Can irisin be developed as the molecular evolutionary clock based on the origin and functions?

Irisin, a myokine identified in 2012, has garnered research interest for its capacity to induce browning of adipocytes and improve metabolic parameters. As such, the potential therapeutic applications of this exercise-induced peptide continue to be explored. Though present across diverse animal species, sequence analysis has revealed subtle variation in the irisin protein. In this review, we consider the effects of irisin on disease states in light of its molecular evolution. We summarize current evidence for irisin's influence on pathologies and discuss how sequence changes may inform development of irisin-based therapies. Furthermore, we propose that the phylogenetic variations in irisin could potentially be leveraged as a molecular clock to elucidate evolutionary relationships.

Expression and Impact of Fibronectin, Tenascin-C, Osteopontin, and Type XIV Collagen in Fuchs Endothelial Corneal Dystrophy.

Purpose: Fuchs endothelial corneal dystrophy (FECD) is characterized by Descemet's membrane (DM) abnormalities, namely an increased thickness and a progressive appearance of guttae and fibrillar membranes. The goal of this study was to identify abnormal extracellular matrix (ECM) proteins expressed in FECD DMs and to evaluate their impact on cell adhesion and migration. Methods: Gene expression profiles from in vitro (GSE112039) and ex vivo (GSE74123) healthy and FECD corneal endothelial cells were analyzed to identify deregulated matrisome genes. Healthy and end-stage FECD DMs were fixed and analyzed for guttae size and height. Immunostaining of fibronectin, tenascin-C, osteopontin, and type XIV collagen was performed on ex vivo specimens, as well as on tissue-engineered corneal endothelium reconstructed using healthy and FECD cells. An analysis of ECM protein expression according to guttae and fibrillar membrane was performed using immunofluorescent staining and phase contrast microscopy. Finally, cell adhesion was evaluated on fibronectin, tenascin-C, and osteopontin, and cell migration was studied on fibronectin and tenascin-C. Results: SPP1 (osteopontin), FN1 (fibronectin), and TNC (tenascin-C) genes were upregulated in FECD ex vivo cells, and SSP1 was upregulated in both in vitro and ex vivo FECD conditions. Osteopontin, fibronectin, tenascin-C, and type XIV collagen were expressed in FECD specimens, with differences in their location. Corneal endothelial cell adhesion was not significantly affected by fibronectin or tenascin-C but was decreased by osteopontin. The combination of fibronectin and tenascin-C significantly increased cell migration. Conclusions: This study highlights new abnormal ECM components in FECD, suggests a certain chronology in their deposition, and demonstrates their impact on cell behavior.

Deletion of FNDC5/irisin modifies murine osteocyte function in a sex-specific manner.

Irisin, released from exercised muscle, has been shown to have beneficial effects on numerous tissues but its effects on bone are unclear. We found significant sex and genotype differences in bone from wildtype (WT) mice compared to mice lacking Fndc5 (knockout [KO]), with and without calcium deficiency. Despite their bone being indistinguishable from WT females, KO female mice were partially protected from osteocytic osteolysis and osteoclastic bone resorption when allowed to lactate or when placed on a low-calcium diet. Male KO mice have more but weaker bone compared to WT males, and when challenged with a low-calcium diet lost more bone than WT males. To begin to understand responsible molecular mechanisms, osteocyte transcriptomics was performed. Osteocytes from WT females had greater expression of genes associated with osteocytic osteolysis and osteoclastic bone resorption compared to WT males which had greater expression of genes associated with steroid and fatty acid metabolism. Few differences were observed between female KO and WT osteocytes, but with a low-calcium diet, the KO females had lower expression of genes responsible for osteocytic osteolysis and osteoclastic resorption than the WT females. Male KO osteocytes had lower expression of genes associated with steroid and fatty acid metabolism, but higher expression of genes associated with bone resorption compared to male WT. In conclusion, irisin plays a critical role in the development of the male but not the female skeleton and protects male but not female bone from calcium deficiency. We propose irisin ensures the survival of offspring by targeting the osteocyte to provide calcium in lactating females, a novel function for this myokine.

Rare genetic variation in fibronectin 1 (FN1) protects against APOEε4 in Alzheimer's disease.

