Oct4 facilitates chondrogenic differentiation of mesenchymal stem cells by mediating CIP2A expression.

Bone development and cartilage formation require strict modulation of gene expression for mesenchymal stem cells (MSCs) to progress through their differentiation stages. Octamer-binding transcription factor 4 (Oct4) expression is generally restricted to developing embryonic pluripotent cells, but its role in chondrogenic differentiation (CD) of MSCs remains unclear. We therefore investigated the role of Oct4 in CD using a microarray, quantitative real-time polymerase chain reaction, and western blotting. The expression of Oct4 was elevated when the CD of cultured MSCs was induced. Silencing Oct4 damaged MSC growth and proliferation and decreased CD, indicated by decreased cartilage matrix formation and the expression of Col2a1, Col10a1, Acan, and Sox9. We found a positive correlation between the expression of CIP2A, a natural inhibitor of protein phosphatase 2A (PP2A) and that of Oct4. Cellular inhibitor of PP2A (CIP2A) expression gradually increased after CD. Overexpression of CIP2A in MSCs with Oct4 depletion promoted cartilage matrix deposition as well as Col2a1, Col10a1, Acan, and Sox9 expression. The chondrogenic induction triggered c-Myc, Akt, ERK, and MEK phosphorylation and upregulated c-Myc and mTOR expression, which was downregulated upon Oct4 knockdown and restored by CIP2A overexpression. These findings indicated that Oct4 functions as an essential chondrogenesis regulator, partly via the CIP2A/PP2A pathway.

MiR-146a-5p promotes IL-1ß-induced chondrocyte apoptosis through the TRAF6-mediated NF-kB pathway.

OBJECTIVE: This study aimed to explore the role of the miR-146a-5p/TRAF6/NF-KB axis in chondrocyte apoptosis. METHODS: Transcriptome sequencing for microRNA expression in control and osteoarthritic cartilage was performed. Bioinformatic analysis was performed to identify the target genes of miR-146a-5p, and subsequently, Gene Ontology (GO) terms and KEGG pathways were identified. Furthermore, protein-protein interactions were analyzed to identify the hub regulatory gene of miR-146a-5p. MiR-146a-5p mimic, inhibitor and the corresponding negative control were constructed, and the apoptosis rates were measured in the transfected groups by flow cytometry, TUNEL staining and Western blot. Potential miRNA-target interactions were identified by dual-luciferase reporter assay. RESULTS: The microRNA array demonstrated that miR-146a-5p was significantly upregulated in osteoarthritic tissues, which was further confirmed by PCR analysis. Compared with the control group, IL-1ß significantly decreased the viability of chondrocytes, while coculture with miR-146a-5p inhibitor rescued the IL-1ß-induced inhibition of chondrocyte viability. Western blot results also identified the proapoptotic effects of miR-146a-5p. Bioinformatic analysis results revealed that miR-146a-5p targeted 159 potential genes, and TRAF6 was the hub gene among the 159 genes. The relative expression of TRAF6 was significantly decreased in the IL-1ß-induced group. When siTRAF6 was added, apoptosis was significantly increased. Luciferase reporter assays showed that luciferase activity of the TRAF6 3'-UTR reporter was decreased in chondrocytes after transfection with the miR-146a-5p mimic. CONCLUSIONS: This work showed that miR-146 induces chondrocyte apoptosis by targeting the TRAF6-mediated NF-KB signaling pathway, and miR-146 may be a potential target for OA treatment.

MicroRNA-218 promotes early chondrogenesis of mesenchymal stem cells and inhibits later chondrocyte maturation.

