Liposome Delivery Enhances Clodronate Nephrotoxicity in Dahl SS Hypertension and Renal Injury
Authors: Fehrenbach Daniel J , Abais‐Battad Justine M , Lund Haley , Dasinger John Henry , Mattson David L 
Published: June 2021

Studies in the Dahl Salt‐Sensitive (SS) rat have shown that infiltrating renal CD3+ T cells and ED‐1+ macrophages are involved in the development of SS hypertension (HTN) and renal damage. We have previously demonstrated the importance of infiltrating cells by pharmacological immunosuppression and genetic deletion of adaptive immune cells, but the role of innate immunity in SS HTN is still unclear. It is practically more difficult to assess the contribution of the innate immune system macrophage to this pathology due its vital role in development and in bone and liver physiology. A proposed method of macrophage depletion is via liposome‐encapsulated Clodronate which has shown mixed results, from attenuation of Ang‐II induced HTN, to inducing salt‐sensitive HTN. This study tests the hypothesis that Clodronate‐Liposome treatment will attenuate Dahl SS HTN and renal damage through macrophage depletion. To do so, animals were administered either Clodronate‐Liposomes (Clod‐Lipo) or PBS‐Liposomes (PBS‐Lipo) as a vehicle control. A third group of animals were treated with Clodronate alone (Clod) to assess direct effects of the drug. 7‐week‐old male Dahl SS rats were implanted with radiotelemeters and after recovery and baseline pressure recordings on a low‐salt (0.4% NaCl) diet, animals received a tail vein injection of either PBS‐Lipo (equivalent volume), Clod‐Lipo (50mg/kg), or Clod (50mg/kg). Booster injections (25mg/kg) were administered every 7 days thereafter. Three days after the initial injection, all animals were switched to a high‐salt (4.0% NaCl) diet. Animals were followed for 21 days of the high‐salt diet with weekly overnight urine collections. Clod treatment alone did not alter circulating or splenic levels of macrophages unlike Clod‐Lipo treatment which depleted macrophages by about 50%. Contrary to our hypothesis, by the end of the study, Clod‐Lipo treated animals developed an exacerbated SS response in respect to blood pressure (Clod‐Lipo 178.7±7.5mmhg, PBS‐Lipo 157.2±3.9mmhg, p<0.01 Two‐way RM‐ANOVA, n=8/7) and albuminuria (Clod‐Lipo 403.6±32.8mg/day, PBS‐Lipo 180.6±22.4mg/day, p<0.001 Two‐way RM‐ANOVA, n=11/7). This exacerbation in the Clod‐Lipo group was accompanied by an increase in infiltration of adaptive T and B cells despite a depletion of circulating macrophages and a reduction in infiltrating macrophages. The third group of animals receiving Clod alone demonstrated a similar, though delayed, exacerbation in the SS increase in blood pressure (181.6±6.2mmhg p<0.05 Two‐way RM‐ANOVA, n=4). In addition, Clod treated animals showed elevated albuminuria (259.2±48.6mg/day, p<0.05 Two‐way RM‐ANOVA, n=5) though not to the same extent as Clod‐Lipo treated animals. These SS phenotypic changes in Clod treated animals were independent of changes to circulating, splenic, and infiltrating renal immune cells. This study demonstrates Clodronate's direct nephrotoxic effects similar to those of other bisphosphonates. Furthermore, liposome delivery of Clodronate further exacerbated the SS renal damage and lymphocyte infiltration, indicating the potential adverse effects of this macrophage depletion strategy.Support or Funding InformationHL‐116264, HL‐137748, and 18PRE3400038This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.