Anti‑inflammatory and anti‑catabolic effect of non‑animal stabilized hyaluronic acid and mesenchymal stem cell‑conditioned medium in an osteoarthritis coculture model

  • Authors:
    • Mario Simental‑Mendía
    • Sonia Amelia Lozano‑Sepúlveda
    • Vanessa Pérez‑Silos
    • Lizeth Fuentes‑Mera
    • Herminia Guadalupe Martínez‑Rodríguez
    • Carlos Alberto Acosta‑Olivo
    • Víctor Manuel Peña‑Martínez
    • Félix Vilchez‑Cavazos
  • View Affiliations

  • Published online on: February 26, 2020
  • Pages: 2243-2250
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Previous clinical studies have reported the clinical effectiveness of non‑animal stabilized hyaluronic acid (NASHA) and adipose‑derived mesenchymal stromal/stem cells (MSC) in the treatment of knee osteoarthritis (OA). Unlike MSC secreted mediators, in vitro anti‑inflammatory effects of NASHA have not been evaluated. We aimed to evaluate and compare the anti‑inflammatory effect of NASHA and MSC conditioned medium (stem cell‑conditioned medium; SC‑CM), in an explant‑based coculture model of OA. Cartilage and synovial membrane from seven patients undergoing total knee arthroplasty were used to create a coculture system. Recombinant IL‑1β was added to the cocultures to induce inflammation. Four experimental groups were generated: i) Basal; ii) IL‑1β; iii) NASHA (NASHA + IL‑1β); and iv) SC‑CM (SC‑CM + IL‑1β). Glycosaminoglycans (GAG) released in the culture medium and of nitric oxide (NO) production were quantified. Gene expression in cartilage and synovium of IL‑1β, matrix metallopeptidase 13 (MMP13), ADAM metallopeptidase with thrombospondin type 1 motif 5 (ADAMTS5) and tissue inhibitor of metalloproteinases 1 (TIMP1) was measured by reverse transcription‑quantitative PCR. Media GAG concentration was decreased in cocultures with NASHA and SC‑CM (48 h, P<0.05; 72 h, P<0.01) compared with IL‑1β. Production of NO was significantly lower only in SC‑CM after 72 h (P<0.01). In cartilage, SC‑CM inhibited the expression of IL‑1β, MMP13 and ADAMTS5, while NASHA had this effect only in MMP13 and ADAMTS5. In synovium, SC‑CM decreased the expression level of MMP13 and ADAMTS5, while NASHA only decreased ADAMTS5 expression. Both NASHA and SC‑CM increased TIMP1 expression in cartilage and synovium. Treatments with NASHA and SC‑CM were shown to be a therapeutic option that may help counteract the catabolism produced by the inflammatory state in knee OA. The anti‑inflammatory mediators produced by MSC promote a lower expression of inflammatory targets in our study model.


Osteoarthritis (OA) is a chronic and progressive disease that affects more than 20% of people over 45 years (1). The pathology of knee OA includes progressive articular cartilage loss, sclerotic changes in subchondral bone, and osteophyte formation. The cells in the OA joint change their phenotype as a result of aging and exposure to mechanical stress, oxidative stress, and inflammation (2,3). It is now well-recognized that there is a major inflammatory component to this disease and it is thought that inflammatory mediators may play an important role in joint degradation (4). Particularly, interleukin-1β (IL-1β) is known as a major inducer of articular cartilage extracellular matrix (ECM) degradation, promoting the production and activation of different factors that act as mediators and/or effectors of progressive cartilage loss (5).

To date, there is no treatment that cures OA. Thus, the development of therapeutic alternatives that can prevent or delay the destruction of cartilage and tissues involved in the knee joint or stimulate its adequate repair is required (6). The usefulness of treatments focused on the repair of focal lesions is very limited in OA, because more generalized cartilage defects and joint inflammation are present. Therefore, intra-articular treatment options seem more viable as therapeutic alternatives for the management of knee OA (7). Some of the advantages of using intra-articular injections are increased local bioavailability, reduced systemic exposure, fewer adverse events, and reduced cost (8).

