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Background:Mesenchymal stem cells (MSCs) are adult stem cells with self-renewal and multi-directional differentiation potential, and possess the functions of immunomodulation, regulation of cell growth, and repair of damage. Over recent years, MSCs have been found to regulate the secretion of inflammatory factors and to exert regulatory effects on various lymphocytes in inflammatory states, and on the subsequent repair of tissue damage caused by inflammation. In the present study, we analyzed the effects of tissue inflammation on the characteristics ofMSCs. Methods: Human fat derived from the infrapatellar fat pad (IPFP) of knees with differing degrees of inflammation were extracted from specimens derived from total knee arthroplasties. HE and immunohistochemical staining was performed to directly observe the evidence and degree of inflammation in human infrapatellar fat pad tissue in order to classify MSCs cells, by their origin, into highly inflamed and lowly inflamed groups, and to study the effect of tissue inflammation on cell acquisition rates via cellular counting data. Flow cytometry assays were performed to investigate the effect of tissue inflammation on MSC surface marker expression. Trilineage differentiation, including osteogenesis, adipogenesis, and chondrogenesis was performed to assess the effect of tissue inflammation on the ability of MSCs to undergo directed differentiation. The effect of tissue inflammation on the ability of MSCs to proliferate was investigated via clone formation studies. RNA-sequencing was performed to evaluate the transcriptomes of MSCs derived from different areas of inflammation. The effect of tissue inflammation on tissue repair capacity and safety of MSCs were investigated via a murine model of acute liver injury. Results: The results of cell count data indicate that a high degree of tissue inflammation significantly decreases the acquisition rate of MSCs, and the proportion of CD34+ and CD146+ cells. The results of our trilineage differentiation assay shows that a higher degree of inflammation decreases osteogenic differentiation, and enhances adipogenic and chondrogenic differentiation of MSCs. Clone formation assays indicates that a higher degree of tissue inflammation at the MSC source increases the proliferative ability of MSCs. The transcriptomes of MSCs remain relatively stable in fat pad tissues derived from both highly and lowly inflamed samples. The results of acute liver injury investigations in mice indicate that MSCs of low inflammatory tissue origin have an enhanced tissue repair capability. Conclusions: High tissue inflammation at the source of MSCs reduces the acquisition rate of MSCs, increases the proliferative capacity of MSCs, inhibits osteogenic differentiation, and enhances adipogenic and chondrogenic differentiation. A higher degree of tissue inflammation reduces the tissue repair capacity of MSCs; however, MSCs obtained from differing source degrees of inflammation retain stable and similar genetic transcriptomic information, and have good tissue repair capability.

Intra-articular injection of mesenchymal stem cells (MSCs) is a promising strategy for osteoarthritis (OA) treatment. However, more and more studies reveal that the injected MSCs have poor adhesion, migration, and survival in the joint cavity. A recent study shows that tropoelastin (TE) regulates adhesion, proliferation and phenotypic maintenance of MSCs as a soluble additive, indicating that TE could promote MSCs-homing in regenerative medicine. In this study, we used TE as injection medium, and compared it with classic media in MSCs intra-articular injection such as normal saline (NS), hyaluronic acid (HA), and platelet-rich plasma (PRP). We found that TE could effectively improve adhesion, migration, chondrogenic differentiation of infrapatellar fat pad MSCs (IPFP-MSCs) and enhance matrix synthesis of osteoarthritic chondrocytes (OACs) in indirect-coculture system. Moreover, TE could significantly enhance IPFP-MSCs adhesion via activation of integrin β1, ERK1/2 and vinculin (VCL) in vitro. In addition, intra-articular injection of TE-IPFP MSCs suspension resulted in a short-term increase in survival rate of IPFP-MSCs and better histology scores of rat joint tissues. Inhibition of integrin β1 or ERK1/2 attenuated the protective effect of TE-IPFP MSCs suspension in vivo. In conclusion, TE promotes performance of IPFP-MSCs and protects knee cartilage from damage in OA through enhancement of cell adhesion and activation of integrin β1/ERK/VCL pathway. Our findings may provide new insights in MSCs intra-articular injection for OA treatment., Graphical abstract Image 1, Highlights • Tropoelastin promotes adhesion and migration of IPFP MSCs. • Tropoelastin promotes IPFP MSCs survival and adhesion in joint cavity. • Injection of IPFP MSCs in tropoelastin reduces OA progression.