The risk of developing Alzheimer's disease (AD) significantly increases in individuals carrying the APOEε4 allele. Elderly cognitively healthy individuals with APOEε4 also exist, suggesting the presence of cellular mechanisms that counteract the pathological effects of APOEε4; however, these mechanisms are unknown. We hypothesized that APOEε4 carriers without dementia might carry genetic variations that could protect them from developing APOEε4-mediated AD pathology. To test this, we leveraged whole-genome sequencing (WGS) data in the National Institute on Aging Alzheimer's Disease Family Based Study (NIA-AD FBS), Washington Heights/Inwood Columbia Aging Project (WHICAP), and Estudio Familiar de Influencia Genetica en Alzheimer (EFIGA) cohorts and identified potentially protective variants segregating exclusively among unaffected APOEε4 carriers. In homozygous unaffected carriers above 70 years old, we identified 510 rare coding variants. Pathway analysis of the genes harboring these variants showed significant enrichment in extracellular matrix (ECM)-related processes, suggesting protective effects of functional modifications in ECM proteins. We prioritized two genes that were highly represented in the ECM-related gene ontology terms, (FN1) and collagen type VI alpha 2 chain (COL6A2) and are known to be expressed at the blood-brain barrier (BBB), for postmortem validation and in vivo functional studies. An independent analysis in a large cohort of 7185 APOEε4 homozygous carriers found that rs140926439 variant in FN1 was protective of AD (OR = 0.29; 95% CI [0.11, 0.78], P = 0.014) and delayed age at onset of disease by 3.37 years (95% CI [0.42, 6.32], P = 0.025). The FN1 and COL6A2 protein levels were increased at the BBB in APOEε4 carriers with AD. Brain expression of cognitively unaffected homozygous APOEε4 carriers had significantly lower FN1 deposition and less reactive gliosis compared to homozygous APOEε4 carriers with AD, suggesting that FN1 might be a downstream driver of APOEε4-mediated AD-related pathology and cognitive decline. To validate our findings, we used zebrafish models with loss-of-function (LOF) mutations in fn1b-the ortholog for human FN1. We found that fibronectin LOF reduced gliosis, enhanced gliovascular remodeling, and potentiated the microglial response, suggesting that pathological accumulation of FN1 could impair toxic protein clearance, which is ameliorated with FN1 LOF. Our study suggests that vascular deposition of FN1 is related to the pathogenicity of APOEε4, and LOF variants in FN1 may reduce APOEε4-related AD risk, providing novel clues to potential therapeutic interventions targeting the ECM to mitigate AD risk.

(-)-epicatechin treatment did not modify the thermogenic pathway in the gastrocnemius muscle of male rat offspring obeses by programming.

The aim of this study was to analyse the expression of genes related to the regulation of energy metabolism in skeletal muscle tissue by comparing male offspring in two age groups [at 110 and 245 postnatal days (pnd)] from a mother with obesity induced by a high-fat diet and (-)-epicatechin (Epi) administration. Four groups of six male offspring from different litters were randomly selected for the control groups [C and offspring of mothers with maternal obesity (MO)] or Epi intervention groups. We evaluated the effect of Epi on gastrocnemius tissue by analysing the mRNA and protein expression levels of Fndc5/irisin, Pgc-1α, Ucp3, and Sln. Epi significantly increased the Pgc-1α protein in the MO group of offspring at 110 pnd (p < 0.036, MO vs. MO+Epi), while at 245 pnd, Epi increased Fndc5/irisin mRNA expression in the MO+Epi group versus the MO group (p = 0.006).No differences were detected in Fndc5/irisin, Ucp3 or Sln mRNA or protein levels (including Pgc-1α mRNA) in the offspring at 110 pnd or in Pgc-1α, Ucp3, or Sln mRNA or protein levels (including Fndc5/irisin protein) at 245 pnd among the experimental groups. In conclusion, (-)-epicatechin treatment increased Fndc5/irisin mRNA expression and Pgc-α protein levels in the gastrocnemius muscle of offspring at postnatal days 110 and 245. Furthermore, it is suggested that the flavonoid effect in a model of obesity and its impact on thermogenesis in skeletal muscle are regulated by a different pathway than Fndc5/irisin.

Fibronectin 1 supports oocyte in vitro maturation in pigs.

