Osteoarthritis is a degenerative joint disease that affects millions of people worldwide. It is characterized by the gradual breakdown of cartilage in the joints, leading to pain, stiffness, and loss of function. Osteoarthritis is characterized by progressive degeneration of articular cartilage and ligaments, as well as chronic synovitis and abnormal bone remodeling (Sellam and Berenbaum, 2010).

While there are several treatment options for osteoarthritis, there is still a great need for more effective and long-lasting interventions. Recent research has shown that regular aerobic activity and stem cell therapy may have promising effects on osteoarthritis management.

Tumor necrosis factor-α (TNFα) is involved in various physiological and pathological processes, including regulation of immune cells, apoptosis (programmed cell death), and granuloma formation (Jang, Lee et al., 2021). TNFα promotes the production of matrix metalloproteinases (MMPs). MMPs are enzymes that break down extracellular matrix components, including cartilage. An increase in the level of TNFα in the joint space leads to an increase in the expression of MMP and, as a result, cartilage degradation (Mukherjee and Das, 2024). MMP13 is an enzyme that plays a vital role in the degradation of extracellular matrix components, especially type II and type I collagen (key structural proteins in cartilage and other connective tissues) (Mukherjee and Das, 2024).

The present study investigates the independent and interactive effects of aerobic exercise and stem cell injection on TNFα gene expression and MMP13 protein expression in knee cartilage tissue in male rats with osteoarthritis model.

METHODS

The current experimental research was conducted on 25 male Wistar rats (age: 8-12 weeks, weight: 180-200 g). The animals were kept in standard and controllable conditions in the research center of laboratory animals in Islamic Azad University, Sari branch. The animals were randomly divided into five groups: healthy, osteoarthritis, osteoarthritis-exercise, osteoarthritis-stem cell, osteoarthritis-stem cell-exercise, and received interventions based on the group name. Endurance training was performed for eight weeks (5 sessions/week, 30-50 minutes/session, and at a speed of 16 meters/minute) on a non-inclined treadmill. Mesenchymal stem cells (1x106 cells/kg) were injected once to the respective groups. Forty-eight hours after the last training session, cartilage tissue was removed and sent to the laboratory. One-way analysis of variance and Tukey's post hoc test were used to determine the difference between groups. All calculations were done with SPSS software, and the significance level was considered as P≤0.05.

RESULTS

The results of the one-way analysis of variance regarding the TNFα variable showed that there was a significant difference between the research groups (P=0.046, F=2.937). TNFα gene expression was significantly higher in the osteoarthritis group than in the healthy group (P=0.027). No other significant changes were observed between the research groups. Analysis of data related to MMP13 protein expression using one-way analysis of variance test showed that there was a significant difference between the research groups (P=0.001, F=124.614). MMP13 protein expression of cartilage tissue in osteoarthritis (P=0.001), osteoarthritis-exercise (P=0.003), osteoarthritis-stem cell (P=0.011), and osteoarthritis-exercise-stem cell (P=0.001) groups were significantly higher than that in the healthy group. MMP13 protein expression of cartilage tissue in osteoarthritis-exercise (P=0.001), osteoarthritis-stem cell (P=0.001), and osteoarthritis-exercise-stem cell (P=0.001) groups was significantly lower than that in the osteoarthritis group. There were no other significant differences between the groups.

The observed increase in TNFα and MMP13 variables supports the common understanding that inflammation and cartilage destruction are key processes involved in the progression of osteoarthritis. TNFα, as a pro-inflammatory cytokine, can initiate and perpetuate inflammatory responses in the joint and lead to cartilage degradation (Wojdasiewicz, Poniatowski et al., 2014). MMP13, as the enzyme responsible for collagen degradation, further contributes to cartilage matrix degradation (Troeberg and Nagase, 2012). Therefore, targeting these molecular signaling pathways, as shown by the effects of physical activity and stem cell therapy on their expression, may have therapeutic potential in the management of osteoarthritis.

In the present study, analyzing the TNFα gene expression results in the treatment groups indicates that the interventions (aerobic exercise, stem cell therapy, and the combination of exercise and stem cells) have partially altered the elevated expression of TNFα caused by osteoarthritis. However, these changes were not statistically significant. The increased protein expression of MMP13 in the cartilage tissue of the induced osteoarthritis groups was modified by exercise, the use of stem cells, and their interaction; nevertheless, there was no difference between the groups with different treatments. TNFα leads to inflammation by causing inflammation, and MMP13 causes cartilage breakdown by breaking down collagen. Past studies showed that inflammatory factors regulate MMPs. Inhibiting the synthesis and/or activity of MMPs is highly important in the development of structural modification therapies for osteoarthritis. Considering the lack of change in TNFα due to the interventions and the change in MMP13 due to the interventions, it seems that inflammatory factors other than TNFα have an effect on the expression of MMP13 protein in the present study.

The results of this study showed that the expressions of the TNFα gene and MMP13 protein increased in knee cartilage tissue in the rat model of arthritis. The use of regular aerobic activity and treatment with stem cells showed promising effects in modulating the expression of MMP13, which can be considered a therapeutic intervention to improve osteoarthritis by suppressing this protein. The results of the present study also showed that inflammatory factors other than TNFα seem to have an effect on the expression of MMP13 protein. Therefore, to find the exact mechanism of MMP13 changes, future studies are needed to investigate the changes of other inflammatory factors, such as interleukins, transcription factors (e.g., NFKB and AKT), and PI3K proteins.

Ethical Considerations

Compliance with ethical guidelines

All principles of working with laboratory animals approved by the Ministry of Health of the Islamic Republic of Iran were observed in this study. The current research has the ethics code number IR.IAU.SDJ.REC.1402.093 from Islamic Azad University, Sanandaj Branch.

Funding

There is no funding support.

Authors’ Contributions

This article is a part of Hamidreza Sharifi's doctoral dissertation, which was approved at Islamic Azad University, Sanandaj Branch, under the guidance and advice of co-authors. All authors contributed to the writing of the article and approved the final version of the article.

Conflict of Interest

The authors declare no conflict of interest.

Acknowledgments

We appreciate everyone who provided scientific consultation for this paper.