The intra-articular injection of hyaluronic acid (HA) for knee OA has been widely studied. One of the most recent available products is non-animal stabilized HA (NASHA, Durolane HA®; Bioventus LLC). NASHA is a type of HA that is produced through bacterial synthesis of HA. It is subsequently purified and stabilized in a carefully controlled process involving molecular cross-linking (9). Available clinical evidence supports the use of NASHA as an effective and safe procedure for symptom relief in knee OA (10). However, the anti-inflammatory effect at the cellular and molecular level of this type of HA in knee OA has not been studied.

On the other hand, results from clinical studies suggest a clinical improvement for intra-articular injection of mesenchymal stromal/stem cells (MSC) in knee OA (11,12). Along with their differentiation potential into several lineages, including chondrogenic lineage, MSC influence their micro-environment by secreting trophic mediators (1315). The paracrine capacities of MSC raise the attractive approach of possibly basing future therapies on their secreted factors rather than on the cells themselves.

In vitro stimulation with inflammatory factors provides an option to further use the MSC trophic effects since they need to be activated to exert such action (1618). The effect of the mediators produced by stimulated MSC on inflammation and catabolic processes involved in OA have been tested before (16,17,19). The aim of this study was to evaluate the anti-inflammatory and anti-catabolic effect of the NASHA and compare it with the conditioned media from adipose-derived MSC in an in vitro coculture model of cartilage and synovium.

Materials and methods

Patients and samples

Cartilage and synovial tissue explants were harvested as surgical waste from patients with OA undergoing total knee arthroplasty (n=7) from April 2017 to December 2018. Usage of human tissues was approved by the Research Ethics Committee of the University Hospital and School of Medicine of Universidad Autonoma de Nuevo Leon (approval no. OR17-00002). All patients had advanced knee OA diagnosis (Kellgren-Lawrence grade IV), disabling knee pain and required total knee prosthesis. The cartilage and synovial explants for the coculture assays were obtained from seven male patients with a median of 71 years and an interquartile range of 60–79 years. Patients with joint infection were excluded and patients in whom the tissue sample was insufficient were eliminated. Informed consent was obtained from all patients. To obtain the explants, the joint tissue from the femoral condyles and synovial capsule was recovered, from which cuts of 6–8 mm in diameter were made by means of the Osteochondral Autograft Transfer System (OATS®, Arthrex Inc.). Efforts were made to obtain cartilage explants from samples without erosion or exposure of the subcondral bone which were identified in non-weight bearing zones and at the periphery of the femoral condyles. Most of synovial samples from which explants were obtained were characterized for presenting friability, redness, hyperemia and synovial hypertrophy. The obtained explants were washed three times with 1X sterile phosphate-buffered saline (PBS; Gibco; Thermo Fisher Scientific, Inc.), once in 70% ethanol and once in sterile PBS 1X with gentamicin 50 µg/ml.

Cartilage-synovial tissue coculture

Cartilage and synovial tissue explants from the same donor were distributed in 12-transwell plates for coculture. In each well, an explant of cartilage and synovial tissue was placed; such explants were divided by a polystyrene membrane transwell insert with a pore diameter of 1.0 µm (Corning Incorporated). Cartilage explant was placed within the insert and the synovial tissue explant was placed at the bottom of the plate. Each coculture was maintained under the following conditions: 1) Basal: Complete DMEM/F12 (Gibco) [added with gentamicin 50 µg/ml and fungizone 0.25 µg/ml (Gibco), supplemented with fetal bovine serum (FBS) 10% (Gibco)], 2) Interleukin-1β (IL-1β): Complete DMEM/F12 with 10 ng/ml of human recombinant protein IL-1β (R&D Systems Inc.), 3) NASHA: Complete DMEM/F12 added with NASHA (hyaluronic acid, 1 mg/ml, Durolane HA) and 10 ng/ml of IL-1β; and 4) Stem cell-conditioned medium (SC-CM): Complete DMEM/F12 added with conditioned stem cell culture media (1 ml) and 10 ng/ml of IL-1β. All the cultures were incubated at 37°C, with 5% CO2 upon 0, 48 and 72 h for further RNA isolation and supernatants were storage to −80°C until required for additional analysis.

