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Molecular detection and quantification of Plasmodium falciparum-infected human hepatocytes in chimeric immune-deficient mice.

Foquet L, Hermsen CC, van Gemert GJ, Libbrecht L, Sauerwein R, Meuleman P, Leroux-Roels G - Malar. J. (2013)

Bottom Line: Adequate assessment of hepatic infections is generally compromised by the limited number of human hepatocytes infected by developing parasites.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.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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Vaccinology, Ghent University and University Hospital, De Pintelaan 185, Ghent 9000, Belgium. geert.lerouxroels@ugent.be.

ABSTRACT

Background: 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. Adequate assessment of hepatic infections is generally compromised by the limited number of human hepatocytes infected by developing parasites.

Methods: A 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.

Results: This 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.

Conclusions: This 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.

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correlation human albumin and percentage repopulation. The calculated percentage repopulation (%huHEP/totHEP) is plotted against the measured human albumin plasma concentrations for the animals described in Additional file 2: Table S2.
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Figure 5: correlation human albumin and percentage repopulation. The calculated percentage repopulation (%huHEP/totHEP) is plotted against the measured human albumin plasma concentrations for the animals described in Additional file 2: Table S2.

Mentions: Quantification of both human and mouse DNA in addition to P. falciparum DNA allowed for the determination of the degree (percentage) of human repopulation of each of the 12 mouse liver fragments (Additional file 1: Table S1). This approach differs from previous studies in which the degree of chimerism was calculated by determining the proportion of the liver parenchyma surface area that consisted of human hepatocytes based on immunohistochemical analyses of a series of liver sections [11,31,36]. Since hepatocytes are much larger than non-parenchymal cells, the area occupied by human hepatocytes will overestimate the fraction represented as cell numbers. The data show that the percentage of repopulation of a humanized mouse in different liver lobes is quite stable and does not significantly influence the infection with P. falciparum sporozoites (Additional file 2: Table S2). FigureĀ 5 shows that the percentage human hepatocytes correlates well with the human albumin concentrations in chimeric mice (R = 0.57; P < 0.0001).


Molecular detection and quantification of Plasmodium falciparum-infected human hepatocytes in chimeric immune-deficient mice.

Foquet L, Hermsen CC, van Gemert GJ, Libbrecht L, Sauerwein R, Meuleman P, Leroux-Roels G - Malar. J. (2013)

correlation human albumin and percentage repopulation. The calculated percentage repopulation (%huHEP/totHEP) is plotted against the measured human albumin plasma concentrations for the animals described in Additional file 2: Table S2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4222492&req=5

Figure 5: correlation human albumin and percentage repopulation. The calculated percentage repopulation (%huHEP/totHEP) is plotted against the measured human albumin plasma concentrations for the animals described in Additional file 2: Table S2.
Mentions: Quantification of both human and mouse DNA in addition to P. falciparum DNA allowed for the determination of the degree (percentage) of human repopulation of each of the 12 mouse liver fragments (Additional file 1: Table S1). This approach differs from previous studies in which the degree of chimerism was calculated by determining the proportion of the liver parenchyma surface area that consisted of human hepatocytes based on immunohistochemical analyses of a series of liver sections [11,31,36]. Since hepatocytes are much larger than non-parenchymal cells, the area occupied by human hepatocytes will overestimate the fraction represented as cell numbers. The data show that the percentage of repopulation of a humanized mouse in different liver lobes is quite stable and does not significantly influence the infection with P. falciparum sporozoites (Additional file 2: Table S2). FigureĀ 5 shows that the percentage human hepatocytes correlates well with the human albumin concentrations in chimeric mice (R = 0.57; P < 0.0001).

Bottom Line: Adequate assessment of hepatic infections is generally compromised by the limited number of human hepatocytes infected by developing parasites.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.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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Vaccinology, Ghent University and University Hospital, De Pintelaan 185, Ghent 9000, Belgium. geert.lerouxroels@ugent.be.

ABSTRACT

Background: 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. Adequate assessment of hepatic infections is generally compromised by the limited number of human hepatocytes infected by developing parasites.

Methods: A 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.

Results: This 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.

Conclusions: This 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.

Show MeSH
Related in: MedlinePlus