Extracellular matrix remodelling in dental pulp tissue of carious human teeth through the prism of single-cell RNA sequencing.

Carious lesions are bacteria-caused destructions of the mineralised dental tissues, marked by the simultaneous activation of immune responses and regenerative events within the soft dental pulp tissue. While major molecular players in tooth decay have been uncovered during the past years, a detailed map of the molecular and cellular landscape of the diseased pulp is still missing. In this study we used single-cell RNA sequencing analysis, supplemented with immunostaining, to generate a comprehensive single-cell atlas of the pulp of carious human teeth. Our data demonstrated modifications in the various cell clusters within the pulp of carious teeth, such as immune cells, mesenchymal stem cells (MSC) and fibroblasts, when compared to the pulp of healthy human teeth. Active immune response in the carious pulp tissue is accompanied by specific changes in the fibroblast and MSC clusters. These changes include the upregulation of genes encoding extracellular matrix (ECM) components, including COL1A1 and Fibronectin (FN1), and the enrichment of the fibroblast cluster with myofibroblasts. The incremental changes in the ECM composition of carious pulp tissues were further confirmed by immunostaining analyses. Assessment of the Fibronectin fibres under mechanical strain conditions showed a significant tension reduction in carious pulp tissues, compared to the healthy ones. The present data demonstrate molecular, cellular and biomechanical alterations in the pulp of human carious teeth, indicative of extensive ECM remodelling, reminiscent of fibrosis observed in other organs. This comprehensive atlas of carious human teeth can facilitate future studies of dental pathologies and enable comparative analyses across diseased organs.

TGF-ß/YB-1/Atg7 axis promotes the proliferation of hepatic progenitor cells and liver fibrogenesis.

Hepatic fibrosis is characterized by excessive extracellular matrix deposition and ductular reactions, manifested as the expansion of hepatic progenitor cells (HPCs). We previously reported that the Y-box binding protein 1 (YB-1) in HPCs is involved in chronic liver injury. In this study, we constructed YB-1f/f Foxl1-Cre mice and investigated the role of YB-1 in HPC expansion in murine choline-deficient, ethionine-supplemented (CDE), and 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) models. Liver injury and fibrosis were measured using hematoxylin and eosin (HE), Masson, and Sirius Red staining. HPC proliferation was detected using EdU and immunofluorescence (IF). Autophagic flow was measured by mCherry-GFP-LC3B staining and transmission electron microscopy (TEM). YB-1 expression was measured by immunofluorescence and western blotting. CUT & Tag analysis, chromatin immunoprecipitation, and RT-PCR were performed to explore the regulation of autophagy-related protein 7 (Atg7) transcription by YB-1. Our results indicated that liver injury was accompanied by high expression of YB-1, proliferative HPCs, and activated autophagy in the CDE and DDC models. YB-1f/f Cre+/- mice displayed less liver injury and fibrosis than YB-1f/f Cre-/- mice in the CDE and DDC models. YB-1 promoted proliferation and autophagy of HPCs in vitro and in vivo. Transforming growth factor-ß (TGF-ß) induced YB-1 nuclear translocation and facilitated the proliferation and autophagy of HPCs. YB-1 nuclear translocation promoted the transcription of Atg7, which is essential for TGF-ß/YB-1 mediated HPCs expansion in vitro and in vivo. In summary, YB-1 nuclear translocation induced by TGF-ß in HPCs promotes the proliferation and autophagy of HPCs and Atg7 participates in YB-1-mediated HPC-expansion and liver fibrosis.

Exploring the mechanism of Danggui Buxue Decoction in regulating atherosclerotic disease network based on integrated pharmacological methods.