MSC derived from adipose tissue

Adipose tissue samples were obtained from total or partial hip arthroplasty surgeries (n=5). The adipose tissue samples used to isolate MSC were derived from five male patients with a median of 70 years and an interquartile range of 67–77 years. MSC isolation was performed following a method previously described (20). Briefly, after the isolation, cells were plated in a 75 cm2 culture flasks, changes of complete culture medium (DMEM/F12 added with gentamicin 50 µg/ml and fungizone 0.25 µg/ml, supplemented with FBS 10%) were made every 72 h until the cells reached 80–90% confluence and were ready to undergo a subculture. The cells from the different donors received a maximum of 2 subcultures, then they were pooled and maintained in new 75 cm2 culture flasks until 80–90% confluency (approximately 2×106 cells), and were stimulated with IL-1β (10 ng/ml) in 5 ml of complete DMEM/F12 medium for 24 h; the culture medium of the stimulated confluent cells (stem-cell conditioned medium, SC-CM) was stored at −80°C and was used to supplement the cocultures of the cartilage and synovial tissue explants.

Immunophenotyping of the adipose-derived MSC by flow cytometry

The immunophenotype of the MSC in culture was evaluated within passages 2 and 3. Adherent cells cultured in 75 cm2 bottles were detached with 0.75% trypsin/0.5 mM EDTA (Gibco). Cell pellet was obtained and resuspended in complete DMEM medium and incubated at 37°C for 20 min. Subsequently, the cells were centrifuged and the cell button was diluted at approximately 200,000 cells per tube in 50 µl of staining buffer (SB, Becton Dickinson Pharmingen) or in a 0.5% bovine serum albumin (BSA, Sigma-Aldrich; Merck KGaA) solution in PBS. The antibody was added following the manufacture's recommendation and then the mix was incubated for 20 min in the dark. Then the samples were washed with 0.5 ml of 0.5% BSA in PBS and centrifuged at 1,800 × g (730 × g), for 1 min at 4°C. The button of stained cells from each tube were resuspended in 250 µl of 0.5% BSA in PBS and analyzed for three-color immunofluorescence by flow cytometry (FACSAria; BD Biosciences).

Differentiation potential of the adipose-derived MSC

To confirm the multilineage differentiation potential of the isolated MSC, we investigated the ability of those cells to differentiate into chondrogenic, adipogenic, and osteogenic lineages (21). Briefly, for adipocyte differentiation, the cells were induced with an adipogenic differentiation medium containing dexamethasone (1 mM), indomethacin (60 mM), 3-isobutyl-1-metyl-xanthine (500 mM; IBMX), and insulin (5 mg/ml) for 3 weeks. Differentiation into adipocytes was demonstrated by Oil-Red-O staining for the presence of lipid droplets. For chondrocyte differentiation, MSC were maintained as micromass cultures; the formed pellets were induced in the presence of a medium containing TGF-β1 (10 ng/ml), dexamethasone (0.1 µM), ascorbic acid (50 µg/ml), and insulin-transferrin-selenium solution (ITS, 10 µg/ml) for 3 weeks. The cells were differentiated toward chondrocytes, as shown by glycosaminoglycan (GAG) deposition and Alcian Blue staining. To evaluate the osteoblastic potential, cells were stained with Alizarin Red to detect mineralization after a cultivation period of 14 days with an osteogenic medium containing dexamethasone (0.1 µM), β-glycerophosphate (10 mM), and ascorbic acid (50 µg/ml).

Total RNA extraction and gene expression analysis by reverse transcription-quantitative (RT-q)PCR

Total RNA was isolated from the cartilage and synovial explants samples at the indicated culture times using the commercial kit RNeasy Lipid Tissue Mini kit (Qiagen GmbH) following the manufacturer's recommendations. The quality of the samples obtained from RNA was evaluated by spectrophotometric quantification in the NanoDrop 2000 equipment (Thermo Fisher Scientific Inc.). From each total RNA sample obtained, complementary DNA synthesis (cDNA) was performed by means of a retrotranscription reaction with the GoTaq Probe 2-Step RT-qPCR System commercial kit (Promega Corporation). From the synthesized cDNA, the expression of different genes related to the inflammatory response and the ECM synthesis in cartilage and synovium was evaluated through the use of TaqMan probes (Applied Biosystems; Thermo Fisher Scientific, Inc.) specific for each gene in each coculture test and in each experimental group (4 groups) in triplicate. The expression assays were performed in the 7500 Fast Real-Time PCR System (Applied Biosystems; Thermo Fisher Scientific, Inc.). The list of genes evaluated is shown in Table I. Data analysis was carried out by means of the ΔΔCq method for relative expression, using β-2-microglobulin (B2M) as endogenous gene to normalization (22).

