preparing paraffin sections of kidney and peyer&#39;s patch isolated from an igan murine model for histological and immunohistochemical analyses

<meta charset="utf8"></head><body>在小鼠模型中用 Dioscin 治疗免疫球蛋白 A 肾病 （IgAN）

IgA Nephropathy (IgAN) is the most common type of primary glomerulonephritis, for which there is no specific treatment, and it remains a significant cause of end-stage renal disease1. Although the pathogenesis of IgAN is still not fully understood, the &#34;multi-hit hypothesis&#34; is generally accepted and supported by a large body of clinical and experimental research evidence2. The pathogenesis of IgAN involves activating B cells and producing Galactose-deficient IgA1 (Gd-IgA1)3. The increase in circulating Gd-IgA1 due to the excessive proliferation and activation of IgA-secreting cells during the mucosal immune response is a critical link in the pathogenesis of IgAN4,5,6. As the central place for the proliferation and activation of B lymphocyte phenotype conversion to IgA-secreting cells, Peyer&#39;s patch is the primary source of IgA secretion, closely related to the occurrence and development of IgAN7,8. In addition, the proliferation of IgA1-secreting cells, as well as expression of Core 1&#946;-1,3-galactosyltransferase(C1GalT1) and C1GalT1-specific molecular chaperone (Cosmc), were associated with abnormal glycosylation of IgA1, which causes GD-IgA1 production in IgAN patients6,9.
Clinical study on IgAN treatment with herbal medicine has progressed in recent years. Yiqi Qingjie Formula is an essential formula for treating IgAN by the Department of Nephrology of Guang&#39;anmen Hospital. The previous study of our group found that Gd-IgA1 decreased in the serum of IgAN patients after treatment with Yiqi Qingjie Formula. As one of the most used herbs in the Yiqi Qingjie Formula, Dioscoreae Nipponicae Rhizoma (DNR) is the dried rhizome of Dioscorea Nipponica Makino, which has various functions such as regulating immunity, suppressing inflammation, relieving cough and asthma10,11. Several scholars treated IgAN with DNR and achieved good results12,13,14. As the main active ingredient in DNR15, Dioscin (DIO) lowers uric acid, inhibits fibrosis, inhibits inflammatory response, and anti-oxidative stress16,17. Therefore, DIO may have a novel action mechanism to inhibit the cellular secretion of excessive Gd-IgA1 and exert specific kidney protection effects. Still, no study has been reported on DIO&#39;s action mechanism for treating IgAN.
To explore the potential therapeutic mechanism of DIO on IgAN and provide a new method for the treatment of IgAN, we carried out experiments for the therapeutic effects of DIO on IgAN in vivo and in vitro.

<meta charset="utf8"></head><body>作者聚焦：研究中草药活性薯片素治疗 IgA 肾病的潜力

Dioscin 通过抑制体内 B 细胞活化和减少体外半乳糖缺乏型 IgA1 的产生来缓解 IgA 肾病

制备从 IgAN 鼠模型中分离的肾脏和 Peyer 贴剂的石蜡切片，用于组织学和免疫组织化学分析

preparing-paraffin-sections-kidney-peyer-s-patch-isolated-from-an

Research

JoVE Journal

Medicine

Preparing Paraffin Sections of Kidney and Peyer&#39;s Patch Isolated from an IgAN Murine Model for Histological and Immunohistochemical Analyses

231 次观看. 要开始准备从适当麻醉的 IgA 神经病变模型小鼠中分离的肾脏和 Peyer 贴片的石蜡切片，首先用 4% 多聚甲醛固定一个 Peyer 贴片或 3 毫米厚的肾脏组织 24 小时。然后使用乙醇和二甲苯的梯度将它们脱水，然后用水清洗它们。将组织浸入蜡中两小时，然后密封和冷冻。接下来，切下 4 微米厚的 Peyer 贴片切片或 2 微米厚的肾脏切片，并将它们放入温?...

