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Abstract：
BACKGROUND: Chronic kidney disease (CKD) and its associated end-stage renal disease (ESRD) are significant health problems that pose a threat to human well-being. Renal fibrosis is a common feature and ultimate pathological outcome of various CKD leading to ESRD. The Astragalus mongholicus Bunge and Panax notoginseng formula (A&P) is a refined compound formulated by our research group, which has been clinically administered for over a decade and has demonstrated the ability to improve the inflammatory state of various acute or chronic kidney diseases. However, the underlying mechanism by which A&P ameliorates renal fibrosis remains unclear.
METHODS: We established a mouse model by surgically ligating the unilateral ureter to induce renal injury in vivo. And we utilized renal in situ electroporation of a plasmid with low LncRNA A33 expression to establish the unilateral ureteral obstruction(UUO)mouse model. In vitro, we stimulated primary tubular epithelial cells(pTEC) injury using TGF-β1, siRNA-A33, and pcDNA3.1-A33 plasmids were transfected into pTECs to respectively knockdown and overexpress LncRNA A33, and both in vitro and in vivo models were intervened with A&P.
RESULTS: The results demonstrated that A&P effectively alleviated renal fibrosis in mice. Subsequent findings indicated high expression of LncRNA A33 in the kidneys of UUO mice and TGF-β1-induced renal tubular cells. In situ, renal electroporation of a plasmid with reduced LncRNA A33 expression revealed that inhibiting LncRNA A33 significantly improved renal fibrosis in UUO mice. Moreover, A&P effectively suppressed LncRNA A33 expression both in vitro and in vivo. Subsequent downregulation of LncRNA A33 in renal tubular epithelial cells resulted in the downregulation of numerous fibrotic markers, a significant inhibition of LncRNA A33, and a notable reduction in downstream ferroptosis signaling. Cell experiments demonstrated that A&P improved renal fibrosis in UUO mice by inhibiting LncRNA A33 and downregulating ferroptosis signaling.
CONCLUSIONS: Through the inhibition of LncRNA A33 and subsequent downregulation of ferroptosis signaling, A&P showed potential as a therapeutic approach for improving renal fibrosis in UUO mice, providing a potential treatment avenue for CKD.
METHODS: We established a mouse model by surgically ligating the unilateral ureter to induce renal injury in vivo. And we utilized renal in situ electroporation of a plasmid with low LncRNA A33 expression to establish the unilateral ureteral obstruction(UUO)mouse model. In vitro, we stimulated primary tubular epithelial cells(pTEC) injury using TGF-β1, siRNA-A33, and pcDNA3.1-A33 plasmids were transfected into pTECs to respectively knockdown and overexpress LncRNA A33, and both in vitro and in vivo models were intervened with A&P.
RESULTS: The results demonstrated that A&P effectively alleviated renal fibrosis in mice. Subsequent findings indicated high expression of LncRNA A33 in the kidneys of UUO mice and TGF-β1-induced renal tubular cells. In situ, renal electroporation of a plasmid with reduced LncRNA A33 expression revealed that inhibiting LncRNA A33 significantly improved renal fibrosis in UUO mice. Moreover, A&P effectively suppressed LncRNA A33 expression both in vitro and in vivo. Subsequent downregulation of LncRNA A33 in renal tubular epithelial cells resulted in the downregulation of numerous fibrotic markers, a significant inhibition of LncRNA A33, and a notable reduction in downstream ferroptosis signaling. Cell experiments demonstrated that A&P improved renal fibrosis in UUO mice by inhibiting LncRNA A33 and downregulating ferroptosis signaling.
CONCLUSIONS: Through the inhibition of LncRNA A33 and subsequent downregulation of ferroptosis signaling, A&P showed potential as a therapeutic approach for improving renal fibrosis in UUO mice, providing a potential treatment avenue for CKD.
摘要：
背景：慢性肾病（CKD）及其相关的终末期肾病（ESRD）是对人类福祉构成威胁的重大健康问题。肾纤维化是导致ESRD的各种CKD的共同特征和最终病理结果。黄芪和三七配方（A＆P）是由我们的研究小组配制的精制化合物，已在临床上给药超过十年，并已证明能够改善各种急性或慢性肾脏疾病的炎症状态。然而,A&P改善肾纤维化的潜在机制尚不清楚.
方法：我们通过手术结扎单侧输尿管以体内诱导肾损伤建立了小鼠模型。我们利用低LncRNAA33表达质粒的肾脏原位电穿孔建立单侧输尿管梗阻（UUO）小鼠模型。体外，我们使用TGF-β1，siRNA-A33和pcDNA3.1-A33质粒刺激原代肾小管上皮细胞（pTEC）损伤，将质粒转染到pTEC中，分别敲低和过表达LncRNAA33，并在体外和体内模型中进行干预。
结果：结果表明A&P能有效缓解小鼠肾纤维化。随后的发现表明LncRNAA33在UUO小鼠的肾脏和TGF-β1诱导的肾小管细胞中高表达。在原地,LncRNAA33表达降低的质粒的肾电穿孔显示,抑制LncRNAA33显著改善UUO小鼠的肾纤维化。此外，A&P在体外和体内均有效抑制LncRNAA33表达。随后LncRNAA33在肾小管上皮细胞中的下调导致许多纤维化标志物的下调,LncRNAA33的显着抑制，以及下游铁凋亡信号的显着减少。细胞实验表明,A&P通过抑制LncRNAA33和下调铁凋亡信号传导来改善UUO小鼠的肾纤维化。
结论：通过抑制LncRNAA33和随后下调铁凋亡信号，A&P显示出作为改善UUO小鼠肾纤维化的治疗方法的潜力，为CKD提供潜在的治疗途径。
方法：我们通过手术结扎单侧输尿管以体内诱导肾损伤建立了小鼠模型。我们利用低LncRNAA33表达质粒的肾脏原位电穿孔建立单侧输尿管梗阻（UUO）小鼠模型。体外，我们使用TGF-β1，siRNA-A33和pcDNA3.1-A33质粒刺激原代肾小管上皮细胞（pTEC）损伤，将质粒转染到pTEC中，分别敲低和过表达LncRNAA33，并在体外和体内模型中进行干预。
结果：结果表明A&P能有效缓解小鼠肾纤维化。随后的发现表明LncRNAA33在UUO小鼠的肾脏和TGF-β1诱导的肾小管细胞中高表达。在原地,LncRNAA33表达降低的质粒的肾电穿孔显示,抑制LncRNAA33显著改善UUO小鼠的肾纤维化。此外，A&P在体外和体内均有效抑制LncRNAA33表达。随后LncRNAA33在肾小管上皮细胞中的下调导致许多纤维化标志物的下调,LncRNAA33的显着抑制，以及下游铁凋亡信号的显着减少。细胞实验表明,A&P通过抑制LncRNAA33和下调铁凋亡信号传导来改善UUO小鼠的肾纤维化。
结论：通过抑制LncRNAA33和随后下调铁凋亡信号，A&P显示出作为改善UUO小鼠肾纤维化的治疗方法的潜力，为CKD提供潜在的治疗途径。
