Extracellular Matrix Production Research Articles

Mesenchymal Cells Retain the Specificity of Embryonal Origin During Osteogenic Differentiation.

Mesenchymal stem cells (MSCs) are widely used in therapy, but the differences between MSCs of various origins and their ability to undergo osteogenic differentiation and produce extracellular matrix are not fully understood. To address this, we conducted a comparative analysis of mesenchymal cell primary cultures from 6 human sources, including osteoblast-like cells from the adult femur, adipose-derived stem cells, Wharton's jelly-derived mesenchymal cells, gingival fibroblasts, dental pulp stem cells, and periodontal ligament stem cells. We analyzed these cells' secretome, proteome, and transcriptome under standard and osteogenic cultivation conditions. Despite the overall similarity in osteogenic differentiation, the cells maintain their embryonic specificity after isolation and differentiation in vitro. Furthermore, we propose classifying mesenchymal cells into 3 groups: dental stem cells of neural crest origin, mesenchymal stem cells, and fetal stem cells. Specifically, fetal stem cells have the most promising secretome for various applications, while mesenchymal stem cells have a specialized secretome optimal for extracellular matrix production. Nevertheless, mesenchymal cells from all sources secreted core bone extracellular matrix-associated proteins. In conclusion, our study illuminates the distinctive characteristics of mesenchymal stem cells from various sources, providing insights into their potential applications in regenerative medicine and enhancing our understanding of the inherent diversity of mesenchymal cells in vivo.

Nε-Carboxymethyl-Lysine Modification of Extracellular Matrix Proteins Augments Fibroblast Activation.

The extracellular matrix (ECM) is a dynamic complex protein network that provides structural integrity and plays an active role in shaping fibroblast behavior both in health and disease. Despite its essential functions, the impact of age-associated post-translational modifications on ECM-driven fibroblast activities such as proliferation, survival, fibroblast-to-myofibroblast transformation (FMT), and extracellular matrix production remains largely unknown. Nε-carboxymethyl-lysine (CML) is one of the well-characterized advanced glycation end-products (AGEs) that can occur on lysine residues within ECM proteins through non-enzymatic glycation. In this study, we determined the accumulation and the effects of the CML-modified ECM (CML-ECM) on fibroblast activation. Immunostainings and immunoblot analysis demonstrated significant increases in CML-AGE content in idiopathic pulmonary fibrosis (IPF) compared to age-matched healthy lungs. Gene expression analysis and fibroblast activation assays collectively implicate the ECM as a negative regulator of fibroblast activation. Notably, the CML modification of the ECM resulted in a significant decrease in its anti-fibrotic effects including proliferation, FMT, apoptosis, and ECM production. Together, the results of this study revealed an unexplored pathological role played by the CML-ECM on fibroblast activation, which has wide implications in IPF and other fibrotic diseases.