Oocyte in vitro maturation (IVM) based on the follicular fluid (FF) environment can exploit untapped resources, however, what FF factors regulate oocyte maturation remains unclear. This work demonstrated that serum and FF significantly promoted oocyte polar body extrusion (PBE) and subsequent embryo development, and FF was especially effective. Fibronectin 1 (FN1) was predicted as one potential candidate to regulate oocyte maturation by proteomics. FN1 transcription obviously decreased, and the protein expression significantly increased and migrated to plasma membrane or even outside during oocyte IVM. Treatment with 10 ng/mL FN1 significantly improved oocyte PBE rate. FN1 significantly upregulated the percentage of regular spindle morphology, downregulated the γ-H2AX level, decreased the levels of ROS and apoptosis, and increased GSH and mitochondrion contents by ameliorating the expression of corresponding genes. Moreover, FN1 significantly increased the p-PI3K level to enhance the activation of PI3K signaling pathway. In conclusion, this study discovers and confirms that FN1 is one factor in FF that significantly enhances oocyte maturation, and the underlying mechanism is that FN1 ameliorates oocyte nuclear and cytoplasmic maturation by promoting the activation of PI3K signaling pathway.

Impact of L-Arginine on diabetes-induced neuropathy and myopathy: Roles of PAI-1, Irisin, oxidative stress, NF-κß, autophagy and microRNA-29a.

BACKGROUND: T2DM is a chronic disorder with progressive neuromuscular alterations. L-arginine (ARG) is the most common semi-essential amino acid having several metabolic functions. AIM: to investigate the impact of L-arginine in combating diabetic-induced neuromyopathy and its possible mechanisms. MATERIALS & METHODS: 24 rats were divided into CON, CON+ARG, DC, DC+ARG. Behavioral tests, Body weight (BW), fasting blood glucose (FBG), insulin, total antioxidant capacity (TAC), malondialdehyde (MDA), plasminogen activator inhibitor-1 (PAI-1), and irisin were done. Creatine kinase-MM (CK-MM), interleukin 4 (IL-4), interleukin 6 (IL-6), TAC, MDA, expression of microRNA-29a mRNA & light chain 3 protein were determined in muscle. Histological and NF-κß immunohistochemical expression in muscle and nerve were assessed. RESULTS: ARG supplementation to diabetic rats improved altered behavior, significantly increased BW, insulin, TAC, irisin and Il-4, decreased levels of glucose, microRNA-29a, NF-κß and LC3 expression, PAI-1, CK-MM and restored the normal histological appearance. CONCLUSIONS: ARG supplementation potently alleviated diabetic-induced neuromuscular alterations.

mir-150-5p inhibits the osteogenic differentiation of bone marrow-derived mesenchymal stem cells by targeting irisin to regulate the p38/MAPK signaling pathway.

PURPOSE: To study the effect of miR-150-5p on the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs), and further explore the relationship between its regulatory mechanism and irisin. METHODS: We isolated mouse BMSCs, and induced osteogenic differentiation by osteogenic induction medium. Using qPCR to detect the expression of osteogenic differentiation-related genes, western blot to detect the expression of osteogenic differentiation-related proteins, and luciferase reporter system to verify that FNDC5 is the target of miR-150-5p. Irisin intraperitoneal injection to treat osteoporosis in mice constructed by subcutaneous injection of dexamethasone. RESULTS: Up-regulation of miR-150-5p inhibited the proliferation of BMSCs, and decreased the content of osteocalcin, ALP activity, calcium deposition, the expression of osteogenic differentiation genes (Runx2, OSX, OCN, OPN, ALP and BMP2) and protein (BMP2, OCN, and Runx2). And down-regulation of miR-150-5p plays the opposite role of up-regulation of miR-150-5p on osteogenic differentiation of BMSCs. Results of luciferase reporter gene assay showed that FNDC5 gene was the target gene of miR-150-5p, and miR-150-5p inhibited the expression of FNDC5 in mouse BMSCs. The expression of osteogenic differentiation genes and protein, the content of osteocalcin, ALP activity and calcium deposition in BMSCs co-overexpressed by miR-150-5p and FNDC5 was significantly higher than that of miR-150-5p overexpressed alone. In addition, the overexpression of FNDC5 reversed the blocked of p38/MAPK pathway by the overexpression of miR-150-5p in BMSCs. Irisin, a protein encoded by FNDC5 gene, improved symptoms in osteoporosis mice through intraperitoneal injection, while the inhibitor of p38/MAPK pathway weakened this function of irisin. CONCLUSION: miR-150-5p inhibits the osteogenic differentiation of BMSCs by targeting irisin to regulate the/p38/MAPK signaling pathway, and miR-150-5p/irisin/p38 pathway is a potential target for treating osteoporosis.

FNDC5/irisin ameliorates bone loss of type 1 diabetes by suppressing endoplasmic reticulum stress­mediated ferroptosis.

