastin fragmentation, and reduction of the FAK/AktThr308/mTORC1 pathway. GLPG0187 treatment reduced the amount and severity of SMC modulation assessed by single-cell RNA sequencing. CONCLUSIONS: The integrin αv-FAK-AktThr308 signaling pathway is activated in iPSC SMCs from MFS patients, specifically from the SHF lineage. Mechanistically, this signaling pathway promotes SMC proliferation and migration in vitro. As biological proof of concept, GLPG0187 treatment slowed aneurysm growth and p-AktThr308 signaling in Fbn1C1039G/+ mice. Integrin αv blockade via GLPG0187 may be a promising therapeutic approach to inhibit MFS aneurysmal growth.

Decreased mitochondrial function in UVA-irradiated dermal fibroblasts causes the insufficient formation of type I collagen and fibrillin-1 fibers.

BACKGROUND: Decreases of collagen fibers and the disappearance of oxytalan fibers are typical symptoms of photoaged skin. Although a low quality of mitochondria (MT) in photoaged skin cells has been observed, it is unknown whether the decreased quality of MT is responsible for the insufficient formation of dermal fibers. OBJECTIVE: To identify the role of mitochondrial quality in skin photoaging focusing on the formation of dermal fibers. METHODS: Type I collagen and fibrillin-1 fibers in normal human dermal fibroblasts (NHDFs) were observed by immunostaining. Type I collagen and fibrillin-1 proteins in NHDFs were quantified by ELISA. Mitochondrial quality was evaluated by measuring levels of intracellular ATP and MITOL, which regulate mitochondrial quality. RESULTS: UVA-irradiated NHDFs formed insufficient type I collagen and fibrillin-1 fibers and had a decreased ratio of extracellular versus intracellular levels of those proteins. Although expression levels of motor proteins that transport those proteins intracellularly were not affected by UVA, intracellular ATP levels, which is the driving force of motor proteins, were decreased by UVA along with decreased MITOL protein. Knockdown of MITOL in NHDFs decreased the level of intracellular ATP and caused the insufficient formation of type I collagen and fibrillin-1 fibers due to interfering with the secretion of those proteins. CONCLUSION: These results indicate that a low quality of MT with ATP depletion in dermal fibroblasts caused by irradiation with UVA induces the insufficient formation of type I collagen and fibrillin-1 fibers due to the decreased extracellular secretion of those proteins.

Deciphering the immune landscape dominated by cancer-associated fibroblasts to investigate their potential in indicating prognosis and guiding therapeutic regimens in high grade serous ovarian carcinoma.

Limited immunotherapeutic effect in high-grade serous ovarian carcinoma (HGSOC) propels exploration of the mechanics behind this resistance, which may be partly elucidated by investigating characters of cancer-associated fibroblasts (CAFs), a significant population in HGSOC involved in shaping tumor immune microenvironment. Herein, leveraging gene expression data of HGSOC samples from The Cancer Genome Atlas and Gene Expression Omnibus datasets, we suggested that CAFs detrimentally affected the outcomes of HGSOC patients. Subsequently, we performed weighted gene co-expression network analysis (WGCNA) to identify a CAFs-related module and screened out seven hub genes from this module, all of which were positively correlated with the infiltration of immunosuppressive macrophages. As one of the hub genes, the expression of fibrillin 1 (FBN1) and its relevance to CD206 were further verified by immunohistochemistry staining in HGSOC samples. Meanwhile, we extracted genes that correlated well with CAF signatures to construct a CAFscore. The capacity of the CAFscore as an independent prognostic factor was validated by Cox regression analyses, and its relevance to components as well as signals in the tumor immune microenvironment was also investigated. Under the evaluation by the CAFscore, HGSOC patients with relatively high CAFscore had worse outcomes, activated mesenchymal signaling pathways, and immune checkpoint blockade (ICB) resistance signatures, which was consistent with the fact that non-responders in anti-PD-1 treatment cohorts tended to have higher CAFscore. Besides, the possibility of CAFscore to guide the selection of sensitive chemotherapeutic agents was explored. In conclusion, individualized assessment of the CAFscore could uncover the extent of stroma activation and immunosuppression and inform therapeutic strategies to improve the benefit of therapies.