Table I.

Analyzed genes by reverse transcription-quantitative PCR. The corresponding code is presented for each of the TaqMan probes used.

Table I.

Analyzed genes by reverse transcription-quantitative PCR. The corresponding code is presented for each of the TaqMan probes used.

Interleukin-1 βIL-1βHs01555410_m1
Matrix metallopeptidase 13MMP13Hs00233992_m1
ADAM metallopeptidase with thrombospondin type 1 motif 5ADAMTS5Hs00199841_m1
Tissue inhibitor of metalloproteinases 1TIMP1Hs00171558_m1

a Applied Biosystems; Thermo Fisher Scientific, Inc.

Release of GAG

The sulfated GAG (chondroitin sulfate, heparan sulfate, keratan sulfate and dermatan sulfate) were quantified by means of a spectrophotometric analysis using the metachromatic probe dimethylmethylene blue (DMB) using the Rheumara® commercial GAG detection kit (Astarte Biologics). The binding of DMB to sulfated GAG induces a change in the absorption spectrum, which is directly proportional to the amount of sulphated GAG. Measurements were performed in 96-well plates in the Cytation3 HT multimodal plate reader (BioTek Instruments Inc.) at a wavelength of 525 nm. The results obtained were normalized with respect to the wet weight of the cartilage explants. The presence of sulphated GAG from each coculture trial (7 trials) and from each experimental group (4 groups) was evaluated 5 times.

Nitric oxide production

Nitric oxide (NO) was determined in the culture supernatants by using the Griess reagent (Sigma-Aldrich), which converts available nitrate into nitrite through nitrite reductase. Absorbance at 540 nm was determined in 96-well plates in the Cytation3 HT multimodal plate reader (BioTek Instruments Inc.). Sodium nitrate was used to generate a standard curve with which the concentrations of the samples to be evaluated were determined. The production of NO from each coculture trial (7 trials) and from each experimental group (4 groups) was evaluated 5 times.

Statistical analysis

This study was an experimental, prospective, in vitro study. Sample size was calculated for the comparison of related means, based on an alpha value of 0.05 in two directions and a β value of 0.08; resulting in 7 trials per experimental group. The data are presented as the mean ± standard deviation (SD) of seven independent experiments performed in triplicate unless otherwise mentioned. The numerical variables were analyzed with the parametric ANOVA test with the Tukey post hoc test for multiple comparisons to observe the possible differences between the experimental groups. P<0.05 was considered to indicate a statistically significant difference. Data were processed with GraphPad Prism software version 5.00 for Windows (GraphPad Software, Inc.).


Immunophenotyping of human adipose-derived MSC

The analysis by flow cytometry of the MSC revealed the surface markers that should be positive for this type of cells (CD73+/CD90+/CD105+). In the histogram of the positive markers, only a well-defined peak is distinguished, which demonstrates the specificity of the test (Fig. 1A). The expression of the markers in the sample analyzed is also plotted; as expected, the expression of the CD105 was the one that varied more between individuals and passages. The markers CD73 and CD90 were the most stable. In Fig. 1A, a representative image of the analysis of the samples is shown, which is expressed as dot-plot and shows the purity of the cell population. Most of the cells are located in a very concentrated area and in the quadrant corresponding to those that are double positive.

Assessment of pluripotency in the human adipose-derived MSC

We assess the ability of isolated human adipose-derived MSC to differentiate into adipocytes, chondrocytes, and osteoblasts. The differentiated MSC to adipocytes underwent production of vacuoles containing lipid droplets detected with Oil-Red-O staining as red areas (Fig. 1B). The induced chondrogenic differentiation was detected by the presence of GAG within the extracellular matrix stained with Alcian Blue (Fig. 1B). Lastly, MSC developed a more round polygonal appearance and the mineralized foci were detected as red spotted areas stained with Alizarin Red after osteogenic differentiation (Fig. 1B). These data confirmed that the isolates of adipose-derived MSC used in this study are efficiently capable to initiate a multilineage differentiation process.