BACKGROUND: Ferroptosis is known to play a crucial role in diabetic osteopathy. However, key genes and molecular mechanisms remain largely unclear. This study aimed to identify a crucial ferroptosis-related differentially expressed gene (FR-DEG) in diabetic osteopathy and investigate its potential mechanism. METHODS: We identified fibronectin type III domain-containing protein 5 (FNDC5)/irisin as an essential FR-DEG in diabetic osteopathy using the Ferroptosis Database (FerrDb) and GSE189112 dataset. Initially, a diabetic mouse model was induced by intraperitoneal injection of streptozotocin (STZ), followed by intraperitoneal injection of irisin. MC3T3-E1 cells treated with high glucose (HG) were used as an in vitro model. FNDC5 overexpression plasmid was used to explore underlying mechanisms in vitro experiments. Femurs were collected for micro-CT scan, histomorphometry, and immunohistochemical analysis. Peripheral serum was collected for ELISA analysis. Cell viability was assessed using a CCK-8 kit. The levels of glutathione (GSH), malondialdehyde (MDA), iron, reactive oxygen species (ROS), and lipid ROS were detected by the corresponding kits. Mitochondria ultrastructure was observed through transmission electron microscopy (TEM). Finally, mRNA and protein expressions were examined by quantitative real-time PCR (qRT-PCR) and western blot analysis. RESULTS: The expression of FNDC5 was found to be significantly decreased in both in vivo and in vitro models. Treatment with irisin significantly suppressed ferroptosis and improved bone loss. This was demonstrated by reduced lipid peroxidation and iron overload, increased antioxidant capability, as well as the inhibition of the ferroptosis pathway in bone tissues. Furthermore, in vitro studies demonstrated that FNDC5 overexpression significantly improved HG-induced ferroptosis and promoted osteogenesis. Mechanistic investigations revealed that FNDC5 overexpression mitigated ferroptosis in osteoblasts by inhibiting the eukaryotic initiation factor 2 alpha (eIF2α)/activated transcription factor 4 (ATF4)/C/EBP-homologous protein (CHOP) pathway. CONCLUSIONS: Collectively, our study uncovered the important role of FNDC5/irisin in regulating ferroptosis of diabetic osteopathy, which might be a potential therapeutic target.

Myocardin regulates fibronectin expression and secretion from human pleural mesothelial cells.

During the progression of pleural fibrosis, pleural mesothelial cells (PMCs) undergo a phenotype switching process known as mesothelial-mesenchymal transition (MesoMT). During MesoMT, transformed PMCs become myofibroblasts that produce increased extracellular matrix (ECM) proteins, including collagen and fibronectin (FN1) that is critical to develop fibrosis. Here, we studied the mechanism that regulates FN1 expression in myofibroblasts derived from human pleural mesothelial cells (HPMCs). We found that myocardin (Myocd), a transcriptional coactivator of serum response factor (SRF) and a master regulator of smooth muscle and cardiac muscle differentiation, strongly controls FN1 gene expression. Myocd gene silencing markedly inhibited FN1 expression. FN1 promoter analysis revealed that deletion of the Smad3-binding element diminished FN1 promoter activity, whereas deletion of the putative SRF-binding element increased FN1 promoter activity. Smad3 gene silencing decreased FN1 expression, whereas SRF gene silencing increased FN1 expression. Moreover, SRF competes with Smad3 for binding to Myocd. These results indicate that Myocd activates FN1 expression through Smad3, whereas SRF inhibits FN1 expression in HPMCs. In HPMCs, TGF-ß induced Smad3 nuclear localization, and the proximity ligation signal between Myocd and Smad3 was markedly increased after TGF-ß stimulation at nucleus, suggesting that TGF-ß facilitates nuclear translocation of Smad3 and interaction between Smad3 and Myocd. Moreover, Myocd and Smad3 were coimmunoprecipitated and isolated Myocd and Smad3 proteins directly bound each other. Chromatin immunoprecipitation assays revealed that Myocd interacts with the FN1 promoter at the Smad3-binding consensus sequence. The results indicate that Myocd regulates FN1 gene activation through interaction and activation of the Smad3 transcription factor.NEW & NOTEWORTHY During phenotype switching from mesothelial to mesenchymal, pleural mesothelial cells (PMCs) produce extracellular matrix (ECM) proteins, including collagen and fibronectin (FN1), critical components in the development of fibrosis. Here, we found that myocardin, a transcriptional coactivator of serum response factor (SRF), strongly activates FN1 expression through Smad3, whereas SRF inhibits FN1 expression. This study provides insights about the regulation of FN1 that could lead to the development of novel interventional approaches to prevent pleural fibrosis.