Succinylation Inhibits the Enzymatic Hydrolysis of the Extracellular Matrix Protein Fibrillin 1 and Promotes Gastric Cancer Progression.

Extracellular matrix (ECM) remodeling is crucial in the regulation of gastric cancer (GC) progression. This work aims to reveal novel posttranslational modifications and their relevant mechanisms in GC. In 3D matrix culture and animal models, it is found that fibrillin 1 (FBN1) expression is increased in advanced GC and has succinylation modification. The succinylation modification of FBN1 blocks its degradation by matrix metalloproteinases (MMPs). The long-term accumulation and deposition of FBN1 enhance tumor progression by activating TGF-ß1 and intracellular PI3K/Akt pathway. The FBN1 succinylation site monoclonal antibody can effectively intervene the effect of succinylation modification and inhibit GC progression. FBN1 is specifically upregulated in the progression of GC compared with other tumors. In conclusion, FBN1 is widely present in the form of K672-succinylated modifications in GC. Besides, the succinyl group of FBN1 blocks its binding to MMP2, inhibits its degradation by MMP2, and leads to the accumulation of FBN1, which poses a long-term risk to the poor prognosis of GC.

Fibrillin-1 mutant mouse captures defining features of human primary open glaucoma including anomalous aqueous humor TGF beta-2.

Primary open angle glaucoma (POAG) features an optic neuropathy, elevated aqueous humor (AH) TGFß2, and major risk factors of central corneal thickness (CCT), increasing age and intraocular pressure (IOP). We examined Tight skin (Tsk) mice to see if mutation of fibrillin-1, a repository for latent TGFß, is associated with characteristics of human POAG. We measured: CCT by ocular coherence tomography (OCT); IOP; retinal ganglion cell (RGC) and optic nerve axon counts by microscopic techniques; visual electrophysiologic scotopic threshold responses (STR) and pattern electroretinogram (PERG); and AH TGFß2 levels and activity by ELISA and MINK epithelial cell-based assays respectively. Tsk mice had open anterior chamber angles and compared with age-matched wild type (WT) mice: 23% thinner CCT (p < 0.003); IOP that was higher (p < 0.0001), more asymmetric (p = 0.047), rose with age (p = 0.04) and had a POAG-like frequency distribution. Tsk mice also had RGCs that were fewer (p < 0.04), declined with age (p = 0.0003) and showed increased apoptosis and glial activity; fewer optic nerve axons (p = 0.02); abnormal axons and glia; reduced STR (p < 0.002) and PERG (p < 0.007) visual responses; and higher AH TGFß2 levels (p = 0.0002) and activity (p = 1E-11) especially with age. Tsk mice showed defining features of POAG, implicating aberrant fibrillin-1 homeostasis as a pathogenic contributor to emergence of a POAG phenotype.

The Fibrillin-1/VEGFR2/STAT2 signaling axis promotes chemoresistance via modulating glycolysis and angiogenesis in ovarian cancer organoids and cells.