Hedgehog Activation for Enhanced Rotator Cuff Tendon-to-Bone Healing

Background: Rotator cuff repair is a common orthopaedic procedure, yet the rate of failure to heal after surgery is high. Repair site rupture is due to poor tendon-to-bone healing and lack of regeneration of the native fibrocartilaginous enthesis. During development, the enthesis is formed and mineralized by a pool of progenitors activated by hedgehog signaling. Furthermore, hedgehog signaling drives regenerative enthesis healing in young animals, in contrast to older animals, in which enthesis injuries heal via fibrovascular scar and without participation of hedgehog signaling. Hypothesis: Hedgehog activation improves tendon-to-bone healing in an animal model of rotator cuff repair. Study Design: Controlled laboratory study. Methods: A total of 78 adult Sprague-Dawley rats were used. Supraspinatus tendon injury and repair were completed bilaterally, with microsphere-encapsulated hedgehog agonist administered to right shoulders and control microspheres administered to left shoulders. Animals were sacrificed after 3, 14, 28, or 56 days. Gene expression and histological, biomechanical, and bone morphometric analyses were conducted. Results: At 3 days, hedgehog signaling pathway genes Gli1 (1.70; P = .029) and Smo (2.06; P = .0173), as well as Runx2 (1.69; P = .0386), a transcription factor of osteogenesis, were upregulated in treated relative to control repairs. At 14 days, transcription factors of tenogenesis, Scx (4.00; P = .041), and chondrogenesis, Sox9 (2.95; P = .010), and mineralized fibrocartilage genes Col2 (3.18; P = .031) and Colx (1.85; P = .006), were upregulated in treated relative to control repairs. Treatment promoted fibrocartilage formation at the healing interface by 28 days, with improvements in tendon-bone maturity, organization, and continuity. Treatment led to improved biomechanical properties. The material property strength (2.43 vs 1.89 N/m2; P = .046) and the structural property work to failure (29.01 vs 18.09 mJ; P = .030) were increased in treated relative to control repairs at 28 days and 56 days, respectively. Treatment had a marginal effect on bone morphometry underlying the repair. Trabecular thickness (0.08 vs 0.07 mm; P = .035) was increased at 28 days. Conclusion: Hedgehog agonist treatment activated hedgehog signaling at the tendon-to-bone repair site and prompted increased mineralized fibrocartilage production. This extracellular matrix production and mineralization resulted in improved biomechanical properties, demonstrating the therapeutic potential of hedgehog agonism for improving tendon-to-bone healing after rotator cuff repair. Clinical Relevance: This study demonstrates the therapeutic potential of hedgehog agonist treatment for improving tendon-to-bone healing after rotator cuff injury and repair.

Human Plaque Myofibroblasts to Study Mechanisms of Atherosclerosis.

Background Plaque myofibroblasts are critical players in the initiation and advancement of atherosclerotic disease. They are involved in the production of extracellular matrix, the formation of the fibrous cap, and the underlying lipidic core via modulation processes in response to different environmental cues. Despite clear phenotypic differences between myofibroblast cells and healthy vascular smooth muscle cells, smooth muscle cells are still widely used as a cellular model in atherosclerotic research. Methods and Results Here, we present a conditioned outgrowth method to isolate and culture myofibroblast cells from plaques. We obtained these cells from 27 donors (24 carotid and 3 femoral endarterectomies). We show that they keep their proliferative capacity for 8 passages, are transcriptionally stable, retain donor-specific gene expression programs, and express extracellular matrix proteins (FN1, COL1A1, and DCN) and smooth muscle cell markers (ACTA2, MYH11, and CNN1). Single-cell transcriptomics reveals that the cells in culture closely resemble the plaque myofibroblasts. Chromatin immunoprecipitation sequencing shows the presence of histone H3 lysine 4 dimethylation at the MYH11 promoter, pointing to their smooth muscle cell origin. Finally, we demonstrated that plaque myofibroblasts can be efficiently transduced (>97%) and are capable of taking up oxidized low-density lipoprotein and undergoing calcification. Conclusions In conclusion, we present a method to isolate and culture cells that retain plaque myofibroblast phenotypical and functional capabilities, making them a suitable in vitro model for studying selected mechanisms of atherosclerosis.

The role of cGAS-STING signaling in pulmonary fibrosis and its therapeutic potential.

Pulmonary fibrosis is a progressive and ultimately fatal lung disease, exhibiting the excessive production of extracellular matrix and aberrant activation of fibroblast. While Pirfenidone and Nintedanib are FDA-approved drugs that can slow down the progression of pulmonary fibrosis, they are unable to reverse the disease. Therefore, there is an urgent demand to develop more efficient therapeutic approaches for pulmonary fibrosis. The intracellular DNA sensor called cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) synthase (cGAS) plays a crucial role in detecting DNA and generating cGAMP, a second messenger. Subsequently, cGAMP triggers the activation of stimulator of interferon genes (STING), initiating a signaling cascade that leads to the stimulation of type I interferons and other signaling molecules involved in immune responses. Recent studies have highlighted the involvement of aberrant activation of cGAS-STING contributes to fibrotic lung diseases. This review aims to provide a comprehensive summary of the current knowledge regarding the role of cGAS-STING pathway in pulmonary fibrosis. Moreover, we discuss the potential therapeutic implications of targeting the cGAS-STING pathway, including the utilization of inhibitors of cGAS and STING.

Role of CD44 expressed in renal tubules during maladaptive repair in renal fibrogenesis in an allopurinol-induced rat model of chronic kidney disease.