Distinguishing characteristics of Staphylococcus schleiferi and Staphylococcus coagulans of human and canine origin.

Staphylococcus schleiferi and Staphylococcus coagulans are opportunistic pathogens of animals and humans. They were previously classified as Staphylococcus schleiferi subs. schleiferi and Staphylococcus schleiferi subs. coagulans, respectively, and recently reclassified as separate species. S. coagulans, is frequently associated with dogs, whereas S. schleiferi is more commonly isolated from humans. Coagulase activity status is a defining characteristic of the otherwise closely related species. However, the use of coagulase tests originally developed to distinguish S. aureus from non-coagulase-producing staphylococci, for this purpose is questionable and the basis for their host preference has not been elucidated. In the current study, a putative coa gene was identified and correlated with coagulase activity measured using a chromogenic assay with human and bovine prothrombin (closely related to canine prothrombin). The results of the tests performed with human prothrombin showed greater reactivity of S. coagulans isolates from humans than isolates obtained from dogs with the same substrate. Our data suggest that unlike S. coagulans isolates from humans, isolates from dogs have more coagulase activity with bovine prothrombin (similar to canine prothrombin) than human prothrombin. Differences in nuc and 16s rRNA genes suggest a divergence in S. coagulans and S. schleiferi. Phenotypic and genotypic variation based on the number of IgG binding domains, and the numbers of tandem repeats in C-terminal fibronectin binding motifs was also found in protein A, and fibronectin-binding protein B respectively. This study identified a coa gene and associated phenotypic activity that differentiates S. coagulans and S. schleiferi and identified key phylogenetic and phenotypic differences between the species.

Live imaging of Fibronectin 1a-mNeonGreen and Fibronectin 1b-mCherry knock-in alleles during early zebrafish development.

Within the developing embryo, cells assemble and remodel their surrounding extracellular matrix during morphogenesis. Fibronectin is an extracellular matrix glycoprotein and is a ligand for several members of the Integrin adhesion receptor family. Here, we compare the expression pattern and loss of function phenotypes of the two zebrafish fibronectin paralogs fn1a and fn1b. We engineered two fluorescently tagged knock-in alleles to facilitate live in vivo imaging of the Fibronectin matrix. Genetic complementation experiments indicate that the knock-in alleles are fully functional. Fn1a-mNeonGreen and Fn1b-mCherry are co-localized in ECM fibers on the surface of the paraxial mesoderm and myotendinous junction. In 5-days old zebrafish larvae, Fn1a-mNeonGreen predominantly localizes to the branchial arches, heart ventricle, olfactory placode and within the otic capsule while Fn1b-mCherry is deposited at the pericardium, proximal convoluted tubule, posterior hindgut and at the ventral mesoderm/cardinal vein. We examined Fn1a-mNeonGreen and Fn1b-mCherry in maternal zygotic integrin α5 mutants and integrin ß1a; ß1b double mutants and find distinct requirements for these Integrins in assembling the two Fibronectins into ECM fibers in different tissues. Rescue experiments via mRNA injection indicate that the two fibronectins are not fully inter-changeable. Lastly, we examined cross-regulation between the two Fibronectins and find fn1a is necessary for normal Fn1b fibrillogenesis in the presomitic mesoderm, but fn1b is dispensable for the normal pattern of Fn1a deposition.

The integrated analysis and underlying mechanisms of FNDC5 on diabetic induced cognitive deficits.

OBJECTIVES: Chronic hyperglycemia is considered as an important factor to promote the neurodegenerative process of brain, and the synaptic plasticity as well as heterogeneity of hippocampal cells are thought to be associated with cognitive dysfunction in the early process of neurodegeneration. To date, fibronectin type III domain-containing protein 5 (FNDC5) has been highlighted its protective role in multiple neurodegenerative diseases. However, the potential molecular and cellular mechanisms of FNDC5 on synaptic plasticity regulation in cognitive impairment (CI) induced by diabetics are still need to known. METHODS/DESIGN: To investigate the heterogeneity and synaptic plasticity of hippocampus in animals with CI state induced by hyperglycemia, and explore the potential role of FNDC5 involved in this process. Firstly, the single cell sequencing was performed based on the hippocampal tissue from db diabetic mice induced CI and normal health control mice by ex vivo experiments; and then the integrated analysis and observations validation using Quantitative Real-time PCR, western blot as well as other in vitro studies. RESULTS: We observed and clarified the sub-cluster of type IC spiral ganglion neurons expressed marker genes as Trmp3 and sub-cluster of astrocytes with marker gene as Atp1a2 in hippocampal cells from diabetic animals induced CI and the effect of those on neuron-glial communication. We also found that FNDC5\BDNF-Trk axis was involved in the synaptic plasticity regulation of hippocampus. In high glucose induced brain injury model in vitro, we investigated that FNDC5 significantly regulates BDNF expression and that over-expression of FNDC5 up-regulated BDNF expression (p < 0.05) and can also significantly increase the expression of synapsin-1 (p < 0.05), which is related to synaptic plasticity, In addition, the unbalanced methylation level between H3K4 and H3K9 in Fndc5 gene promoter correlated with significantly down-regulated expression of FNDC5 (p < 0.05) in the hyperglycemia state. CONCLUSION: The current study revealed that the synaptic plasticity of hippocampal cells in hyperglycemia might be regulated by FNDC5\BDNF-Trk axis, playing the protective role in the process of CI induced by hyperglycemia and providing a target for the early treatment of hyperglycemia induced cognitive dysfunction in clinic.

