Diabetic nephropathy (DN) ranks among the most detrimental long-term effects of diabetes, affecting more than 30% of all patients. Within the diseased kidney, intraglomerular mesangial cells play a key role in facilitating the pro-fibrotic turnover of extracellular matrix components and a progredient glomerular hyperproliferation. These pathological effects are in part caused by an impaired functionality of soluble guanylate cyclase (sGC) and a consequentially reduced synthesis of anti-fibrotic messenger 3′,5′-cyclic guanosine monophosphate (cGMP). Bay 58-2667 (cinaciguat) is able to re-activate defective sGC; however, the drug suffers from poor bioavailability and its systemic administration is linked to adverse events such as severe hypotension, which can hamper the therapeutic effect. In this study, cinaciguat was therefore efficiently encapsulated into virus-mimetic nanoparticles (NPs) that are able to specifically target renal mesangial cells and therefore increase the intracellular drug accumulation. NP-assisted drug delivery thereby increased in vitro potency of cinaciguat-induced sGC stabilization and activation, as well as the related downstream signaling 4- to 5-fold. Additionally, administration of drug-loaded NPs provided a considerable suppression of the non-canonical transforming growth factor β (TGF-β) signaling pathway and the resulting pro-fibrotic remodeling by 50–100%, making the system a promising tool for a more refined therapy of DN and other related kidney pathologies.

We thank Astrid Seefeld, Simon Kerler, Frieder Kees and Annette Garbe for their excellent technical assistance and Frank Schweda (University of Regensburg) for providing primary antibodies. This work was financially supported by the Bavarian State and Sonderforschungsbereich SFB699. CONFLICT OF INTEREST: All authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. DECLARATION OF TRANSPARENCY AND SCIENTIFIC RIGOUR This Declaration acknowledges that this paper adheres to the principles for transparent reporting and scientific rigour of preclinical research as stated in the BJP guidelines for Natural Prod-This article is protected by copyright. All rights reserved. ucts Research, Design and Analysis, Immunoblotting and Immunochemistry, and Animal Experimentation, and as recommended by funding agencies, publishers and other organisations engaged with supporting research. AUTHOR CONTRIBUTIONS: MH and JS were involved in the conception, design and interpretation of the experiments; MH, SP, RS and JS performed and analyzed the experiments; MH and JS wrote the manuscript; all authors were involved in the critical revision of the manuscript for important intellectual content.

Nitric oxide (NO/cyclic guanosine monophosphate (cGMP)-regulated cellular mechanisms are involved in a variety of (patho-) physiological processes. One of the main effector molecules in this system, proteinkinase G (PKG), serves as a molecular switch by phosphorylating different target proteins and thereby turning them on or off. To date, only a few interaction partners of PKG have been described although the identification of protein–protein interactions (PPI) is indispensable for the understanding of cellular processes and diseases. Conventionally used methods to detect PPIs exhibit several disadvantages, e.g., co-immunoprecipitations, which depend on suitable high-affinity antibodies. Therefore, we established a cell-based protein-fragment complementation assay (PCA) for the identification of PKG target proteins. Here, a reporter protein (click beetle luciferase) is split into two fragments and fused to two different possible interaction partners. If interaction occurs, the reporter protein is functionally complemented and the catalyzed reaction can then be quantitatively measured. By using this technique, we confirmed the regulator of G-Protein signaling 2 (RGS2) as an interaction partner of PKGIα (a PKG-isoform) following stimulation with 8-Br-cGMP and 8-pCPT-cGMP. Hence, our results support the conclusion that the established approach could serve as a novel tool for the rapid, easy and cost-efficient detection of novel PKG target proteins.

Clinical presentation of scurvy, magnetic resonance tomography (MRI) imaging of thighs, cytology of bone marrow aspiration in scurvy (A) purpura/panniculitis of the right thigh (left) and diffuse ecchymosis of the left forefoot (middle) and lower leg (right); (B) MRI-imaging of the lower legs with diffuse subcutaneous edema (white arrow-heads) and MRI-signs interpreted as potential myositis (white arrows); (C) hypoplastic cytology in bone marrow aspirate (magnification: 100×, Pappenheim-staining); (D-F) dysplasia of haematopoiesis as described in the manuscript (black arrows) (all magnification 1000×, all Pappenheim-staining) [Colour figure can be viewed at wileyonlinelibrary.com]

No abstract