Release of GAG to the culture medium

The presence of sulfated GAG was detected in the culture media of all four experimental groups at 48 and 72 h (Fig. 2). A greater release of GAG was detected in the treatment group stimulated with IL-1β with respect to the basal, NASHA and SC-CM group at 48 h (P<0.05) and 72 h (P<0.01). It was observed that the level of GAG released to the culture medium was maintained at the level of the basal group (68.3±5.1 µg/g of cartilage) after supplementation with NASHA (76.9±1.5 µg/g of cartilage) or with SC-CM (70.6±2.9 µg/g of cartilage) at 72 h, with no significant changes between these two last groups.

Quantification of NO

The concentrations of nitrite, as an index of NO production, increased markedly in the groups in which inflammation with IL-1β was induced (IL-1β, NASHA and SC-CM) compared with the basal group at 48 and 72 h (P<0.001; Fig. 3). A decrease in the production of NO was observed in the groups treated with NASHA and MSC with respect to the IL-1β group at 72 h; nevertheless, only in the SC-CM group a significant decrease in the NO levels was detected (IL-1β, 33.1±9.8 µM vs. SC-CM, 23.9±5.4 µM; P<0.05).

Expression of genes related to inflammation and matrix turnover in cartilage and synovial membrane explants

The recombinant IL-1β markedly increased the expression of IL-1β in the IL-1β, NASHA, and SC-CM groups, both in cartilage (P<0.001) and in the synovium (P<0.05) showing a clearly inflammatory effect (Fig. 4A, B). Treatments with NASHA and SC-CM decreased the expression of IL-1β at 72 h in both cartilage and synovial membrane. In cartilage, only the treatment with the conditioned culture medium of MSC significantly decreased the expression of IL-1β (P<0.05) at 72 h. In synovial membrane, the treatment with SC-CM also decreased the expression level of IL-1β compared to the treatment with NASHA. Despite this, the difference shown was not statistically significant (NASHA, 5.2±5.7 vs. SC-CM, 3.1±3.1; P>0.05).


The expression of MMP13 was significantly increased in the IL-1β group compared with its basal expression levels after exposure to recombinant IL-1β in cartilage and synovial explants (P<0.05; Fig. 4C, D). However, the expression of MMP13 significantly decreased in both cartilage and synovium (P<0.05) after 72 h of treatment with SC-CM. Treatment with NASHA exhibited a significantly downregulation of MMP13 only in cartilage at 72 h (P<0.05).


In the cartilage explants, both treatments significantly decreased the expression of ADAMTS5 at 48 and 72 h (P<0.05 and P<0.01, respectively) despite the inflammatory influence of recombinant IL-1β (Fig. 4E). In explants of synovial tissue, the same behavior was detected only at 72 h (P<0.05; Fig. 4F).


Conversely to the effect of IL-1β in the aforementioned genes in cartilage explants, the expression of TIMP1 was decreased as a consequence of the inflammatory stimulation compared with basal level (Fig. 4G). It can be distinguished how in both cartilage and synovial membrane explants, treatments with NASHA and SC-CM recovered the expression level of TIMP1 (Fig. 4G, H), and the difference was significantly higher than in the IL-1β group at 48 and 72 h (P<0.01).


Exposure of synovial and cartilage explants in coculture to NASHA and SC-CM, resulted in positive changes in gene expression profiles and release of agents consistent with an anti-inflammatory and anti-catabolic activity in articular tissues. However, only the mediators produced by the MSC previously stimulated with IL-1β were able to significantly decrease a greater number of inflammation and matrix degradation targets studied in the in vitro model used.