BACKGROUND: Chemotherapy resistance is a primary reason of ovarian cancer therapy failure; hence it is important to investigate the underlying mechanisms of chemotherapy resistance and develop novel potential therapeutic targets. METHODS: RNA sequencing of cisplatin-resistant and -sensitive (chemoresistant and chemosensitive, respectively) ovarian cancer organoids was performed, followed by detection of the expression level of fibrillin-1 (FBN1) in organoids and clinical specimens of ovarian cancer. Subsequently, glucose metabolism, angiogenesis, and chemosensitivity were analyzed in structural glycoprotein FBN1-knockout cisplatin-resistant ovarian cancer organoids and cell lines. To gain insights into the specific functions and mechanisms of action of FBN1 in ovarian cancer, immunoprecipitation, silver nitrate staining, mass spectrometry, immunofluorescence, Western blotting, and FÓ§rster resonance energy transfer-fluorescence lifetime imaging analyses were performed, followed by in vivo assays using vertebrate model systems of nude mice and zebrafish. RESULTS: FBN1 expression was significantly enhanced in cisplatin-resistant ovarian cancer organoids and tissues, indicating that FBN1 might be a key factor in chemoresistance of ovarian cancer. We also discovered that FBN1 sustained the energy stress and induced angiogenesis in vitro and in vivo, which promoted the cisplatin-resistance of ovarian cancer. Knockout of FBN1 combined with treatment of the antiangiogenic drug apatinib improved the cisplatin-sensitivity of ovarian cancer cells. Mechanistically, FBN1 mediated the phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2) at the Tyr1054 residue, which activated its downstream focal adhesion kinase (FAK)/protein kinase B (PKB or AKT) pathway, induced the phosphorylation of signal transducer and activator of transcription 2 (STAT2) at the tyrosine residue 690 (Tyr690), promoted the nuclear translocation of STAT2, and ultimately altered the expression of genes associated with STAT2-mediated angiogenesis and glycolysis. CONCLUSIONS: The FBN1/VEGFR2/STAT2 signaling axis may induce chemoresistance of ovarian cancer cells by participating in the process of glycolysis and angiogenesis. The present study suggested a novel FBN1-targeted therapy and/or combination of FBN1 inhibition and antiangiogenic drug for treating ovarian cancer.

Protein tyrosine phosphatase receptor δ serves as the orexigenic asprosin receptor.

Asprosin is a fasting-induced glucogenic and centrally acting orexigenic hormone. The olfactory receptor Olfr734 is known to be the hepatic receptor for asprosin that mediates its effects on glucose production, but the receptor for asprosin's orexigenic function has been unclear. Here, we have identified protein tyrosine phosphatase receptor δ (Ptprd) as the orexigenic receptor for asprosin. Asprosin functions as a high-affinity Ptprd ligand in hypothalamic AgRP neurons, regulating the activity of this circuit in a cell-autonomous manner. Genetic ablation of Ptprd results in a strong loss of appetite, leanness, and an inability to respond to the orexigenic effects of asprosin. Ablation of Ptprd specifically in AgRP neurons causes resistance to diet-induced obesity. Introduction of the soluble Ptprd ligand-binding domain in the circulation of mice suppresses appetite and blood glucose levels by sequestering plasma asprosin. Identification of Ptprd as the orexigenic asprosin receptor creates a new avenue for the development of anti-obesity therapeutics.

Sensitive asprosin detection in clinical samples reveals serum/saliva correlation and indicates cartilage as source for serum asprosin.

The C-terminal pro-fibrillin-1 propeptide asprosin is described as white adipose tissue derived hormone that stimulates rapid hepatic glucose release and activates hunger-promoting hypothalamic neurons. Numerous studies proposed correlations of asprosin levels with clinical parameters. However, the enormous variability of reported serum and plasma asprosin levels illustrates the need for sensitive and reliable detection methods in clinical samples. Here we report on newly developed biochemical methods for asprosin concentration and detection in several body fluids including serum, plasma, saliva, breast milk, and urine. Since we found that glycosylation impacts human asprosin detection we analyzed its glycosylation profile. Employing a new sandwich ELISA revealed that serum and saliva asprosin correlate strongly, depend on biological sex, and feeding status. To investigate the contribution of connective tissue-derived asprosin to serum levels we screened two cohorts with described cartilage turnover. Serum asprosin correlated with COMP, a marker for cartilage degradation upon running exercise and after total hip replacement surgery. This together with our finding that asprosin is produced by primary human chondrocytes and expressed in human cartilage suggests a contribution of cartilage to serum asprosin. Furthermore, we determined asprosin levels in breast milk, and urine, for the first time, and propose saliva asprosin as an accessible clinical marker for future studies.

