Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.
BMC Complement Altern Med. 2013 Aug 23;13:213. doi: 10.1186/1472-6882-13-213.
Hyaluronidases have been found as the target enzymes in the development of osteoarthritis (OA) disease. While there is still no curative treatment for this disease, recent studies on the treatment of OA were focused on the effectiveness of natural products which are expected to improve the symptoms with minimal side effects. The aim of this study was to screen selected Malaysian plants on their anti-hyaluronidase activity as well as to evaluate the active plant and its derived fractions on its potential anti-arthritic and antioxidant activities.
A total of 20 methanolic crude extracts (bark and leaf) from ten different plants were screened using a colorimetric hyaluronidase enzymatic assay. The active plant extract (Payena dasyphylla) was then studied for its hyaluronidase inhibitory activity in the interleukin-1β (IL-1β) stimulated human chondrocytes cell line (NHAC-kn) using zymography method. The Payena dasyphylla methanolic bark extract was then fractionated into several fractions in where the ethyl acetate (EA) fraction was evaluated for its inhibitory effects on the HYAL1 and HYAL2 gene expressions using reverse transcription-polymerase chain reaction (RT-PCR) technique. While the MMP-3 and MMP-13 protein expressions were evaluated using western blot method. The phenolic and flavonoid contents of the three fractions as well as the antioxidant property of the EA fraction were also evaluated.
Bark extract of Payena dasyphylla (100 μg/ml) showed the highest inhibitory activity against bovine testicular hyaluronidase with 91.63%. The plant extract also inhibited hyaluronidase expression in the cultured human chondrocyte cells in response to IL-1β (100 ng/ml). Similarly, treatment with Payena dasyphylla ethyl acetate (EA) fraction (100 μg/ml) inhibited the HYAL1 and HYAL2 mRNA gene expressions as well as MMP-3 and MMP-13 protein expression in a dose dependent manner. Payena dasyphylla EA fraction has demonstrated the highest amount of phenolic and flavonoid content with 168.62 ± 10.93 mg GAE/g and 95.96 ± 2.96 mg RE/g respectively as compared to water and hexane fractions. In addition, the Payena dasyphylla EA fraction showed strong antioxidant activity with IC₅₀ value of 11.64 ± 1.69 μg/mL.
These findings have shown that Payena dasyphylla might contained potential phenolic compounds that inhibiting the key enzyme in osteoarthritis development, which is the hyaluronidase enzyme through interruption of HYAL1 and HYAL1 gene expressions. The degradation of cartilage could also be inhibited by the plant through suppression of MMP-3 and MMP-13 protein expressions. We also reported that the inhibitory effect of Payena dasyphylla on hyaluronidase activity and expression might be due to its anti-oxidant property.
透明质酸酶已被发现为骨关节炎（OA）疾病发展的靶酶。虽然这种疾病仍然没有治愈的方法，但最近对 OA 的治疗研究集中在天然产物的有效性上，这些天然产物有望在最小副作用的情况下改善症状。本研究的目的是筛选选定的马来西亚植物对透明质酸酶的活性，以及评估活性植物及其衍生部分对潜在的抗关节炎和抗氧化活性。
用比色法透明质酸酶酶法对来自十种不同植物的 20 种甲醇粗提取物（树皮和叶）进行筛选。然后，用白细胞介素-1β（IL-1β）刺激的人软骨细胞系（NHAC-kn）中的酶谱法研究活性植物提取物（Payena dasyphylla）对透明质酸酶的抑制活性。然后，将 Payena dasyphylla 甲醇树皮提取物分成几个部分，其中乙酸乙酯（EA）部分用逆转录-聚合酶链反应（RT-PCR）技术评估对 HYAL1 和 HYAL2 基因表达的抑制作用。同时，用 Western blot 法评估 MMP-3 和 MMP-13 蛋白的表达。还评估了三个部分的酚类和类黄酮含量以及 EA 部分的抗氧化特性。
Payena dasyphylla（100μg/ml）树皮提取物对牛睾丸透明质酸酶的抑制活性最高，为 91.63%。该植物提取物还抑制了培养的人软骨细胞对 IL-1β（100ng/ml）的透明质酸酶表达。同样，Payena dasyphylla 乙酸乙酯（EA）部分（100μg/ml）以剂量依赖性方式抑制 HYAL1 和 HYAL2 mRNA 基因表达以及 MMP-3 和 MMP-13 蛋白表达。与水和己烷部分相比，Payena dasyphylla EA 部分表现出最高量的酚类和类黄酮含量，分别为 168.62±10.93mgGAE/g和 95.96±2.96mgRE/g。此外，Payena dasyphylla EA 部分表现出较强的抗氧化活性，IC₅₀值为 11.64±1.69μg/ml。
这些发现表明，Payena dasyphylla 可能含有潜在的酚类化合物，通过阻断 HYAL1 和 HYAL1 基因表达，抑制骨关节炎发展的关键酶，即透明质酸酶。通过抑制 MMP-3 和 MMP-13 蛋白的表达，植物也可以抑制软骨的降解。我们还报告说，Payena dasyphylla 对透明质酸酶活性和表达的抑制作用可能归因于其抗氧化特性。
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