OBJECTIVE: To explore the mechanism of Danggui Buxue Decoction (DGBXD) in regulating Atherosclerosis (AS) network based on integrated pharmacological methods. METHODS: The active ingredients and targets of DGBXD are obtained from TCMSP database and ETCM. AS-related targets were collected from the Genecards and OMIM databases. The drug-disease protein interaction (PPI) networks were constructed by Cytoscape. Meanwhile, it was used to screen out densely interacting regions, namely clusters. Finally, Gene Ontology (GO) annotations are performed on the targets and genes in the cluster to obtain biological processes, and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations are performed on the targets of the PPI network to obtain signaling pathways. RESULTS: A total of 212 known targets, 265 potential targets and 229 AS genes were obtained. The 'DGBXD known-AS PPI network' and 'DGBXD-AS PPI Network' were constructed and analyzed. DGBXD can regulate inflammation, platelet activation, endothelial cell apoptosis, oxidative stress, lipid metabolism, vascular smooth muscle proliferation, angiogenesis, TNF, HIF-1, FoxO signaling pathway, etc. The experimental data showed that compared with the model group, the expressions of ICAM-1, VCAM-1, and interleukin (IL)-1ß protein and mRNA in the DGBXD group decreased (P<0.05). However, plasma IL-1ß, TNF-α, and MCP-1 in the DGBXD group were not significantly different from the model group (P>0.05). CONCLUSION: The mechanism of DGBXD in the treatment of AS may be related to the improvement of extracellular matrix (ECM) deposition in the blood vessel wall and the anti-vascular local inflammatory response, which may provide a reference for the study of the mechanism of DGBXD.

Avicularin suppresses cartilage extracellular matrix degradation and inflammation via TRAF6/MAPK activation.

BACKGROUND: Osteoarthritis (OA) is an intractable degenerative disease of the whole joint, which is characterized by synovitis inflammation, cartilage damage, and chronic pain. Tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) performs an important role in OA. PURPOSE: We aim to investigate avicularin to protect cartilage extracellular matrix degradation (ECM) and suppresses inflammation both in rat and human chondrocytes. METHODS: 5-Ethynyl-2'-deoxyuridine (EdU) staining, Quantitative real-time PCR, TRAF6 plasmid transfection, Western blot, Measurement of nitric oxide (NO), ROS detection and Immunofluorescence were utilized in vitro. micro-CT scanning, Safranin O-Fast Green, toluidine blue and immunohistochemistry staining were performed in vivo. RESULTS: In vitro, avicularin attenuates the degradation of ECM and inflammation, which could inhibit the activation of TRAF6/MAPK pathway via targeting TRAF6. Increased MMP3 and MMP13 expressions and decreased Aggrecan and Collagen Ⅱ levels were observed in anterior cruciate ligament transection (ACLT) induced osteoarthritic rats. Interestingly, intra-articular injection of avicularin attenuates this phenomenon. CONCLUSIONS: Taken together, our results indicate that avicularin suppresses cartilage extracellular matrix degradation and inflammation via TRAF6/MAPK activation by targeting TRAF6. These observations identify TRAF6 as a relevant drug target, and avicularin may as a potential therapeutic agent in osteoarthritis.

α-Cyperone (CYP) down-regulates NF-κB and MAPKs signaling, attenuating inflammation and extracellular matrix degradation in chondrocytes, to ameliorate osteoarthritis in mice.

Inflammation and extracellular matrix (ECM) degradation have been implicated in the pathological process of osteoarthritis (OA). α-Cyperone is the main active component of the traditional Chinese medicine Cyperus rotundus L. In this study, we found that α-Cyperone abolished the IL-1ß-induced production of inflammatory cytokines in isolated rat chondrocytes, such as cyclooxygenase-2 (COX-2), tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6) and inducible nitric oxide synthase (iNOS), in a dose-dependent manner (0.75, 1.5 or 3 µM). Also, the results showed that α-Cyperone downregulated the expression of metalloproteinases (MMPs) and thrombospondin motifs 5 (ADAMTS5), and upregulated the expression of type-2 collagen. Mechanistically, molecular docking tests revealed that α-Cyperone stably and effectively binds to p65, p38, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK). α-Cyperone inhibited NF-κB activation by blocking its nuclear transfer, and decreasing the phosphorylation of mitogen-activated protein kinase (MAPKs). In addition, in vivo studies based on a mouse model of arthritis showed that α-Cyperone prevented the development of osteoarthritis. Therefore, α-Cyperone may be a potential anti-OA drug.