Insufficient vascularization of grafts often leads to delayed tissue ingrowth and impaired tissue function in tissue engineering. The surface topography of grafts plays critical roles in angiogenesis. In the present study, we prepared silk fibroin (SF)-based microtopography films with the number of convex dots ranging from 37 to 4835/mm2. The convex dot-featured topography surfaces were characterized by scanning electron microscopy, a Profilm3D optical profilometer, atomic force microscopy, and a contact angle goniometer. The effect of microtopographic films on the proliferation, adhesion, and expression of angiogenic factors of human umbilical vein endothelial cells (HUVECs) was investigated. Our results demonstrated that the SF film surface with 2899 convex dots/mm2 significantly enhanced adhesion, viability, and levels of vascular endothelial growth factors and basic fibroblast growth factors of HUVECs and significantly downregulated the level of α-SMA in human aortic smooth muscle cells, indicating that the microtopographic films could promote angiogenesis. Furthermore, in vitro results showed that HUVEC proliferation was positively correlated with yes-associated protein (YAP) activation, suggesting that the enhanced angiogenesis was mediated via the YAP pathway. Finally, mice subcutaneous embedding model results indicated that the SF film surface with 2899 convex dots/mm2 could significantly enhance angiogenesis in vivo. Altogether, our results showed that the SF film surface with 2899 convex dots/mm2 promoted the angiogenesis of HUVECs and offered a novel angiogenesis-promoting strategy of implant surface design for tissue engineering.

Osteoarthritis (OA) is the most common disabling joint disease and its pathological process is closely related to oxidative stress. Recent studies have shown that antioxidants allicin, sulforaphane, and lycopene derived from natural ingredients garlic, broccoli, and tomato can reduce the degree of oxidative stress and the expression of inflammatory markers, indicating that theses antioxidants might be helpful for OA treatment. In this study, we investigated the effects of allicin, sulforaphane, and lycopene on H2O2-stimulated human osteochondral samples and osteoarthritic chondrocytes. Our results revealed that allicin, sulforaphane, and lycopene effectively reduced the oxidative stress-induced cell apoptosis, and increased gene expression of antioxidant enzymes. Besides, these natural ingredients-derived antioxidants reduced the expression of inflammatory factors, enhanced the chondrogenic matrix synthesis, and reduced the hypertrophic differentiation of osteoarthritic chondrocytes. These regulations were mainly through the activation of Keap1/Nrf2 pathway. Our findings suggest that these antioxidants might be a potential therapeutic strategy for OA.

Osteoarthritis (OA) is the most common disabling joint disease throughout the world and its therapeutic effect is still not satisfactory in clinic nowadays. Recent studies showed that the exosomes derived from several types of mesenchymal stem cells (MSCs) could maintain chondrocyte homeostasis and ameliorate the pathological severity of OA in animal models, indicating that MSCs-derived exosomes could be a novel promising strategy for treating OA. In this study, we investigated the role and underlying mechanisms of infrapatellar fat pad (IPFP) MSCs-derived exosomes (MSCIPFP-Exos) on OA in vitro and in vivo. Our data revealed that MSCIPFP could produce amounts of MSCIPFP-Exos, which exhibited the typical morphological features of exosomes. The MSCIPFP-Exos ameliorated the OA severity in vivo and inhibited cell apoptosis, enhanced matrix synthesis and reduced the expression of catabolic factor in vitro. Moreover, MSCIPFP-Exos could significantly enhance autophagy level in chondrocytes partially via mTOR inhibition. Exosomal RNA-seq showed that the level of miR-100-5p that could bind to the 3'-untranslated region (3'UTR) of mTOR was the highest among microRNAs. MSCIPFP-Exos decreased the luciferase activity of mTOR 3'UTR, while inhibition of miR-100-5p could reverse the MSCIPFP-Exos-decreased mTOR signaling pathway. Intra-articular injection of antagomir-miR-100-5p dramatically attenuated MSCIPFP-Exos-mediated protective effect on articular cartilage in vivo. In brief, MSCIPFP-derived exosomes protect articular cartilage from damage and ameliorate gait abnormality in OA mice by maintaining cartilage homeostasis, the mechanism of which may be related to miR100-5p-regulated inhibition of mTOR-autophagy pathway. As it is relatively feasible to obtain human IPFP from OA patients by arthroscopic operation in clinic, MSCIPFP-derived exosomes may be a potential therapy for OA in the future.