The kidney is a major target organ for the adverse effects of pharmaceuticals; renal tubular epithelial cells (TECs) are particularly vulnerable to drug-induced toxicity. TECs have regenerative capacity; however, maladaptive repair of TECs after injury leads to renal fibrosis, resulting in chronic kidney disease (CKD). We previously reported the specific expression of CD44 in failed-repair TECs of rat CKD model induced by ischemia reperfusion injury. Here, we investigated the pathophysiological role of CD44 in renal fibrogenesis in allopurinol-treated rat CKD model. Dilated or atrophic TECs expressing CD44 in fibrotic areas were collected by laser microdissection and subjected to microarray analysis. Gene ontology showed that extracellular matrix (ECM)-related genes were upregulated and differentiation-related genes were downregulated in dilated/atrophic TECs. Ingenuity Pathway Analysis identified CD44 as an upstream regulator of fibrosis-related genes, including Fn1, which encodes fibronectin. Immunohistochemistry demonstrated that dilated/atrophic TECs expressing CD44 showed decreases in differentiation markers of TECs and clear expression of mesenchymal markers during basement membrane attachment. In situ hybridization revealed an increase in Fn1 mRNA in the cytoplasm of dilated/atrophic TECs, whereas fibronectin was localized in the stroma around these TECs, supporting the production/secretion of ECM by dilated/atrophic TECs. Overall, these data indicated that dilated/atrophic TECs underwent a partial epithelial-mesenchymal transition (pEMT) and that CD44 promoted renal fibrogenesis via induction of ECM production in failed-repair TECs exhibiting pEMT. CD44 was detected in the urine and serum of APL-treated rats, which may reflect the expression of CD44 in the kidney.

Survivin regulates intracellular stiffness and extracellular matrix production in vascular smooth muscle cells.

Vascular dysfunction is a common cause of cardiovascular diseases characterized by the narrowing and stiffening of arteries, such as atherosclerosis, restenosis, and hypertension. Arterial narrowing results from the aberrant proliferation of vascular smooth muscle cells (VSMCs) and their increased synthesis and deposition of extracellular matrix (ECM) proteins. These, in turn, are modulated by arterial stiffness, but the mechanism for this is not fully understood. We found that survivin is an important regulator of stiffness-mediated ECM synthesis and intracellular stiffness in VSMCs. Whole-transcriptome analysis and cell culture experiments showed that survivin expression is upregulated in injured femoral arteries in mice and in human VSMCs cultured on stiff fibronectin-coated hydrogels. Suppressed expression of survivin in human VSMCs significantly decreased the stiffness-mediated expression of ECM components related to arterial stiffening, such as collagen-I, fibronectin, and lysyl oxidase. By contrast, expression of these ECM proteins was rescued by ectopic expression of survivin in human VSMCs cultured on soft hydrogels. Interestingly, atomic force microscopy analysis showed that suppressed or ectopic expression of survivin decreases or increases intracellular stiffness, respectively. Furthermore, we observed that inhibiting Rac and Rho reduces survivin expression, elucidating a mechanical pathway connecting intracellular tension, mediated by Rac and Rho, to survivin induction. Finally, we found that survivin inhibition decreases FAK phosphorylation, indicating that survivin-dependent intracellular tension feeds back to maintain signaling through FAK. These findings suggest a novel mechanism by which survivin potentially modulates arterial stiffness.

Cartilage-inspired self-assembly glycopeptide hydrogels for cartilage regeneration via ROS scavenging

