Physalin A Inhibits MAPK and NF-κB Signal Transduction Through Integrin αVβ3 and Exerts Chondroprotective Effect

Osteoarthritis (OA) is a common articular ailment presented with cartilage loss and destruction that is common observed in the elderly population. Physalin A (PA), a natural bioactive withanolide, exerts anti-inflammatory residences in more than a few diseases; however, little is known about its efficacy for OA treatment. Here, we explored the therapeutic effects and potential mechanism of PA in mouse OA. After the in vitro administration of PA, the expression of inflammation indicators including inducible nitric oxide synthase and cyclooxygenase-2 was low, indicating that PA could alleviate the IL-1β-induced chondrocyte inflammation response. Moreover, PA reduced IL-1β-induced destruction of the extracellular matrix by upregulating the gene expression of anabolism factors, including collagen II, aggrecan, and sry-box transcription factor 9, and downregulating the gene expression of catabolic factors, including thrombospondin motif 5 and matrix metalloproteinases. In addition, the chondroprotective effect of PA was credited to the inhibition of mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) signaling pathways. Furthermore, in vivo experiments showed that intra-articular injection of PA could alleviate cartilage destruction in a mouse OA model. However, the anti-inflammatory, anabolism enhancing, catabolism inhibiting, and MAPK and NF-κB signaling pathway inhibiting properties of PA on IL-1β-induced chondrocytes could be reversed when integrin αVβ3 is knocked down by siRNA. In conclusion, our work demonstrates that PA exhibits a chondroprotective effect that may be mediated by integrin αVβ3. Thus, PA or integrin αVβ3 might be a promising agent or molecular target for the treatment of OA.

The Long-term Chondroprotective Effect of Meniscal Allograft Transplant: A 10- to 14-Year Follow-up Study

Background: The long-term chondroprotective effect of meniscal allograft transplant (MAT) and its superiority over meniscectomy have rarely been reported. Hypothesis: MAT would reduce osteoarthritis (OA) progression when compared with the meniscus-deficient knee. Graft extrusion distance would strongly affect the chondroprotective effect of the MAT. Study Design: Cohort study; Level of evidence, 3. Methods: A total of 17 knees receiving MAT were followed up as the MAT group. The MAT group was further divided into the nonextrusion subgroup (n = 9) and the extrusion subgroup (n = 8) according to 3-mm extrusion on the magnetic resonance imaging (MRI) coronal section. A further 26 consecutive patients receiving meniscectomy in the same period were followed up as the ME group. The healthy control group consisted of healthy contralateral legs chosen from the MAT and ME groups (n = 27). Joint space width (JSW) narrowing was measured on radiographs. Three-dimensional MRI with a T2 mapping sequence was used to quantitatively analyze cartilage degeneration and meniscal allograft extrusion in 5 directions (0°, 45°, 90°, 135°, and 180°). The cartilage degeneration index (CDI) was calculated according to the size and degree of the chondral lesions on MRI scans. The correlation between the CDI increase and the extrusion distance was analyzed. Results: The mean follow-up time was 11.3 years (range, 10-14 years). The MAT group had moderate superiority in chondral protection with less JSW narrowing (0.58 ± 0.66 mm) and CDI increase (1132 ± 1589) compared with the ME group (JSW narrowing: 1.26 ± 1.13 mm, P = .025; CDI increase: 2182 ± 1958, P = .079). The JSW narrowing (0.71 ± 0.80 mm; P = .186) and CDI increase (2004 ± 1965; P = .830) of the extrusion subgroup were close to those of the ME group, demonstrating that a 3-mm extrusion led to complete loss of the meniscal chondroprotective effect. The nonextrusion group had significantly less JSW narrowing (0.48 ± 0.48 mm; P = .042) and CDI increase (358 ± 249; P = .011) than the ME group. The JSW narrowing of the healthy control group was 0.22 ± 0.27 mm. The cartilage T2 values of the extrusion subgroup were similar to those of the ME group, with more OA features, whereas the T2 values of the nonextrusion subgroup were closer to those of the healthy control group. The extrusion distance in the 90° direction ( P = .002) and the follow-up time ( P = .019) significantly affected the CDI increase in the multivariate regression model. The average extrusion distance in the 45°, 90°, and 135° directions better predicted chondroprotection compared with the other individual directions. Conclusion: MAT had moderate advantages in chondroprotection compared with meniscectomy in the long term. Graft extrusion distance strongly affected the chondroprotective effect of MAT. The chondroprotective effect of the nonextruded meniscal allograft was close to that of the native meniscus, whereas the allografts with an extrusion >3 mm completely lost their function after meniscectomy.