BACKGROUND: MicroRNAs (miRNAs) reportedly participate in the mesenchymal stem cell (MSC) chondrogenic differentiation regulation. We objected to examine how miR-218 regulate chondrogenic differentiation of synovium-derived MSCs (SDSCs) and the maturation of RCJ3.1C5.1 chondrocytes. SDSCs were sourced from the knee joint synovium of New Zealand white rabbits, and their multilineage differentiation potentials were examined. The level of miR-218 was measured during SDSC chondrogenic differentiation, together with determination of SDSCs chondrogenic markers and RCJ3.1C5.1 chondrocytes maturation markers expression level after transfection of miR-218 mimics/inhibitor. RESULTS: miR-218 expression was notably upregulated in early chondrogenesis but mostly ceased during the maturation phases of chondrogenic differentiation in SDSCs. The transfection of miR-218 mimics notably enhanced SDSCs chondrocytes differentiation, as evidenced by augmented expressions of chondrogenic markers (SOX9, COL2A1, ACAN, GAG, and COMP) in terms of mRNA and protein level, and the inhibition of miR-218 yielded opposite resutls. Additionally, miR-218 overexpression substantially suppressed the expression of osteogenic markers (ALP, BSP, COL1A1, OCN and OPN) during the early stage of chondrogenesis while increasing that of chondrogenic markers (SOX9, COL2A1, ACAN, GAG and COMP). However, miR-218 mimics notably suppressed maturation markers (CMP, COL10A1, MMP-13 and VEGF) expression in RCJ3.1C5.18 chondrocytes, and the miR-218 inhibitor promoted the expression of these maturation markers. We proposed miR-218 plays a regulatory role on 15-hydroxyprostaglandin dehydrogenase (HPGD), which plays a key role in chondrogenic differentiation, and this finding indicates that miR-218 directly regulates HPGD expression in SDSCs. CONCLUSION: Our study suggests that miR-218 contributes to early chondrogenesis while suppressing later chondrocyte maturation. The miR-218-HPGD pathway offers us a perspective into how SDSCs differentiate into chondrogenic cells.

A New Insight of Kartogenin Induced the Mesenchymal Stem Cells (MSCs) Selectively Differentiate into Chondrocytes by Activating the Bone Morphogenetic Protein 7 (BMP-7)/Smad5 Pathway.

BACKGROUND Rotator cuff injury is the most common cause of shoulder disability, and although the repair technique has improved, the rate of rotator cuff reduction after repair is still high. The fibrocartilage region, which appears to be histologically inserted, cannot be regenerated. In recent years, studies have reported that mesenchymal stem cells (MSCs) have enhanced cartilage regeneration in the tendon and bone interface after rotator cuff repair, which has become a hot topic of research. MATERIAL AND METHODS Two mesenchymal stem cell types, SMSC (synovial-derived mesenchymal stem cells) and BMSC (bone marrow-derived mesenchymal stem cells) were intervened using kartogenin (KGN). The cytotoxicity was evaluated and the proliferation of the 2 cells was observed. Four commonly used cartilage phenotype genes were detected by quantitative real-time polymerase chain reaction, and the cartilage differentiation of MSCs induced by KGN was explored. The bidirectional regulation of the expression of BMP-7 and the downstream gene Smad5 was observed by constructing a lentiviral overexpression vector containing the target gene BMP-7. To explore whether BMP-7/Smad5 pathway activation promotes differentiation of SMSCs into chondrocytes. RESULTS KGN can induce the selective differentiation of endogenous MSCs into chondrocytes by activating the BMP-7/Smad5 pathway, which promotes the regeneration of interfacial cartilage, and improves the quality of tendon healing of the tendon after rotator cuff repair. CONCLUSIONS This study found a new biological intervention method to promote the effect of tendon on bone healing after rotator cuff repair.

Clinical Efficacy of Intra-Articular Injection with P-PRP Versus that of L-PRP in Treating Knee Cartilage Lesion: A Randomized Controlled Trial.

OBJECTIVE: Platelet-rich plasma（PRP), with different concentration of leukocytes, may lead to varying effects in the treatment of cartilage lesions. So far, current research has not shown enough evidence on this. To evaluate the clinical efficacy and safety of intra-articular injection with pure platelet-rich plasma (P-PRP) versus those of leukocyte platelet-rich plasma (L-PRP) in treating knee cartilage lesions, we conducted a double-blind, randomized controlled clinical trial with a larger sample and longer follow-up period. METHODS: From October 2019 to October 2020, 95 patients were invited to participate in our study, and 60 (63.2%) were randomized to P-PRP (n = 30) or L-PRP (n = 30) groups. Patients from the two groups were treated with knee intra-articular injections of P-PRP or L-PRP. Visual analog scale (VAS) and Western Ontario and McMaster Universities Arthritis Index (WOMAC) scores were assessed using an unpaired t-test for independent samples preoperatively and at 6 weeks, 12 weeks, 6 months, and 12 months after intervention. RESULTS: We followed up 27 cases in the P-PRP group and 26 cases in the L-PRP group. No significant differences in VAS and WOMAC scores were found between the two groups before the intervention (p > 0.05). The WOMAC Pain and VAS-Motions scores of the P-PRP group were significantly lower than those of the L-PRP group at 6 weeks after the intervention (p < 0.05). While the long-term clinical efficacy of both injections was similar and weakened after 12 months, more adverse events were found in the L-PRP group. CONCLUSIONS: The short-term results demonstrate a positive effect in reducing pain and improving function in patients with knee cartilage lesions in the two groups. While the P-PRP injection showed better clinical efficacy in the early phase of postoperative rehabilitation and resulted in fewer adverse events, long-term follow-up showed similar and weakened efficacy after 12 months. TRIAL REGISTRATION: ChiCTR1900026365. Registered on October 3, 2019, http://www.chictr.org.cn/showproj.aspx?proj=43911.

