Department of Molecular and Cell Biology, Boston University Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA.
Department of Periodontology, Boston University Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA.
Int J Mol Sci. 2019 Sep 27;20(19):4798. doi: 10.3390/ijms20194798.
The goal of this study was to determine if adenovirus-delivered LOXL2 protects against progressive knee osteoarthritis (OA), assess its specific mechanism of action; and determine if the overexpression of LOXL2 in transgenic mice can protect against the development of OA-related cartilage damage and joint disability.
Four-month-old Cho/+ male and female mice were intraperitoneally injected with either Adv-RFP-LOXL2 or an empty vector twice a month for four months. The proteoglycan levels and the expression of anabolic and catabolic genes were examined by immunostaining and qRT-PCR. The effect of LOXL2 expression on signaling was tested via the pro-inflammatory cytokine IL1β in the cartilage cell line ATDC5. Finally; the OA by monosodium iodoacetate (MIA) injection was also induced in transgenic mice with systemic overexpression of LOXL2 and examined gene expression and joint function by treadmill tests and assessment of allodynia.
The adenovirus treatment upregulated LOXL2; Sox9; Acan and Runx2 expression in both males and females. The Adv-RFP-LOXL2 injection; but not the empty vector injection increased proteoglycan staining and aggrecan expression but reduced MMP13 expression. LOXL2 attenuated IL-1β-induced phospho-NF-κB/p65 and rescued chondrogenic lineage-related genes in ATDC5 cells; demonstrating one potential protective mechanism. LOXL2 attenuated phospho-NF-κB independent of its enzymatic activity. Finally; LOXL2-overexpressing transgenic mice were protected from MIA-induced OA-related functional changes; including the time and distance traveled on the treadmill and allodynia.
Our study demonstrates that systemic LOXL2 adenovirus or LOXL2 genetic overexpression in mice can protect against OA. These findings demonstrate the potential for LOXL2 gene therapy for knee-OA clinical treatment in the future.
本研究旨在确定腺病毒递送的 LOXL2 是否能预防进行性膝骨关节炎（OA），评估其具体的作用机制，并确定 LOXL2 在转基因小鼠中的过表达是否能预防 OA 相关软骨损伤和关节失能。
4 月龄 Cho/+ 雄性和雌性小鼠每月两次经腹腔注射 Adv-RFP-LOXL2 或空载体共 4 个月。通过免疫染色和 qRT-PCR 检测蛋白聚糖水平和合成代谢与分解代谢基因的表达。通过 IL1β 刺激软骨细胞系 ATDC5 来测试 LOXL2 表达对信号的影响。最后，通过系统性过表达 LOXL2 在转基因小鼠中诱导单碘乙酸（MIA）注射 OA，并通过跑步机试验和评估痛觉过敏来检测基因表达和关节功能。
腺病毒处理上调了雄性和雌性小鼠 LOXL2、Sox9、Acan 和 Runx2 的表达。Adv-RFP-LOXL2 注射而非空载体注射增加了蛋白聚糖染色和聚集蛋白聚糖表达，但减少了 MMP13 表达。LOXL2 减弱了 IL-1β 诱导的磷酸化 NF-κB/p65，并在 ATDC5 细胞中挽救了软骨形成相关基因，表明了一种潜在的保护机制。LOXL2 减弱了磷酸化 NF-κB 而不依赖其酶活性。最后，LOXL2 过表达转基因小鼠免受 MIA 诱导的 OA 相关功能变化的影响，包括在跑步机上的时间和距离以及痛觉过敏。
我们的研究表明，全身 LOXL2 腺病毒或 LOXL2 基因过表达在小鼠中可预防 OA。这些发现为未来 LOXL2 基因治疗膝关节 OA 的临床治疗提供了潜力。
