Otto Warburg在1926年提出，發現腫瘤即使在有氧情況下會提高醣解作用效率獲得ATP。近年的研究指出腫瘤的某些致癌基因可以提高醣解酵素的象，解釋了Warburg所發現的現象。果醣雙磷酸醛縮酶是醣解作用第四個酵素，存在三種同功異構酶A、B和C型，分別由三個不同基因所表現，這三型在不同組織與胚胎發育有不同表現量。過去研究指出果醣雙磷酸縮醛酶在腫瘤細胞會被正調控表現促進醣解作用，但是有關於果醣雙磷酸醛縮酶在腫瘤研究是很少。在我的實驗中，我首先要利用RNAi技術建立穩定沉默aldolase A的子宮頸癌細胞株HeLa。接下來，分析shAldoA與野生型HeLa的外型和生長，兩者間沒有明顯差異。我們也利用西方式點墨法分析醣解與檸檬酸循環酵素之間也沒顯著差異，但進行免疫螢光染色發現aldolaseA也存在細胞核中，而核質分離實驗也證實我們看到的現象。雖然在正常環境看不到差異，但是給於一個壓力(stress)在shAldoA細胞株是否能看到差異之處？這次設計三種壓力，分別是缺氧環境和兩種電子傳遞鏈抑制劑。在缺氧環境實驗裡，細胞存活情形、細胞週期分析以及西方點墨法分析沒有很顯著差異。在電子傳遞鏈的抑制劑實驗上，我們使用的是Rotenone和TTFA。在Rotenone和TTFA的細胞群落分析發現，發現shAldoA細胞株對這兩種藥物有很高的敏感性，尤其在Rotenone的最低濃度對於shAldoA細胞有很強烈致死效果。接下來也使用了臨床的抗癌藥物2-Deoxyglucose，也觀察到在Colony formation存活能力和Annexin-FITC實驗偵測其死亡情形，shAldoA細胞株對於此藥物具有稍高的敏感性。我們也使用另外一種臨床治癌用藥Etoposide，這是ㄧ種拓樸酶二型(Topoisomerase II)抑制劑，會導致細胞DNA雙股螺旋斷裂，但是沒有發現跟先前藥物處理類似的結果，可能其作用機制或許跟前三者不同，前三者是跟阻斷能量生成相關，而Etoposide直接是影響DNA結構穩定性。在這三種有效藥物中，兩個作用在粒腺體ㄧ個則在醣解代謝，其實不太一樣，未來需要更深入研究才能釐清Aldoalse A缺陷與細胞存活有何關聯。

Otto Warburg addressed a phenomenon that cancer cells prefer the glycolysis pathway to the TCA cycle for obtaining ATP even in aerobic conditions in 1926. Later researches found that some excessive activated oncogenes could mediate the activity of the glycolysis pathway through modulate glycolytic enzymes. Three aldolase isozymes (A, B, and C), encoded by three different genes, are differentially expressed during development. In past suduy, aldolase was known to be upregulated in tumor cells to promote the glycolysis,but reports about aldolase were rare.I used the RNAi-mediated technique to stably knock down the endogenous aldolase A in cervical HeLa cancer model successfully. Analysis of the growth and morphology between wild type and Aldolase A deficient HeLa cells, there does not a difference exist. In immunostaining experiment,although alsdolase is a glycolytic enzyme in the cytoplasma,but I found that Aldolase A also existed in nuclei. Although in normal condition there don’t exist any differences between HeLa-shAldoA and wild cells, we tried give the stress on HeLa-shAldoA cell line. There are some designs: hypoxia and two ETC inhibitors. Because aldolase was induced in hypoxia condition, we treated the cell in hypoxia. In hypoxia experiments, there don’t exist the difference between them. In ETC inhibitors experiments, we found the HeLa-shAldoA was more sensitive than wild cells.We also used two clinical drugs: 2-deoxyglucose and etoposide,the inhibitor of glucose metabolism and topoisomeriase II,respectively.In the treatment of 2-deoxyglucose, shAldoA cells were more sensitive,but not sensitive to etoposide.We speculated that etoposode resulted in the cell death through the genotoxic damage and others are related with the ATP biosynthesis.Three effective drugs,in fact,2-DG inhibits glycolysis and others inhibits ETC,the mechanism involed in cell death of shAldoA was different.In future,we need time to explore what the role aldolase A play on the cell survival.

Ambrus, A., Tretter, L., and Adam-Vizi, V. (2009). Inhibition of the alpha-
ketoglutarate dehydrogenase-mediated reactive oxygen species generation by lipoic
acid. J Neurochem 109 Suppl 1, 222-229.