Clinicopathological Significance and Molecular Mechanism of AKT/GSK3ß Pathway-based Fibrillin-1 Expression in Gastric Cancer Progression.

This research was developed to explore the significance of fibrillin-1 (FBN1) in the progression of gastric cancer and its relationship with the activation of the AKT/glycogen synthase kinase-3beta (GSK3ß) pathway. For this aim, immunohistochemical assays were adopted to detect FBN1 expression in chronic superficial gastritis, chronic atrophic gastritis, gastric cancer, and normal mucosa. The expression of FBN1 in gastric cancer and adjacent tissue samples was detected by reverse transcription-quantitative (RT-q) PCR and Western blot, and the relationship between FBN1 and the clinicopathological features of gastric cancer patients was analyzed. Lentivirus was utilized to construct SGC-7901 gastric cancer cell lines stably overexpressing and silencing FBN1, and the effects on cell proliferation, colony formation, and apoptosis were analyzed. AKT, GSK3ß, and their phosphorylated proteins were detected by Western blot. Results showed that the positive expression rate of FBN1 increased successively in chronic superficial gastritis, chronic atrophic gastritis, and gastric cancer. FBN1 was up-regulated in gastric cancer tissues and correlated with the depth of tumor invasion. Overexpression of FBN1 promoted the proliferation and colony formation of gastric cancer cells, inhibited apoptosis, and promoted the phosphorylation of AKT and GSK3ß. Silencing FBN1 expression inhibited the proliferation and clonal formation of gastric cancer cells, promoted apoptosis, and inhibited the phosphorylation of AKT and GSK3ß. In conclusion, FBN1 was up-regulated in gastric cancer tissues and correlated with the depth of gastric tumor invasion. FBN1 silencing inhibited the progression of gastric cancer through the AKT/GSK3ß pathway.

Arabidopsis fibrillin 1-2 subfamily members exert their functions via specific protein-protein interactions.

Fibrillins (FBNs) are plastidial proteins found in photosynthetic organisms from cyanobacteria to higher plants. The function of most FBNs remains unknown. Here, we focused on members of the FBN subgroup comprising FBN1a, FBN1b, and FBN2. We show that these three polypeptides interact between each other, potentially forming a network around the plastoglobule surface. Both FBN2 and FBN1s interact with allene oxide synthase, and the elimination of any of these FBNs results in a delay in jasmonate-mediated anthocyanin accumulation in response to a combination of moderate high light and low temperature. Mutations in the genes encoding FBN1s or FBN2 also affect the protection of PSII under the combination of these stresses. Fully developed leaves of these mutants have lower maximum quantum efficiency of PSII (Fv/Fm) and higher oxidative stress than wild-type plants. These effects are additive, and the fbn1a-1b-2 triple mutant shows a stronger decrease in Fv/Fm and a greater increase in oxidative stress than fbn1a-1b or fbn2 mutants. Co-immunoprecipitation analysis indicated that FBN2 also interacts with other proteins involved in different metabolic processes. We propose that these fibrillins facilitate accurate positioning of different proteins involved in distinct metabolic processes, and that their elimination leads to dysfunction of those proteins.

Polymethoxyflavones from Kaempferia parviflora ameliorate skin aging in primary human dermal fibroblasts and ex vivo human skin.

Skin aging is accompanied by an increase in the number of senescent cells, resulting in various pathological outcomes. These include inflammation, impaired barrier function, and susceptibility to skin disorders such as cancer. Kaempferia parviflora (Thai black ginger), a medicinal plant native to Thailand, has been shown to counteract inflammation, cancer, and senescence. This study demonstrates that polymethoxyflavones (5,7-dimethoxyflavone, 5,7,4'-trimethoxyflavone, and 3,5,7,3',4'-pentamethoxyflavone) purified from K. parviflora rhizomes suppressed cellular senescence, reactive oxygen species, and the senescence-associated secretory phenotype in primary human dermal fibroblasts. In addition, they increased tropocollagen synthesis and alleviated free radical-induced cellular and mitochondrial damage. Moreover, the compounds mitigated chronological aging in a human ex vivo skin model by attenuating senescence and restoring expression of essential components of the extracellular matrix, including collagen type I, fibrillin-1, and hyaluronic acid. Finally, we report that polymethoxyflavones enhanced epidermal thickness and epidermal-dermal stability, while blocking age-related inflammation in skin explants. Our findings support the use of polymethoxyflavones from K. parviflora as natural anti-aging agents, highlighting their potential as active ingredients in cosmeceutical and nutraceutical products.

