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Oleylphosphocholine (OlPC) arrests Cryptosporidium parvum growth in vitro and prevents lethal infection in interferon gamma receptor knock-out mice.

Sonzogni-Desautels K, Renteria AE, Camargo FV, Di Lenardo TZ, Mikhail A, Arrowood MJ, Fortin A, Ndao M - Front Microbiol (2015)

Bottom Line: Treatment with OlPC, at 40 mg/kg/day resulted in 100% survival, complete clearance of parasite in stools and a 99.9% parasite burden reduction in the intestines at Day 30.Doses of 30 and 20 mg/kg/day also demonstrated an increased survival rate and a dose-dependent parasite burden reduction.Together, our results strongly support that OlPC represents a potential candidate for the treatment of C. parvum infections in immunocompromised patients.

View Article: PubMed Central - PubMed

Affiliation: National Reference Centre for Parasitology, Research Institute of the McGill University Health Centre, Montreal QC, Canada ; Institute of Parasitology, Macdonald Campus, McGill University, Montreal QC, Canada.

ABSTRACT
Cryptosporidium parvum is a species of protozoa that causes cryptosporidiosis, an intestinal disease affecting many mammals including humans. Typically, in healthy individuals, cryptosporidiosis is a self-limiting disease. However, C. parvum can cause a severe and persistent infection that can be life-threatening for immunocompromised individuals, such as AIDS patients. As there are no available treatments for these patients that can cure the disease, there is an urgent need to identify treatment options. We tested the anti-parasitic activity of the alkylphosphocholine oleylphosphocholine (OlPC), an analog of miltefosine, against C. parvum in in vitro and in vivo studies. In vitro experiments using C. parvum infected human ileocecal adenocarcinoma cells (HCT-8 cells) showed that OlPC has an EC50 of 18.84 nM. Moreover, no cell toxicity has been seen at concentrations ≤50 μM. C57BL/6 interferon gamma receptor knock-out mice, were infected by gavage with 4000 C. parvum oocysts on Day 0. Oral treatments, with OlPC, miltefosine, paromomycin or PBS, began on Day 3 post-infection for 10 days. Treatment with OlPC, at 40 mg/kg/day resulted in 100% survival, complete clearance of parasite in stools and a 99.9% parasite burden reduction in the intestines at Day 30. Doses of 30 and 20 mg/kg/day also demonstrated an increased survival rate and a dose-dependent parasite burden reduction. Mice treated with 10 mg/kg/day of miltefosine resulted in 50% survival at Day 30. In contrast, control mice, treated with PBS or 100 mg/kg/day of paromomycin, died or had to be euthanized between Days 6 and 13 due to severe illness. Results of parasite burden were obtained by qPCR and cross-validated by both flow cytometry of stool oocysts and histological sections of the ileum. Together, our results strongly support that OlPC represents a potential candidate for the treatment of C. parvum infections in immunocompromised patients.

No MeSH data available.


Related in: MedlinePlus

Parasite and oocyst burden in C57BL/6 IFNγR-KO mice infected with C. parvum. All intestinal samples were analyzed by qPCR represented by the clear bars and by flow cytometry represented by the checkered bars. qPCR results were normalized to parasites per gram of intestines and flow cytometry results were normalized to oocysts per gram of intestines. (A) Parasite and oocyst burden in mice at Day 10. (B) Parasite and oocyst burden in mice at Day 30. Error bars were calculated as ±SEM.
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Figure 4: Parasite and oocyst burden in C57BL/6 IFNγR-KO mice infected with C. parvum. All intestinal samples were analyzed by qPCR represented by the clear bars and by flow cytometry represented by the checkered bars. qPCR results were normalized to parasites per gram of intestines and flow cytometry results were normalized to oocysts per gram of intestines. (A) Parasite and oocyst burden in mice at Day 10. (B) Parasite and oocyst burden in mice at Day 30. Error bars were calculated as ±SEM.

Mentions: To quantitate parasite and oocyst burden in the intestines and assess disease progression of infected mice during and after their respective treatments, mice were euthanized and intestines were collected at two time points: Days 10 or 30. Intestinal samples from every mouse were processed individually to purify C. parvum parasites, analyzed using qPCR and normalized to parasites per gram of intestines (p/g of intestines). On Day 10, infected control groups revealed massive levels of C. parvum in the gut. Mice treated with PBS had the highest parasite burden with 4.77 × 107 p/g of intestines followed by mice treated with 100 mg/kg/day of paromomycin with 5.01 × 106 p/g of intestines (Figure 4A). Mice treated with 10 mg/kg/day of miltefosine also had very high parasite burden with 7.94 × 106 p/g of intestines (Figure 4A). In mice treated with OlPC, parasite burden was significantly lower compared to the infected PBS control group. After 7 days of treatment (Day 10), 20 mg/kg/day OlPC-treated mice showed a parasite burden of 3.44 × 106 p/g of intestines (P < 0.01; Figure 4A). In mice treated with 30 mg/kg/day of OlPC the parasite burden was even lower with 1.81 × 105 p/g of intestines (P < 0.001) corresponding to a 99.6% (2-log reduction) parasite burden reduction in comparison with the PBS control group (Figure 4A). Similarly, mice treated with 40 mg/kg/day of OlPC also revealed a 99.6% reduction in parasite burden with 2.25 × 105 p/g of intestines (P < 0.001) when compared to the infected PBS control mice (Figure 4A).


