Sains Malaysiana 38(5)(2009): 761–766

Infeksi Plasmodium berghei dan Kesannya ke atas Pengisyaratan MAP Kinase Eritrosit Perumah

(Plasmodium berghei Infection and its Effect on MAP Kinase Signaling in its Erythrocyte Host)

Mohd Fakharul Zaman Raja Yahya

Pusat Pengajian Bioperubatan & Kesihatan

Kolej Universiti Kejururawatan & Kesihatan Masterskill

43200 Cheras, Selangor Darul Ehsan, Malaysia

Hasidah Mohd Sidek*

Pusat Pengajian Biosains & Bioteknologi

Fakulti Sains & Teknologi, Universiti Kebangsaan Malaysia

43600 UKM Bangi, Selangor Darul Ehsan, Malaysia

Diserahkan: 24 Jun 2008 / Diterima: 12 Mac 2009

ABSTRAK

Kajian ini melibatkan pemantauan perkembangan parasitemia dan taburan morfologi Plasmodium berghei sewaktu infeksi parasit dalam mencit, serta penentuan kesan infeksi P. berghei ke atas pengisyaratan MAP kinase eritrosit perumah. Analisis mikroskop ke atas slaid calitan darah terwarna-Giemsa yang disediakan daripada mencit terinfeksi-P. berghei (strain PZZ1/00) menunjukkan darjah parasitemia mencapai sehingga 70% dalam masa dua minggu selepas penyuntikan parasit. Morfologi cecincin dan trofozoit parasit dicerap dengan jelas sepanjang tempoh infeksi manakala morfologi skizon parasit hanya dicerap dengan ketara selepas hari ketiga selepas penyuntikan parasit. Pemblotan Western [antibodi primer: anti-MAP kinase (ERK-1/2 tak terfosfat) monoklon; antibodi sekunder: anti-IgG, poliklon terkonjugat-HRP] ke atas protein sitosol eritrosit terinfeksi-P. berghei (70% parasitemia) susulan pemisahan SDS-PAGE menunjukkan bahawa keamatan protein imunoreaktif-MAP kinase eritrosit berberat molekul 42 dan 44 kDa didapati meningkat secara signifikan (p<0.05) pada 70% iaitu peningkatan sebanyak 21.5% dan 22.3% masing-masing berbanding sampel kawalan tanpa infeksi. Samada kesan infeksi P. berghei (70% parasitemia) ke atas pengisyaratan MAP kinase perumah ini berkaitan dengan pengaktifan enzim ini perlu dikaji dengan lebih lanjut.

Kata kunci: MAP kinase; malaria; Plasmodium berghei; transduksi isyarat

ABSTRACT

The present investigation involves monitoring the development of parasitemia and the distribution of Plasmodium berghei morphologies during parasite infection in mice as well as the effect P. berghei infection on MAP kinase signaling in its erythrocyte host. Microscopic analyses of Giemsa-stained blood films prepared from P. berghei (strain PZZ1/00)-infected mice showed that the level of parasitemia reached up to 70% two weeks after inoculation of the parasite. Parasite ring and trophozoite forms were clearly detected in the blood films throughout the study period while the schizont form was visibly observed on day 3 post-inoculation. Western blotting [primary antibody: monoclonal anti-MAP kinase (non-phosphorylated ERK-1/2); secondary antibody: HRP-conjugated polyclonal anti-IgG] carried out on SDS-PAGE-separated cytosolic protein samples from P. berghei-infected (70% parasitemia) erythrocytes showed that the levels of both 42 and 44 kDa MAP kinase-immunoreactive proteins increased significantly (p<0.05) at 70% parasitemia, by up to 21.5% and 22.3% respectively as compared to non-infected control samples. Whether the effect of P. berghei infection at 70% parasitemia on host MAP kinase signaling is related to the MAP kinase activation remains to be further investigated.

Keyword: Malaria; MAP kinase; Plasmodium berghei; signal transduction

RUJUKAN

Bavil, A.A., Hayes, S., Goretzki, L., Kroger, M., Anders, J. & Hendriks, R. 2004. Convenient and versatile subcellular extraction procedure, that facilitates classical protein expression profiling and functional protein analysis. Proteomics 4: 1397-1405.

Berman, K., Cobb, M.H., Gibson, T.B., Karandikar, M., Pearson, G., Robinson, F. & Xu, B.E. 2001. Mitogen-activated protein (MAP) kinase pathways: Regulation and physiological functions. Endocrine Reviews 22: 153-183.

Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein – dye binding. Analytical Biochemistry 72: 248-254 .