Periplocymarin protects against myocardial fibrosis induced by ß-adrenergic activation in mice.

Periplocymarin is an effective component of Periplocae Cortex, which was wildly used as an ingredient in Traditional Chinese Medicine. Our group previously reported that periplocymarin exerted cardiotonic role via promoting calcium influx. However, its exact role in the pathogenesis of myocardial fibrosis has not been elucidated yet. The present study was aimed at determining the potential effect and underlying mechanism of periplocymarin in isoproterenol (ISO)-induced myocardial fibrosis. C57BL/6 mice were subcutaneously injected with ISO (5 mg/kg/day) or saline for 1 week. The early-to-atrial wave ratio (E/A ratio) measured by echocardiography revealed that ISO-induced heart stiffness was remarkably reversed by administration of periplocymarin (5 mg/kg/day). Masson trichrome staining exhibited that treatment of periplocymarin reduced the excessive deposition of extracellular matrix (ECM). Further investigations employing real-time PCR and western blot demonstrated that periplocymarin suppressed the expression of fibrosis related genes (Col1a1, Col3a1, Acta2 and Tgfb1) and proteins (Collagen I, Collagen III, α-SMA and TGF-ß1) induced by ISO. Metabolomics analysis demonstrated that periplocymarin ameliorated the disorders triggered by ISO and many of the differential metabolic substances were involved in amino acid, glucose and lipid metabolism. Further analysis using network pharmacology revealed that three key genes, namely NOS2, NOS3 and Ptgs2, may be the potential targets of periplocymarin and responsible for the disorders. Validation using heart tissues showed that the mRNA expression of NOS3 was decreased while Ptgs2 was increased upon ISO treatment, which were reversed by periplocymarin. Moreover, the expression of COX-2 (Ptgs2 encoded protein) was consistent with the aspect of Ptgs2 mRNA, while eNOS (NOS3 encoded protein) expression was unchanged. In vitro studies exhibited that periplocymarin exerts anti-fibrotic function via regulating at least eNOS and COX-2 in cardiomyocyte. Taken together, periplocymarin protects against myocardial fibrosis induced by ß-adrenergic activation, the potential mechanism was that periplocymarin targeted on, at least eNOS and COX-2, to improve the metabolic processes of cardiomyocyte and thus attenuated the myocardial fibrosis. Our study highlighted that periplocymarin is a potential therapeutic agent for the prevention of myocardial fibrosis.

ECM-regulation of autophagy: The yin and the yang of autophagy during wound healing.

Wound healing is a complex sequence of tissue protection, replacement, and reorganization leading to regenerated tissue. Disruption of any of these steps results in the process being incomplete as an ulcer or over-exuberant as a hypertrophic scar. Over the past decade, it has become evident that the extracellular matrix and associated components orchestrate this process. However, the cellular events that are induced by the extracellular matrix to accomplish wound healing remain to be defined. Herein we propose that matrix-regulated cellular macro-autophagy is key to both the tissue replacement and resolution stages of healing by directing cellular function or apoptosis. Further, disruptions in matrix turnover alter autophagic function leading to chronic wounds or scarring. While the literature that directly investigates autophagy during wound healing is sparse, the emerging picture supports our proposing a model of the centrality of the matrix-autophagy modulation as central to physiologic and pathologic healing.

Differential regulation of extracellular matrix proteins in three recurrent liver metastases of a single patient with colorectal cancer.