Cartilage injury represents a frequent dilemma in clinical practice owing to its inherently limited self-renewal capacity. Biomimetic strategy-based engineered biomaterial, capable of coordinated regulation for cellular and microenvironmental crosstalk, provides an adequate avenue to boost cartilage regeneration. The level of oxidative stress in microenvironments is verified to be vital for tissue regeneration, yet it is often overlooked in engineered biomaterials for cartilage regeneration. Herein, inspired by natural cartilage architecture, a fibril-network glycopeptide hydrogel (Nap-FFGRGD@FU), composed of marine-derived polysaccharide fucoidan (FU) and naphthalenephenylalanine-phenylalanine-glycine-arginine-glycine-aspartic peptide (Nap-FFGRGD), was presented through a simple supramolecular self-assembly approach. The Nap-FFGRGD@FU hydrogels exhibit a native cartilage-like architecture, characterized by interwoven collagen fibers and attached proteoglycans. Beyond structural simulation, fucoidan-exerted robust biological effects and Arg-Gly-Asp (RGD) sequence-provided cell attachment sites realized functional reinforcement, synergistically promoted extracellular matrix (ECM) production and reactive oxygen species (ROS) elimination, thus contributing to chondrocytes-ECM harmony. In vitro co-culture with glycopeptide hydrogels not only facilitated cartilage ECM anabolic metabolism but also scavenged ROS accumulation in chondrocytes. Mechanistically, the chondro-protective effects induced by glycopeptide hydrogels rely on the activation of endogenous antioxidant pathways associated with nuclear factor erythroid 2-related factor 2 (NRF2). In vivo implantation of glycopeptide hydrogels successfully improved the de novo cartilage generation by 1.65-fold, concomitant with coordinately restructured subchondral bone structure. Collectively, our ingeniously crafted bionic glycopeptide hydrogels simultaneously rewired chondrocytes’ function by augmenting anabolic metabolism and rebuilt ECM microenvironment via preserving redox equilibrium, holding great potential for cartilage tissue engineering.

Morphogenesis and development of human telencephalic organoids in the absence and presence of exogenous extracellular matrix.

The establishment and maintenance of apical-basal polarity is a fundamental step in brain development, instructing the organization of neural progenitor cells (NPCs) and the developing cerebral cortex. Particularly, basally located extracellular matrix (ECM) is crucial for this process. In vitro, epithelial polarization can be achieved via endogenous ECM production, or exogenous ECM supplementation. While neuroepithelial development is recapitulated in neural organoids, the effects of different ECM sources in tissue morphogenesis remain underexplored. Here, we show that exposure to a solubilized basement membrane matrix substrate, Matrigel, at early neuroepithelial stages causes rapid tissue polarization and rearrangement of neuroepithelial architecture. In cultures exposed to pure ECM components or unexposed to any exogenous ECM, polarity acquisition is slower and driven by endogenous ECM production. After the onset of neurogenesis, tissue architecture and neuronal differentiation are largely independent of the initial ECM source, but Matrigel exposure has long-lasting effects on tissue patterning. These results advance the knowledge on mechanisms of exogenously and endogenously guided morphogenesis, demonstrating the self-sustainability of neuroepithelial cultures by endogenous processes.

Engineering Stem Cell Fate Controlling Biomaterials to Develop Muscle Connective Tissue Layered Myofibers.

Skeletal muscle connective tissue (MCT) surrounds myofiber bundles to provide structural support, produce force transduction from tendons, and regulate satellite cell differentiation during muscle regeneration. Engineered muscle tissue composed of myofibers layered within MCT has not yet been developed. Herein, a bioengineering strategy to create MCT-layered myofibers through the development of stem cell fate-controlling biomaterials that achieve both myogenesis and fibroblast differentiation in a locally controlled manner at the single construct is introduced. The reciprocal role of transforming growth factor-beta 1 (TGF-β1) and its inhibitor as well as 3D matrix stiffness to achieve co-differentiation of MCT fibroblasts and myofibers from a human-induced pluripotent stem cell (hiPSC)-derived paraxial mesoderm is studied. To avoid myogenic inhibition, TGF-β1 is conjugated on the gelatin-based hydrogel to control the fibroblasts' populations locally; the TGF-β1 degrades after 2 weeks, resulting in increased MCT-specific extracellular matrix (ECM) production. The locations of myofibers and fibroblasts are precisely controlled by using photolithography and co-axial wet spinning techniques, which results in the formation of MCT-layered functional myofibers in 3D constructs. This advanced engineering strategy is envisioned as a possible method for obtaining biomimetic human muscle grafts for various biomedical applications.

