Immunofluorescent staining and mirna expression of HCT-8 and HT-29 cell lines upon cryptosporidium infection
The protozoan Cryptosporidium mainly infects epithelial cells of the colorectal region. The infection of this species can be observed and confirmed by immunofluorescent staining method in vitro. In addition, the onco-microRNA (miRNA) and tumour suppressor miRNA expression pattern of the epithelial...
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| Main Authors: | , , |
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| Format: | Article |
| Language: | English English |
| Published: |
Malaysian Society of Applied Biology
2018
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| Subjects: | |
| Online Access: | http://irep.iium.edu.my/70601/1/70601_IMMUNOFLUORESCENT%20STAINING%20AND%20miRNA%20EXPRESSION_article.pdf http://irep.iium.edu.my/70601/2/70601_IMMUNOFLUORESCENT%20STAINING%20AND%20miRNA%20EXPRESSION_scopus.pdf http://irep.iium.edu.my/70601/ |
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| Summary: | The protozoan Cryptosporidium mainly infects epithelial cells of the colorectal region. The infection of this species can be
observed and confirmed by immunofluorescent staining method in vitro. In addition, the onco-microRNA (miRNA) and
tumour suppressor miRNA expression pattern of the epithelial cells upon Cryptosporidium infection may reveal possible
effect of Cryptosporidium in the onset and progression of cancer. In this study, two epithelial colorectal cancer cell lines
HCT8 and HT29 were infected with Cryptosporidium parvum (C. parvum) and observed using direct immunofluorescent
technique via fluorescein conjugated Vicia Villosa Lectin (VVL) staining. Real Time-quantitative Polymerase Chain Reaction
(RT-qPCR) was performed to determine the expression of miR-21 and miR-145 in infected cell lines. Data normalisation of
the miRNA expression was carried out using reference gene RNU44. The immunofluorescent micrographs exhibited the
Cryptosporidium infection on both HCT-8 and HT-29 cell lines with green fluorescence. Upregulation of miR-21 and
downregulation of normalised expression of miR-145 was observed in both of the cell lines upon Cryptosporidium infection.
We found the expression of miR-21 to be increased and miR-145 greatly decreased in both cell lines upon infection. Our
current observation suggests that, C. parvum can potentially infect the colorectal cancer cell lines and play critical role in
regulating onco-miRNAs and tumor suppressor miRNAs. |
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