Colorectal cancer (CRC) patients suffer from the second highest mortality among all cancer entities. In half of all CRC patients, colorectal cancer liver metastases (CRLM) can be observed. Metastatic colorectal cancer is associated with poor overall survival and limited treatment options. Even after successful surgical resection of the primary tumor, metachronous liver metastases occur in one out of eight cases. The only available curative intended treatment is hepatic resection, but metachronous CRLM frequently recur after approximately 1 year. In this study, we performed a proteome analysis of three recurrent liver metastases of a single CRC patient by mass spectrometry. Despite surgical resection of the primary CRC and adjuvant chemotherapy plus cetuximab treatment, the patient developed three metachronous CRLM which occurred consecutively after 9, 21 and 31 months. We identified a set of 1132 proteins expressed in the three metachronous CRLM, of which 481 were differentially regulated, including 81 proteins that were associated with the extracellular matrix (ECM). 56 ECM associated proteins were identified as upregulated in the third metastasis, 26 (46%) of which were previously described as negative prognostic markers in CRC, including tenascin C, nidogen 1, fibulin 1 and vitronectin. These data may reflect an ascending trend of malignancy from the first to the third metachronous colorectal cancer liver metastasis. Additionally, the results indicate different ECM phenotypes for recurrent metachronous metastasis, associated with different grades of malignancy and highlights the importance of individual analysis of molecular features in different, consecutive metastatic events in a single patient.

Biochanin-A ameliorates pulmonary fibrosis by suppressing the TGF-ß mediated EMT, myofibroblasts differentiation and collagen deposition in in vitro and in vivo systems.

BACKGROUND: Idiopathic Pulmonary Fibrosis (IPF) is a progressive inflammatory disorder driven by a fibrotic cascade of events such as epithelial to mesenchymal transition, extracellular matrix production and collagen formation in the lungs in a sequential manner. IPF incidences were raising rapidly across the world. FDA approved pirfenidone and nintedanib (tyrosine kinase inhibitors) are being used as a first-line treatment drugs for IPF, however, neither the quality of life nor survival rates have been improved because of patient noncompliance due to multiple side effects. Thus, the development of novel therapeutic approaches targeting TGF-ß mediated cascade of fibrotic events is urgently needed to improve the survival of the patients suffering from devastating disease. PURPOSE: The aim of this study was to investigate and validate the anti-fibrotic properties of Biochanin-A (isoflavone) against TGF-ß mediated fibrosis in in vitro, ex vivo, in vivo models and to determine the molecular mechanisms that mediate these anti-fibrotic effects. METHODS: The therapeutic activity of BCA was determined in in vitro/ex vivo models. Cells were pre-treated with BCA and incubated in presence or absence of recombinant-TGF-ß to stimulate the fibrotic cascade of events. Pulmonary fibrosis was developed by intratracheal administration of bleomycin in rats. BCA treatment was given for 14 days from post bleomycin instillation and then various investigations (collagen content, fibrosis gene/protein expression and histopathological changes) were performed to assess the anti-fibrotic activity of BCA. RESULTS: In vitro/ex vivo (Primary normal, IPF cell line and primary IPF cells/ Precision cut mouse lung slices) experiments revealed that, BCA treatment significantly (p < 0.001) reduced the expression of TGF-ß modulated fibrotic genes/protein expressions (including their functions) which are involved in the cascade of fibrotic events. BCA treatment significantly (p < 0.01) reduced the bleomycin-induced inflammatory cell-infiltration, inflammatory markers expression, collagen deposition and expression of fibrotic markers in lung tissues equivalent or better than pirfenidone treatment. In addition, BCA treatment significantly (p < 0.001) attenuated the TGF-ß1/BLM-mediated increase of TGF-ß/Smad2/3 phosphorylation and resulted in the reduction of pathological abnormalities in lung tissues determined by histopathology observations. CONCLUSION: Collectively, BCA treatment demonstrated the remarkable therapeutic effects on TGF-ß/BLM mediated pulmonary fibrosis using IPF cells and rodent models. This current study may offer a novel treatment approach to halt and may be even rescue the devastating lung scarring of IPF.

The Inter-α-Trypsin Inhibitor Family: Versatile Molecules in Biology and Pathology.