Estrogen receptors differentially modifies lamellipodial and focal adhesion dynamics in airway smooth muscle cell migration

Sex-steroid signaling, especially estrogen, has a paradoxical impact on regulating airway remodeling. In our previous studies, we demonstrated differential effects of 17β-estradiol (E2) towards estrogen receptors (ERs: α and β) in regulating airway smooth muscle (ASM) cell proliferation and extracellular matrix (ECM) production. However, the role of ERs and their signaling on ASM migration is still unexplored. In this study, we examined how ERα versus ERβ affects the mitogen (Platelet-derived growth factor, PDGF)-induced human ASM cell migration as well as the underlying mechanisms involved. We used Lionheart-FX automated microscopy and transwell assays to measure cell migration and found that activating specific ERs had differential effects on PDGF-induced ASM cell migration. Pharmacological activation of ERβ or shRNA mediated knockdown of ERα and specific activation of ERβ blunted PDGF-induced cell migration. Furthermore, specific ERβ activation showed inhibition of actin polymerization by reducing the F/G-actin ratio. Using Zeiss confocal microscopy coupled with three-dimensional algorithmic ZEN-image analysis showed an ERβ-mediated reduction in PDGF-induced expressions of neural Wiskott-Aldrich syndrome protein (N-WASP) and actin-related proteins-2/3 (Arp2/3) complex, thereby inhibiting actin-branching and lamellipodia. In addition, ERβ activation also reduces the clustering of actin-binding proteins (vinculin and paxillin) at the leading edge of ASM cells. However, cells treated with E2 or ERα agonists do not show significant changes in actin/lamellipodial dynamics. Overall, these findings unveil the significance of ERβ activation in regulating lamellipodial and focal adhesion dynamics to regulate ASM cell migration and could be a novel target to blunt airway remodeling.

An overview of the production of tissue extracellular matrix and decellularization process.

Thousands of patients need an organ transplant yearly, while only a tiny percentage have this chance to receive a tissue/organ transplant. Nowadays, decellularized animal tissue is one of the most widely used methods to produce engineered scaffolds for transplantation. Decellularization is defined as physically or chemically removing cellular components from tissues while retaining structural and functional extracellular matrix (ECM) components and creating an ECM-derived scaffold. Then, decellularized scaffolds could be reseeded with different cells to fabricate an autologous graft. Effective decellularization methods preserve ECM structure and bioactivity through the application of the agents and techniques used throughout the process. The most valuable agents for the decellularization process depend on biological properties, cellular density, and the thickness of the desired tissue. ECM-derived scaffolds from various mammalian tissues have been recently used in research and preclinical applications in tissue engineering. Many studies have shown that decellularized ECM-derived scaffolds could be obtained from tissues and organs such as the liver, cartilage, bone, kidney, lung, and skin. This review addresses the significance of ECM in organisms and various decellularization agents utilized to prepare the ECM. Also, we describe the current knowledge of the decellularization of different tissues and their applications.

Blockade of TGF-β signalling alleviates human adipose stem cell senescence induced by native ECM in obesity visceral white adipose tissue

BackgroundAbdominal obesity is appreciated as a major player in insulin resistance and metabolically dysfunctional adipose tissue. Inappropriate extracellular matrix (ECM) remodelling and functional alterations in human adipose stromal/stem cells (hASCs) have been linked with visceral white adipose tissue (vWAT) dysfunction in obesity. Understanding the interactions between hASCs and the native ECM environment in obese vWAT is required for the development of future therapeutic approaches for obesity-associated metabolic complications.MethodsThe phenotypes and transcriptome properties of hASCs from the vWAT of obese patients and lean donors were assessed. The hASC-derived matrix from vWAT of obese or lean patients was generated in vitro using a decellularized method. The topography and the major components of the hASC-derived matrix were determined. The effects of the obese hASC-derived matrix on cell senescence and mitochondrial function were further determined.ResultsWe showed that hASCs derived from the vWAT of obese patients exhibited senescence and were accompanied by the increased production of ECM. The matrix secreted by obese hASCs formed a fibrillar suprastructure with an abundance of fibronectin, type I collagen, and transforming growth factor beta 1 (TGF-β1), which resembles the native matrix microenvironment of hASCs in vWAT derived from obese patients. Furthermore, the obese hASC-derived matrix promoted lean hASC ageing and induced mitochondrial dysfunction compared to the lean hASC-derived matrix. Blockade of TGF-β1 signalling using an anti-TGF-β1 neutralizing antibody alleviated the lean hASC senescence and mitochondrial dysfunction induced by the obese hASC-derived matrix.ConclusionsNative ECM in obesity vWAT initiates hASC senescence through TGF-β1-mediated mitochondrial dysfunction. These data provide a key mechanism for understanding the importance of cell-ECM interactions in hASCs senescence in obesity.

