CRISPR/dCas9-Mediated Parkin Inhibition Impairs Mitophagy and Aggravates Apoptosis of Rat Nucleus Pulposus Cells Under Oxidative Stress.

OBJECTIVE: The aim of this study is to explore the role of Parkin in intervertebral disk degeneration (IDD) and its mitophagy regulation mechanism. STUDY DESIGN AND METHODS: Rat nucleus pulposus (NP) cells were stimulated with hydrogen peroxide (H2O2) to a mimic pathological condition. Apoptosis and mitophagy were assessed by Western blot, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and immunofluorescence staining. The CRISPR-dCas9-KRAB system was used to silence the expression of Parkin. RESULT: In this study, we found that Parkin was downregulated in rat NP cells under oxidative stress. In addition, treatment with H2O2 resulted in mitochondrial dysfunction, autophagy inhibition, and a significant increase in the rate of apoptosis of NP cells. Meanwhile, mitophagy inhibition enhanced H2O2-induced apoptosis. Furthermore, repression of Parkin significantly attenuated mitophagy and exacerbated apoptosis. CONCLUSION: These results suggested that Parkin may play a protective role in alleviating the apoptosis of NP cells via mitophagy, and that targeting Parkin may provide a promising therapeutic strategy for the prevention of IDD.

[Long-term clinical studies on the treatment of lumbosacral spinal tuberculosis with CT-guided minimally invasive surgery].

OBJECTIVE: To evaluate the clinical effects of CT-guided percutaneous puncture and local chemotherapy for lumbosacral spinal tuberculosis. METHODS: From January 2002 to March 2013, 145 patients (84 males and 61 females) with lumbosacral spinal tuberculosis underwent failed conservative treatment were treated with CT-guided minimally invasive surgery. Their clinical data were retrospectively analyzed. Among them, 143 patients were followed up, aged from 2.5 to 81 years with an average of (42.60 +/- 17.14) years. Fourteen cases (6 cases with internal fixation and 8 cases without internal fixation) recurred and 32 cases complicated with paraspinal abscess after operation. Preoperatively 1 case complicated with lower limb weakness and superficial sensation worse, 1 case with muscle strength decrease, this 2 cases were grade D of Frankle, other cases were grade E. And 1 patient underwent fenestration operation and local chemotherapy, 144 cases with CT-guided percutaneous puncture and local chemotherapy. ESR and lumbar lordosis angle of all patients were observed preoperatively and final follow-up. RESULTS: The mean follow-up time was 67 months (ranged, 21 to 149 months) in 143 cases, and 73 cases more than 5 years. All patients obtained clinical healing. ESR was (44.96 +/- 12.41) mm/h before operation and (7.25 +/- 3.43) mm/h at final follow-up, there was significant difference between preoperative and postoperative (t=35.06, P=0.000). Lumbar lordosis angle was (36.32 +/- 8.55) degrees before operation and (33.35 +/- 8.16) at final follow-up, there was significant difference between preoperative and postoperative (t=13.90, P=0.000). CONCLUSION: When conservative treatment fails for 3 months or more, the patients have good spinal stabilization, nerve function is more than grade D of Frankel, CT-guided percutaneous puncture and local chemotherapy can get satisfactory outcomes for lumbosacral spinal tuberculosis.