Oleylphosphocholine (OlPC) arrests Cryptosporidium parvum growth in vitro and prevents lethal infection in interferon gamma receptor knock-out mice.

Sonzogni-Desautels K, Renteria AE, Camargo FV, Di Lenardo TZ, Mikhail A, Arrowood MJ, Fortin A, Ndao M - Front Microbiol (2015)

Parasite and oocyst burden in C57BL/6 IFNγR-KO mice infected with C. parvum. All intestinal samples were analyzed by qPCR represented by the clear bars and by flow cytometry represented by the checkered bars. qPCR results were normalized to parasites per gram of intestines and flow cytometry results were normalized to oocysts per gram of intestines. (A) Parasite and oocyst burden in mice at Day 10. (B) Parasite and oocyst burden in mice at Day 30. Error bars were calculated as ±SEM.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585137&req=5

Figure 4: Parasite and oocyst burden in C57BL/6 IFNγR-KO mice infected with C. parvum. All intestinal samples were analyzed by qPCR represented by the clear bars and by flow cytometry represented by the checkered bars. qPCR results were normalized to parasites per gram of intestines and flow cytometry results were normalized to oocysts per gram of intestines. (A) Parasite and oocyst burden in mice at Day 10. (B) Parasite and oocyst burden in mice at Day 30. Error bars were calculated as ±SEM.
Mentions: To quantitate parasite and oocyst burden in the intestines and assess disease progression of infected mice during and after their respective treatments, mice were euthanized and intestines were collected at two time points: Days 10 or 30. Intestinal samples from every mouse were processed individually to purify C. parvum parasites, analyzed using qPCR and normalized to parasites per gram of intestines (p/g of intestines). On Day 10, infected control groups revealed massive levels of C. parvum in the gut. Mice treated with PBS had the highest parasite burden with 4.77 × 107 p/g of intestines followed by mice treated with 100 mg/kg/day of paromomycin with 5.01 × 106 p/g of intestines (Figure 4A). Mice treated with 10 mg/kg/day of miltefosine also had very high parasite burden with 7.94 × 106 p/g of intestines (Figure 4A). In mice treated with OlPC, parasite burden was significantly lower compared to the infected PBS control group. After 7 days of treatment (Day 10), 20 mg/kg/day OlPC-treated mice showed a parasite burden of 3.44 × 106 p/g of intestines (P < 0.01; Figure 4A). In mice treated with 30 mg/kg/day of OlPC the parasite burden was even lower with 1.81 × 105 p/g of intestines (P < 0.001) corresponding to a 99.6% (2-log reduction) parasite burden reduction in comparison with the PBS control group (Figure 4A). Similarly, mice treated with 40 mg/kg/day of OlPC also revealed a 99.6% reduction in parasite burden with 2.25 × 105 p/g of intestines (P < 0.001) when compared to the infected PBS control mice (Figure 4A).

Bottom Line: Treatment with OlPC, at 40 mg/kg/day resulted in 100% survival, complete clearance of parasite in stools and a 99.9% parasite burden reduction in the intestines at Day 30.Doses of 30 and 20 mg/kg/day also demonstrated an increased survival rate and a dose-dependent parasite burden reduction.Together, our results strongly support that OlPC represents a potential candidate for the treatment of C. parvum infections in immunocompromised patients.

View Article: PubMed Central - PubMed

Affiliation: National Reference Centre for Parasitology, Research Institute of the McGill University Health Centre, Montreal QC, Canada ; Institute of Parasitology, Macdonald Campus, McGill University, Montreal QC, Canada.

ABSTRACT
Cryptosporidium parvum is a species of protozoa that causes cryptosporidiosis, an intestinal disease affecting many mammals including humans. Typically, in healthy individuals, cryptosporidiosis is a self-limiting disease. However, C. parvum can cause a severe and persistent infection that can be life-threatening for immunocompromised individuals, such as AIDS patients. As there are no available treatments for these patients that can cure the disease, there is an urgent need to identify treatment options. We tested the anti-parasitic activity of the alkylphosphocholine oleylphosphocholine (OlPC), an analog of miltefosine, against C. parvum in in vitro and in vivo studies. In vitro experiments using C. parvum infected human ileocecal adenocarcinoma cells (HCT-8 cells) showed that OlPC has an EC50 of 18.84 nM. Moreover, no cell toxicity has been seen at concentrations ≤50 μM. C57BL/6 interferon gamma receptor knock-out mice, were infected by gavage with 4000 C. parvum oocysts on Day 0. Oral treatments, with OlPC, miltefosine, paromomycin or PBS, began on Day 3 post-infection for 10 days. Treatment with OlPC, at 40 mg/kg/day resulted in 100% survival, complete clearance of parasite in stools and a 99.9% parasite burden reduction in the intestines at Day 30. Doses of 30 and 20 mg/kg/day also demonstrated an increased survival rate and a dose-dependent parasite burden reduction. Mice treated with 10 mg/kg/day of miltefosine resulted in 50% survival at Day 30. In contrast, control mice, treated with PBS or 100 mg/kg/day of paromomycin, died or had to be euthanized between Days 6 and 13 due to severe illness. Results of parasite burden were obtained by qPCR and cross-validated by both flow cytometry of stool oocysts and histological sections of the ileum. Together, our results strongly support that OlPC represents a potential candidate for the treatment of C. parvum infections in immunocompromised patients.

No MeSH data available.


Related in: MedlinePlus