Chaussepied, M., Lallemand, D., Moreau, M.F., Adamson, R., Hall, R. & Langsley, G. 1998. Upregulation of Jun and Fos family members and permanent JNK activity lead to constitutive AP-1 activation in Theileria-transformed leukocytes. Molecular & Biochemical Parasitology 94: 215-226.

David, A.F., Phillips, J.R., Simon, L.C., Reto, B. & Solomon, N. 2004. Antimalarial drug discovery: Efficacy models for compound screening. Nature Reviews 3: 509-520.

Denkers, E.Y., Butcher, B.A., Del Rio, L. & Kim, L. 2004. Manipulation of mitogen-activated protein kinase/nuclear factor-kappaβ-signaling cascades during intracellular Toxoplasma gondii infection. Immunology Review 201: 191-205.

Dessauge, F., Hilaly, S., Baumgartner, M., Blumen, B., Werling, D. & Langsley, G. 2005. c-Myc activation by Theileria parasites promotes survival of infected B-lymphocytes. Oncogene 24: 1075-1083.

Dobbelaere, D.A. & Kuenzi, P. 2004. The strategies of Thieleria parasite: A new twist in host-pathogen interactions. Current Opinion in Immunology 16: 524-530.

Doerig, C. 2004. Protein kinases as targets for anti-parasitic chemotherapy. Biochemica et Biophysica Acta 1697: 155-168.

Evans, J.L., Goldfine, I.D., Maddux, B.A. & Grodsky, G.M. 2002. Oxidative stress and stress-activated signaling pathways: a unifying hypothesis of type 2 diabetes. Endocrine Review 23: 599-622.

Field, J.W. & Shute, P.G. 1955. The microscopy diagnosis of human malaria: II-A morphological study of the erythrocytic parasites. Studies from the Institute For Medical Research Federation of Malaya (4). Hlm. 93-129. Kuala Lumpur: The Government Press.

Frantzen, F., Grimsrud, K., Heggli, D.E. & Sundrehagen, E. 1997. Selective precipitation of human hemoglobin by organic solvents and metal cations. Hemoglobin 21: 155-172.

Hall, B.S., Daramola O.O., Barden, G. & Targett, G.A. 1997. Modulation of protein kinase C activity in Plasmodium falciparum-infected erythrocytes. Blood 89: 1770-1778.

Jiang, L., He, L. & Fountoulakis, M. 2004. Comparison of protein precipitation methods for sample preparation prior to proteomic analysis. Journal of Chromatography A 1023: 317-320.

Johnson, G.L. & Lapadat, R. 2002. Mitogen-activated protein kinase pathways mediated by ERK, JNK and p38 protein kinases. Science 298: 1911-1912.

Jones, G. L. & Edmundson, H.M. 1990. Protein phosphorylation during asexual life cycle of the human malarial parasite Plasmodium falciparum. Biochemica et Biophysica Acta 1053: 118-124.

Laemmli, U.K. 1970. Cleavage of structure proteins during the assembly of the head of bacteriophage T4. Nature 341: 152-154.

Laurent, D. 2006. Antimalarial potential of xestoquinone, a protein kinase inhibitor isolated from a Vanuatu marine sponge Xestospongia. Bioorganic and Medical Chemistry 14: 4477-4482.

Liao, P., Georgakopoulos, D. & Kovacs, A. 2001. The in vivo role of p38 MAP kinases in cardiac remodeling and restrictive cardiomyopathy Proceeding of National Academy Science United States of America 98: 12283-12288

Macpherson, G.G., Warrel, M.J., White, N.J., Looareesuwan, S. & Warrell, D.A. 1985. Human cerebral malaria: A quantitative ultrastructural analysis of parasitized erythrocyte sequestration. American Journal of Pathology 119: 385-401.

Phillips, R.S. 1983. Studies in biology no. 152 malaria. First edition. Edward Arnold (Publisher) Limited.

Roberts, P.J. & Der, C.J. 2007. Targeting the Raf-MEK-ERK mitogen-activated protein kinase cascade for the treatment of cancer. Oncogene 26: 3291-3310.

Thomas, V. 1983. Parasitologi perubatan. First Edition. Kuala Lumpur: Dewan Bahasa Pustaka.

Thurston, J.P. 1953. Parasitological reviews: Plasmodium berghei. Experimental Parasitology 2: 311-332.

Towbin, H., Staehelin, T. & Gordon, J. 1979. Electrophoretic transfer of proteins from polyacrylamide to nitrocellulose sheets: Procedure and some applications. Proceedings of the National Academy of Sciences 76: 4350-4354.

*Pengarang untuk surat-menyurat; email: hasidah@ukm.my