视频: 制备从 IgAN 鼠模型中分离的肾脏和 Peyer 贴剂的石蜡切片，用于组织学和免疫组织化学分析

Dioscin Mediated IgA Nephropathy Alleviation by Inhibiting B Cell Activation In Vivo and Decreasing Galactose-Deficient IgA1 Production In Vitro

The increase of circulating galactose-deficient IgA1 (Gd-IgA1) is caused by excessive activation of IgA-positive secretory cells in the process of mucosal immune responses, which is a critical link in the pathogenesis of IgA nephropathy (IgAN). Peyer&#39;s patch, the prominent place where B lymphocytes are transformed into IgA-secreting plasma cells, is the primary source of IgA. In addition, the lower expression of core 1&#946;-1,3-galactosyltransferase (C1GalT1) and its molecular chaperone, C1GalT1-specific molecular chaperone (Cosmc), is related to abnormal glycosylation of IgA1 in IgAN patients. Our clinical experience shows that Dioscoreae Nipponicae Rhizoma&#39;s (DNR) herbal medicine can relieve proteinuria and hematuria and improve renal function in IgAN patients. Dioscin (DIO) is one of the main active ingredients of DNR, which has various pharmacological activities. This study explores DIO&#39;s possible mechanism in treating IgAN.The IgAN model mouse was established by mucosal immune induction. The mice were divided into the control, model, and DIO gavage groups. The glomerular IgA deposition in mice, renal pathological changes, and B cell markers CD20 and CXCR5 expression in Peyer&#39;s patch were detected by immunofluorescence and immunohistochemistry. After lipopolysaccharide (LPS) stimulation, DIO&#39;s effects on DAKIKI cells proliferation, IgA and Gd-IgA1 secretion, C1GalT1, and Cosmc expression were studied by cell counting kit-8 (CCK-8) assay, enzyme-linked immunosorbent assay (ELISA) test, quantitative real-time polymerase chain reaction (QRT-PCR), and western blotting (WB). In in vivo studies, IgA deposition accompanied by glomerular mesangial hyperplasia and increased expression of CD20 and CXCR5 in Peyer&#39;s patch in the IgAN model mouse was alleviated by DIO. In vitro studies showed 0.25 &#181;g/mL to 1.0 &#181;g/mL DIO inhibited LPS-induced DAKIKI cell proliferation, IgA and Gd-IgA1 secretion, and up-regulated the mRNA and protein expression of C1GalT1 and Cosmc. This study demonstrates that DIO may reduce Gd-IgA1 production by inhibiting excessive activation of IgA-secreting cells and up-regulating C1GALT1/Cosmc expression.

This study provides experimental data for treating immunoglobulin A nephropathy (IgAN) with Dioscin (DIO), the active ingredient of Dioscoreae Nipponicae Rhizoma (DNR), and a paradigm for studying herbal medicine's effects and underlying mechanisms in vivo and in vitro.

The increase of circulating galactose-deficient IgA1 (Gd-IgA1) is caused by excessive activation of IgA-positive secretory cells in the process of mucosal ...

科研

医学

Preparing IgAN Model Mice for the Dioscin-Mediated IgA Nephropathy Alleviation Experiment

为 Dioscin 介导的 IgA 肾病缓解实验准备 IgAN 模型小鼠

Histological Analysis of Kidney Tissue and Peyer&#39;s Patch Isolated from an IgAN Murine Model

从 IgAN 小鼠模型中分离的肾组织和 Peyer 斑的组织学分析

Immunohistochemical Analysis of Peyer&#39;s Patch Isolated from an IgAN Murine Model

从 IgAN 鼠模型中分离的 Peyer 贴片的免疫组织化学分析

Assays for Screening Safe Concentrations and Effects of DIO on Normal DAKIKI Cells

筛选 DIO 对正常 DAKIKI 细胞的安全浓度和影响的测定

Detecting the Effect of DIO on the mRNA and Protein Levels of C1GALT1 and Cosmc in DAKIKI Cells

检测 DIO 对 DAKIKI 细胞中 C1GALT1 和 Cosmc 的 mRNA 和蛋白水平的影响