Scientists discovered that blocking a tiny RNA molecule can help rejuvenate aged mesenchymal stem cells (MSCs) — which have the ability to give rise to specialized cells — derived from people with idiopathic pulmonary fibrosis (IPF).
Those rejuvenated stem cells were found to have improved therapeutic efficacy when used to treat the disease in a mouse model of induced pulmonary fibrosis (PF).
“Our study provides a novel candidate target to enhance the therapeutic efficacy of MSC-based therapy for pulmonary fibrosis-related disease,” the researchers wrote.
Their findings were reported in a study titled “Inhibition of miR-199a-5p rejuvenates aged mesenchymal stem cells derived from patients with idiopathic pulmonary fibrosis and improves their therapeutic efficacy in experimental pulmonary fibrosis,” published in the journal Stem Cell Research & Therapy.
Patient-derived MSC-based therapy is a potential treatment for IPF that uses adult stem cells found in several locations, including the bone marrow, skin, and fat tissue. Because they are stem cells, MSCs can transform into nearly all types of cells found in the body, including those making up the bones, cartilage, and fatty tissue.
MSCs have been shown to exert protective effects in diseased lungs that include reducing inflammation, transforming into local cell types, activating resident stem cells, and decreasing collagen deposit buildup — the process that leads to scar tissue formation, called fibrosis.
However, MSC function declines with age — a process known as senescence — along with the cells’ ability to repair tissue. Furthermore, compared with those from age-matched healthy controls, MSCs isolated from IPF patients (IPF-MSCs) exhibit more senescence and lower efficacy in IPF animal models.
Studies suggest MSC senescence is mediated by microRNAs (miRNAs), which are tiny RNA molecules that repress gene activity. Additionally, it has been reported that the level of one miRNA in particular, called miR-199a-5p, is elevated in people with IPF and in mice with induced PF.
These observations prompted researchers at the Southern Medical University, in Guangzhou, China, and their colleagues to investigate whether miR-199a-5p regulates senescence in IPF-MSCs. Their goal was to determine if blocking this miRNA can rejuvenate IPF-MSCs and improve their efficacy in an IPF mouse model.
Initial experiments compared control-MSCs isolated from healthy donors with IPF-MSCs, both derived from fatty (adipose) tissue. The team found IPF-MSCs were able to transform into fat cells more easily than control-MSCs. However, at the same time, they were less able to give rise to bone and cartilage cells, suggesting their ability to grow into different cell types was altered.
Next, testing showed that the levels of cellular senescence markers in MSC cells isolated from six IPF patients were significantly higher than those of seven healthy individuals. The investigators also observed that IPF-MSCs were quicker to slow and stop growing, confirming “IPF-MSCs showed more cellular senescence.”
In blood samples collected from healthy donors and IPF patients, the levels of miR-199a-5p were significantly higher in those with idiopathic pulmonary fibrosis compared with those from controls.
Supporting these results, investigators demonstrated that when healthy cells were treated with a miR-199a-5p mimic, they ceased growing and their levels of senescence markers increased. In contrast, blocking miR-199a-5p led to a significant reduction in senescence markers and increased cell growth.
“Collectively, these results indicate that miR-199a-5p mediated the cellular senescence of IPF-MSCs,” the team wrote.
The team then investigated whether miR-199a-5p induced MSC senescence by regulating autophagy — the process by which cells destroy and recycle components they no longer need.
Compared with control-MSCs, there was a decrease in IPF-MSCs in the number of autophagosomes, which are the vesicles that contain cellular components waiting to be degraded by autophagy. Also, markers for autophagy were reduced in IPF-MSCs compared with controls.
Again, the scientists observed that control cells treated with a miR-199a-5p mimic had a lower number of autophagosomes compared with control cells not treated with the miRNA mimic. Moreover, treated control cells had increased levels of senescence markers and decreased cell growth. Conversely, blocking miR-199a-5p increased the number of autophagosomes and markers for autophagy, and reduced senescence markers, demonstrating that “miR-199a-5p induces cellular senescence of MSCs by regulating autophagy.”
To determine the impact of blocking miR-199a-5p on the therapeutic efficacy of IPF-MSCs, investigators performed a series of experiments in a mouse model of induced PF. First, they exposed mice to bleomycin to induce the onset of the disease. Then, control-MSCs, IPF-MSCs, and IPF-MSCs specifically treated to have low miR-199a-5p levels (anti-miR-199a-5p-IPF-MSCs), were injected into the animals’ tail vein. The animals’ lung tissues were collected and examined 14 days later.
As expected, bleomycin-treated mice showed signs of lung damage and high levels of inflammatory cells. In contrast, the amount of damaged tissue was significantly reduced in all MSC-treated groups, with the most significant improvement seen in mice treated with control-MSCs. Notably, the integrity of lung tissue in the anti-miR-199a-5p-IPF-MSC group was better than that of IPF-MSC mice.
While the number of surviving MSCs was highest in tissue isolated from the control-MSC group, MSC survival was much higher in the anti-miR-199a-5p-IPF-MSC group compared with the IPF-MSC group.
A more detailed analysis revealed the expression (production) of fibrosis-related proteins collagen I and alpha-SMA was significantly increased in bleomycin-treated mice, but dramatically reduced in the MSC-treated animals. Again, the production of these proteins was lower in the anti-miR-199a-5p-IPF-MSC group than in IPF-MSC mice. In addition, the expression level of IPF-associated inflammatory signaling proteins followed the same trend.
“These results indicate that anti-miR-199a-5p-IPF-MSCs were superior to IPF-MSCs in attenuation of fibrosis formation and inflammation in the mouse model of pulmonary fibrosis,” the researchers wrote.
“These results reveal that miR-199a-5p inhibits autophagy … leading to MSC senescence in patients with IPF,” the researchers concluded. “The downregulation [inhibition] of miR-199a-5p could rejuvenate IPF-MSC senescence and improve the therapeutic potency of MSC therapy for pulmonary fibrosis.”
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