Culture of Mesenchymal Stem Cells Derived From the Infrapatellar Fat Pad Without Enzyme and Preliminary Study on the Repair of Articular Cartilage Defects in Rabbits.

Objective: The aim of the study was to evaluate the advantages of without enzyme isolating patellar fat pad-derived mesenchymal stem cells (IPFP-SCs) and the feasibility of cartilage repair. Methods: The IPFP-SCs were isolated using the without enzyme method and compared with the IPFP-SCs obtained by the traditional enzyme digestion method in terms of cell proliferation ability, characterization, and differentiation ability, and the differences in chondrogenic induction and differentiation between the two groups were compared. Twenty-four New Zealand rabbits were randomly divided into four groups (n = 6). After the articular cartilage defects were modeled, different preparations were injected into the joint cavity. The rabbits in the group A were injected with the mixture of IPFP-SCs and pure PRP (P-PRP), separated using the without enzyme method, while those in the group B were injected with the mixture of IPFP-SCs and P-PRP separated with the digestion method, while those in the group C were injected with SVF separated using the without enzyme method, and those in the group D were injected with normal saline. At 6 weeks and 12 weeks after operation, the cartilage repair of rabbit joint specimens was observed and evaluated by gross observation and histological staining, and the effects of different IPFP-SCs application forms in repairing cartilage defects were compared. Results: The time required to obtain IPFP-SCs by enzyme-free isolation was significantly less than that by enzyme digestion, while the acquisition rate of primary cells was significantly lower than that by enzyme digestion. After culture and amplification, the two IPFP-SCs from different sources did not show significant differences in cell proliferation, cell phenotype, and differentiation ability. In animal experiments, groups A and B had the best effect on the repair of cartilage defects, and there was no significant difference between the two groups. The repair effect in group C was weaker than that in the former two groups, but it was relatively better than that in group D. Conclusion: It is more time-saving to obtain IPFP-SCs by the without enzyme method than by enzymatic digestion, and there is no significant difference in cell identification and differentiation potential between the two methods. However, the rate of obtaining primary cells was significantly lower than that with the enzyme digestion method. IPFP-SCs showed good repair effect in the rabbit animal cartilage defect model, providing ideas and reference for the clinical application of stem cells in repairing articular cartilage.

Regulating effect of Circ_ATRNL1 on the promotion of SOX9 expression to promote chondrogenic differentiation of hAMSCs mediated by MiR-145-5p.

Circ_ATRNL1 is significantly highly expressed in cartilage tissues of patients with osteoarthritis. This study explored the role and mechanism of circ_ATRNL1 in cartilage differentiation of human adipose-derived mesenchymal stem cells (hAMSCs). hAMSCs were isolated and identified by flow cytometry. The degree of chondrocyte and adipogenic differentiation was assessed using Alcian blue staining and Oil Red O staining, respectively. The expressions of cartilage- and adipogenic-related genes, circ_ATRNL1, and SOX9 were detected by reverse transcription quantitative polymerase chain reaction. The correlation between SOX9 and circ_ATRNL1 was analyzed using Pearson test. Bioinformatics and luciferase analysis were used to detect the overlapped target miRNAs of circ_ATRNL1 and SOX9. The role of circ_ATRNL1/miRNA/SOX9 was examined using functional rescue assays. hAMSCs were identified as CD90-, CD105-, and CD44-positive. The degree of cartilage differentiation of hAMSCs was significantly enhanced after 2 weeks. Cartilage-related genes, circ_ATRNL1 and SOX9, were significantly upregulated, and positively correlated with each other. Circ_ATRNL1 overexpression enhanced hAMSC proliferation and differentiation into chondrogenesis, and promoted the expressions of COL2, Aggrecan, and SOX9. Overexpression of circ_ATRNL1 inhibited the adipogenic differentiation of hAMSCs and the expressions of adipogenic-related genes. miR-145-5p was a target miRNA for circ_ATRNL1 and SOX9. miR-145-5p mimic inhibited hAMSC differentiation toward cartilage, and inhibited the expression of cartilage-related factors. miR-145-5p mimic effectively reversed the regulating effect of circ_ATRNL1 on hAMSCs. Circ_ATRNL1 regulates the promotion of SOX9 expression to promote chondrogenic differentiation of hAMSCs mediated by miR-145-5p.