Compositional Analysis of Extracellular Aggregates in the Eyes of Patients With Exfoliation Syndrome and Exfoliation Glaucoma.

Purpose: Exfoliation syndrome (XFS) is a condition characterized by the production of insoluble fibrillar aggregates (exfoliation material; XFM) in the eye and elsewhere. Many patients with XFS progress to exfoliation glaucoma (XFG), a significant cause of global blindness. We used quantitative mass spectrometry to analyze the composition of XFM in lens capsule specimens and in aqueous humor (AH) samples from patients with XFS, patients with XFG and unaffected individuals. Methods: Pieces of lens capsule and samples of AH were obtained with consent from patients undergoing cataract surgery. Tryptic digests of capsule or AH were analyzed by high-performance liquid chromatography-mass spectrometry and relative differences between samples were quantified using the tandem mass tag technique. The distribution of XFM on the capsular surface was visualized by SEM and super-resolution light microscopy. Results: A small set of proteins was consistently upregulated in capsule samples from patients with XFS and patients with XFG, including microfibril components fibrillin-1, latent transforming growth factor-ß-binding protein-2 and latent transforming growth factor-ß-binding protein-3. Lysyl oxidase-like 1, a cross-linking enzyme associated with XFS in genetic studies, was an abundant XFM constituent. Ligands of the transforming growth factor-ß superfamily were prominent, including LEFTY2, a protein best known for its role in establishing the embryonic body axis. Elevated levels of LEFTY2 were also detected in AH from patients with XFG, a finding confirmed subsequently by ELISA. Conclusions: This analysis verified the presence of suspected XFM proteins and identified novel components. Quantitative comparisons between patient samples revealed a consistent XFM proteome characterized by strong expression of fibrillin-1, lysyl oxidase-like-1, and LEFTY2. Elevated levels of LEFTY2 in the AH of patients with XFG may serve as a biomarker for the disease.

Intracerebroventricular asprosin administration strongly stimulates hypothalamic-pituitary-testicular axis in rats.

Asprosin, a protein-based secretary product of white adipose tissue, stimulates appetite hepatic glucose production. It crosses blood-brain barrier and stimulates appetite center and causes sperm chemotaxis but exact role of this endogenous agent is not completely known. This study was conducted to investigate possible effects of central asprosin infusion on the hormones involved in the hypothalamic-pituitary-testicular (HPT) axis and sperm cells. Spraque Dawley male rats were divided into four groups; control, sham, low asprosin (34) and high asprosin (68 nM) groups, (n = 10 for each group). Control group remain intact while a brain infusion kit was placed in the lateral ventricles of the rats in the sham group (artificial cerebrospinal fluid) and asprosin (34 and 68 nM) was infused for 14 days. At the end of the experiment, the hypothalamus, blood, and epididymis tissues of the rats were collected. Gonadotropin-releasing hormone (GnRH) mRNA and tissue protein levels were determined in the hypothalamus tissue by RT-PCR and Western Blot methods. Serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone levels were examined using the ELISA method from blood samples and sperm cells were examined in the epididymis tissue. GnRH mRNA and protein expressions of asprosin administered groups were higher than control and sham groups (p < 0.05). Asprosin infusion was also found to increase serum FSH, LH, and testosterone levels (p < 0.05). In addition, sperm density, motility, and progressive movement were observed to increase in asprosin administered groups (p < 0.05). This study suggests that central asprosin stimulate the HPT axis and also epididymis tissue. Our results implicates potential role for asprosin in male infertility.