Wnt4作为新的急性肾损伤早期组织和尿生物标记物及相关机制研究

The mortality of acute kidney injury (AKI) remains high, it mainly due to the lack of sensitive and specific biomarkers for early detection. At the same time, new therapeutic targets still need be further investigated. Therefore, it is very important to find the new biomarkers for early detection and improve therapy and prognosis in AKI. Our previous study discovered that the kidney expression of Wnt4, a member of the Wnt family, increased significantly in ischemia/reperfusion injury (IRI) mouse model. Furthermore, Wnt4 was detected in both kidney and urine in 3 hours post IRI and closely correlated with tubular injury. The increased Wnt4 expression obviously responded earlier than serum creatinine. In the present study, three different kinds of AKI models will be established to observe the expression of Wnt4 in these AKI models induced by various causes. We will investigate the relationship between Wnt4 expression and kidney tubular injury and define whether Wnt4 could be a novel biomarker for early detection of AKI. Primary tubular epithelial cells of these models will be cultured to determine whether epithelial cell injury could be reduced by down regulating PAX2 gene and blocking Wnt/β-catenin and Wnt4/JNK1 pathway．Meanwhile, the injury and repair of epithelial cells after Wnt4 intervention also needed be explored. Furthermore, we will verify whether urinary Wnt4 can also be detected in AKI patients induced by different clinical causes. We hypothesis that Wnt4 could be a new biomarker for early detection and a new target for the therapy of AKI in the future.

急性肾损伤（AKI）死亡率居高不下，主要因为缺乏敏感特异早期诊断指标和新治疗靶点。因此寻找AKI早期诊断生物标记物对诊疗及预后有重要意义。我们前期研究发现肾缺血再灌注肾损伤后Wnt4表达明显增加，肾损伤3小时血肌酐尚未改变，即可于肾组织及尿液中检测到Wnt4，并与肾小管损伤密切相关。本课题拟建立三种不同AKI模型验证Wnt4在不同病因所致AKI中表达情况及Wnt4表达变化与肾小管损伤关系，验证早于血肌酐变化的Wnt4可否作为AKI早期诊断生物标记物。体外培养原代肾小管上皮细胞，采取Wnt4抑制剂进行干预，阐明下调PAX2基因，阻断Wnt/β-catenin和Wnt4/JNK1通路是否可保护肾小管上皮细胞并揭示Wnt4在肾小管上皮细胞损伤中作用机制。同时收集不同病因所致AKI患者血尿标本，从临床研究验证Wnt4是否可作为新的AKI诊断、治疗和预后的生物标记物，为临床AKI早期诊疗提供新靶点。

急性肾损伤（acute kidney injury, AKI）因其高发病率、高死亡率和高治疗成本，已经成为世界性的健康问题。目前AKI的临床诊断主要依赖于血肌酐的升高，然而血肌酐却是一种不敏感且非特异的肾脏损伤指标。因此，我们急需寻找更敏感、更可靠的肾小管损伤标志物，以促进AKI的早期诊治，降低AKI死亡率。Wnt4是一种与肾小管形成相关的分泌性生长因子，于胚胎发育期表达，在成熟肾单位中消失。我们的前期研究发现，肾脏损伤后Wnt4在小管上皮细胞中重新表达，但其发挥的作用及相关机制仍不清楚。本研究中，我们利用缺血再灌注损伤模型及顺铂诱导的AKI模型，证实Wnt4在损伤早期的肾脏及尿液中表达上调，其升高明显早于血肌酐变化，且Wnt4表达水平与肾小管损伤程度成正相关，与肾损伤分子-1（kidney injury molecule-1，KIM-1）表达趋势相似。另外，Wnt4与近、远端肾小管标志物共染，提示Wnt4表达于各个受损的肾小管节段。多数TUNEL阳性及Ki67阳性细胞与Wnt4共定位，说明肾损伤后Wnt4快速活化并参与了肾小管修复及再生。随后我们进一步在临床试验中验证上述结果，发现单纯微小病变性肾病（minimal change disease, MCD）和MCD合并肾小管损伤两组患者的血肌酐及肾小球滤过率（estimated glomerular filtration rate，eGFR）虽然没有明显差异，但在MCD合并肾小管损伤患者的肾脏组织和尿液中，Wnt4表达明显增加，且与肾组织病理改变及KIM-1的表达呈正相关。另外，我们还收集了造影剂致急性肾损伤（contrast-induced acute kidney injury， CI-AKI）患者的尿液标本，发现CI-AKI患者尿Wnt4升高明显早于血肌酐、尿素氮及eGFR的变化。因此，动物实验和临床试验均表明，肾小管损伤后肾组织及尿液中Wnt4的表达明显早于血肌酐的变化，提示尿液Wnt4具有作为无创生物学标志物的潜力，用于临床检测早期AKI，促进急性肾小管损伤的早期诊治，改善患者预后。