Swansea University Medical School, Swansea University, Swansea, UK.
Department of Animal Sciences, University of Florida, Gainesville, Florida, USA.
Reproduction. 2022 Aug 5;164(3):109-123. doi: 10.1530/REP-22-0032. Print 2022 Sep 1.
Bovine granulosa cells need to be cultured with serum to generate inflammation in response to bacterial lipopolysaccharide. This study shows that it is cholesterol that facilitates this lipopolysaccharide-stimulated cytokine secretion.
During bacterial infections of the bovine uterus or mammary gland, ovarian granulosa cells mount inflammatory responses to lipopolysaccharide (LPS). In vitro, LPS stimulates granulosa cell secretion of the cytokines IL-1α and IL-1β and the chemokine IL-8. These LPS-stimulated inflammatory responses depend on culturing granulosa cells with serum, but the mechanism is unclear. Here, we tested the hypothesis that cholesterol supports inflammatory responses to LPS in bovine granulosa cells. We used granulosa cells isolated from 4 to 8 mm and >8.5 mm diameter ovarian follicles and manipulated the availability of cholesterol. We found that serum or follicular fluid containing cholesterol increased LPS-stimulated secretion of IL-1α and IL-1β from granulosa cells. Conversely, depleting cholesterol using methyl-β-cyclodextrin diminished LPS-stimulated secretion of IL-1α, IL-1β and IL-8 from granulosa cells cultured in serum. Follicular fluid contained more high-density lipoprotein cholesterol than low-density lipoprotein cholesterol, and granulosa cells expressed the receptor for high-density lipoprotein, scavenger receptor class B member 1 (SCARB1). Furthermore, culturing granulosa cells with high-density lipoprotein cholesterol, but not low-density lipoprotein or very low-density lipoprotein cholesterol, increased LPS-stimulated inflammation in granulosa cells. Cholesterol biosynthesis also played a role in granulosa cell inflammation because RNAi of mevalonate pathway enzymes inhibited LPS-stimulated inflammation. Finally, treatment with follicle-stimulating hormone, but not luteinising hormone, increased LPS-stimulated granulosa cell inflammation, and follicle-stimulating hormone increased SCARB1 protein. However, changes in inflammation were not associated with changes in oestradiol or progesterone secretion. Taken together, these findings imply that cholesterol supports inflammatory responses to LPS in granulosa cells.
牛颗粒细胞需要在血清中培养才能对细菌脂多糖产生炎症反应。本研究表明，正是胆固醇促进了脂多糖刺激的细胞因子分泌。
在牛子宫或乳腺的细菌感染中，卵巢颗粒细胞对脂多糖（LPS）产生炎症反应。在体外，LPS 刺激颗粒细胞分泌白细胞介素-1α（IL-1α）和白细胞介素-1β（IL-1β）以及趋化因子白细胞介素-8（IL-8）。这些 LPS 刺激的炎症反应依赖于颗粒细胞在血清中的培养，但机制尚不清楚。在这里，我们检验了胆固醇支持牛颗粒细胞对 LPS 炎症反应的假设。我们使用了从 4 至 8 毫米和> 8.5 毫米直径的卵泡中分离出的颗粒细胞，并操纵了胆固醇的可用性。我们发现，含有胆固醇的血清或卵泡液增加了 LPS 刺激的颗粒细胞分泌的 IL-1α 和 IL-1β。相反，使用甲基-β-环糊精消耗胆固醇会减少在含血清的培养基中 LPS 刺激的 IL-1α、IL-1β 和 IL-8 的分泌。卵泡液中的高密度脂蛋白胆固醇含量高于低密度脂蛋白胆固醇，颗粒细胞表达高密度脂蛋白受体，即清道夫受体 B 类成员 1（SCARB1）。此外，用高密度脂蛋白胆固醇而不是低密度脂蛋白或极低密度脂蛋白胆固醇培养颗粒细胞会增加 LPS 刺激的颗粒细胞炎症。胆固醇生物合成在颗粒细胞炎症中也发挥了作用，因为甲羟戊酸途径酶的 RNAi 抑制了 LPS 刺激的炎症。最后，卵泡刺激素（FSH）处理而不是黄体生成素（LH）处理会增加 LPS 刺激的颗粒细胞炎症，FSH 增加了 SCARB1 蛋白。然而，炎症的变化与雌二醇或孕酮分泌的变化无关。总之，这些发现表明胆固醇支持颗粒细胞对 LPS 的炎症反应。