It is known that the tissues involved in the OA joint produce a wide variety of catabolic and proinflammatory mediators that lead to the destruction of the extracellular matrix of these structures (23); therefore, is important to evaluate these processes in tissues other than articular cartilage. While in vitro inflammation models exist for some inflammatory diseases, to date, there is no simply and readily available OA model. As earlier mentioned, we used an ex vivo coculture system of cartilage and synovial membrane explants using inserts that allowed the interaction of the two types of tissue through different molecule production to better represent the joint environment, unlike what happens in an isolated culture of cartilage or synovium (16,24). Additionally, models based on explant culture have the main advantage that they can be used to examine the response of cells in their natural extracellular matrix, without affecting the cell phenotype (25).

The release of GAG has been reported to be useful as a measure of ECM degradation in cultures of cartilage explants after an inflammatory stimulus (e.g., IL-1β) (2628). In our model, we detected an increase of GAG release induced by IL-1β (10 ng/ml) and both NASHA and SC-CM were able to protect articular tissues against ECM degradation. This effect might be directly related with the reduction of gene expression of ECM degrading enzymes such as MMP13 and ADAMTS5.

The inflammatory effects of IL-1β include triggering a signaling pathway that stimulates the release of NO. As consequence, cartilage degradation is produced due to an increased synthesis of proteolytic enzymes such as MMP13 and ADAMTS5 (29). In our study, both NASHA and SC-CM reduced the IL-1β-induced production of the inflammatory mediator NO but only the SC-CM showed a meaningful significantly difference. This outcome could be a consequence of the decreased gene expression of MMP13 and ADAMTS5 in our cartilage and synovial explant coculture model. It has been reported that human SC-CM might produce this protective action as a consequence of iNOS downregulation (17), since NO inhibits ECM synthesis (30).

In OA, anabolic and catabolic processes are significantly affected as a result of a homeostasis imbalance. Important markers of catabolism in OA are MMP13 and ADAMTS5, which are commonly elevated. We found an increased IL-1β-induced expression of MMP13 in cartilage and synovial explants. This effect has been previously reported over MMP13 and other MMP in similar OA explant-based culture models (16,31). On the other hand, ADAMTS5 was not upregulated by IL-1β in explant tissues. These is in agreement with previous investigations who have found that ADAMTS5 is not affected by inflammatory cytokines (i.e., IL-1β, TNF-α) especially in human chondrocytes and cartilage (3234). Our results showed that SC-CM inhibited the expression of IL-1β, MMP13 and ADAMTS5 in cartilage, whereas NASHA produced a similar effect only in MMP13 and ADAMTS5. Moreover, synovial tissues had also different decreased expression levels of catabolic markers after SC-CM (MMP13 and ADAMTS5) and NASHA (ADAMTS5) treatments. We also found a significant overexpression of TIMP1 in IL-1β-treated cartilage and synovial explants after exposure to NASHA and SC-CM. TIMP1 is a natural inhibitor of different MMP, including MMP13 (35), and is thought to play a role in regulating ECM turnover by modulating the degradative capacity of these MMP. A higher TIPM1 gene expression might indicate a protective effect against ECM degradation in articular tissues. Together, these results suggest that both NASHA and SC-CM exerts an anti-inflammatory and anti-catabolic effect by partially counteract the catabolic effect of proinflammatory cytokines through different pathways.

Several clinical studies have shown that the use of NASHA is well tolerated in patients in whom it has been administered intra-articularly, and that its clinical effectiveness remains for several weeks in the treatment of knee OA (3638). However, to our knowledge, the ability of NASHA to reduce the inflammatory process associated with the development and progression of OA has not been evaluated. On the other hand, it has been shown that much of the chondrogenic potential of MSC is due to the secretion of soluble factors that act in a paracrine fashion and that play an important role in immunomodulation and tissue regeneration (17,39). Considering this characteristic, we could compare the anti-inflammatory effect of these two types of therapies in the same culture system.

A systematic review about the anti-inflammatory effect of intra-articular HA reported that high molecular weight HA can promote an anti-inflammatory response within the cell through binding sites of CD44, TLR-2 and TLR-4 receptors (40). The binding of the high molecular weight HA to CD44 receptor downregulates the expression of IL-8, IL-33, MMP, proteoglycans, and PGE2 and suppresses NF-κB activation (40). It would be important to determine if NASHA exerts its anti-inflammatory effect over some of the targets reported in this study through the aforementioned mechanism.

