Inhibition of LRRK2 restores parkin-mediated mitophagy and attenuates intervertebral disc degeneration.

OBJECTIVE: To elucidate the role of LRRK2 in intervertebral disc degeneration (IDD) as well as its mitophagy regulation mechanism. METHODS: The expression of LRRK2 in human degenerative nucleus pulposus tissues as well as in oxidative stress-induced rat nucleus pulposus cells (NPCs) was detected by western blot. LRRK2 was knocked down in NPCs by lentivirus (LV)-shLRRK2 transfection; apoptosis and mitophagy were assessed by western blot, TUNEL assay, immunofluorescence staining and mitophagy detection assay in LRRK2-deficient NPCs under oxidative stress. After knockdown of Parkin in NPCs with siRNA transfection, apoptosis and mitophagy were further assessed. In puncture-induced rat IDD model, X-ray, MRI, hematoxylin-eosin (HE) and Safranin O-Fast green (SO) staining were performed to evaluate the therapeutic effects of LV-shLRRK2 on IDD. RESULTS: We found that the expression of LRRK2 was increased in degenerative NPCs both in vivo and in vitro. LRRK2 deficiency significantly suppressed oxidative stress-induced mitochondria-dependent apoptosis in NPCs; meanwhile, mitophagy was promoted. However, these effects were abolished by the mitophagy inhibitor, suggesting the effect of LRRK2 on apoptosis in NPCs is mitophagy-dependent. Furthermore, Parkin knockdown study showed that LRRK2 deficiency activated mitophagy by recruiting Parkin. In vivo study demonstrated that LRRK2 inhibition ameliorated IDD in rats. CONCLUSIONS: The results revealed that LRRK2 is involved in the pathogenesis of IDD, while knockdown of LRRK2 inhibits oxidative stress-induced apoptosis through mitophagy. Thus, inhibition of LRRK2 may be a promising therapeutic strategy for IDD.

Parental preferences for the procedural sedation of children in dentistry: a discrete choice experiment.

Purpose: The aim of this study was to explore parental preferences for the procedural sedation of children in dentistry through a discrete choice experiment (DCE) to inform clinical decisions and oral health management. Methods: Based on literature reviews, interviews with parents of pediatric dental patients, and expert consultation, six attributes, including fasting time, recovery time, sedative administration routes, adverse reactions, sedation depth and procedure cost, were incorporated into the DCE questionnaire. The DCE questionnaire collected data on parental preferences for pediatric dental sedation treatment from June to August 2022. A conditional logit model was used to analyze preference and willingness to pay (WTP) for each attribute and its level. Subgroup analyses assessing the impact of parents' dental anxiety on procedural sedation preferences were also conducted using conditional logit models. Results: A total of 186 valid questionnaires were gathered. Parents' preferences for fewer adverse reactions, a milder sedation depth, lower out-of-pocket cost, shorter fasting and recovery times and administration by inhalation were significantly associated with their choice of sedation model. The conditional logit model showed that parents were most interested in treatments with no adverse reactions (0% vs. 15%) (Coef, 1.033; 95% CI, 0.833-1.233), followed by those providing minimal sedation (vs. deep sedation) (Coef, 0.609; 95% CI, 0.448-0.769). Moreover, the relative importance of adverse reactions and fasting time was higher among anxious than nonanxious parents. The study found a WTP threshold of ¥1,538 for reducing adverse reactions (15% to 0%). The WTP threshold for the best sedation procedure scenario (no fasting requirement, 10 min recovery time, administration by inhalation, 0% adverse reaction incidence and minimal sedation) was ¥3,830. Conclusion: Reducing the adverse reactions and depth of sedation are predominant considerations for parents regarding procedural sedation in pediatric dentistry, followed by lower cost, shorter fasting and recovery times and inhalation sedation. Parents with dental anxiety had a stronger preference for options with a lower incidence of adverse reactions and shorter fasting time than parents without dental anxiety. This discovery is helpful for doctors and can promote collaborative decision-making among parents and doctors.

Urolithin A-induced mitophagy suppresses apoptosis and attenuates intervertebral disc degeneration via the AMPK signaling pathway.

Intervertebral disc degeneration (IDD) is a major cause of low back pain (LBP), and effective therapies are still lacking. Previous studies reported that mitochondrial dysfunction contributes to apoptosis, and urolithin A (UA) specifically induces mitophagy. Herein, we aimed to investigate the protective effect of UA-induced mitophagy on tert-butyl hydroperoxide (TBHP)-induced apoptosis in nucleus pulposus (NP) cells in vitro and a rat model of IDD in vivo. Mitochondrial function, apoptosis, and mitophagy were measured in UA-treated NP cells by western blotting and immunofluorescence; the therapeutic effects of UA on IDD were assessed in rats with puncture-induced IDD. The results showed that UA could activate mitophagy in primary NP cells, and UA treatment inhibited TBHP-induced mitochondrial dysfunction and the intrinsic apoptosis pathway. Mechanistically, we revealed that UA promoted mitophagy by activating AMPK signaling in TBHP-induced NP cells. In vivo, UA was shown to effectively alleviate the progression of puncture-induced IDD in rats. Taken together, our results suggest that UA could be a novel and effective therapeutic strategy for IDD.