Chondrogenic Differentiation of Mouse Bone Marrow Mesenchymal Stem Cells Induced by Cartilage-derived Morphogenetic Protein-2 In Vitro

To study the cartilage differentiation of mouse mesenchymal stem cells （MSCs） induced by cartilage-derived morphogenetic proteins-2 in vitro, the MSCs were isolated from mouse bone marrow and cultured in vitro. The cells in passage 3 were induced into chondrogenic differentiation with different concentrations of recombinant human cartilage-derived morphogenetic proteins-2 （0, 10, 20, 50 and 100 ng/mL）. After 14 days of induction, morphology of cells was observed under phase-contrast microscope. Collagen Ⅱ mRNA and protein were examined with RT-PCR, Western blotting and immunocytochemistry respectively and the sulfate glycosaminoglycan was measured by Alcian blue staining. RT-PCR showed that CDMP-2 could promote expression of collagen Ⅱ mRNA in an dose-dependant manner, especially at the concentration of 50 ng/mL and 100 ng/mL. Immunocytochemistry and Western blotting revealed a similar change. Alcian blue staining exhibited deposition of typical cartilage extracellular matrix. Our results suggest that mouse bone marrow mesencymal stem cells can differentiate into chondrogenic phonotype with the induction of CDMP-2 in vitro, which provides a basis for further research on the role of CDMP-2 in chondrogenesis.

Stromal cell-derived factor-1α regulates chondrogenic differentiation via activation of the Wnt/β-catenin pathway in mesenchymal stem cells

BACKGROUND Mesenchymal stem cells(MSCs)have been applied to treat degenerative articular diseases,and stromal cell-derived factor-1α(SDF-1α)may enhance their therapeutic efficacy.However,the regulatory effects of SDF-1αon cartilage differentiation remain largely unknown.Identifying the specific regulatory effects of SDF-1αon MSCs will provide a useful target for the treatment of degenerative articular diseases.AIM To explore the role and mechanism of SDF-1αin cartilage differentiation of MSCs and primary chondrocytes.METHODS The expression level of C-X-C chemokine receptor 4(CXCR4)in MSCs was assessed by immunofluorescence.MSCs treated with SDF-1αwere stained for alkaline phosphatase(ALP)and with Alcian blue to observe differentiation.Western blot analysis was used to examine the expression of SRY-box transcription factor 9,aggrecan,collagen II,runt-related transcription factor 2,collagen X,and matrix metalloproteinase(MMP)13 in untreated MSCs,of aggrecan,collagen II,collagen X,and MMP13 in SDF-1α-treated primary chondrocytes,of glycogen synthase kinase 3β(GSK3β)p-GSK3βandβ-catenin expression in SDF-1α-treated MSCs,and of aggrecan,collagen X,and MMP13 in SDF-1α-treated MSCs in the presence or absence of ICG-001(SDF-1αinhibitor).RESULTS Immunofluorescence showed CXCR4 expression in the membranes of MSCs.ALP stain was intensified in MSCs treated with SDF-1αfor 14 d.The SDF-1αtreatment promoted expression of collagen X and MMP13 during cartilage differentiation,whereas it had no effect on the expression of collagen II or aggrecan nor on the formation of cartilage matrix in MSCs.Further,those SDF-1α-mediated effects on MSCs were validated in primary chondrocytes.SDF-1αpromoted the expression of p-GSK3βandβ-catenin in MSCs.And,finally,inhibition of this pathway by ICG-001(5μmol/L)neutralized the SDF-1α-mediated up-regulation of collagen X and MMP13 expression in MSCs.CONCLUSION SDF-1αmay promote hypertrophic cartilage differentiation in MSCs by activating the Wnt/β-catenin pathway.These findings provide further evidence for the use of MSCs and SDF-1αin the treatment of cartilage degeneration and osteoarthritis.

