Laboratory of Cardiovascular Research and Integrative Pharmacology, Department of Pharmacology, College of Pharmacy, University of Sargodha, Sargodha 40100, Pakistan.
Laboratory of Cardiovascular Research and Integrative Pharmacology, Department of Pharmacology, College of Pharmacy, University of Sargodha, Sargodha 40100, Pakistan.
J Ethnopharmacol. 2019 Jun 12;237:92-107. doi: 10.1016/j.jep.2019.03.019. Epub 2019 Mar 11.
The roots of Ribes orientale (Family Grossulariaceae) have long been used as a folk remedy to treat rheumatism and joints pain in Northern Areas of Pakistan.
The purpose of study was to observe the preventive efficacy of roots of Ribes orientale (RO) aqueous ethanolic extract (30:70) and its aqueous and n-butanol fractions in treating rheumatoid arthritis and to determine its possible mechanism of action.
Arthritis was evaluated in vitro using heat induced bovine serum albumin and egg albumin denaturation and membrane stabilizing assays at 50-6400 μg/ml concentration of extract/fractions whereas, in vivo arthritis was evaluated at 50, 100, 200 mg/kg doses of extract/fractions in formaldehyde model by measuring rat paw volume/diameter. Moreover, highest effective dose (200 mg/kg) of extract/fractions was evaluated in Freünd complete adjuvant (FCA) model. Arthritis was induced in Sprague Dawley rats by immunization with 0.1 ml FCA in left footpad. RO extract/fractions at 200 mg/kg were orally administered from day 0, 30 min prior to adjuvant injection and sustained for 28 days. Paw volume/diameter, arthritic score, body weight, and hematological (WBC, RBC, ESR, Hb and Platelet count) and biochemical (AST, ALT, ALP, urea, creatinine, CRP and RF) parameters were observed. The mRNA expression levels of COX-2, IL-1β, IL-6, NF-kB, TNF-α, IL-4 and IL-10 were measured by real time reverse transcription polymerase chain reaction (RT-PCR) whereas, PGE2 and TNF-α levels in serum samples were measured by Enzyme linked immunosorbent assay (ELISA). Furthermore, radiographs of hind paws and histological changes in ankle joint were analyzed in adjuvant injected rats. The anti-oxidant activity of plant extract and fractions was evaluated using DPPH and reducing power assays. In addition, phytochemistry, total phenolic and flavonoid contents, and HPLC analysis of most active fraction (aqueous fraction) were performed.
Results showed that RO extract and fractions (notably aqueous fraction) significantly reduced protein denaturation and protected erythrocyte membrane in concentration dependent manner. Similarly, extract/fractions induced dose-dependent decrease in paw volume/diameter in the formaldehyde model. Plant extract and fractions significantly suppressed paw swelling and arthritic score, prevented cachexia and remarkably ameliorated hematological and biochemical changes. Furthermore, RO extract/fractions downregulated gene expression levels of PGE2, COX-2, IL-1β, IL-6, NF-kB and TNF-α whereas, upregulated those of IL-4 and IL-10, compared with FCA control rats. The radiographic and histopathologic improvement in joint architecture was also observed in RO treated rats. Piroxicam, used as reference drug, also significantly suppressed arthritis. Additionally, plant exhibited notable anti-oxidant activity and phytochemical analysis revealed the presence of flavonoids and polyphenols.
Results indicated that suppression of pro-inflammatory enzymes/cytokines, inhibition of protein denaturation, lysosomal membrane stabilizing abilities, and redox/free radical scavenging properties of RO extract and fractions support anti-arthritic and immunomodulatory property of Ribes orientale that might be due to its polyphenolic and flavonoid constituents. This suggests that Ribes orientale roots may be used as a therapeutic agent for treating human arthritis.
东北茶藨子（虎耳草科）的根一直被用作民间疗法，用于治疗巴基斯坦北部地区的风湿病和关节疼痛。
本研究的目的是观察东北茶藨子（RO）水醇提取物（30:70）及其水和正丁醇级分在治疗类风湿关节炎中的预防功效，并确定其可能的作用机制。
在 50-6400μg/ml 浓度的提取物/级分的体外条件下，使用热诱导牛血清白蛋白和卵白蛋白变性和膜稳定测定法评估关节炎，而在体内关节炎则使用 50、100、200mg/kg 剂量的提取物/级分在甲醛模型中通过测量大鼠爪体积/直径来评估。此外，在 Freund 完全佐剂（FCA）模型中评估了提取物/级分的最高有效剂量（200mg/kg）。通过在左足底垫中免疫 0.1ml FCA 在 Sprague Dawley 大鼠中诱导关节炎。RO 提取物/级分以 200mg/kg 的剂量从第 0 天开始口服，在佐剂注射前 30 分钟给药，并持续 28 天。测量爪体积/直径、关节炎评分、体重以及血液学（WBC、RBC、ESR、Hb 和血小板计数）和生化参数（AST、ALT、ALP、尿素、肌酐、CRP 和 RF）。通过实时逆转录聚合酶链反应（RT-PCR）测量 COX-2、IL-1β、IL-6、NF-kB、TNF-α、IL-4 和 IL-10 的 mRNA 表达水平，通过酶联免疫吸附测定（ELISA）测量血清样品中的 PGE2 和 TNF-α 水平。此外，分析了佐剂注射大鼠的后爪 X 光片和踝关节的组织学变化。使用 DPPH 和还原能力测定法评估植物提取物和级分的抗氧化活性。此外，进行了植物提取物和级分的植物化学、总酚和类黄酮含量以及最活跃级分（水级分）的 HPLC 分析。
结果表明，RO 提取物和级分（特别是水级分）显著降低了蛋白变性并以浓度依赖的方式保护了红细胞膜。同样，提取物/级分在甲醛模型中诱导了爪体积/直径的剂量依赖性降低。植物提取物和级分显著抑制了爪肿胀和关节炎评分，防止了消瘦，并显著改善了血液学和生化变化。此外，与 FCA 对照组大鼠相比，RO 提取物/级分下调了 PGE2、COX-2、IL-1β、IL-6、NF-kB 和 TNF-α的基因表达水平，而上调了 IL-4 和 IL-10 的基因表达水平。在 RO 治疗的大鼠中也观察到关节结构的放射照相和组织病理学改善。用作参考药物的吡罗昔康也显著抑制了关节炎。此外，植物表现出显著的抗氧化活性，植物化学分析显示存在类黄酮和多酚。
结果表明，RO 提取物和级分抑制致炎酶/细胞因子、抑制蛋白变性、溶酶体膜稳定能力以及还原/自由基清除特性，支持东北茶藨子的抗关节炎和免疫调节特性，这可能是由于其多酚和类黄酮成分。这表明东北茶藨子根可能被用作治疗人类关节炎的治疗剂。