BackgroundOsteoarthritis (OA), the most common joint disorder, is characterized by a progressive degradation of articular cartilage. Increasing evidence suggests that OA is closely associated with cartilage pathologies including chondrocyte hypertrophy and fibrosis.MethodsIn this study, we showed that asiatic acid (AA) treatment reduced chondrocyte hypertrophy and fibrosis. First, the cytotoxicity of AA (0, 5, 10, and 20 μM) to chondrocytes was evaluated, and 5 μM was selected for subsequent experiments. Then, we detected the gene and protein level of chondrocyte hypertrophic markers including type X collagen (COL-X), matrix metalloproteinase-13 (MMP-13), alkaline phosphatase (ALP), and runt-related transcription factor 2 (Runx2); chondrocyte fibrosis markers including type I collagen (COL-Ι) and alpha-smooth muscle actin (α-SMA); and chondrogenic markers including SRY-related HMG box 9 (SOX9), type II collagen (COL-II), and aggrecan (ACAN). Further, we tested the mechanism of AA on inhibiting chondrocyte hypertrophy and fibrosis. Finally, we verified the results in an anterior cruciate ligament transection (ACLT) rat OA model.ResultsWe found that AA treatment inhibited the hypertrophic and fibrotic phenotype of chondrocytes, without affecting the chondrogenic phenotype. Moreover, we found that AA treatment activated AMP-activated protein kinase (AMPK) and inhibited phosphoinositide-3 kinase/protein kinase B (PI3K/AKT) signaling pathway in vitro. The results in an ACLT rat OA model also indicated that AA significantly attenuated chondrocyte hypertrophy and fibrosis.ConclusionAA treatment could reduce hypertrophic and fibrotic differentiation and maintain the chondrogenic phenotype of articular chondrocytes by targeting the AMPK/PI3K/AKT signaling pathway. Our study suggested that AA might be a prospective drug component that targets hypertrophic and fibrotic chondrocytes for OA treatment.

Osteoarthritis (OA) is the most common disabling joint disease and its pathological process is closely related to oxidative stress. Recent studies have shown that antioxidants allicin, sulforaphane, and lycopene derived from natural ingredients garlic, broccoli, and tomato can reduce the degree of oxidative stress and the expression of inflammatory markers, indicating that theses antioxidants might be helpful for OA treatment. In this study, we investigated the effects of allicin, sulforaphane, and lycopene on H

Background: Mesenchymal stromal cell (MSC)–based therapies have emerged as a promising strategy for osteoarthritis (OA) treatment. In particular, infrapatellar fat pad (IPFP)–derived MSCs have become a good option to treat knee OA. Purpose: To investigate the influence of the local microenvironment of the knee joint, especially OA cartilage, on the bioactivities of injected/implanted IPFP MSCs. Study Design: Controlled laboratory study. Methods: Conditioned medium (CM) derived from OA cartilage fragments was collected and characterized. Donor-matched IPFP MSCs were treated with control medium (Dulbecco’s modified Eagle medium (DMEM)/F-12 or chondrogenic medium), control medium + CM, or CM alone; and a series of behaviors including the viability, migration, chondrogenic and hypertrophic differentiation, and catabolic activity of IPFP MSCs were evaluated among groups. Results: There were 14 cytokines detected in CM. CM treatment improved the viability of IPFP MSCs. CM hindered the migration of IPFP MSCs. In chondrogenic differentiation, the presence of CM increased the expression of chondrogenic markers but also enhanced the state of hypertrophy and catabolism. Conclusion: OA cartilage–secreted factors could induce chondrogenic differentiation but also resulted in negative effects including the weakened migration, increased hypertrophy, and catabolism of IPFP MSCs in vitro. Clinical Relevance: These findings provide an insight on the fate of IPFP MSCs after intra-articular injections.

Ca2+ has been recognized as a key molecule for chondrocytes, however, the role and mechanism of spontaneous [Ca 2+ ] i signaling in cartilaginous extracellular matrix (ECM) metabolism regulation are unclear. Here we found that spontaneous Ca 2+ signal of in-situ porcine chondrocytes was [Ca 2+ ] o dependent, and mediated by [Ca 2+ ] i store release. T-type voltage-dependent calcium channel (T-VDCC) mediated [Ca 2+ ] o influx was associated with decreased cell viability and expression levels of ECM deposition genes. Further analysis revealed that chondrocytes expressed both inositol 1,4,5-trisphosphate receptor (InsP3R) and Orai isoforms. Inhibition of endoplasmic reticulum (ER) Ca 2+ release and store-operated calcium entry significantly abolished spontaneous [Ca 2+ ] i signaling of in-situ chondrocytes. Moreover, blocking ER Ca 2+ release with InsP3R inhibitors significantly upregulated ECM degradation enzymes production, and was accompanied by decreased proteoglycan and collagen type II intensity. Taken together, our data provided evidence that spontaneous [Ca 2+ ] i signaling of in-situ porcine chondrocytes was tightly regulated by [Ca 2+ ] o influx, InsP3Rs mediated [Ca 2+ ] i store release, and Orais mediated calcium release-activated calcium channels activation. Both T-VDCC mediated [Ca 2+ ] o influx and InsP3Rs mediated ER Ca 2+ release were found crucial to cartilaginous ECM metabolism through distinct regulatory mechanisms.