Stereotactic ablative radiotherapy and FAPα-based cancer vaccine suppresses metastatic tumor growth in 4T1 mouse breast cancer

Background and purposeThis study tested the hypothesis that a novel combination of stereotactic ablation radiotherapy (SABR) and a cancer vaccine against fibroblast activation protein-alpha (FAPα) can suppress established tumor growth and impede potential metastasis. MethodsThe poorly immunogenic metastatic mouse mammary carcinoma 4T1 was used as a model. Mice were randomly assigned to five treatment groups: (1) untreated control, (2) FAPα-based cancer vaccine, (3) SABR, (4) SABR + pCDH (lentiviral control vector), (5) SABR + FAPα-based cancer vaccine (SABR/FAPα-Vax). FAPα-based cancer vaccine were administered subcutaneously every week for a total of three treatments. SABR was delivered to the primary tumor by 3 × 8 Gy after the first vaccination. ResultsConsistent with the poorly immunogenic nature of 4T1, tumor-bearing mice receiving FAPα-based cancer vaccine or SABR monotherapy showed a modest reduction in tumor volume and increased animal lifespan. In contrast, SABR/FAPα-Vax was well-tolerated, significantly reduced tumor burden, and increased survival compared to monotherapy. The increased survival correlated with inhibition of extracellular matrix (ECM) production, tumor vascularization and lymphangiogenesis. SABR/FAPα-Vax also resulted in an abscopal effect capable of eliminating lung metastases. SABR/FAPα-Vax recruited and activated CD8 + T cells to attack tumor cells and FAPα + stromal cells, and initiated suppressor cell reprogramming, including facilitating macrophage polarization toward an anti-tumor (M1) state, as well as depleting myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs). ConclusionThese findings provide a novel therapeutic combination of radiation and FAPα-based cancer vaccine with promising results against poorly immunogenic metastatic cancer. This study may pave the way to overcome the therapeutic resistance caused by FAPα + CAFs.

The Effects of Fibroblast Growth Factor-2 (FGF-2) on Achilles Tendon Injury

Category: Basic Sciences/Biologics; Ankle Introduction/Purpose: Achilles tendon defects are a challenging problem, and there is a need for new treatments. We investigated the use of Platelet-rich fibrin (PRF), a dense fibrin scaffold containing many growth factors and cytokines, for Achilles tendon injuries. The study found that PRF accelerates healing by promoting the proliferation and activation of tenocytes via FGF receptor/AKT signaling and TGF-β/SMAD3 signaling. While FGF-2, a growth factor in PRF, has been shown to promote tenocyte differentiation and proliferation, its effect on Achilles tendon injuries has not been reported. Therefore, this study aimed to investigate the effect of FGF-2 on Achilles tendon healing when administered alone and to determine its specific contribution to the healing process. Methods: We used a rat model of Achilles tendon defect. A 4-mm portion of the right Achilles tendon, 4 mm proximal to the calcaneal insertion, was resected to create a tendon defect. Gelatin hydrogel sheet (MedGel PI5) as a scaffold for PBS, FGF-2 (Rigroth®), and FGF receptor inhibitor (FGFRI: Pemigatinib) was placed into the gap before suturing it with nylon sutures. We observed HE-stained specimens at 200x and assessed histological healing by counting cells and vessels and grading cell morphology. The motor function was evaluated by the Basso, Beattie, and Bresnahan (BBB) score and the treadmill test. We compared the effects of FGF-2 on tenocytes isolated from rat Achilles tendons in the control, FGF2, and FGFRI groups. The proliferation and migration of tenocytes were measured by MTS assay and wound closure assay, respectively. Protein expression, such as Tenomodulin (Tnmd), Collagen Ⅰ, Ⅲ, and Screlaxis-A in cells, were evaluated by Western Blotting. Results: In the rat model, the number of cells, vessels and cell morphology scores at the defect sites increased in the FGF-2 group and decreased in the FGFRI group at 2 and 4 weeks postoperatively. In the motor functional evaluation, the FGF-2 group showed earlier improvement in the BBB score and the treadmill test compared to the control and FGFRI groups. The MTS assay showed that the number of viable cells was higher in the FGF-2 group and lower in the FGFRI group than in the control. Wound closure assay showed tenocyte migration was increased in the FGF-2 group and decreased in the FGFRI group. Regarding protein expression, Tnmd, Collagen I, III, and Screlaxis-A were increased in the FGF-2 group and decreased in the FGFRI group. Conclusion: In vivo, FGF-2 promoted healing of Achilles tendon injury, and FGFRI impeded it in terms of motor function and histology. Additionally, in vitro, FGF-2 promoted the proliferation and migration of tenocytes and enhanced extracellular matrix production, while FGFRI diminished these effects. Although native FGF-2 is produced at the injured site, further administration of FGF-2 accelerated Achilles tendon injury healing, while inhibiting the FGF receptor delayed recovery in this study. Therefore, FGF- 2 is very important in Achilles tendon injury healing and may promote recovery by enhancing angiogenesis, proliferation, and extracellular matrix production of tenocytes.

