Hanson Kari, Isder Carly, Shogren Kristen, Mikula Anthony L, Lu Lichun, Yaszemski Michael J, Elder Benjamin D
Departments of1Neurologic Surgery.
2Orthopedic Surgery, and.
J Neurosurg Spine. 2021 Apr 2;34(6):931-935. doi: 10.3171/2020.10.SPINE201511. Print 2021 Jun 1.
The use of intrawound vancomycin powder in spine surgery has been shown to decrease the rate of surgical site infections; however, the optimal dose is unknown. High-dose vancomycin inhibits osteoblast proliferation in vitro and may decrease the rate of solid arthrodesis. Bone marrow-derived mesenchymal stem cells (BMSCs) are multipotent cells that are a source of osteogenesis in spine fusions. The purpose of this study was to determine the effects of vancomycin on rat BMSC viability and differentiation in vitro.
BMSCs were isolated from the femurs of immature female rats, cultured, and then split into two equal groups; half were treated to stimulate osteoblastic differentiation and half were not. Osteogenesis was stimulated by the addition of 50 µg/mL l-ascorbic acid, 10 mM β-glycerol phosphate, and 0.1 µM dexamethasone. Vancomycin was added to cell culture medium at concentrations of 0, 0.04, 0.4, or 4 mg/mL. Early differentiation was determined by alkaline phosphatase activity (4 days posttreatment) and late differentiation by alizarin red staining for mineralization (9 days posttreatment). Cell viability was determined at both the early and late time points by measurement of formazan colorimetric product.
Viability within the first 4 days decreased with high-dose vancomycin treatment, with cells receiving 4 mg/mL vancomycin having 40%-60% viability compared to the control. A gradual decrease in alizarin red staining and nodule formation was observed with increasing vancomycin doses. In the presence of the osteogenic factors, vancomycin did not have deleterious effects on alkaline phosphatase activity, whereas a trend toward reduced activity was seen in the absence of osteogenic factors when compared to osteogenically treated cells.
Vancomycin reduced BMSC viability and impaired late osteogenic differentiation with high-dose treatment. Therefore, the inhibitory effects of high-dose vancomycin on spinal fusion may result from both reduced BMSC viability and some impairment of osteogenic differentiation.
脊柱手术中使用伤口内万古霉素粉末已被证明可降低手术部位感染率；然而，最佳剂量尚不清楚。高剂量万古霉素在体外抑制成骨细胞增殖，并可能降低坚固性骨融合的发生率。骨髓间充质干细胞（BMSC）是多能细胞，是脊柱融合中成骨的来源。本研究的目的是确定万古霉素对大鼠BMSC体外活力和分化的影响。
从未成年雌性大鼠的股骨中分离出BMSC，进行培养，然后分成两组；一半进行处理以刺激成骨细胞分化，另一半不进行处理。通过添加50μg/mL l-抗坏血酸、10mMβ-甘油磷酸和0.1μM地塞米松来刺激成骨。将万古霉素以0、0.04、0.4或4mg/mL的浓度添加到细胞培养基中。通过碱性磷酸酶活性（处理后4天）确定早期分化，通过茜素红染色矿化（处理后9天）确定晚期分化。在早期和晚期时间点通过测量甲臜比色产物来确定细胞活力。
高剂量万古霉素处理后，前4天内细胞活力下降，接受4mg/mL万古霉素的细胞与对照组相比活力为40%-60%。随着万古霉素剂量增加，观察到茜素红染色和结节形成逐渐减少。在存在成骨因子的情况下，万古霉素对碱性磷酸酶活性没有有害影响，而与经成骨处理的细胞相比，在不存在成骨因子的情况下观察到活性有降低趋势。
高剂量处理时，万古霉素降低了BMSC活力并损害了晚期成骨分化。因此，高剂量万古霉素对脊柱融合的抑制作用可能是由于BMSC活力降低和成骨分化的某些损害共同导致的。
