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The Role of Trypsin in The Internalization Process of Influenza H1N1 Virus into Vero and MDCK Cells
Fawzi Rahmadiyan Zuhairi1, Maharani2 & Marselina I. Tan1
1Research Group of Physiology, Developmental Biology, and Biomedical Science, School of Life Science and Technology, Institut Teknologi Bandung
Jalan Ganesa No. 10, Bandung 40132, Indonesia
2Department of Research and Development, BioFarma Co, Ltd.
Jalan Pasteur No. 28, Bandung 40161, Indonesia
Email: fawzi.zuhairi@gmail.com
Abstract. Trypsin is supposed to play an important role in the internalization of the influenza virus into host cells in vitro because of its protease activity. The purpose of this study is to determine the role of trypsin in the internalization process of the influenza H1N1 virus in Vero and MDCK (Madin-Darby Canine Kidney) cells; to see whether trypsin can trigger the endocytosis route via receptor-mediated endocytosis or non-receptor-mediated endocytosis. The viruses were inoculated into Vero and MDCK cells under three conditions: (1) without trypsin in the medium (VVi and MVi); (2) with medium containing trypsin (VTVi1 and MTVi1); and (3) cells washed with trypsin prior to virus inoculation and incubated with medium containing trypsin (VTVi2 and MTVi2). Advance cytopathic effects (CPE) could only be observed in the MDCK cells. HA (hemagglutination) titer on the MDCK cells was higher than that on the Vero cells. Moreover, HA titer on MTVi1 was higher than MVi, and the lowest was found on MTVi2. Trypsin supported the internalization of the influenza virus by the cleavage of HA glycoprotein into HA1 and HA2. However, trypsin could halt the internalization of the influenza virus when the MDCK cells were washed with trypsin before virus inoculation. Trypsin could digest transmembrane protein containing sialic acid that is important for the virus internalization. Conversely, trypsin did not affect the internalization of the influenza virus in the Vero cells based on HA titer. This result was also supported by TEM observation, which showed that the ability of Vero cellsto internalize the influenza virus is very low. We suggest that a limited amount of sialic acid (α-2,6)–containing receptor on the membrane of the Vero cells can halt influenza virus internalization. On the other hand, active internalization of the virus was seen in all groups of MDCK cells (MVi, MTVi1, and MTVi2). However, trypsin could not trigger the internalization of the influenza virus in Vero and MDCK cells via non-receptor-mediated endocytosis. From this study it can be concluded that trypsin enhanced the internalization of influenza virus into MDCK cells by cleavage of the hemagglutinin (HA) of the virus, but did not improve the ability of the host cells to internalize the virus, especially via non-receptor-mediated endocytosis.
Keywords: influenza; MDCK cell; trypsin; vero cell.
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