Search

Additional file 1: Figure S1. FTIR spectra assay of ALN, DSPE-PEG-NHS and DSPE-PEG-ALN. A FTIR spectra result of ALN. B FTIR spectra result of DSPE-PEG-NHS. The intensity of peak at 1,733 cm-1, correspondent to C=O stretch of NHS groups. C FTIR spectra result of DSPE-PEG-ALN. The characteristic peak of NHS groups is strongly reduced in the DSPE-PEG-ALN, which is consistent with nucleophilic substitution of NHS for alendronate. Figure S2. 1H-NMR spectroscopy result of ALN. A Chemical structure of ALN. B 1H-NMR results of ALN in the range of 1.2ppm to 3.2 ppm. The characteristic signal at 1.88 and 2.93 ppm corresponding to CH2 protons. Figure S3. 1H-NMR spectroscopy result of DSPE-PEG-NHS. A Chemical structure of DSPE-PEG-NHS. B 1H-NMR results of DSPE-PEG-NHS in the range of 0 ppm to 10 ppm, evidencing the main peak of PEG chain (3.64 ppm). C 1H-NMR results of DSPE-PEG-NHS in the range of 0 ppm to 4.6 ppm, evidencing the carbon lateral chains (1.25 ppm), terminal methyl groups (0.80 ppm) and NHS (2.67 ppm, arrow). Figure S4. 1H-NMR spectroscopy result of DSPE-PEG-ALN.A Chemical structure of DSPE-PEG-ALN. B 1H-NMR results of DSPE-PEG-ALN in the range of 0 ppm to 10 ppm, evidencing the main peak of PEG chain (3.64 ppm). C1H-NMR results of DSPE-PEG-ALN in the range of 0 ppm to 3.7 ppm showed a reduction of NHS group signal (2.67 ppm) and the appearance of ALN characteristic signal (1.98 and 3.13 ppm). Figure S5. H&E staining of heart, liver, spleen, lung and kidney in five groups. Figure S6. A Zeta potential value of PL, PL-exo and PL-exo-ALN. B The cell viability of BMSCs treated as above was detected by CCK-8. C H&E staining of liver in the above groups. D The osteogenesis- and angiogenesis-associated growth factors in PL and PL-exo was determined by ELISA. All results are presented as the means ± SDs, *P < 0.05, **P < 0.01. Figure S7. Effects of PL-exo on osteogenic and angiogenic differentiation of BMSCs and EPCs. A–C Early osteogenic differentiation was determined by ALP staining and ALP activity assays after 5 days of induction. Late osteogenic differentiation was determined by Alizarin Red staining and the calcium deposition was quantified by measuring the optical density, after 14 days of induction. D–E In vitro tube formation assay of EPCs treated as indicated (scale bar: 150 μm). F–G The expression levels of p-AKT, AKT, Col1 and β-catenin in BMSCs treated as indicated for 36 h. Exosomes (50 µg/mL) and MK-2206 (5μM, a selective inhibitor of AKT) were added to the related medium in the experimental groups, while the control group was cultured in osteogenic induction medium without any addition. H–I The expression levels of p-AKT, AKT, VEGF and Hif-α in EPCs treated as indicated for 24 h. Exosomes (50 µg/mL) and MK-2206 (5μM) were added to the related medium in the experimental groups, while the control group was cultured in EGM-2 complete medium without any addition. All results are presented as the means ± SDs, *P < 0.05, **P < 0.01. Figure S8. Effects of PL, PL-exo and PL-exo-ALN on RANKL-induced osteoclastogenesis in BMMs. A Representative images of TRAcP-stained osteoclasts after treated with PL, PL-exo and PL-exo-ALN. B The quantitative analysis of TRAcP-positive multinucleated cells (>3 nuclei) per well (96-well plate). All results are presented as the means ± SDs, **P < 0.01. Table S1. Post hoc power analyses of the different experiments (Unpaired t-tests). Table S2. Post hoc power analyses of the different experiments (One-way ANOVA).

The prevalence of cervical disc disease is high, and the traditional surgical method of anterior cervical discectomy and fusion (ACDF) carries with it the disadvantages of motion loss at the operated level, and accelerated adjacent level disc degeneration. Preliminary results of the efficacy and reoperative rate comparing TDA versus ACDF have been reported; however, the long-term outcomes of TDA versus ACDF still remain a topic of debate. This review was prepared following the standard procedures set forth by the Cochrane Collaboration organization, and preferred reporting items for systematic reviews and meta-analyses (PRISMA). The only studies included were randomized controlled trials with a minimum of 4 years of follow-up data. The meta-analysis included the neck disability index (NDI), visual analog scale (VAS) of neck and arm pain, SF-36 physical component scores (SF-36 PCS), over success, neurological success, work status, implant-related complications, and secondary surgery events. Four randomized controlled trials meet the inclusion criteria. The long-term improvement of NDI, VAS of neck and arm pain, SF-36 PCS, over success, and neurological success favored the TDA group. The TDA group also had a lower incidence of secondary surgery for both the index level (RR: 0.45 [0.28, 0.72]) and adjacent level (RR: 0.53 [0.33, 0.88]). In this meta-analysis of 4 included RCTs with a minimum 4 years of follow-ups, total disc arthroplasty showed improvements over ACDF as measured by the NDI, VAS of neck and arm pain, and SF-36 PCS.