Pharmacodynamic study of chitosan in combination with salvianolic acid B in the treatment of CC14 -induced liver fibrosis in mice.

Liver fibrosis is a chronic liver lesion caused by excessive deposition of the extracellular matrix after liver damage, resulting in fibrous scarring of liver tissue. The progression of liver fibrosis is partially influenced by the gut microbiota. Chitosan can play a therapeutic role in liver fibrosis by regulating the gut microbiota based on the 'gut-liver axis' theory. Salvianolic acid B can inhibit the development of liver fibrosis by inhibiting the activation of hepatic stellate cells and reducing the production of extracellular matrix. In this study, the therapeutic effect of chitosan in combination with salvianolic acid B on liver fibrosis was investigated in a mouse liver fibrosis model. The results showed that the combination of chitosan and salvianolic acid B was better than the drug alone, improving AST/ALT levels and reducing the expression of α-SAM, COL I, IL-6 and other related genes. It improved the structure of gut microbiota and increased the abundance of beneficial bacteria such as Lactobacillus. The above results could provide new ideas for the clinical treatment of liver fibrosis.

Involvement of Epithelial-Mesenchymal Transition (EMT) in Autoimmune Diseases.

Epithelial-mesenchymal transition (EMT) is a complex reversible biological process characterized by the loss of epithelial features and the acquisition of mesenchymal features. EMT was initially described in developmental processes and was further associated with pathological conditions including metastatic cascade arising in neoplastic progression and organ fibrosis. Fibrosis is delineated by an excessive number of myofibroblasts, resulting in exuberant production of extracellular matrix (ECM) proteins, thereby compromising organ function and ultimately leading to its failure. It is now well acknowledged that a significant number of myofibroblasts result from the conversion of epithelial cells via EMT. Over the past two decades, evidence has accrued linking fibrosis to many chronic autoimmune and inflammatory diseases, including systemic sclerosis (SSc), rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), Sjögren's syndrome (SS), and inflammatory bowel diseases (IBD). In addition, chronic inflammatory states observed in most autoimmune and inflammatory diseases can act as a potent trigger of EMT, leading to the development of a pathological fibrotic state. In the present review, we aim to describe the current state of knowledge regarding the contribution of EMT to the pathophysiological processes of various rheumatic conditions.

Fibronectin-coated polyurethane meniscal scaffolding supplemented with MSCs improves scaffold integration and proteoglycan production in a rabbit model.