Comparison of Chondrogenic Ability between Mesenchymal Stem Cells and Buffy Coat <i>in Vitro</i>

Objective: In vitro comparison of chondrogenic differentiation ability of bone marrow extracted mesenchymal stem cells (MSCs) and buffy coat. Methods: MSCs of New Zealand white rabbits were cultured in vitro and adherent cells were passaged. The cells were inoculated on polyglycolic acid (PGA) scaffold (3 mm in diameter and 2 mm in height) liquid for 21 days. Under the same conditions, without subculture, buffy coat was directly inoculated into the cell scaffold. The degree of chondrogenic differentiation was compared by Safranin-Ostaining, histological scoring and biochemical functional detection. Results: The chondrogenic differentiation ability of the buffy coat group was better than that of the MSC group. Safranin-Ostain was stronger in buffy coat group than in MSC group. The Bern Score was also higher in the buffy coat group than in the MSC group and the total amount of sulfated glycosaminoglycans (GAGs) in buffy coat group was higher than that in MSC group. Conclusion: The chondrogenic differentiation ability of buffy coat is higher than that of MSC. Through this result, it was found buffy coat can also differentiate into sub-cartilage without stimulating growth factor. Thus, buffy coat can make a great source in artificial cartilage engineering.

Chondrogenic differentiation of rat bone marrow mesenchymal stem cells induced by puerarin and tetrandrine

Objective: This study aims to clarify the effect of the active components puerarin and tetrandrine on the chondrogenic differentiation of bone marrow mesenchymal stem cells(BMSCs).Methods: Using network pharmacology, protein targets of puerarin and tetrandrine were predicted, and a database of cartilage formation targets was established. The protein target information related to disease was then collected, and the drug-targeting network was constructed by analyzing the protein–protein interactions. Genes related to chondrogenesis induced by puerarin and tetrandrine and chondroblast differentiation signaling pathways were searched. Finally, potential drug-and disease-related genes,as well as proteins, were screened and verified using real-time RT-PCR and western blotting.Results: Network pharmacological studies have shown that puerarin and tetrandrine are involved in BMSCs cartilage differentiation. The experimental results showed that puerarin and tetrandrine could regulate the expression of cartilage differentiation-related genes and proteins. Puerarin increased the protein expression of COL2 A1, COL10 A1, MMP13, and SOX-9,as well as the gene expression of Col2 a1, Mmp13, Tgfb1, and Sox-9. Tetrandrine increased the protein expression of COL2 A1,COL10 A1, MMP13, and SOX-9, as well as the gene expression of Col10 a1, Tgfb1, Sox-9, and Acan. The combination of puerarin and tetrandrine increased the protein expression of COL2 A1, COL10 A1, MMP13, and SOX-9 and the gene expression of Col2 a1,Col10 a1, Sox-9, and Acan.Conclusions: Puerarin, tetrandrine, and their combination can promote the proliferation of BMSCs and induce their differentiation into chondrocytes, and they are thus expected to be inducers of chondrogenic differentiation. These results suggest that puerarin and tetrandrine have potential therapeutic effects on osteoarthritis.

Stimulating factors for regulation of osteogenic and chondrogenic differentiation of mesenchymal stem cells

Mesenchymal stem cells(MSCs),distributed in many tissues in the human body,are multipotent cells capable of differentiating in specific directions.It is usually considered that the differentiation process of MSCs depends on specialized external stimulating factors,including cell signaling pathways,cytokines,and other physical stimuli.Recent findings have revealed other underrated roles in the differentiation process of MSCs,such as material morphology and exosomes.Although relevant achievements have substantially advanced the applicability of MSCs,some of these regulatory mechanisms still need to be better understood.Moreover,limitations such as long-term survival in vivo hinder the clinical application of MSCs therapy.This review article summarizes current knowledge regarding the differentiation patterns of MSCs under specific stimulating factors.

