Simulated microgravity-induced oxidative stress and loss of osteogenic potential of osteoblasts can be prevented by protection of primary cilia

by Selleckchem | Dec 09 2023

Oxidative stress has been considered to be closely related to spaceflight-induced bone loss; however, mechanism is elusive and there are no effective countermeasures. Using cultured rat calvarial osteoblasts exposed to microgravity simulated by a random positioning machine, this study addressed the hypotheses that microgravity-induced shortening of primary cilia leads to oxidative stress and that primary cilium protection prevents oxidative stress and osteogenesis loss. Microgravity was found to induce oxidative stress (as represented by increased levels of reactive oxygen species (ROS) and malondialdehyde production, and decreased activities of antioxidant enzymes), which was perfectly replicated in osteoblasts growing in NG with abrogated primary cilia (created by transfection of an interfering RNA), suggesting the possibility that shortening of primary cilia leads to oxidative stress. Oxidative stress was accompanied by mitochondrial dysfunction (represented by increased mitochondrial ROS and decreased mitochondrial membrane potential) and intracellular Ca2+ overload, and the latter was found to be caused by increased activity of Ca2+ channel transient receptor potential vanilloid 4 (TRPV4), as also evidenced by TRPV4 agonist GSK1016790A-elicited Ca2+ influx. Supplementation of HC-067047, a specific antagonist of TRPV4, attenuated microgravity-induced mitochondrial dysfunction, oxidative stress, and osteogenesis loss. Although TRPV4 was found localized in primary cilia and expressed at low levels in NG, microgravity-induced shortening of primary cilia led to increased TRPV4 levels and Ca2+ influx. When primary cilia were protected by miR-129-3p overexpression or supplementation with a natural flavonoid moslosooflavone, microgravity-induced increased TRPV4 expression, mitochondrial dysfunction, oxidative stress, and osteogenesis loss were all prevented. Our data revealed a new mechanism that primary cilia function as a controller for TRPV4 expression. Microgravity-induced injury on primary cilia leads to increased expression and overactive channel of TRPV4, causing intracellular Ca2+ overload and oxidative stress, and primary cilium protection could be an effective countermeasure against microgravity-induced oxidative stress and loss of osteogenic potential of osteoblasts.

Comments:

The passage you provided describes a study conducted on rat calvarial osteoblasts to investigate the relationship between microgravity (simulated using a random positioning machine) and oxidative stress, as well as the role of primary cilia in this process. Here's a breakdown of the findings and the proposed mechanism:

### Study Findings:

1. **Microgravity-Induced Oxidative Stress:**
- Exposure to microgravity conditions led to increased levels of reactive oxygen species (ROS) and malondialdehyde production.
- Antioxidant enzyme activities decreased, indicating oxidative stress in osteoblasts.

2. **Primary Cilia Shortening and Oxidative Stress:**
- Microgravity-induced shortening of primary cilia was observed.
- Osteoblasts with abrogated primary cilia (achieved through interfering RNA transfection) exhibited oxidative stress, suggesting a link between cilia shortening and oxidative stress.

3. **Mitochondrial Dysfunction and Ca2+ Overload:**
- Oxidative stress was accompanied by mitochondrial dysfunction, including increased mitochondrial ROS and decreased mitochondrial membrane potential.
- Intracellular Ca2+ overload occurred, attributed to the increased activity of the Ca2+ channel transient receptor potential vanilloid 4 (TRPV4).

4. **Role of TRPV4:**
- TRPV4 activity, specifically its overexpression and overactive channel, contributed to mitochondrial dysfunction, oxidative stress, and loss of osteogenic potential.
- The TRPV4 antagonist HC-067047 attenuated microgravity-induced mitochondrial dysfunction, oxidative stress, and osteogenesis loss.

5. **Primary Cilia Protection as a Countermeasure:**
- Protection of primary cilia, achieved through miR-129-3p overexpression or supplementation with moslosooflavone, prevented microgravity-induced TRPV4 expression, mitochondrial dysfunction, oxidative stress, and osteogenesis loss.

### Proposed Mechanism:

1. **Primary Cilia as Controllers for TRPV4 Expression:**
- The study revealed a novel mechanism where primary cilia act as controllers for TRPV4 expression.
- Microgravity-induced damage to primary cilia led to increased expression and overactive channels of TRPV4.

2. **TRPV4-Mediated Effects:**
- Increased TRPV4 expression and activity caused intracellular Ca2+ overload and oxidative stress in osteoblasts under microgravity conditions.

3. **Primary Cilium Protection as a Countermeasure:**
- Protecting primary cilia prevented the overexpression and overactivity of TRPV4, thereby mitigating intracellular Ca2+ overload and oxidative stress.
- This protection strategy proved effective against microgravity-induced oxidative stress and loss of osteogenic potential in osteoblasts.

In summary, the study suggests that microgravity-induced shortening of primary cilia leads to increased TRPV4 expression and activity, causing intracellular Ca2+ overload and oxidative stress in osteoblasts. Protecting primary cilia could serve as an effective countermeasure against these effects, potentially preserving osteogenic potential in conditions of reduced gravity.