Our findings are in line with those of previous research, which found that conditioned media from MSC stimulated with pro-inflammatory mediators are able to inhibit inflammatory processes by targeting catabolic and inflammatory factors (16,17). One of these reports suggest that the inhibitory effects of SC-CM could be partially supported by the fact that mediators produced by MSC stimulated with proinflamatory agents induce a reduction of NF-κB activation in OA chondrocytes (17).

Some of the limitations of this study include that we only show changes in gene expression of markers related to inflammation and matrix turnover. It would be important to confirm these changes at the protein level and with other specific markers of main ECM components such as collagen type II and aggrecan at both gene and protein level. Moreover, a further characterization of specific markers related to degradation and inflammation of cartilage and synovium would yield more information about these processes since only the GAG and NO were quantified.

This study evaluated the anti-inflammatory and anti-catabolic properties of NASHA compared to those of SC-CM. Both treatments showed similar favorable effects; however, SC-CM was able to counteract inflammatory effects over a higher number of the evaluated targets. The wide number of bioactive molecules probably works in favor of the MSC to achieve the positive effects observed. Further studies are needed to elucidate the exact mechanism by which each therapy exerts its positive effects.


The authors would like to thank Dr Sergio Lozano (member of the American Translators Association and the American Medical Writers Association) for editing the English of the manuscript.


No funding was received.

Availability of data and materials

All data generated or analyzed during this study are included in this published article.

Authors' contributions

MSM, SALS, FVC, VMPM and LFM conceived the original idea and designed the experiments. MSM, SALS and VPS performed the experiments. HGMR, FVC and CAAO analyzed the data. VPS, FVC and VMPM interpreted the data. MSM and VMPM wrote the manuscript. FVC and LFM edited the original article. FVC, LFM and VMPM critically revised the manuscript. All authors read and approved the final manuscript.

Ethics approval and consent to participate

The present study was approved (approval no. OR17-00002) by The Ethical Committee and Research Review Board (Research Ethics Committee) of the University Hospital and School of Medicine of Universidad Autonoma de Nuevo Leon. Informed consent was obtained from all patients.

Patient consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.



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Simental‑Mendía M, Lozano‑Sepúlveda SA, Pérez‑Silos V, Fuentes‑Mera L, Martínez‑Rodríguez HG, Acosta‑Olivo CA, Peña‑Martínez VM and Vilchez‑Cavazos F: Anti‑inflammatory and anti‑catabolic effect of non‑animal stabilized hyaluronic acid and mesenchymal stem cell‑conditioned medium in an osteoarthritis coculture model. Mol Med Rep 21: 2243-2250, 2020
Simental‑Mendía, M., Lozano‑Sepúlveda, S.A., Pérez‑Silos, V., Fuentes‑Mera, L., Martínez‑Rodríguez, H.G., Acosta‑Olivo, C.A. ... Vilchez‑Cavazos, F. (2020). Anti‑inflammatory and anti‑catabolic effect of non‑animal stabilized hyaluronic acid and mesenchymal stem cell‑conditioned medium in an osteoarthritis coculture model. Molecular Medicine Reports, 21, 2243-2250.
Simental‑Mendía, M., Lozano‑Sepúlveda, S. A., Pérez‑Silos, V., Fuentes‑Mera, L., Martínez‑Rodríguez, H. G., Acosta‑Olivo, C. A., Peña‑Martínez, V. M., Vilchez‑Cavazos, F."Anti‑inflammatory and anti‑catabolic effect of non‑animal stabilized hyaluronic acid and mesenchymal stem cell‑conditioned medium in an osteoarthritis coculture model". Molecular Medicine Reports 21.5 (2020): 2243-2250.
Simental‑Mendía, M., Lozano‑Sepúlveda, S. A., Pérez‑Silos, V., Fuentes‑Mera, L., Martínez‑Rodríguez, H. G., Acosta‑Olivo, C. A., Peña‑Martínez, V. M., Vilchez‑Cavazos, F."Anti‑inflammatory and anti‑catabolic effect of non‑animal stabilized hyaluronic acid and mesenchymal stem cell‑conditioned medium in an osteoarthritis coculture model". Molecular Medicine Reports 21, no. 5 (2020): 2243-2250.