Chondrogenic differentiation of human bone mesenchymal stem cells treated with growth differentiation factor 5 under hypoxia

Objective To explore the feasibility and effectiveness of the self-assembly cartilage tissue engineered with chondrogenically differentiated human bone mesenchymal stem cells (hBMCs) induced by growth differentiation factor-5 (GDF-5)

Tumor necrosis factor-αinhibition restores matrix formation by human adipose-derived stem cells in the late stage of chondrogenic differentiation

BACKGROUND Cartilage tissue engineering is a promising strategy for treating cartilage damage.Matrix formation by adipose-derived stem cells(ADSCs),which are one type of seed cell used for cartilage tissue engineering,decreases in the late stage of induced chondrogenic differentiation in vitro,which seriously limits research on ADSCs and their application.AIM To improve the chondrogenic differentiation efficiency of ADSCs in vitro,and optimize the existing chondrogenic induction protocol.METHODS Tumor necrosis factor-alpha(TNF-α)inhibitor was added to chondrogenic culture medium,and then Western blotting,enzyme linked immunosorbent assay,immunofluorescence and toluidine blue staining were used to detect the cartilage matrix secretion and the expression of key proteins of nuclear factor kappa-B(NF-κB)signaling pathway.RESULTS In this study,we found that the levels of TNF-αand matrix metalloproteinase 3 were increased during the chondrogenic differentiation of ADSCs.TNF-αthen bound to its receptor and activated the NF-κB pathway,leading to a decrease in cartilage matrix synthesis and secretion.Blocking TNF-αwith its inhibitors etanercept(1μg/mL)or infliximab(10μg/mL)significantly restored matrix formation.CONCLUSION Therefore,this study developed a combination of ADSC therapy and targeted anti-inflammatory drugs to optimize the chondrogenesis of ADSCs,and this approach could be very beneficial for translating ADSC-based approaches to treat cartilage damage.

Chondrogenic commitment of human umbilical cord blood and umbilical cord-derived mesenchymal stem cells induced by the supernatant of chondrocytes:A comparison study

Background:Native cartilage has low capacity for regeneration because it has very few progenitor cells.Human umbilical cord blood-derived mesenchymal stem cells(hUCB-MSCs)and human umbilical cord-derived MSCs(hUC-MSCs)have been employed as promising sources of stem cells for cartilage injury repair.Reproduction of hyaline cartilage from MSCs remains a challenging endeavor.The paracrine factors secreted by chondrocytes possess the capability to induce chondrogenesis from MSCs.Methods:The conditioned medium derived from chondrocytes was utilized to induce chondrogenic differentiation of hUCB-MSCs and hUC-MSCs.The expression levels of collagen type I alpha 1 chain(Col1a1),collagen type II alpha 1 chain(Col2a1),and SRY-box transcription factor 9(SOX9)were assessed through quantitative real-time polymerase chain reaction(qRT-PCR),Western blot(WB),and immunofluorescence(IF)assays.To elucidate the mechanism of differentiation,the concentration of transforming growth factor-β1(TGF-β1)in the conditioned medium of chondrocytes was quantified using enzyme-l inked immunosorbent assay(ELISA).Meanwhile,the viability of cells was assessed using Cell Counting Kit-8(CCK-8)assays.Results:The expression levels of Col2a1 and SOX9 were found to be higher in induced hUC-MSCs compared to those in induced hUCB-MSCs.The conditioned medium of chondrocytes contained TGF-β1.The CCK-8 assays revealed that the proliferation rate of hUC-MSCs was significantly higher compared to that of hUCB-MSCs.Conclusions:The chondrogenic potential and proliferation capacity of hUC-MSCs surpass those of hUCB-MSCs,thereby establishing hUC-MSCs as a superior source of seed cells for cartilage tissue engineering.

Role of integrin b1 and tenascin C mediate TGF-SMAD2/3 signaling in chondrogenic differentiation of BMSCs induced by type I collagen hydrogel