Over-expression of MEG3 promotes differentiation of bone marrow mesenchymal stem cells into chondrocytes by regulating miR-129-5p/RUNX1 axis.

This study explored the role of MEG3 in the cartilage differentiation of bone marrow mesenchymal stem cells (BMSCs). We investigated the effects of over-expression and knockdown of MEG3 on cell viability, cell differentiation, and the expressions of MEG3, miR-129-5p, COL2, chondrocyte differentiation-related genes (sry-type high-mobility-group box 9 (SOX9), SOX5, Aggrecan, silent information regulator 1 (SIRT1), and Cartilage oligomeric matrix protein (COMP)). The targeting relationship between MEG3 and miR-129-5p and the target gene of miR-129-5p was confirmed through Starbase, TargetScan and luciferase experiments. Finally, a series of rescue experiments were conducted to study the regulatory effects of MEG3 and miR-129-5p. BMSCs were identified as CD29+ and CD44+ positive, and their differentiation was time-dependent. As BMSCs differentiated, MEG3 expression was up-regulated, but miR-129-5p was down-regulated. Over-expressed MEG3 promoted the viability and differentiation of BMSCs, up-regulated the expressions of COL2 and chondrocyte differentiation-related genes, and inhibited miR-129-5p. Runt-related transcription factor 1 (RUNX1) was negatively regulated as a target gene of miR-129-5p. Results of rescue experiments showed that the inhibitory effect of miR-129-5p mimic on BMSCs could be partially reversed by MEG3. Over-expression of MEG3 regulated the miR-129-5p/RUNX1 axis to promote the differentiation of BMSCs into chondrocytes. This study provides a reliable basis for the application of lncRNA in articular cartilage injury.

The clinical efficacy of arthroscopic therapy with knee infrapatellar fat pad cell concentrates in treating knee cartilage lesion: a prospective, randomized, and controlled study.

INTRODUCTION: To evaluate the clinical efficacy of arthroscopic therapy with infrapatellar fat pad cell concentrates in treating knee cartilage lesions, we conducted a prospective randomized single-blind clinical study of controlled method. METHODS: Sixty cases from Shanghai Changzheng Hospital from April 2018 to December 2019 were chosen and randomly divided into 2 groups equally. Patients in the experiment group were treated through knee arthroscopy with knee infrapatellar fat pad cell concentrates containing mesenchymal stromal cells, while patients in the control group were treated through regular knee arthroscopic therapy. VAS and WOMAC scores were assessed at pre-operation, and 6 weeks, 12 weeks, 6 months, and 12 months after intervention. MORCART scores were assessed at pre-operation and 12 months after intervention. RESULTS: Twenty-nine cases in the experiment group and 28 cases in the control group were followed up. No significant difference in VAS, WOMAC, and MOCART scores were found between the two groups before surgery (P > 0.05). The WOMAC total and WOMAC function scores of the experiment group were significantly lower than those of the control group 6 months and 12 months after surgery (P < 0.05). The VAS rest and VAS motion scores of the experiment group were found significantly lower than those of the control group 12 months after surgery (P < 0.05). The MOCART scores of the experiment group were found significantly higher compared with the control group 12 months after surgery (P < 0.05). No significant difference in WOMAC stiffness scores were found between the two groups. CONCLUSIONS: The short-term results of our study are encouraging and demonstrate that knee arthroscopy with infrapatellar fat pad cell concentrates containing mesenchymal stromal cells is safe and provides assistance in reducing pain and improving function in patients with knee cartilage lesions. TRIAL REGISTRATION: ChiCTR1800015379. Registered on 27 March 2018, http://www.chictr.org.cn/showproj.aspx?proj=25901 .