Astrocytic TIMP-1 regulates production of Anastellin, an inhibitor of oligodendrocyte differentiation and FTY720 responses.

Astrocyte activation is associated with neuropathology and the production of tissue inhibitor of metalloproteinase-1 (TIMP1). TIMP1 is a pleiotropic extracellular protein that functions both as a protease inhibitor and as a growth factor. Astrocytes that lack expression of Timp1 do not support rat oligodendrocyte progenitor cell (rOPC) differentiation, and adult global Timp1 knockout (Timp1KO) mice do not efficiently remyelinate following a demyelinating injury. Here, we performed an unbiased proteomic analysis and identified a fibronectin-derived peptide called Anastellin (Ana) that was unique to the Timp1KO astrocyte secretome. Ana was found to block rOPC differentiation in vitro and enhanced the inhibitory influence of fibronectin on rOPC differentiation. Ana is known to act upon the sphingosine-1-phosphate receptor 1, and we determined that Ana also blocked the pro-myelinating effect of FTY720 (or fingolimod) on rOPC differentiation in vitro. Administration of FTY720 to wild-type C57BL/6 mice during MOG35-55-experimental autoimmune encephalomyelitis ameliorated clinical disability while FTY720 administered to mice lacking expression of Timp1 (Timp1KO) had no effect. Analysis of Timp1 and fibronectin (FN1) transcripts from primary human astrocytes from healthy and multiple sclerosis (MS) donors revealed lower TIMP1 expression was coincident with elevated FN1 in MS astrocytes. Last, analyses of proteomic databases of MS samples identified Ana peptides to be more abundant in the cerebrospinal fluid (CSF) of human MS patients with high disease activity. A role for Ana in MS as a consequence of a lack of astrocytic TIMP-1 production could influence both the efficacy of fingolimod responses and innate remyelination potential in the MS brain.

Anti-Fibrosis Effect of Panax ginseng and Inula japonica Formula in Human Pulmonary Fibroblasts.

Panax ginseng Meyer and Inula japonica Thunb. are well established in traditional medicine and are known for their therapeutic properties in managing a range of ailments such as diabetes, asthma, and cancer. Although P. ginseng and I. japonica can alleviate pulmonary fibrosis (PF), the anti-fibrosis effect on PF by the combination of two herbal medicines remains unexplored. Therefore, this study explores this combined effect. In conditions that were not cytotoxic, MRC-5 cells underwent treatment using the formula combining P. ginseng and I. japonica (ISE081), followed by stimulation with transforming growth factor (TGF)-ß1, to explore the fibroblast-to-myofibroblast transition (FMT). After harvesting the cells, mRNA levels and protein expressions associated with inflammation and FMT-related markers were determined to evaluate the antiinflammation activities and antifibrosis effect of ISE081. Additionally, the anti-migratory effects of ISE081 were validated through a wound-healing assay. ISE081 remarkably reduced the mRNA levels of interleukin (IL)-6, IL-8, α-smooth muscle actin (SMA), and TGF-ß1 in MRC-5 cells and suppressed the α-SMA and fibronectin expressions, respectively. Furthermore, ISE081 inhibited Smad2/3 phosphorylation and wound migration of MRC-5 cells. Under the same conditions, comparing those of ISE081, P. ginseng did not affect the expression of α-SMA, fibronectin, and Smad2/3 phosphorylation, whereas I. japonica significantly inhibited them but with cytotoxicity. The results indicate that the synergistic application of P. ginseng and I. japonica enhances the anti-fibrotic properties in pulmonary fibroblasts and concurrently diminishes toxicity. Therefore, ISE081 has the potential as a prevention and treatment herbal medicine for PF.