Cartilage defects may lead to severe degenerative joint diseases.Tissue engineering based on type I collagen hydrogel that has chondrogenic potential is ideal for cartilage repair.However,the underlying mechanisms of chondrogenic differentiation driven by type I collagen hydrogel have not been fully clarified.Herein,we explored potential collagen receptors and chondrogenic signaling pathways through bioinformatical analysis to investigate the mechanism of collagen-induced chondrogenesis.Results showed that the super enhancer-related genes induced by collagen hydrogel were significantly enriched in the TGF-b signaling pathway,and integrin-b1(ITGB1),a receptor of collagen,was highly expressed in bone marrow mesenchymal stem cells(BMSCs).Further analysis showed genes such as COL2A1 and Tenascin C(TNC)that interacted with ITGB1 were significantly enriched in extracellular matrix(ECM)structural constituents in the chondrogenic induction group.Knockdown of ITGB1 led to the downregulation of cartilage-specific genes(SOX9,ACAN,COL2A1),SMAD2 and TNC,as well as the downregulation of phosphorylation of SMAD2/3.Knockdown of TNC also resulted in the decrease of cartilage markers,ITGB1 and the SMAD2/3 phosphorylation but overexpression of TNC showed the opposite trend.Finally,in vitro and in vivo experiments confirmed the involvement of ITGB1 and TNC in collagen-mediated chondrogenic differentiation and cartilage regeneration.In summary,we demonstrated that ITGB1 was a crucial receptor for chondrogenic differentiation of BMSCs induced by collagen hydrogel.It can activate TGF-SMAD2/3 signaling,followed by impacting TNC expression,which in turn promotes the interaction of ITGB1 and TGF-SMAD2/3 signaling to enhance chondrogenesis.These may provide concernful support for cartilage tissue engineering and biomaterials development.

Cartilage-like protein-polysaccharide hybrid hydrogel for enhancing chondrogenic differentiation of bone marrow mesenchymal stem cells

The regeneration of articular cartilage posed a formidable challenge due to the restricted treatment efficacy of exist-ing therapies.Scaffold-based tissue engineering emerges as a promising avenue for cartilage reconstitution.However,most scaffolds exhibit inadequate mechanical characteristics,poor biocompatibility,or absent cell adhesion sites.In this study,cartilage-like protein-polysaccharide hybrid hydrogel based on DOPA-modified hyaluronic acid,bovine type Ⅰ collagen(Col Ⅰ),and recombinant humanized type Ⅱ collagen(rhCol Ⅱ),denoted as HDCR.HDCR hydrogels possessed the advantage of injectability and in situ crosslinking through pH adjustment.Moreover,HDCR hydrogels exhibited a manipulable degradation rate and favorable biocompatibility.Notably,HDCR hydrogels significantly induced chondrogenic differentiation of rabbit bone marrow mesenchymal stem cells in vitro,as demonstrated by the upregulation of crucial chondrogenic genes(type Ⅱ collagen,aggrecan)and the abundant accumulation of glycosaminoglycan.This approach presented a strategy to manufacture injectable,biodegradable scaffolds based on cartilage-like protein-polysaccharide polymers,offering a minimally invasive solution for cartilage repair.

Sox9 augments BMP2-induced chondrogenic differentiation by downregulating Smad7 in mesenchymal stem cells(MSCs)

Cartilage injuries caused by arthritis or trauma pose formidable challenges for effective clinical management due to the limited intrinsic proliferative capability of chondrocytes.Autologous stem cell-based therapies and transgene-enhanced cartilage tissue engineering may open new avenues for the treatment of cartilage injuries.Bone morphogenetic protein 2(BMP2)induces effective chondrogenesis of mesenchymal stem cells(MSCs)and can thus be explored as a potential therapeutic agent for cartilage defect repair.However,BMP2 also induces robust endochondral ossification.Although the precise mechanisms through which BMP2 governs the divergence of chondrogenesis and osteogenesis remain to be fully understood,blocking endochondral ossification during BMP2-induced cartilage formation may have practical significance for cartilage tissue engineering.Here,we investigate the role of Sox9-donwregulated Smad7 in BMP2-induced chondrogenic differentiation of MSCs.We find that overexpression of Sox9 leads to a decrease in BMP2-induced Smad7 expression in MSCs.Sox9 inhibits BMP2-induced expression of osteopontin while enhancing the expression of chondrogenic marker Col2a1 in MSCs.Forced expression of Sox9 in MSCs promotes BMP2-induced chondrogenesis and suppresses BMP2-induced endochondral ossification.Constitutive Smad7 expression inhibits BMP2-induced chondrogenesis in stem cell implantation assay.Mouse limb explant assay reveals that Sox9 expands BMP2-stimulated chondrocyte proliferating zone while Smad7 promotes BMP2-intitated hypertrophic zone of the growth plate.Cell cycle analysis indicates that Smad7 induces significant early apoptosis in BMP2-stimulated MSCs.Taken together,our results strongly suggest that Sox9 may facilitate BMP2-induced chondrogenesis by downregulating Smad7,which can be exploited for effective cartilage tissue engineering.

