Switching cell penetrating and CXCR4-binding activities of nanoscale-organized arginine-rich peptides

Arginine-rich protein motifs have been described as potent cell-penetrating peptides (CPPs) but also as rather specific ligands of the cell surface chemokine receptor CXCR4, involved in the infection by the human immunodeficiency virus (HIV). Polyarginines are commonly used to functionalize nanoscale vehicles for gene therapy and drug delivery, aimed to enhance cell penetrability of the therapeutic cargo. However, under which conditions these peptides do act as either unspecific or specific ligands is unknown. We have here explored the cell penetrability of differently charged polyarginines in two alternative presentations, namely as unassembled fusion proteins or assembled in multimeric protein nanoparticles. By this, we have observed that arginine-rich peptides switch between receptor-mediated and receptor-independent mechanisms of cell penetration. The relative weight of these activities is determined by the electrostatic charge of the construct and the oligomerization status of the nanoscale material, both regulatable by conventional protein engineering approaches.     

Nephrotoxicity and Kidney Transport Assessment on 3D Perfused Proximal Tubules

Proximal tubules in the kidney play a crucial role in reabsorbing and eliminating substrates from the body into the urine, leading to high local concentrations of xenobiotics. This makes the proximal tubule a major target for drug toxicity that needs to be evaluated during the drug development process. Here, we describe an advanced in vitro model consisting of fully polarized renal proximal tubular epithelial cells cultured in a microfluidic system. Up to 40 leak-tight tubules were cultured on this platform that provides access to the basolateral as well as the apical side of the epithelial cells. Exposure to the nephrotoxicant cisplatin caused a dose-dependent disruption of the epithelial barrier, a decrease in viability, an increase in effluent LDH activity, and changes in expression of tight-junction marker zona-occludence 1, actin, and DNA-damage marker H2A.X, as detected by immunostaining. Activity and inhibition of the efflux pumps P-glycoprotein (P-gp) and multidrug resistance protein (MRP) were demonstrated using fluorescence-based transporter assays. In addition, the transepithelial transport function from the basolateral to the apical side of the proximal tubule was studied. The apparent permeability of the fluorescent P-gp substrate rhodamine 123 was decreased by 35% by co-incubation with cyclosporin A. Furthermore, the activity of the glucose transporter SGLT2 was demonstrated using the fluorescent glucose analog 6-NBDG which was sensitive to inhibition by phlorizin. Our results demonstrate that we developed a functional 3D perfused proximal tubule model with advanced renal epithelial characteristics that can be used for drug screening studies.     

Impact of Hematopoietic Stem Cell Transplantation on Dental Development

To investigate dental development in patients treated with a hematopoietic stem cell transplantation (HSCT), 42 children and young adults who were under 12 years old at time of HSCT were examined for dental agenesis, microdontia, and root-to-crown ratio. Conditioning regimens were total body irradiation (TBI) based in 12 patients, busulfan based in 21 patients, and 9 patients had other chemotherapeutic agents. Sixteen patients were <3 years old, 9 patients were 3 to 6 years old, and 17 patients were 6 to 12 years old at HSCT. Prevalence of agenesis and microdontia of at least 1 permanent tooth were, respectively, 51.3% and 46.2% in the study population, and 76.3% had an aberrant root-to-crown ratio. All these results were highly different from the prevalence in the healthy population. Patients treated before the age of 3 years had more microdontia (76.9%) and agenesis (92.3%) compared with patients treated at an older age. In the subgroup of patients treated after 6 years, there was more microdontia when treated with busulfan (50%) compared with treatment with TBI (0%) (P?=?.044). Patients treated with HSCT had many disturbances in dental development. Age at HSCT and possibly also the conditioning regimen used had an effect on their type and prevalence. Dental follow-up should be incorporated in the multidisciplinary follow-up program of these patients.