Mechanisms Up- and Down-Stream of Hypoxia-Inducible Factor in Acute Kidney Injury
Final Report Abstract
According to the unifying hypothesis of Research Unit 1368, local renal hypoxia is a hallmark in acute kidney injury (AKI). Remarkably, adaptation to hypoxia via activation of hypoxia-inducible factors (HIFs) remains incomplete. HIF transcription factors are master regulators of the cellular adaptation to hypoxia. HIF target genes are involved in a complex regulatory network governing metabolism, cell cycle, vascular tone, pH regulation, erythropoiesis, scavenging of free radicals, angiogenesis etc.. The mechanisms behind insufficient renal HIF activation in AKI remained to be unravelled. According to the reviewer comments, our main hypothesis/ aim was to investigate miR-22 as a putative inhibitor of renal HIF-1 activity. In our study, we established a couple of techniques to address our aims. For instance, we successfully established a pmirGLO Dual-Luciferase miR22 Target Expression vector for in vitro test of miR-22 activity in DCT cells. Moreover, in cooperation with Exiqon Inc., we established “miRCURY LNA™ microRNA Inhibitors and Target Site Blocker” for renal miR-22 knockdown in vivo. During our study, we further established protocols to induce more or less severe AKI (indicated by plasma creatinine and molecular kidney injury marker) by rhabdomyolysis. Our results show that, in line with our project hypothesis, HIF-1α mRNA indeed is a miR-22 target. Overexpression of miR-22 as well as inhibition of miR-22 strongly correlated with HIF-1α protein level and HIF-1 activity, demonstrating that miR-22 inhibits HIF-1α expression in vitro and in vivo. However, HIF activation by inhibition of miR-22 fails to protect from rhabdomyolysis-induced AKI. With regard to publication, this in vivo result markedly lowered the impact of our findings. We invested much more time as initially assumed in the project proposal to establish a marked renal miR-22 knockdown in vivo and to explore the miR-22 effect during AKI. The latter was crucial to explain the missing positive influence of HIF activation by miR-22 during AKI. It turned out that inhibition of miR-22 has profound effects on renal gene expression in general. We identified a plethora of strongly regulated genes: in control mice anti-miR-22 molecules up-regulated 998 genes, and down-regulated 915 genes; in rhabdomyolysisinduced AKI, anti-miR-22 molecules up-regulated 1582 genes, and down-regulated 1804 genes. This strong influence on gene expression dominated the HIF response in rhabdomyolysis-induced AKI. Since miR-22 inhibition significantly enhances the HIF response in both controls and AKI, we conclude that indeed, miR-22 acts as a renal HIF repressor. However, suppressing the renal HIF response in kidney injury does not seem to be the leading function of miR-22. Moreover, we observed that inhibition of miR-22 has opposite effects on gene expression in control and AKI mice. Nevertheless, Transcription Factor Pathway Analysis indicated that miR-22 preferentially regulates genes involved in development and differentiation. Based on our data we assume that the miR-22/ HIF axis, thus, might be more relevant during kidney development and in renal cancer, which needs detailed investigation in future studies. In sum, we confirmed that, according to our hypothesis, miR-22 inhibits the renal HIF response. Although we demonstrated that miR-22 is not a useful target to prevent kidney injury, we present a couple of new and unexpected findings regarding renal miR-22 function.
Publications
- Selective endothelin inhibition in diabetic nephropathy: is it the icing on the cake? Acta Physiol (Oxf) 212(1): 1-4, 2014
Rosenberger C, Fähling M
(See online at https://doi.org/10.1111/apha.12332) - Urinary calprotectin: a new diagnostic marker in urothelial carcinoma of the bladder. World J Urol 32(6): 1485-1492, 2014
Ebbing J, Mathia S, Seibert FS, Pagonas N, Bauer F, Erber B, Günzel K, Kilic E, Kempkensteffen C, Miller K, Bachmann A, Rosenberger C, Zidek W, Westhoff TH
(See online at https://doi.org/10.1007/s00345-013-1227-8) - Endothelin-converting enzyme is a plausible target gene for hypoxia-inducible factor. Kidney Int 87(4): 761-770, 2015
Khamaisi M, Toukan H, Axelrod JH, Rosenberger C, Skarzinski G, Shina A, Meidan R, Koesters R, Rosen S, Walkinshaw G, Mimura I, Nangaku M, Heyman SN
(See online at https://doi.org/10.1038/ki.2014.362) - Cyclosporin a induces renal episodic hypoxia. Acta Physiol (Oxf) 219(3): 625-639, 2017
Fähling M, Mathia S, Scheidl J, Abramovitch R, Milman Z, Paliege A, Peters H, Persson PB, Heyman SN, Rosenberger C
(See online at https://doi.org/10.1111/apha.12811) - Myoglobin facilitates angiotensin II-induced constriction of renal afferent arterioles. Am J Physiol Renal Physiol 312(5): F908-F916, 2017
Liu ZZ, Mathia S, Pahlitzsch T, Wennysia IC, Persson PB, Lai EY, Högner A, Xu MZ, Schubert R, Rosenberger C, Patzak A
(See online at https://doi.org/10.1152/ajprenal.00394.2016) - Potential Hypoxic Renal Injury in Patients With Diabetes on SGLT2 Inhibitors: Caution Regarding Concomitant Use of NSAIDs and Iodinated Contrast Media. Diabetes Care 40(4): e40-e41, 2017
Heyman SN, Khamaisi M, Rosen S, Rosenberger C, Abassi Z
(See online at https://doi.org/10.2337/dc16-2200) - Understanding and preventing contrast-induced acute kidney injury. Nat Rev Nephrol 13(3): 169-180, 2017
Fähling M, Seeliger E, Patzak A, Persson PB
(See online at https://doi.org/10.1038/nrneph.2016.196) - Urinary Calprotectin Differentiates Between Prerenal and Intrinsic Acute Renal Allograft Failure. Transplantation 101(2): 387-394, 2017
Seibert FS, Rosenberger C, Mathia S, Arndt R, Arns W, Andrea H, Pagonas N, Bauer F, Zidek W, Westhoff TH
(See online at https://doi.org/10.1097/TP.0000000000001124) - A dual role of miR-22 in rhabdomyolysis-induced acute kidney injury. Acta Physiol (Oxf) 224(3): e13102, 2018
Mathia S, Rudigier LJ, Kasim M, Kirschner KM, Persson PB, Eckardt KU, Rosenberger C, Fähling M
(See online at https://doi.org/10.1111/apha.13102) - Cyclosporine A induces endothelin-converting enzyme-1: Studies in vivo and in vitro. Acta Physiol (Oxf) 223(1): e13033, 2018
Heyman SN, Abassi Z, Rosenberger C, Yaseen H, Skarjinski G, Shina A, Mathia S, Krits N, Khamaisi M
(See online at https://doi.org/10.1111/apha.13033)