The role of mesenchymal stem cells (MSC) in supporting the formation of new meniscal tissue in a meniscal scaffold is not well understood. The objective of this study was to assess the quality of the meniscal tissue produced in a fibronectin (FN)-coated polyurethane (PU) meniscal scaffold after a meniscal injury was made in an experimental rabbit model. Twelve New Zealand white rabbits were divided in two groups after performing a medial meniscectomy of the anterior horn. In group 1, the meniscal defect was reconstructed with a non-MSC supplemented FN-coated PU scaffold. On the other hand, the same scaffold supplemented with MSCs was used in group 2. The animals were sacrificed at 12week after index surgery. A modified scoring system was used for histological assessment. This new scoring (ranging from 0 to 15) includes a structural evaluation (meniscal scaffold interface and extracellular matrix production) and tissue quality evaluation (proteoglycan and type I-collagen content). The meniscal scaffold was found loose in the joint in three cases, corresponding to two cases in group 1 and 1 case in group 2. No differences were observed between the groups in terms of the total score (7.0 ± 0.9 vs. 9.4 ± 2.6, p = 0.09). However, differences were observed in group 2 in which 2 out of the 5 scored items, scaffold integration (1 ± 0.0 vs. 1.9 ± 0.6, p = 0.03) and proteoglycan production (1.2 ± 0.3 vs. 2.4 ± 0.2, p = 0.001). A trend to a higher production of Type I-Collagen production was also observed in group 2 (1.1 ± 0.4 vs. 1.4 ± 0.7, p = 0.05). In a rabbit model at 12weeks, the adhesion of MSCs to a FN-coated PU scaffold improves scaffold integration, proteoglycan production and the characteristics of the new meniscal-like tissue obtained when compared to a non-supplemented scaffold. This fact could be a major step toward improving the adhesion of the MSCs to meniscal scaffolds and, consequently, the obtention of better quality meniscal tissue.

Magnetic-Responsive Carbon Nanotubes Composite Scaffolds for Chondrogenic Tissue Engineering.

The demand for engineered scaffolds capable of delivering multiple cues to cells continues to grow as the interplay between cell fate with microenvironmental and external cues is revealed. Emphasis has been given to develop stimuli-responsive scaffolds. These scaffolds are designed to sense an external stimulus triggering a specific response (e.g., change in the microenvironment, release therapeutics, etc.) and then initiate/modulate a desired biofunction. Here, magnetic-responsive carboxylated multi-walled carbon nanotubes (cMWCNTs) are integrated into 3D collagen/polylactic acid (PLA) scaffold via a reproducible filtration-based method. The integrity and biomechanical performance of the collagen/PLA scaffolds are preserved after cMWCNT integration. In vitro safety assessment of cMWCNT/collagen/PLA scaffolds shows neither cytotoxicity effects nor macrophage pro-inflammatory response, supporting further in vitro studies. The cMWCNT/collagen/PLA scaffolds enhance chondrocytes metabolic activity while maintaining high cell viability and extracellular matrix (i.e., type II collagen and aggrecan) production. Comprehensive in vitro study applying static and pulsed magnetic field on seeded scaffolds shows no specific cell response in dependence with the applied field. This result is independent of the presence or absence of cMWCNT into the collagen/PLA scaffolds. Taken together, these findings provide additional evidence of the benefits to exploit the CNTs outstanding properties in the design of stimuli-responsive scaffolds.

Mesenchymal stromal cells confer breast cancer doxorubicin resistance by producing hyaluronan.

Chemotherapy resistance represents a major cause of therapeutic failure and mortality in cancer patients. Mesenchymal stromal cells (MSCs), an integral component of tumor microenvironment, are known to promote drug resistance. However, the detailed mechanisms remain to be elucidated. Here, we found that MSCs confer breast cancer resistance to doxorubicin by diminishing its intratumoral accumulation. Hyaluronan (HA), a major extracellular matrix (ECM) product of MSCs, was found to mediate the chemoresistant effect. The chemoresistant effect of MSCs was abrogated when hyaluronic acid synthase 2 (HAS2) was depleted or inhibited. Exogenous HA also protected tumor grafts from doxorubicin. Molecular dynamics simulation analysis indicates that HA can bind with doxorubicin, mainly via hydrophobic and hydrogen bonds, and thus reduce its entry into breast cancer cells. This mechanism is distinct from the reported chemoresistant effect of HA via its receptor on cell surface. High HA serum levels were also found to be positively associated with chemoresistance in breast cancer patients. Our findings indicate that the HA-doxorubicin binding dynamics can confer cancer cells chemoresistance. Reducing HA may enhance chemotherapy efficacy.