Decellularized extracellular matrix loaded with IPFP-SC for repairing rabbit osteochondral defects.

BACKGROUND: Tissue engineering is widely applied to treat osteochondral damage in osteoarthritis (OA). However, the superposition of seed cells, material scaffolds, inducing factors, and microenvironmental factors limit their practical application. We intended to develop a novel tissue engineering method for improving the repairment of osteochondral damage and to discuss its effect on repairing osteochondral defects. METHODS: The combined decellularization methods of physics, chemistry and enzymes were used to decellularize rabbit rib cartilage and articular cartilage, and rabbit decellularizated osteochondral composite scaffolds were prepared. The structure and organization of the scaffolds were analyzed. We extracted and identified infrapatellar fat pad stem cells (IPFP-SCs) from healthy rabbits and OA rabbit, which were different in viability, migration, osteogenic and chondrogenic differentiation. Finally, a variety of decellularizated bone cartilage composite scaffolds were loaded with rabbit IPFP-SC for in vitro and in vivo studies. RESULTS: The decellularization effect was strong, and the organic ingredients were lost. The layered scaffold showed lower density, greater porosity, larger pore size and water absorption than the whole scaffold, but the mechanical properties of the two scaffolds were low. IPFP-SCs were successfully extracted, and the migration and cartilage ability of IPFP-SCs in OA group were weak. The decellularized scaffold showed a high biocompatibility. The structure and composition of osteochondral promoted osteogenic differentiation and chondrogenic differentiation of IPFP-SCs. Moreover, the decellularized extracellular matrix loaded with IPFP-SC had the strongest repairing effect. CONCLUSION: The decellularized extracellular matrix loaded with IPFP-SC showed a better repair effect on rabbit osteochondral defects.

Fibrin Glue-Kartogenin Complex Promotes the Regeneration of the Tendon-Bone Interface in Rotator Cuff Injury.

OBJECTIVE: Rotator cuff injury healing is problematic because the tendon-bone junction often forms cicatricial tissues, rather than fibrocartilage, which leads to mechanical impairment and is prone to redamage. Kartogenin (KGN) is a newly discovered small molecule compound which can induce cartilage formation through chondrogenesis of endogenous mesenchymal stem cells. METHODS: In this study, we used KGN with fibrin glue (FG) to repair the rotator cuff injury by promoting the formation of fibrocartilage at the tendon to bone interface. Firstly, we assessed the release rate of KGN from the FG-KGN complex and then created a rabbit rotator cuff tendon graft-bone tunnel model. The rabbits received saline, FG-KGN, or FG injections onto the tendon to bone interface after injury. Shoulder tissues were harvested at 6 and 12 weeks, and the sections were stained with HE and Safranin O/Fast green. The samples were assessed by histologic evaluation and biomechanical testing. Synovial mesenchymal stem cells derived from the synovial tissue around the rotator cuff were harvested for western blotting and qRT-PCR analysis. RESULTS: KGN was released rapidly from the FG-KGN complex during first 4 hrs and followed by a slow release until 7 days. The tendon graft-bone interface in the control (saline) group and the FG group was filled with scar tissue, rather than cartilage-like tissue, and only a small number of chondrocytes were found at the adjacent bone surface. In the FG-KGN group, the tendon to bone interface was fully integrated and populated by chondrocytes with proteoglycan deposition, indicating the formation of fibrocartilage-like tissues. At 12 weeks, the maximum tensile strength of the FG-KGN group was significantly higher than that of the FG and control groups (P < 0.01). The RNA expression levels of tendinous genes such as Tenascin C and the chondrogenic gene Sox-9 were substantially elevated in SMSCs treated with the FG-KGN complex compared to the other two groups. CONCLUSION: These results indicated that fibrin glue is an effective carrier for KGN, allowing for the sustained release of KGN. The FG-KGN complex could effectively promote the regeneration and formation of fibrocartilage tissue of the tendon-bone interface in the rabbit rotator cuff tendon graft-bone tunnel model.

Exosomes from Kartogenin-Pretreated Infrapatellar Fat Pad Mesenchymal Stem Cells Enhance Chondrocyte Anabolism and Articular Cartilage Regeneration.