Tetrahedral framework nucleic acids enhance the chondrogenic potential of human umbilical cord mesenchymal stem cells via the PI3K/AKT axis

The field of regenerative medicine faces a notable challenge in terms of the regeneration of articular cartilage.Without proper treatment,it can lead to osteoarthritis.Based on the research findings,human umbilical cord mesenchymal stem cells(hUMSCs)are considered an excellent choice for regenerating cartilage.However,there is still a lack of suitable biomaterials to control their ability to self-renew and differentiate.To address this issue,in this study using tetrahedral framework nucleic acids(tFNAs)as a new method in an in vitro culture setting to manage the behaviour of hUMSCs was proposed.Then,the influence of tFNAs on hUMSC proliferation,migration and chondrogenic differentiation was explored by combining bioinformatics methods.In addition,a variety of molecular biology techniques have been used to investigate deep molecular mechanisms.Relevant results demonstrated that tFNAs can affect the transcriptome and multiple signalling pathways of hUMSCs,among which the PI3K/Akt pathway is significantly activated.Furthermore,tFNAs can regulate the expression levels of multiple proteins(GSK3β,RhoA and mTOR)downstream of the PI3K-Akt axis to further enhance cell proliferation,migration and hUMSC chondrogenic differentiation.tFNAs provide new insight into enhancing the chondrogenic potential of hUMSCs,which exhibits promising potential for future utilization within the domains of AC regeneration and clinical treatment.

Ion elemental-optimized layered double hydroxide nanoparticles promote chondrogenic differentiation and intervertebral disc regeneration of mesenchymal stem cells through focal adhesion signaling pathway

Chronic low back pain and dyskinesia caused by intervertebral disc degeneration(IDD)are seriously aggravated and become more prevalent with age.Current clinical treatments do not restore the biological structure and inherent function of the disc.The emergence of tissue engineering and regenerative medicine has provided new insights into the treatment of IDD.We synthesized biocompatible layered double hydroxide(LDH)nanoparticles and optimized their ion elemental compositions to promote chondrogenic differentiation of human umbilical cord mesenchymal stem cells(hUC-MSCs).The chondrogenic differentiation of LDH-treated MSCs was validated using Alcian blue staining,qPCR,and immunofluorescence analyses.LDH-pretreated hUC-MSCs were differentiated prior to transplantation into the degenerative site of a needle puncture IDD rat model.Repair and regeneration evaluated using X-ray,magnetic resonance imaging,and tissue immunostaining 4-12 weeks after transplantation showed recovery of the disc space height and integrated tissue structure.Transcriptome sequencing revealed significant regulatory roles of the extracellular matrix(ECM)and integrin receptors of focal adhesion signaling pathway in enhancing chondrogenic differentiation and thus prompting tissue regeneration.The construction of ion-specific LDH nanomaterials for in situ intervertebral disc regeneration through the focal adhesion signaling pathway provides theoretical basis for clinical transformation in IDD treatment.

Chondrogenic medium in combination with a c-Jun N-terminal kinase inhibitor mediates engineered cartilage regeneration by regulating matrix metabolism and cell proliferation

Cartilage tissue engineering is a promising strategy for repairing cartilage defects.However,achieving satisfactory cartilage regeneration in vitro and maintaining its stability in vivo remains a challenge.The key to achieving this goal is establishing an efficient cartilage regeneration culture system to retain sufficient active cells with physiological functions,generate abundant cartilage extracellular matrix(ECM)and maintain a low level of cartilage ECM degradation.The current chondrogenic medium(CM)can effectively promote cartilage ECM production;however,it has a negative effect on cell proliferation.Meanwhile,the specific c-Jun N-terminal kinase pathway inhibitor SP600125 promotes chondrocyte proliferation but inhibits ECM synthesis.Here,we aimed to construct a three-dimensional cartilage regeneration model using a polyglycolic acid/polylactic acid scaffold in combination with chondrocytes to investigate the effect of different culture modes with CM and SP600125 on in vitro cartilage regeneration and their long-term outcomes in vivo systematically.Our results demonstrate that the long-term combination of CM and SP600125 made up for each other and maximized their respective advantages to obtain optimal cartilage regeneration in vitro.Moreover,the long-term combination achieved stable cartilage regeneration after implantation in vivo with a relatively low initial cell-seeding concentration.Therefore,the long-term combination of CM and SP600125 enhanced in vitro and in vivo cartilage regeneration stability with fewer initial seeding cells and thus optimized the cartilage regeneration culture system.

