Abstract
The application of innovative three-dimensional (3D) spheroids cell culture strategy to Parasitology offers the opportunity to closely explore host–parasite interactions. Here we present a first report on the application of 3D hepatic spheroids to unravel the immune response of canine hepatocytes exposed to Leishmania infantum. The liver, usually considered a major metabolic organ, also performs several important immunological functions and constitutes a target organ for L. infantum infection, the etiological agent of canine leishmaniasis (CanL), and a parasitic disease of major veterinary and public health concern. 3D hepatic spheroids were able to sense and immunologically react to L. infantum parasites, generating an innate immune response by increasing nitric oxide (NO) production and enhancing toll-like receptor (TLR) 2 and interleukin-10 gene expression. The immune response orchestrated by canine hepatocytes also lead to the impairment of several cytochrome P450 (CYP450) with possible implications for liver natural xenobiotic metabolization capacity. The application of meglumine antimoniate (MgA) increased the inflammatory response of 3D hepatic spheroids by inducing the expression of Nucleotide oligomerization domain (NOD) -like receptors 1 and NOD2 and TLR2, TLR4, and TLR9 and enhancing gene expression of tumour necrosis factor α. It is therefore suggested that hepatocytes are key effector cells and can activate and orchestrate the immune response to L. infantum parasites.
Highlights
In living tissue, cells exist in three dimensional (3D) microenvironments, with intricate cell-to-cell and cell-to-matrix interactions, creating a dynamic and complex network of nutrient transport, waste removal, and cell signaling pathways
The amounts of TLR4 and TLR9 mRNA did not reveal major changes in spheroids exposed to parasites, during the entire observation period (Figure 4D,E). These results indicate that dog hepatocytes can recognize L. infantum antigens through TLR2, where other toll-like receptor (TLR) and Nucleotide oligomerization domain (NOD), evaluated in the present study may not be activated by parasite PAMPs
Cell-to-cell and cell-to-matrix interactions, recapitulating many in vivo tissue structures and Hepatocytes rapidly recover from isolation trauma in the 3D system, as indicated by CYPs activity functions
Summary
Cells exist in three dimensional (3D) microenvironments, with intricate cell-to-cell and cell-to-matrix interactions, creating a dynamic and complex network of nutrient transport, waste removal, and cell signaling pathways The mimicking of this microenvironment has long been a major concern in research as standard two dimensional (2D) or monolayer cell cultures may inadequately represent cells’ complex interrelations, which make these culture systems less reliable predictors of in vivo cell behavior [1]. The application of innovative cell culture methods has been mainly driven by the pharmaceutical industry and its need to test new drugs for toxicity [2,5] The translation of this innovation to parasitological studies has been slower and here is presented as a first report on the application of 3D spheroids to unravel the immune response of canine hepatocytes exposed to Leishmania infantum
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