Arora, K.K., and Pedersen, P.L. (1988). Functional significance of mitochondrial bound
hexokinase in tumor cell metabolism. Evidence for preferential phosphorylation of
glucose by intramitochondrially generated ATP. J Biol Chem 263, 17422-17428.

Bustamante, E., and Pedersen, P.L. (1977). High aerobic glycolysis of rat hepatoma cells
in culture: role of mitochondrial hexokinase. Proc Natl Acad Sci U S A 74,
3735-3739.

Bustamante, E., Morris, H.P., and Pedersen, P.L. (1981). Energy metabolism of tumor
cells. Requirement for a form of hexokinase with a propensity for mitochondrial
binding. J Biol Chem 256, 8699-8704.

Bensaad, K., Tsuruta, A., Selak, M.A., Vidal, M.N., Nakano, K., Bartrons, R., Gottlieb
, E., and Vousden, K.H. (2006). TIGAR, a p53-inducible regulator of glycolysis and
apoptosis. Cell 126, 107-120.

Bensaad, K., and Vousden, K.H. (2007). p53: new roles in metabolism. Trends Cell
Biol 17, 286-291.

Boldogh, I.R., and Pon, L.A. (2006). Interactions of mitochondria with the actin cyto-
skeleton. Biochim Biophys Acta 1763, 450-462.

Breitenbach, M., Laun, P., and Gimona, M. (2005). The actin cytoskeleton,Ras-cAMP
signaling and mitochondrial ROS in yeast apoptosis. Trends Cell Biol 15, 637-639.

BeltrandelRio, H., and Wilson, J.E. (1992). Coordinated regulation of cerebral glycolytic
and oxidative metabolism, mediated by mitochondrially bound hexokinase dependent
on intramitochondrially generated ATP. Arch Biochem Biophys 296, 667-677.

Corcoran, C.A., Huang, Y., and Sheikh, M.S. (2006). The regulation of energy genert-
ating metabolic pathways by p53. Cancer Biol Ther 5, 1610-1613.

Czyzewska J, Guzińska-Ustymowicz K, Kemona A, Bandurski R.(2008). The expres-
sion of matrix metalloproteinase 9 and cathepsin B in gastric carcinoma is associated
with lymph node metastasis, but not with postoperative survival. Folia Histochem
Cytobiol 46, 57-64.

Chiquete-Felix, N., Hernandez, J.M., Mendez, J.A., Zepeda-Bastida, A., Chagolla-
Lopez, A., and Mujica, A. (2009). In guinea pig sperm, aldolase A forms a complex
with actin, WAS, and Arp2/3 that plays a role in actin polymerization. Reproduction
137, 669-678.

Di Lisa, F., Kaludercic, N., Carpi, A., Menabo, R., and Giorgio, M. (2009). Mitochon-
drial pathways for ROS formation and myocardial injury: the relevance of p66(Shc)
and monoamine oxidase. Basic Res Cardiol 104, 131-139.

Ehsani-Zonouz, A., Golestani, A., and Nemat-Gorgani, M. (2001). Interaction of
hexokinase with the outer mitochondrial membrane and a hydrophobic matrix. Mol
Cell Biochem 223, 81-87.

Fiek, C., Benz, R., Roos, N., and Brdiczka, D. (1982). Evidence for identity between the
hexokinase-binding protein and the mitochondrial porin in the outer membrane of rat
liver mitochondria. Biochim Biophys Acta 688, 429-440.

Feron, O. (2009). Pyruvate into lactate and back: From the Warburg effect tosymbio-
tic energy fuel exchange in cancer cells. Radiother Oncol.

Funasaka, T., Hogan, V., and Raz, A. (2009). Phosphoglucose isomerase/autocrine
motility factor mediates epithelial and mesenchymal phenotype conversions in breast
cancer. Cancer Res 69, 5349-5356.

Funasaka, T., Haga, A., Raz, A., and Nagase, H. (2001). Tumor autocrine motility factor
is an angiogenic factor that stimulates endothelial cell motility. Biochem Biophys Res
Commun 285, 118-128.

Funasaka, T., Haga, A., Raz, A., and Nagase, H. (2002). Autocrine motility factor
secreted by tumor cells upregulates vascular endothelial growth factor receptor (Flt-1)
expression in endothelial cells. Int J Cancer 101, 217-223.

Gogvadze, V., Orrenius, S., and Zhivotovsky, B. (2008). Mitochondria in cancer cells:
what is so special about them? Trends Cell Biol 18, 165-173.