Sulfated GAG mimetic peptide nanofibers enhance chondrogenic differentiation of mesenchymal stem cells in 3D in vitro models

Articular cartilage,which is exposed to continuous repetitive compressive stress,has limited self-healing capacity in the case of trauma.Thus,it is crucial to develop new treatment options for the effective regeneration of the cartilage tissue.Current cellular therapy treatment options are microfracture and autologous chondrocyte implantation;however,these treatments induce the formation of fibrous cartilage,which degenerates over time,rather than functional hyaline cartilage tissue.Tissue engineering studies using biodegradable scaffolds and autologous cells are vital for developing an effective long-term treatment option.3D scaffolds composed of glycosaminoglycan-like peptide nanofibers are synthetic,bioactive,biocompatible,and biodegradable and trigger cell-cell interactions that enhance chondrogenic differentiation of cells without using any growth factors.We showed differentiation of mesenchymal stem cells into chondrocytes in both 2D and 3D culture,which produce a functional cartilage extracellular matrix,employing bioactive cues integrated into the peptide nanofiber scaffold without adding exogenous growth factors.

The effect of reduced glutathione on the chondrogenesis of human umbilical cord mesenchymal stem cells

It has been discussed whether reduced glutathione (GSH) could promote the chondrogenic differentiation ability of human umbilical cord mesenchymal stem cells (hUC-MSCs). hUC-MSCs were isolated from human umbilical cord and their specificity was identified, then induced into cartilage-like cells in chondrogenic induction medium with transforming growth factor beta 1 (TGF-β1), especially with GSH. The morphological change before and after induction was observed through inverted phase contrast microscope, Type II collagen (COL2-A1) and glycosaminoglycan (GAG) were analyzed qualitatively by Toluidine blue and immunofluorescence technique, respectively, the contents of COL2-A1 and GAG were estimated from the determination of hydroxyproline content and Alcian Blue method separately. The mRNA expressions of GAG and COL2-A1 were assayed by real-time fluorescence quantitative PCR. After continuously cultured for 21 days with GSH, Toluidine blue staining and immunofluorescence reaction were all positive in basic induction medium group (group B), basic induction medium +0.5% dimethylsulfoxide (DMSO) group (group BD) and basic induction medium +0.5% DMSO +500 μM GSH group (group BDG). Moreover, compared with group B and group BD, the contents of COL2-A1 and GAG in group BDG relatively increased and the mRNA expression level of COL2-A1 and GAG also comparatively increased (P < 0.05) and both had a significant statistical significance (P < 0.05). So GSH might promote the induction of hUC-MSCs to differentiate into cartilage-like cells.

Chondrosarcoma of the Hand: A Literature Review

Chondrosarcoma (CS) is a malignant neoplasm of mesenchymal origin characterized by the formation of cartilaginous matrix by neoplastic cells. It is very variable in presentation, clinical, genetic and anatomo-pathological features. CS is more often found in the pelvis and the proximal long bones (femur, humerus). Localization to the small bones of the hand is very rare and it represents a diagnostic and therapeutic challenge. CSs are malignancies resistant to both radiation and chemotherapy . Surgical treatment is the only mean available. As opposed to CSs located elsewhere, CS of the hand is characterized by local recurrence and very low metastatic potential. In order to definitely set a diagnosis of chondrosarcoma in this region, comparison of histological and radiological findings is paramount. The principle goal of surgery shall be minimizing functional impairment. This provides the rationale for performing curettage, local adjuvant therapy and bone grafting in low-grade lesions.