Exercise ameliorating myocardial injury in type 2 diabetic rats by inhibiting excessive mitochondrial fission involving increased irisin expression and AMP-activated protein kinase phosphorylation.

PURPOSE: Though exercise generates beneficial effects on diabetes-associated cardiac damage, the underlying mechanism is largely unclear. Therefore, we prescribed a program of 8-week treadmill training for type 2 diabetes mellitus (T2DM) rats and determined the role of irisin signaling, via interacting with AMP-activated protein kinase (AMPK), in mediating the effects of exercise on myocardial injuries and mitochondrial fission. METHODS: Forty 8-week-old male Wistar rats were randomly divided into groups of control (Con), diabetes mellitus (DM), diabetes plus exercise (Ex), and diabetes plus exercise and Cyclo RGDyk (ExRg). Ex and ExRg rats received 8 weeks of treadmill running, and the rats in the ExRg group additionally were treated with a twice weekly injection of Cyclo RGDyk, an irisin receptor-αV/ß5 antagonist. At the end of the experiment, murine blood samples and heart tissues were collected and analyzed with methods of ELISA, Western blot, real-time quantitative polymerase chain reaction, as well as immunofluorescence staining. RESULTS: Exercise effectively mitigated T2DM-related hyperglycemia, hyperinsulinemia, lipid dysmetabolism, and inflammation, which could be diminished by Cyclo RGDyk treatment. Additionally, exercise alleviated T2DM-induced myocardial injury and excessive mitochondrial fission, whereas the beneficial effects were blocked by the administration of Cyclo RGDyk. T2DM significantly decreased serum irisin concentrations and fibronectin type III domain-containing protein 5 (FNDC5)/irisin gene and protein expression levels in the rat heart, whereas exercise could rescue T2DM-reduced FNDC5/irisin expression. Blocking irisin receptor signaling diminished the exercise-alleviated mitochondrial fission protein expression and elevated AMPK phosphorylation. CONCLUSION: Exercise is effective in mitigating diabetes-related insulin resistance, metabolic dysfunction, and inflammation. Irisin signaling engages in exercise-associated beneficial effects on myocardial injury and excessive mitochondrial fission in diabetes rats involving elevated AMPK phosphorylation.

Irisin inhibits CCK-8-induced TNF-α production via integrin αVß5-NF-κB signaling pathways in Nile tilapia (Oreochromis niloticus).

Irisin, a secreted myokine generated by fibronectin type III domain-containing protein 5, has recently shown the potential to alleviate inflammation. Cholecystokinin-octapeptide (CCK-8) is closely associated with the inflammatory factor TNF-α, a central cytokine in inflammatory reactions. However, the interactions between irisin and CCK-8 in regulating TNF-α production and the underlying mechanism have not yet been elucidated. In the present study, irisin treatment inhibited the basal and the CCK-8-induced TNF-α production in vivo. Additionally, neutralizing circulating irisin using an irisin antiserum significantly augmented the CCK-8-induced stimulation of TNF-α levels. Moreover, the incubation of head kidney cells with irisin or CCK-8 has opposite effects on TNF-α secretion. Notably, irisin treatment inhibited basal and CCK-8-stimulated TNF-α release and gene transcription in head kidney cells. Mechanistically, the inhibitory actions of irisin on basal and CCK-8-induced TNF-α production could be negated by co-administered with the selective integrin αVß5 inhibitor cilengitide. In addition, the inhibitory effect of irisin on basal and CCK-8-triggered TNF-α production could be abolished by the inhibition of the nuclear factor-kappa B (NF-κB) signaling pathway. Furthermore, irisin impeded CCK-8-induced phosphorylation and degradation of IκBα, simultaneously inhibiting NF-κB phosphorylation, preventing its translocation into the nucleus, and suppressing its DNA-binding activity induced by CCK-8. Collectively, these results suggest that the inhibitory effect of irisin on TNF-α production caused by CCK-8 is mediated via the integrin αVß5-NF-κB signaling pathways in tilapia.

Pathologic-genomic correlation identified a novel variant in FN1 and established the diagnosis of recurrent fibronectin glomerulopathy in the kidney allograft.