Inter-α-trypsin inhibitor (IαI) family members are ancient and unique molecules that have evolved over several hundred million years of vertebrate evolution. IαI is a complex containing the proteoglycan bikunin to which heavy chain proteins are covalently attached to the chondroitin sulfate chain. Besides its matrix protective activity through protease inhibitory action, IαI family members interact with extracellular matrix molecules and most notably hyaluronan, inhibit complement, and provide cell regulatory functions. Recent evidence for the diverse roles of the IαI family in both biology and pathology is reviewed and gives insight into their pivotal roles in tissue homeostasis. In addition, the clinical uses of these molecules are explored, such as in the treatment of inflammatory conditions including sepsis and Kawasaki disease, which has recently been associated with severe acute respiratory syndrome coronavirus 2 infection in children.

Poricoic acid A activates AMPK to attenuate fibroblast activation and abnormal extracellular matrix remodelling in renal fibrosis.

BACKGROUND: In chronic kidney disease, although fibrosis prevention is beneficial, few interventions are available that specifically target fibrogenesis. Poricoic acid A (PAA) isolated from Poria cocos exhibits anti-fibrotic effects in the kidney, however the underlying mechanisms remain obscure. PURPOSE: We isolated PAA and investigated its effects and the underlying mechanisms in renal fibrosis. STUDY DESIGN: Unilateral ureteral obstruction (UUO) and 5/6 nephrectomy (Nx) animal models and TGF-ß1-induced renal fibroblasts (NRK-49F) were used to investigate the anti-fibrotic activity of PAA and its underlying mechanisms. METHODS: Western blots, qRT-PCR, immunofluorescence staining, co-immunoprecipitation and molecular docking methods were used. Knock-down and knock-in of adenosine monophosphate-activated protein kinase (AMPK) in the UUO model and cultured NRK-49F cells were employed to verify the mechanisms of action of PAA. RESULTS: PAA improved renal function and alleviated fibrosis by stimulating AMPK and inhibiting Smad3 specifically in Nx and UUO models. Reduced AMPK activity was associated with Smad3 induction, fibroblast activation, and the accumulation and aberrant remodelling of extracellular matrix (ECM) in human renal puncture samples and cultured NRK-49F cells. PAA stimulated AMPK activity and decreased fibrosis in a dose-dependent manner, thus showing that AMPK was essential for PAA to exert its anti-fibrotic effects. AMPK deficiency reduced the anti-fibrotic effects of PAA, while AMPK overexpression enhanced its effect. CONCLUSION: PAA activated AMPK and further inhibited Smad3 specifically to suppress fibrosis by preventing aberrant ECM accumulation and remodelling and facilitating the deactivation of fibroblasts.

Reduction of osteoarthritis severity in the temporomandibular joint of rabbits treated with chondroitin sulfate and glucosamine.

Osteoarthritis is a degenerative disease that causes substantial changes in joint tissues, such as cartilage degeneration and subchondral bone sclerosis. Chondroitin sulfate and glucosamine are commonly used products for the symptomatic treatment of osteoarthritis. The aim of the present study was to investigate the effects of these products when used as structure-modifying drugs on the progression of osteoarthritis in the rabbit temporomandibular joint. Thirty-six New Zealand rabbits were divided into 3 groups (n = 12/group): control (no disease); osteoarthritis (disease induction); and treatment (disease induction and administration of chondroitin sulfate and glucosamine). Osteoarthritis was induced by intra-articular injection of monosodium iodoacetate. Animals were killed at 30 and 90 days after initiation of therapy. The treatment was effective in reducing disease severity, with late effects and changes in the concentration of glycosaminoglycans in the articular disc. The results indicate that chondroitin sulfate and glucosamine may have a structure-modifying effect on the tissues of rabbit temporomandibular joints altered by osteoarthritis.