Nanosecond pulsed electric fields prime mesenchymal stem cells to peptide ghrelin and enhance chondrogenesis and osteochondral defect repair in vivo

Mesenchymal stem cells(MSCs) are important cell sources in cartilage tissue development and homeostasis,and multiple strategies have been developed to improve MSCs chondrogenic differentiation with an aim of promoting cartilage regeneration.Here we report the effects of combining nanosecond pulsed electric fields(ns PEFs) followed by treatment with ghrelin(a hormone that stimulates release of growth hormone) to regulate chondrogenesis of MSCs.ns PEFs and ghrelin were observed to separately enhance the chondrogenesis of MSCs,and the effects were significantly enhanced when the bioelectric stimulation and hormone were combined,which in turn improved osteochondral tissue repair of these cells within Sprague Dawley rats.We further found that ns PEFs can prime MSCs to be more receptive to subsequent stimuli of differentiation by upregulated Oct4/Nanog and activated JNK signaling pathway.Ghrelin initiated chondrogenic differentiation by activation of ERK1/2 signaling pathway,and RNA-seq results indicated 243 genes were regulated,and JAK-STAT signaling pathway was involved.Interestingly,the sequential order of applying these two stimuli is critical,with ns PEFs pretreatment followed by ghrelin enhanced chondrogenesis of MSCs in vitro and subsequent cartilage regeneration in vivo,but not vice versa.This synergistic prochondrogenic effects provide us new insights and strategies for future cell-based therapies.

Tetrahedral framework nucleic acids promote the biological functions and related mechanism of synovium-derived mesenchymal stem cells and show improved articular cartilage regeneration activity in situ

Many recent studies have shown that joint-resident mesenchymal stem cells(MSCs)play a vital role in articular cartilage(AC)in situ regeneration.Specifically,synovium-derived MSCs(SMSCs),which have strong chondrogenic differentiation potential,may be the main driver of cartilage repair.However,both the insufficient number of MSCs and the lack of an ideal regenerative microenvironment in the defect area will seriously affect the regeneration of AC.Tetrahedral framework nucleic acids(tFNAs),notable novel nanomaterials,are considered prospective biological regulators in biomedical engineering.Here,we aimed to explore whether tFNAs have positive effects on AC in situ regeneration and to investigate the related mechanism.The results of in vitro experiments showed that the proliferation and migration of SMSCs were significantly enhanced by tFNAs.In addition,tFNAs,which were added to chondrogenic induction medium,were shown to promote the chondrogenic capacity of SMSCs by increasing the phosphorylation of Smad2/3.In animal models,the injection of tFNAs improved the therapeutic outcome of cartilage defects compared with that of the control treatments without tFNAs.In conclusion,this is the first report to demonstrate that tFNAs can promote the chondrogenic differentiation of SMSCs in vitro and enhance AC regeneration in vivo,indicating that tFNAs may become a promising therapeutic for AC regeneration.

Synthesis of photo-reactive poly(vinyl alcohol)and construction of scaffold-free cartilage like pellets in vitro

Photo-reactive poly(vinyl alcohol)(PRPVA)was synthesized by introduction of phenyl azido groups into poly(vinyl alcohol)(PVA)and applied for surface modification.PRPVA was grafted onto cell culture plate surface homogeneously or in a micropattern.Human mesenchymal stem cells(hMSCs)cultured on cell culture plate surface and PVA-modified surface showed different behaviors.Cells adhered and spread well on cell culture plate surface,while they did not adhere on PVA-grafted surface at all.When hMSCs were cultured on PVA-micropatterned surface,they formed a cell micropattern.Cells formed pellets after cultured on PVA homogeneously modified surface in chondrogenic induction medium for 2 weeks.The pellets were positively stained by hematoxylin/eosin,safranin-O/fast green and toluidin blue,and they were also stained brown by Type II collagen and proteoglycan immunohistological staining.Real-time PCR analysis was conducted to investigate the expression of colI,colII,colX,aggrecan and sox9 mRNA.Results of gene expression were in agreement with those of histological and immunohistological observations.These results indicated that hMSCs cultured on PVA-modified surface performed chondrogenic differentiation,and it was possible to construct scaffold-free cartilage like pellets with PVA-modified surface in vitro.