Work described in this thesis was aimed to further understand the role of macrophages in acute cellular rejection of transplanted kidneys, and perform a pre-clinical assessment of the localization... Show moreWork described in this thesis was aimed to further understand the role of macrophages in acute cellular rejection of transplanted kidneys, and perform a pre-clinical assessment of the localization and efficacy of liposomal prednisolone. First, we explored the role of anti-inflammatory macrophages in the onset of acute cellular rejection using human biopsy samples. Here, we discovered that the presence of high levels of CD163+ macrophages was associated with a lower risk of rejection, in a population of patients with a high rate of delayed graft function. Subsequently, we identified the ability of liposomal prednisolone to target macrophages in the kidney using in vitro models of human macrophages, and their ability to accumulate in inflamed kidney tissue in vivo rat models of ischemia reperfusion injury in the kidney. Finally, we evaluated the treatment efficacy of liposomal prednisolone in a mouse model of cellular rejection following kidney transplantation. Here, we were able to show an improved efficacy of liposomal prednisolone in treating rejection, when compared to treatment with free prednisolone. Show less
Alem, C.M.A. van; Bank, J.R.; Vries, D.K. de; Bajema, I.M.; Mallat, M.J.K.; Fijter, J.W. de; ... ; Kooten, C. van 2022
Acute cellular rejection (ACR) occurs in 10% of renal allograft recipients and is characterized by leukocyte infiltration as observed in needle biopsies. ACR onset is subject to several risk... Show moreAcute cellular rejection (ACR) occurs in 10% of renal allograft recipients and is characterized by leukocyte infiltration as observed in needle biopsies. ACR onset is subject to several risk factors, including delayed graft function (DGF). As the impact of DGF on the etiology of ACR remains unclear, this study analyzed the association between presence of leukocyte subsets and ACR onset, in DCD kidney biopsies with extensive DGF following transplantation. Immunohistochemical analysis of protocol biopsies taken 10 days after kidney transplantation revealed that patients with high levels of renal CD163+ macrophages have a decreased risk (OR = 0.021, P = 0.008) for ACR in the first 6 months after transplantation. In pre-transplant biopsies of a comparable DCD cohort, with >80% DGF, presence of donor CD163+ macrophages showed no effect on ACR risk. Therefore, leukocyte infiltrate present during the inflammatory response at the time of DGF may contain anti-inflammatory macrophages that exert a protective effect against ACR development. Show less
Liposomes can be seen as ideal carriers for anti-inflammatory drugs as their ability to (passively) target sites of inflammation and release their content to inflammatory target cells enables them... Show moreLiposomes can be seen as ideal carriers for anti-inflammatory drugs as their ability to (passively) target sites of inflammation and release their content to inflammatory target cells enables them to increase local efficacy with only limited systemic exposure and adverse effects. Nonetheless, few liposomal formulations seem to reach the clinic. The current review provides an overview of the more recent innovations in liposomal treatment of rheumatoid arthritis, psoriasis, vascular inflammation, and transplantation. Cutting edge developments include the liposomal delivery of gene and RNA therapeutics and the use of hybrid systems where several liposomal bilayer features, or several drugs, are combined in a single formulation. The majority of the articles reviewed here focus on preclinical animal studies where proof-of-principle of an improved efficacy-safety ratio is observed when using liposomal formulations. A few clinical studies are included as well, which brings us to a discussion about the challenges of clinical translation of liposomal nanomedicines in the field of inflammatory diseases. Show less
Background.Systemic exposure to high-dose corticosteroids effectively combats acute rejection after kidney transplantation, but at the cost of substantial side effects. In this study, a murine... Show moreBackground.Systemic exposure to high-dose corticosteroids effectively combats acute rejection after kidney transplantation, but at the cost of substantial side effects. In this study, a murine acute renal allograft rejection model was used to investigate whether liposomal-encapsulated prednisolone (LP) facilitates local exposure to enhance its therapeutic effect.Methods.Male BalbC recipients received renal allografts from male C57BL/6J donors. Recipients were injected daily with 5 mg/kg cyclosporine A and received either 10 mg/kg prednisolone (P), or LP intravenously on day 0, 3, and 6, or no additional treatment. Functional magnetic resonance imaging (fMRI) was performed on day 6 to study allograft perfusion and organs were retrieved on day 7 for further analysis.Results.Staining of polyethylene-glycol-labeled liposomes and high performance liquid chromatography analysis revealed accumulation in the LP treated allograft. LP treatment induced the expression of glucocorticoid responsive gene Fkbp5 in the allograft. Flow-cytometry of allografts revealed liposome presence in CD45(+) cells, and reduced numbers of F4/80(+) macrophages, and CD3(+) T-lymphocytes upon LP treatment. Banff scoring showed reduced interstitial inflammation and tubulitis and fMRI analysis revealed improved allograft perfusion in LP versus NA mice.Conclusions.Liposomal delivery of prednisolone improved renal bio-availability, increased perfusion and reduced cellular infiltrate in the allograft, when compared with conventional prednisolone. Clinical studies should reveal if treatment with LP results in improved efficacy and reduced side effects in patients with renal allograft rejection. Show less
BACKGROUND:Treatment of inflammatory kidney diseases with systemic high-dose glucocorticoids (GCs) has severe side effects. Liposomal encapsulation could facilitate local delivery of GCs to the... Show moreBACKGROUND:Treatment of inflammatory kidney diseases with systemic high-dose glucocorticoids (GCs) has severe side effects. Liposomal encapsulation could facilitate local delivery of GCs to the inflamed kidney, as liposomes encapsulate their payload until extravasation at sites of inflammation, potentially resulting in local bioactivity. Our aim was to evaluate the ability of liposomes to accumulate locally after renal ischaemia-reperfusion injury in the rat and to study its effect on macrophages.METHODS:In vitro, human macrophages were incubated with fluorescent liposomes, liposomal prednisolone, prednisolone, empty liposomes or saline. Uptake was studied microscopically and treatment effect was assessed by interkeukin 6 (IL-6) enzyme-linked immunosorbent assay. The mechanism of action was evaluated by analysing GC receptor activation by microscopy and quantitative polymerase chain reaction (qPCR). In vivo, rats were subjected to ischaemia-reperfusion injury and were injected intravenously with fluorescent liposomes, liposomal prednisolone, prednisolone, empty liposomes or saline. Uptake was measured by the FLARE camera and the treatment effect by immunohistochemistry for myeloid cells and qPCR for inflammatory markers.RESULTS:In vitro, macrophages internalized liposomes after 8 hours. Prednisolone or liposomal prednisolone treatment reduced IL-6 production and both compounds induced translocation of the GC receptor to the nucleus and upregulation of PER1 messenger RNA (mRNA), indicating a similar mechanism of action. In vivo, fluorescent liposomes accumulated in the inflamed kidney. Liposomal prednisolone treatment increased the presence of ED2-positive anti-inflammatory macrophages and both prednisolone and liposomal prednisolone reduced monocyte chemoattractant protein-1 (MCP-1) mRNA production, indicating a reduced pro-inflammatory profile in the kidney.CONCLUSIONS:Liposomal encapsulation is a promising strategy for local delivery of glucocorticoids to the inflamed kidney. Show less