Andrographolide ameliorates bleomycin-induced pulmonary fibrosis by suppressing cell proliferation and myofibroblast differentiation of fibroblasts via the TGF-ß1-mediated Smad-dependent and -independent pathways.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with no effective medication. Andrographolide (Andro), extracted from Chinese herbal Andrographis paniculata, could attenuate bleomycin (BLM)-induced pulmonary fibrosis via inhibition of inflammation and oxidative stress, however, the anti-fibrotic mechanisms have not been clarified. Myofibroblasts are the primary cell types responsible for the accumulation of extracellular matrix (ECM) in fibrotic diseases, and targeting fibroblast proliferation and differentiation is an important therapeutic strategy for the treatment of IPF. Hence, this study aimed to investigate the effects of Andro on the fibroblast proliferation and differentiation in the in vivo and in vitro models. The results showed that Andro improved pulmonary function and inhibited BLM-induced fibroblast proliferation and differentiation and ECM deposition in the lungs. In vitro, Andro inhibited proliferation and induced apoptosis of TGF-ß1-stimulated NIH 3T3 fibroblasts and primary lung fibroblasts (PLFs). Andro also inhibited TGF-ß1-induced myofibroblast differentiation and ECM deposition in both cells. We also found that Andro suppressed TGF-ß1-induced Smad2/3 and Erk1/2 activation, suggesting that Smad2/3 and Erk1/2 inactivation mediates Andro-induced effects on TGF-ß1-induced fibroblast proliferation and differentiation. These results indicated that Andro has novel and potent anti-fibrotic effects in lung fibroblasts via inhibition of the proliferation and myofibroblast differentiation of fibroblasts and subsequent ECM deposition, which are modulated by TGF-ß1-mediated Smad-dependent and -independent pathways.

Methyl helicterate inhibits hepatic stellate cell activation through downregulating the ERK1/2 signaling pathway.

The present study was to investigate the inhibitory effect of methyl helicterate (MH) on hepatic stellate cells (HSC-T6), primarily elucidating the underlying mechanism of MH against liver fibrosis. HSC-T6 cells were activated by platelet-derived growth factor (PDGF) stimulation, and then the effects of MH on cell viability, cytomembrane integrity, colony, migration, apoptosis, and cell cycle were detected. Moreover, the regulative mechanism of MH on HSCs was investigated by detecting the activation of the extracellular signal-regulated kinase (ERK1/2) signaling pathway. The results showed that MH significantly inhibited HSC-T6 cell viability and proliferation in a concentration-dependent manner. It notably promoted the release of lactate dehydrogenase, destroying cell membrane integrity. MH also markedly inhibited HSC-T6 cell clonogenicity and migration. Moreover, MH treatment significantly induced cell apoptosis and arrested cell cycle at the G2 phase. The further study showed that MH inhibited the expression of ERK1, ERK2, c-fos, c-myc, and Ets-1, blocking the ERK1/2 pathway. In conclusion, this study demonstrates that MH significantly inhibits HSC activation and promotes cell apoptosis via downregulation of the ERK1/2 signaling pathway.

Chronic kidney disease induced by an adenine rich diet upregulates integrin linked kinase (ILK) and its depletion prevents the disease progression.

Kidney fibrosis is one of the main pathological findings of progressive chronic kidney disease (CKD) although the pathogenesis of renal scar formation remains incompletely explained. Integrin-linked kinase (ILK), a major scaffold protein between the extracellular matrix (ECM) and intracellular signaling pathways, is involved in several pathophysiological processes during renal damage. However, ILK contribution in the CKD progress remains to be fully elucidated. In the present work, we studied 1) the renal functional and structural consequences of CKD genesis and progression when ILK is depleted and 2) the potential of ILK depletion as a therapeutic approach to delay CKD progression. We induced an experimental CKD model, based on an adenine-supplemented diet on adult wild-type (WT) and ILK-depleted mice, with a tubulointerstitial damage profile resembling that is observed in human CKD. The adenine diet induced in WT mice a progressive increase in plasma creatinine and urea concentrations. In the renal cortex it was also observed tubular damage, interstitial fibrosis and progressive increased ECM components, pro-inflammatory and chemo-attractant cytokines, EMT markers and TGF-ß1 expressions. These observations were highly correlated to a simultaneous increase of ILK expression and activity. In adenine-fed transgenic ILK-depleted mice, all these changes were prevented. Additionally, we evaluated the potential role of ILK depletion to be applied after the disease induction, as an effective approach to interventions in human CKD subjects. In this scenario, two weeks after the establishment of adenine-induced CKD, ILK was abrogated in WT mice and stabilized renal damage, avoiding CKD progression. We propose ILK to be a potential target to delay renal disease progression.