OBJECTIVE: To evaluate the effect of Kartogenin-pretreated exosomes derived from infrapatellar fat pad mesenchymal stem cells on chondrocyte in vitro and articular cartilage regeneration in vivo. METHODS: Infrapatellar fat pad mesenchymal stem cells (IPFP-MSCs) were isolated from rabbits to harvest exosomes. After identification of mesenchymal stem cells and exosomes, rabbit chondrocytes were divided into three groups for further treatment: the EXO group (chondrocytes treated with exosomes isolated from infrapatellar fat pad mesenchymal stem cells), KGN-EXO group (chondrocytes treated with exosomes isolated from infrapatellar fat pad mesenchymal stem cells pretreated with KGN), and control group. After processing and proliferation, phenotypic changes of chondrocytes were measured. In the in vivo study, 4 groups of rabbits with articular cartilage injury were treated with KGN-EXO, EXO, IPFP-MSCs, and control. Macroscopic evaluation and histological evaluation were made to figure out the different effects of the 4 groups on cartilage regeneration in vivo. RESULTS: The proliferation rate of chondrocytes in the EXO or KGN-EXO group was significantly higher than that in the control group (P < 0.05). The qRT-PCR results showed that the expression of Sox-9, Aggrecan, and Col II was the highest in the KGN-EXO group compared with the EXO group and the control group (P < 0.05). The results of Western blot were consistent with the results of qRT-PCR. In vivo, the cartilage defects in the KGN-EXO group showed better gross appearance and improved histological score than those in IPFP-MSC groups, EXO groups, and control groups (P < 0.05). At 12 weeks, the defect site in the KGN-EXO group was almost completely repaired with a flat and smooth surface, while a large amount of hyaline cartilage-like structures and no obvious cracks were observed. CONCLUSION: Our study demonstrates that the exosomes isolated from infrapatellar fat pad mesenchymal stem cells pretreated with KGN have potent ability to induce chondrogenic differentiation of stem cells, effectively promoting the proliferation and the expression of chondrogenic proteins and genes of chondrocytes. The KGN-EXO can also promote the repair of articular cartilage defects more effectively, which can be used as a potential therapeutic method in the future.

Comparison between Intra-Articular Injection of Infrapatellar Fat Pad (IPFP) Cell Concentrates and IPFP-Mesenchymal Stem Cells (MSCs) for Cartilage Defect Repair of the Knee Joint in Rabbits.

Mesenchymal stem cells (MSCs) have emerged as a promising therapeutic method in regenerative medicine. Our previous research adopted a simple nonenzymatic strategy for the preparation of a new type of ready-to-use infrapatellar fat pad (IPFP) cell concentrates. The aim of this study was to compare the therapeutic efficacy of intra-articular (IA) injection of autologous IPFP cell concentrates and allogeneic IPFP-MSCs obtained from these concentrates in a rabbit articular cartilage defect model. IPFP-MSCs sprouting from the IPFP cell concentrates were characterized via flow cytometry as well as based on their potential for differentiation into adipocytes, osteoblasts, and chondrocytes. In the rabbit model, cartilage defects were created on the trochlear groove, followed by treatment with IPFP cell concentrates, IPFP-MSCs, or normal saline IA injection. Distal femur samples were evaluated at 6 and 12 weeks posttreatment via macroscopic observation and histological assessment based on the International Cartilage Repair Society (ICRS) macroscopic scoring system as well as the ICRS visual histological assessment scale. The macroscopic score and histological score were significantly higher in the IPFP-MSC group compared to the IPFP cell concentrate group at 12 weeks. Further, both treatment groups had higher scores compared to the normal saline group. In comparison to the latter, the groups treated with IPFP-MSCs and IPFP cell concentrates showed considerably better cartilage regeneration. Overall, IPFP-MSCs represent an effective therapeutic strategy for stimulating articular cartilage regeneration. Further, due to the simple, cost-effective, nonenzymatic, and safe preparation process, IPFP cell concentrates may represent an effective alternative to stem cell-based therapy in the clinic.

Improved accumulation of TGF-ß by photopolymerized chitosan/silk protein bio-hydrogel matrix to improve differentiations of mesenchymal stem cells in articular cartilage tissue regeneration.

