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目的通过考查甘肃习用药材黄管秦艽栽培种和野生种中总裂环烯醚萜苷对白细胞介素-1β(IL-1β)诱导的大鼠软骨细胞的影响,探讨栽培种和野生种黄管秦艽中总裂环烯醚萜苷治疗骨关节炎(osteoarthritis,OA)的可行性。方法取大鼠关节软骨细胞第4代,将第4代软骨细胞分为4组:空白组(A组)软骨细胞用完全培养基培养;白细胞介素-1β诱导组(B组)软骨细胞以10ng·mL-1重组大鼠白细胞介素-1β培养24 h;白细胞介素-1β和栽培种黄管秦艽总裂环烯醚萜苷组(C组:分为C1组50μg·mL-1;C2组200μg·mL-1;C3组400μg·mL-1)软骨细胞以10 ng·mL-1重组大鼠白细胞介素-1β和栽培种黄管秦艽总裂环烯醚萜苷(以下称栽培组)培养24 h;白细胞介素-1β和野生种黄管秦艽总裂环烯醚萜苷组(D组:分为D1组50μg·mL-1;D2组100μg·mL-1;D3组200μg·mL-1)软骨细胞以10 ng·mL-1重组大鼠白细胞介素-1β和野生种黄管秦艽总裂环烯醚萜苷(以下称野生组)培养24 h。培养后取各组细胞进行免疫细胞化学染色,收集细胞培养液,检测其上清液中表达前列腺素E2的情况。结果四甲基偶氮唑蓝法结果显示,栽培组(50、200、400μg·mL-1)对软骨细胞作用24 h时无毒性作用,野生组(50、100、200μg·mL-1)对软骨细胞作用24 h时无毒性作用。免疫细胞化学染色显示,栽培组和野生组与诱导组相比Ⅱ型胶原阳性表达明显增强,但弱于空白组,并且随着质量浓度的增大Ⅱ型胶原阳性表达逐渐增强,差异有统计学意义(P<0.05)。酶联免疫法结果显示,栽培组和野生组与诱导组相比前列腺素E2含量明显降低,但高于空白组,并且随着质量浓度的增大前列腺素E2含量越低,差异有统计学意义(P<0.05)。结论栽培种和野生种黄管秦艽中的总裂环烯醚萜苷均对炎症软骨细胞具有一定的保护作用。
Objective To investigate the effect of total iridoid glycosides of genus Cynanchum cv. Gansu on the chondrocytes induced by interleukin-1β (IL-1β) Feasibility of total iridoid glycosides in treatment of osteoarthritis (OA) in Gentiana macrophylla. Methods The 4th passage of rat articular chondrocytes was divided into 4 groups: the chondrocytes of the blank group (group A) were cultured in complete medium; the chondrocytes of the group of interleukin-1β (group B) The recombinant interleukin-1β of 10ng · mL-1 was cultured for 24 h. Interleukin-1β and the cultivated species of total iridoidoside of Cynanchum paniculata (C group: divided into C1 group, 50μg · mL-1; C2 group 200μg · mL-1; C3 group 400μg · mL-1) chondrocytes with 10 ng · mL-1 recombinant rat interleukin-1β and cultivated species of yellow tube Gentiana selengeridum glycosides (hereinafter referred to as cultivation (D group: divided into D1 group 50μg · mL-1; D2 group 100μg · mL-1; D3 group 200μg · ML-1) chondrocytes were cultured for 24 h with 10 ng · mL-1 recombinant rat interleukin-1β and wild-type O.aureus total iridoid glycoside (hereinafter referred to as wild group). After culturing, the cells in each group were subjected to immunocytochemical staining, and the cell culture fluid was collected to detect the expression of prostaglandin E2 in the supernatant. Results The results of MTT assay showed that the culture group (50, 200, 400 μg · mL-1) had no toxic effect on chondrocytes for 24 h, while the wild group (50, 100, 200 μg · mL- Chondrocytes 24 h without toxic effect. Immunocytochemical staining showed that compared with the blank group, the positive expression of type Ⅱ collagen in the cultivated group and the wild group was significantly increased compared with the induction group, and the positive expression of type Ⅱ collagen gradually increased with the increase of mass concentration, the difference was statistically significant Significance (P <0.05). Enzyme-linked immunosorbent assay showed that prostaglandin E2 levels in cultivated and wild groups were significantly lower than those in induction group, but higher than that in blank group. The lower the content of prostaglandin E2 was, the higher the concentration was and the difference was statistically significant (P <0.05). CONCLUSIONS The total iridoid glycosides in cultivated and wild species of Brachiaria ficuum have protective effects on inflammatory chondrocytes.