Aucubin inhibits IL-1ß- or TNF-α-induced extracellular matrix degradation in nucleus pulposus cell through blocking the miR-140-5p/CREB1 axis.

In intervertebral disc degeneration (IDD), increased proinflammatory molecules secreted by human nucleus pulposus cells (HNPCs) could promote the expression of extracellular matrix (ECM)-degrading enzymes. IDD could be affected by both genetic and environmental factors, including microRNAs (miRNAs). Aucubin, the active ingredient of a traditional Chinese medicine herb Du Zhong, has been reported to promote osteogenic differentiation; however, the role of aucubin in IDD and the underlying mechanism remain unclear. Herein, we evaluated the effect of aucubin on TNF-α- or IL-1ß-induced ECM degradation in HNPCs. By using online tools, miR-140 was selected as a candidate miRNA that is related to TNF-α or IL-1ß signaling. Overexpression of miR-140 enhanced the effect of aucubin on ECM degradation. Moreover, cAMP responsive element binding protein 1 (CREB1), a major transcriptional factor in immune-related signaling, was a direct downstream target of miR-140. CREB1 knockdown mimicked the function of miR-140 overexpression on ECM degradation. In summary, aucubin might ameliorate IL-1ß- or TNF-α-induced ECM degradation in HNPCs through regulating miR-140/CREB1.

Dual Role of Magnetic Nanoparticles as Intracellular Hotspots and Extracellular Matrix Disruptors Triggered by Magnetic Hyperthermia in 3D Cell Culture Models.

Magnetic hyperthermia is a promising therapy for the localized treatment of cancer based on the exposure of magnetic nanoparticles to an external alternating magnetic field. In order to evaluate some of the mechanisms involved in the cellular damage caused by this treatment, two different 3D cell culture models were prepared using collagen, which is the most abundant protein of the extracellular matrix. The same amount of nanoparticles was added to cells either before or after their incorporation into the 3D structure. Therefore, in one model, particles were located only inside cells (In model), while the other one had particles both inside and outside cells (In&Out model). In the In&Out model, the hyperthermia treatment facilitated the migration of the particles from the outer areas of the 3D structure to the inner parts, achieving a faster homogeneous distribution throughout the whole structure and allowing the particles to gain access to the inner cells. The cell death mechanism activated by the magnetic hyperthermia treatment was different in both models. Necrosis was observed in the In model and apoptosis in the In&Out model 24 h after the hyperthermia application. This was clearly correlated with the amount of nanoparticles located inside the cells. Thus, the combination of both 3D models allowed us to demonstrate two different roles of the magnetic particles during the hyperthermia treatment: (i) The modulation of the cell death mechanism depending on the amount of intracellular particles and (ii) the disruption of the collagen matrix caused by the extracellular nanoparticles.

Is skin microneedling a good alternative method of various skin defects removal.

Skin microneedling accelerates the process of skin regeneration through the creation of numerous microinjuries which emerge when skin is deeply punctured with very thin needles. The whole procedure evokes various reactions which can be divided into three major phases: inflammation, proliferation, and remodeling. It activates platelet growth factors which are responsible for the stimulation of fibroblasts to produce collagen and elastin. Moreover, skin breakdown enhances penetration of active ingredients. Treatment can be performed with the use of different devices, all equipped with needles of various lengths. Due to the fact that skin microneedling stimulates the synthesis of significant rebuilding and structural skin elements (collagen, elastin, proteoglycan), it is used in the treatment of many skin defects of different etiologies (e.g., photoaging, wrinkles, loss of elasticity, hypo- or hypertrophic scars, pigmentation changes, infraorbital dark circles, teleangiectasia, stretch marks, cellulite, alopecia, and vitiligo). In order to accelerate postsurgical regeneration and/or to enhance effects, microneedling is combined with the application of UV light (photodynamic therapy with ALA), LED light, platelet-rich plasma, chemical peels, stem cells, retinoids and other pharmaceuticals, and vitamins. High effectiveness, limited number of side effects, and short recovery time, make skin microneedling a popular cosmetic, and medical treatment.