Fibronectin glomerulopathy is a rare inherited kidney disease, characterized by abnormal accumulation of fibronectin in the glomeruli. We report an exceptional case of recurrent fibronectin glomerulopathy first diagnosed in the kidney allograft. The presence of IgA staining in the native kidney biopsy and the reported family history of IgA nephropathy had led to initial pretransplant diagnosis of IgA nephropathy. At 4.5 years posttransplant, the patient presented with kidney insufficiency and minimal proteinuria. The allograft biopsy revealed glomerular deposits with very weak staining for immunoglobulins and vague filamentous material. Immunostaining for fibronectin was positive, and genetic studies showed a variant of unknown significance in the fibronectin 1 gene. Proteomic analyses of the glomeruli in the native kidney biopsy demonstrated large amount of fibronectin with abundant accumulation of the peptide synthesized by the detected variant. These findings established the diagnosis of recurrent fibronectin glomerulopathy secondary to a novel variant in the fibronectin 1 gene. This report sheds light on recurrent fibronectin glomerulopathy in the allograft, highlights the diagnostic pitfalls of the disease, and underscores the importance of pathologic-genomic correlation to establish the correct diagnosis.

Exercise enhances placental labyrinth trophoblast development by activation of PGC-1α and FNDC5/irisin†.

Placental chorion/labyrinth trophoblasts are energy demanding which is met by the mitochondrial oxidative phosphorylation. Exercise enhances placental development and mitochondrial biogenesis, but the underlying mechanisms remain poorly understood. To address, female C57BL/6 J mice were randomly assigned into two groups: a control group and an exercise (EX) group. All animals were acclimated to treadmill exercise for 1 week before mating, but only the EX group was subjected to daily exercise during pregnancy from embryonic day (E) 1.5 to E16.5. Placenta were collected at E18.5 for biochemical and histochemical analyses, and primary trophoblast cells were isolated from the E18.5 placenta for further analyses. The data showed that exercise during pregnancy promoted the expression of syncytiotrophoblast cell markers, indicating trophoblast cell differentiation, which was closely associated with elevated mitochondrial biogenesis and oxidative metabolism in the E18.5 placenta. In addition, exercise during pregnancy activated peroxisome proliferator-activated receptor γ coactivator-1 α (PGC-1α), which was associated with upregulated placental α-ketoglutarate and the expression of isocitrate dehydrogenases and ten-eleven translocations, facilitating DNA demethylation of the Pgc1a promoter. Furthermore, exercise upregulated fibronectin type III domain containing 5 expression and the secretion of its cleaved form, irisin, which is known to activate PGC-1α. These data suggest that exercise-induced activation of PGC-1α, via epigenetic modifications, is responsible for promoting mitochondrial energy metabolism and chorion/labyrinth trophoblast development.

SNHG16/miR-205/HDAC5 is involved in the progression of renal fibrosis.

Renal interstitial fibrosis (RIF) represents an irreversible and progressive pathological manifestation of chronic renal disease, which ultimately leads to end-stage renal disease. Long noncoding RNAs (lncRNAs) have been suggested to be involved in the progression of RIF. Small nucleolar RNA host gene 16 (SNHG16), a member of lncRNAs, has been found to be involved in the progression of pulmonary fibrosis. This paper first researched the effect of SNHG16 on renal fibrosis. We established a unilateral ureteral obstruction (UUO)-induced mouse RIF model by ligation of the left ureter to evaluate the biological function of SNHG16 in RIF. As a result, SNHG16 was upregulated in UUO-induced renal fibrotic tissues. Knockdown of SNHG16 inhibited RIF and reduced alpha-smooth muscle actin (α-SMA), fibronectin, and college IV expression. miR-205 was a target of SNHG16, and downregulated in UUO-induced renal fibrotic tissues. Inhibition of miR-205 promoted RIF and increased the expression of α-SMA, college IV, and fibronectin. Overexpression of SNHG16 promoted the UUO-induced RIF, but miR-205 abrogated this effect of SNHG16. Histone deacetylase 5 (HDAC5) showed high expression in UUO-induced renal fibrotic tissues. Knockdown of HDAC5 significantly reduced α-SMA, fibronectin, and college IV expression in renal tissues of UUO-induced mice. Inhibition of miR-205 promoted HDAC5 expression, but knockdown of SNHG16 inhibited HDAC5 expression in renal tissues of UUO-induced mice. In conclusion, SHNG16 is highly expressed in renal fibrotic tissues of UUO-induced mice. Knockdown of SHNG16 may prevent UUO-induced RIF by indirectly upregulating HDAC5 via targeting miR-205. SHNG16 may be novel target for treating renal fibrosis.