POS0374 SENESCENCE DID NOT ALTER THE CHONDROPROTECTIVE EFFECT OF EXTRACELLULAR VESICLES FROM MESENCHYMAL STROMAL CELLS

Background:Age is the most important risk factor in degenerative osteoarthritis (OA) and is associated with the accumulation of senescent cells that contribute to functional decline of joint. We previously demonstrated that extracellular vesicles (EVs) from mesenchymal stromal cells (MSCs) largely mediate the therapeutic effect of parental cells in OA.Objectives:Here, we assessed the impact of senescence on the characteristics of EVs from adipose tissue-derived MSCs (ASC-EVs) and their properties in an in vitro model of OAMethods:ASCs were induced to senescence using 25µM etoposide for 24 hours. Senescence was assessed by quantifying proliferation rate, SA-βGal activity, nuclear γH2AX foci number, phalloidin staining and expression of cyclin dependent kinase inhibitors (CDKI) (RT-qPCR). ASC-EVs were isolated by differential ultracentrifugation and characterized by size, concentration, total protein content, structure (cryo-TEM) and immunophenotype. In vitro OA model used chondrocytes isolated from OA patients, which were stimulated with IL1β for 48h before culture with ASCs or ASC-EVs for 7 days. Expression of chondrocytic and inflammatory markers was quantified by RT-qPCR and SASP factors were quantified by ELISA in supernatants.Results:Senescence-induced ASCs experienced growth arrest and increase of SA-βGal staining, of p21 CDKI expression, of nuclear γH2AX foci, of stress fibers and of several SASP factors (IL6, IL8, MMP3) confirming the expression of main senescence features. Senescent ASCs produced 4-fold more EVs than healthy ASCs and senescent ASC-EVs were larger. In vitro, both healthy and senescent ASCs decreased fibrotic markers (type III COLLAGEN), catabolic and hypertrophic markers (MMP3, MMP13, AP) and increased COX2 expression in OA chondrocytes. By contrast, healthy ASCs decreased the expression of IL6 while senescent ASCs highly increased IL6. Looking at the role of ASC-EVs on OA chondrocytes, we found out that both healthy and senescent ASC-EVs were able to increase the expression of AGG and type II COLLAGEN while they decreased the expression of MMP13, AP, type X COLLAGEN, HMOX1 and IL6. Finally, healthy and senescent ASC-EVs decreased the number of SA-βGal positive chondrocytes but did not impact the expression of p21 in IL1β-induced chondrocytes.Conclusion:Our results indicated a chondroprotective effect of ASC-EVs, independently of the senescent state of parental cells and suggested that EVs might act through different mechanisms than ASCs, which warrants further investigationDisclosure of Interests:Jérémy Boulestreau: None declared, Marie Maumus Employee of: Bauerfeind France, Pauline Rozier: None declared, Christian Jorgensen Shareholder of: Medxcell sciences, Consultant of: Medxcell sciences, Daniele Noel Shareholder of: Medxcell sciences, Consultant of: Medxcell sciences

The Role of PPARγ in Hyperglycemia-induced Deleterious Effect on Chondrocytes

Aims: There is a well-established link between OA and diabetes, and study have shown that hyperglycemia might play an important role in the occurrence and development of OA. Accumulative evidence suggested that PPARγ was involved in AGEs-related disease, including diabetes and OA. The study was designed to investigate the effects of hyperglycemia on the expression of PPARγ in chondrocytes and whether PPARγ agonist pioglitazone had a chondroprotective effect.Main methods: Primary human chondrocytes were incubated with different concentration of glucose medium(5.5mM-30mM) in the presence or absence of PPARγ agonist pioglitazone. The AGEs formation level in chondrocytes culture medium was detected by AGEs specific ELISA kits. The expression of IL-1, MMP-13, TNF-α, PPARγ was determined by western blotting and real-time PCR.Key findings: The AGEs fomation level was time-dependently and dose-dependently increased in chondrocyte culture media. Hyperglycemia could enhance the expression of IL-1β, TNF-α, MMP-13, but the level of PPARγ was decreased in a time-dependent and dose-dependent manner, which was inhibited by PPARγ agonist pioglitazone. Noteworthy, the maximum effect was found to at 20mM glucose medium for 24h.Significance: Hyperglycemia could increase the AGEs formation level and induce inflammatory response and matrix degradation reaction in chondrocytes. PPARγ agonists pioglitazone had a chondroprotective effect via inhibit inflammatory response and matrix degradation reaction. PPARγ could be a potential target for pharmacologic intervention in the treatment of diabetic-induced OA.