Abstract
BackgroundChimeric mice with humanized livers represent a promising tool for infections with Plasmodium falciparum to evaluate novel methods for prevention and treatment of pre-erythrocytic stages. Adequate assessment of hepatic infections is generally compromised by the limited number of human hepatocytes infected by developing parasites.MethodsA qPCR-based method has been developed that sensitively and reliably detects P. falciparum liver stage infection of humanized mice and quantitatively expresses the results as the number of parasites per human hepatocyte.ResultsThis assay allows for detection of liver stage parasites after challenging humanized mice with infected mosquito bites or after intravenous injection with sporozoites. The sensitivity of the protocol, which comprises approximately 25% of the total chimeric liver, allows for the detection of a single infected hepatocyte in the analysed tissue.ConclusionsThis method allows for the detection and quantification of P. falciparum parasites in chimeric mice repopulated with human hepatocytes. It will be a useful tool when studying the in vivo therapeutic and/or prophylactic qualities of novel compounds, small molecules or antibodies directed against the liver stage of P. falciparum infections.
Highlights
Chimeric mice with humanized livers represent a promising tool for infections with Plasmodium falciparum to evaluate novel methods for prevention and treatment of pre-erythrocytic stages
Standardizing the sampling of humanized liver fragments Injection of P. falciparum sporozoites in a chimeric mouse will result in infection of an unknown and low number of infected human hepatocytes distributed in a large background of non-infected mouse and human cells (Figure 4)
Five μL of each extract was used in duplicate to measure the number of parasites by Quantitative polymerase chain reaction (qPCR) and 1 μL to determine the fraction of human hepatocytes in each fragment (Figure 2)
Summary
Chimeric mice with humanized livers represent a promising tool for infections with Plasmodium falciparum to evaluate novel methods for prevention and treatment of pre-erythrocytic stages. Humanized mouse models have been developed to study either blood stage or liver stage of malaria. By engrafting human red blood cells (huRBC) into immunodeficient mice, a high percentage of huRBC in mouse peripheral human hepatocytes can be achieved, but emerging parasites cannot successfully infect murine erythrocytes [16]. The Center for Vaccinology (CEVAC) has been successful in producing chimeric uPA+/+-SCID mice with a high degree of repopulation with human hepatocytes to study hepatitis B and C virus (HBV and HCV) infections [17,18,19,20]. Transfer of human hepatocytes was started in FRG mice that originate from Marcus Grompe’s laboratory [10]
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