Articular cartilage regeneration is a challenging process due to its inadequate ability of self-recovering biological mechanisms. The progresses of cartilage tissue engineering is supported to overwhelmed the repairing difficulties and degenerative diseases. The main goal of the present study is to design biomaterials with suitable physico-chemical, mechanical and biological properties for the carrier of growth factor and improving differentiation of mesenchymal stem cell into damaged cartilage tissues. Herein, TGF-ß loaded hydrogel network was prepared through the chemical interactions between vinyl group of natural polymers. Fourier-transform infrared spectroscopy results show the characteristic peaks at 3074 cm-1, 1713 cm-1, and 810 cm-1, which confirm the existence of the vinyl group and successful formation of maleoyl functionalized Chitosan (MCh). The obtained MCh was freely dissolved in the distilled water up to 8% (w/v). X-ray photoelectron spectroscopy survey spectral results show a peak at 289.0 eV which revealed that the OCO and DS were 1.2% and also evidenced the methacryl substitution of Silk fibroin (SF) nanoformulations. The weight loss and mechanical test were analyzed and the results showed that MSF acts as a foremost crosslinking point with MCh through the reaction between the methacrylate groups of MSF and maleoyl groups of MCh which led to enhancing the density and improved the compressive strength. The maximum drug release activity was recorded in the TGF-ß loaded MCh@MSF hydrogel compared to bare MCh hydrogel. Further, the TGF-ß loaded MCh@ MSF hydrogel exhibited the cell viability percentage nearly at 79-102% for MC3T3-E1 and 88-104% for BMDSCs. Similarly, the TGF-ß loaded MCh@MSF exhibited the highest inhibitory activity against E. coli (83%) than S. aureus (67%). Overall, this study concluded the TGF-ß loaded MCh@MSF showed better biocompatibility and could be utilized in the field of cartilage tissue engineering.

Extracellular HMGB-1 activates inflammatory signaling in tendon cells and tissues.

BACKGROUND: Increasing evidence indicates that secretion of high mobility group box 1 protein (HMGB-1) is functionally associated with tendinopathy development. However, the underlying effect and mechanism of extracellular HMGB-1 on tendon cells are unclear. METHODS: We tested the effect of exogenous HMGB-1 on cell growth, migration, and inflammatory signaling responses with isolated rat Achilles tendon cells. Also, we studied the role of extracellular HMGB-1, when administrated alone or in combination with mechanical overloading induced by intensive treadmill running (ITR), in stimulating inflammatory effects in tendon tissues. RESULTS: By using in vitro and in vivo models, we show for the first time that exogenous HMGB-1 dose-dependently induces inflammatory reactions in tendon cells and tendon tissue. Extracellular HMGB-1 promoted redistribution of HMGB-1 from the nucleus to the cytoplasm, and activated canonical nuclear factor kappa B (NF-κB) signaling and mitogen-activated protein kinase (MAPK) signaling. Short-term administration of HMGB-1 induced hyper-cellularity of rat Achilles tendon tissues, accompanied with enhanced immune cell infiltration. Additional ITR to HMGB-1 treatment worsens these responses, and application of HMGB-1 specific inhibitor glycyrrhizin (GL) completely abolishes such inflammatory effects in tendon tissues. CONCLUSION: Collectively, these results confirm that HMGB-1 plays key roles in the induction of tendinopathy. Our findings improve the understanding of the molecular and cellular mechanisms during tendinopathy development, and provide essential information for potential targeted treatments of tendinopathy.

The mineralization, drug release and in vivo bone defect repair properties of calcium phosphates/PLA modified tantalum scaffolds.

Calcium phosphate based biomaterials have been widely studied in biomedical areas. Herein, amorphous calcium phosphate (ACP) nanospheres and hydroxyapatite (HA) nanorods were separately prepared and used for coating tantalum (Ta) scaffolds with a polymer of polylactide (PLA). We have found that different crystal phases of calcium phosphate coated on Ta scaffolds displayed different effects on the surface morphologies, mineralization and bovine serum albumin (BSA) release. The ACP-PLA and HA-PLA coated on Ta scaffold were more favorable for in vitro mineralization than bare and PLA coated Ta scaffolds, and resulted in a highly hydrophilic surfaces. Meanwhile, the osteoblast-like cells (MG63) showed favorable properties of adhesion and spreading on both ACP-PLA and HA-PLA coated Ta scaffolds. The ACP-PLA and HA-PLA coated Ta scaffolds showed a high biocompatibility and potential applications for in vivo bone defect repair.