Live imaging of intracellular parasites reveals changes in host metabolism

PArasitic infections pose challenges to the development of effective treatments because these microorganisms are eukaryotic, like the people and animals they infect. In addition, species such as Toxoplasma gondii live inside cells, making them difficult to study and complicating the development of treatments.

“To (discover) better targets, we need to study in detail the changes that this parasite causes in host cells,” he said. Gina Gallego-Lopezpostdoctoral fellow and parasitologist at the Morgridge Research Institute and the University of Wisconsin-Madison. Many amino acids and lipids that T. gondii metabolic products of the host necessary for its survival, therefore, understanding how the parasite manipulates host metabolic pathways may suggest new strategies to defeat this invader.¹

Recently, Gallego-Lopez and her colleagues developed a new approach to studying intracellular infections, in which they used two-photon microscopy for non-invasive research. measure metabolic changes V T. gondii-infected cells.² This study published in mBio, demonstrated that T. gondii altered the metabolic activity of human fibroblasts, shedding light on the metabolic rearrangement caused by parasites.

“It is very difficult to separate host cell metabolism from parasite metabolism,” he said. Laura Nollparasitologist at the University of Wisconsin-Madison and co-author of the study. To overcome this, the team used T. gondii which expressed the red fluorophore mCherry to track the location of parasites. They then used the natural fluorescence of cellular respiration metabolites to track changes in the amounts of these products during infection.

The team began their research with a less virulent strain of the virus. T. gondii they reproduced more slowly, which allowed them to study changes in metabolism over time. They gained insight into cellular metabolism by measuring nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide phosphate, together called NAD(P)H, two metabolites that accept or donate electrons during cellular respiration. Thus, the change in the proportion of NAD(P)H bound to proteins compared to the free enzyme serves as a reliable indicator of cellular metabolism. They noticed that T. gondii infection increased the ratio of bound enzymes and created a more oxidized intracellular environment, indicating increased metabolism.

“We had no idea about the redox changes caused by Toxoplasma gondii in the host,” Gallego-Lopez said. “This is the first time anyone has been able to measure this in living cells.”

To further study the metabolic effects T. gondii infection, the team studied changes in glucose and lactate concentrations. They showed that glucose levels gradually increased in the first nine hours after infection and then decreased over the remaining 48 hours. Meanwhile, cellular lactate concentrations began to decline six hours after infection. This was consistent with increased glycolysis early in the infection and then decreased glycolysis after 24 hours, in parallel with greater intracellular oxidation, suggesting that the parasite alters glucose metabolism to support its growth.

The team compared these results with previous ones RNA sequencing results.³ They found that the increased gene expression they had previously observed in host cells corresponded to enzymes involved in cellular respiration. In addition, they observed increased expression of reactive oxygen species metabolizing enzymes in T. gondii as well as other metabolically important enzymes.

The present study, and its agreement with the team’s previous gene expression data, shows that T. gondii changes cellular metabolism. Noll’s team suspects this could provide clues about viable therapeutic strategies. The group previously demonstrated that cancer drug candidate decreased T. gondii cell growth.⁴ “Cancer is the rapid proliferation of eukaryotic cells that escape the immune response, and so are (intracellular parasites),” Noll said. “So we need to rethink the drugs that we already have, and some of them may be very useful (against T. gondii)”.

Zhicheng Doumolecular geneticist who studies T. gondii from Clemson University and was not involved in the study, said the work was interesting and that the imaging technique will be useful in the field once labs apply it and demonstrate its effectiveness in more models. He is also interested in learning more about the mechanisms behind metabolic changes.

“This is something we probably need to understand in the future,” Dow said, adding that the field could figure out where these changes come from. “Host cells can change their metabolism to resist infection.”