A novel FGFR1-binding peptide attenuates the degeneration of articular cartilage in adult mice.

OBJECTIVE: We previously reported that genetic ablation of (Fibroblast Growth Factors Receptors) FGFR1 in knee cartilage attenuates the degeneration of articular cartilage in adult mice, which suggests that FGFR1 is a potential targeting molecule for osteoarthritis (OA). Here, we identified R1-P1, an inhibitory peptide for FGFR1 and investigated its effect on the pathogenesis of OA in mice induced by destabilization of medial meniscus (DMM). DESIGN: Binding ability between R1-P1 and FGFR1 protein was evaluated by enzyme-linked immuno sorbent assay (ELISA) and molecular docking. Alterations in cartilage were evaluated histologically. The expression levels of molecules associated with articular cartilage homeostasis and FGFR1 signaling were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting and immunohistochemistry (IHC). The chondrocyte apoptosis was detected by terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) assay. RESULTS: R1-P1 had highly binding affinities to human FGFR1 protein, and efficiently inhibited extracellular signal-regulated kinase (ERK)1/2 pathway in mouse primary chondrocytes. In addition, R1-P1 attenuated the IL-1ß induced significant loss of proteoglycan in full-thickness cartilage tissue from human femur head. Moreover, this peptide can significantly restore the IL-1ß mediated loss of proteoglycan and type II collagen (Col II) and attenuate the expression of matrix metalloproteinase-13 (MMP13) in mouse primary chondrocytes. Finally, intra-articular injection of R1-P1 remarkably attenuated the loss of proteoglycan and the destruction of articular cartilage and decreased the expressions of extracellular matrix (ECM) degrading enzymes and apoptosis in articular chondrocytes of mice underwent DMM surgery. CONCLUSIONS: R1-P1, a novel inhibitory peptide for FGFR1, attenuates the degeneration of articular cartilage in adult mice, which is a potential leading molecule for the treatment of OA.

Yu Gan Long reduces rat liver fibrosis by blocking TGF-ß1/Smad pathway and modulating the immunity.

Yu Gan Long (YGL) is a Chinese traditional herbal medicine that has been used in the treatment of liver fibrosis for many years in clinical practice. However, its anti-hepatofibrotic mechanism has not been studied yet. In this study, the effect and mechanism of YGL in reducing liver fibrosis was demonstrated in vivo. Our results showed that liver fibrosis biomarkers collagen IV (Col IV), type III precollagen (PCIII), hyaluronuc acid (HA) and laminin (LN), were increased after CCl4 treatment and decreased by YGL. Among the liver fibrosis indicators, α-smooth muscle actin (α-SMA) was decreased by YGL in the CCl4-treated rats, while MMP2 and MMP9 was upregulated followed by TIMP1 downregulation. Proteins involved in liver fibrosis such as p-Smad2, p-Smad3 and Smad4 were down-regulated, while Smad7 protein was up-regulated by YGL after CCl4-induced liver damage. YGL also suppressed the increase of TGF-ß1, TNF-α, IL-1ß, IL-6, IL-4 and IL-17 A induced by CCl4 treatment, while promoted IFN-γ expression. Finally, the transcription factors ROR-γt and GATA3 were decreased, while T-bet was increased after YGL treatment. These results suggested that YGL attenuated CCl4-induced hepatic fibrosis by accelerating the extracellular matrix degradation, blocking the TGF-ß1/Smad signaling pathway and modulating the balance among IL-4, IL-17 A and IFN-γ, demonstrating YGL protective effect and its potential mechanisms in treating liver fibrosis.