Gourlay, C.W., and Ayscough, K.R. (2005). The actin cytoskeleton: a key regulator of
apoptosis and ageing? Nat Rev Mol Cell Biol 6, 583-589.

Hamaguchi, T., Iizuka, N., Tsunedomi, R., Hamamoto, Y., Miyamoto, T., Iida, M.,
Tokuhisa, Y., Sakamoto, K., Takashima, M., Tamesa, T., et al. (2008). Glycolysis
module activated by hypoxia-inducible factor 1alpha is related to the aggressive
phenotype of hepatocellular carcinoma. Int J Oncol 33, 725-731.

Hatzivassiliou, G., Zhao, F., Bauer, D.E., Andreadis, C., Shaw, A.N., Dhanak, D.,
Hingorani, S.R., Tuveson, D.A., and Thompson, C.B. (2005). ATP citrate lyase
inhibition can suppress tumor cell growth. Cancer Cell 8, 311-321.

Hirono, Y., Fushida, S., Yonemura, Y., Yamamoto, H., Watanabe, H., and Raz, A. (1996).
Expression of autocrine motility factor receptor correlates with disease progression in
human gastric cancer. Br J Cancer 74, 2003-2007.

Ishikawa, K., Takenaga, K., Akimoto, M., Koshikawa, N., Yamaguchi, A., Imanishi,
H., Nakada, K., Honma, Y., and Hayashi, J. (2008). ROS-generating mitochondrial
DNA mutations can regulate tumor cell metastasis. Science 320, 661-664.

Kaelin, W.G., Jr. (2008). The von Hippel-Lindau tumour suppressor protein: O2
sensing and cancer. Nat Rev Cancer 8, 865-873.

King, A., Selak, M.A., and Gottlieb, E. (2006). Succinate dehydrogenase and
fumarate hydratase: linking mitochondrial dysfunction and cancer. Oncogene
25, 4675-4682.

Knott, A.B., Perkins, G., Schwarzenbacher, R., and Bossy-Wetzel, E. (2008).
Mitochondrial fragmentation in neurodegeneration. Nat Rev Neurosci 9, 505-518.

Kim, J.W., and Dang, C.V. (2005). Multifaceted roles of glycolytic enzymes. Trends
Biochem Sci 30, 142-150.

Linden, M., Gellerfors, P., and Nelson, B.D. (1982). Pore protein and the
hexokinase-binding protein from the outer membrane of rat liver mitochondria are
identical. FEBS Lett 141, 189-192.

Lee, H.C., and Wei, Y.H. (2009). Mitochondrial DNA instability and metabolic
shift in human cancers. Int J Mol Sci 10, 674-701.

Lu, J., Suzuki, T., Satoh, M., Chen, S., Tomonaga, T., Nomura, F., and Suzuki, N. (2008).
Involvement of aldolase A in X-ray resistance of human HeLa and UV(r)-1 cells.
Biochem Biophys Res Commun 369, 948-952.

Liotta, L.A., Mandler, R., Murano, G., Katz, D.A., Gordon, R.K., Chiang, P.K., and
Schiffmann, E. (1986). Tumor cell autocrine motility factor. Proc Natl Acad Sci U S
A 83, 3302-3306.

Matoba, S., Kang, J.G., Patino, W.D., Wragg, A., Boehm, M., Gavrilova, O., Hurley,
P.J., Bunz, F., and Hwang, P.M. (2006). p53 regulates mitochondrial respiration.
Science 312, 1650-1653.

Majewski, N., Nogueira, V., Bhaskar, P., Coy, P.E., Skeen, J.E., Gottlob, K., Chandel,
N.S., Thompson, C.B., Robey, R.B., and Hay, N. (2004). Hexokinas mitochondria
interaction mediated by Akt is required to inhibit apoptosis in the presence or absence
of Bax and Bak. Mol Cell 16, 819-830.

Morioka, M. (1992). [Clinical significance of aldolase A in sera of patients with
leukemia]. Rinsho Ketsueki 33, 1191-1198

Musolino, C., Alonci, A., Allegra, A., Di Cesare, E., Orlando, A., Grosso, P., Buda,
G., and Squadrito, G. (1992). Intracellular and serum levels of aldolase activity in
B chronic lymphocytic leukemia. Int J Hematol 56, 213-217.

Mathupala, S.P., Rempel, A., and Pedersen, P.L. (1997). Aberrant glycolytic metabolism
of cancer cells: a remarkable coordination of genetic, transcriptional,post-translastion,
and mutational events that lead to a critical role for type II hexokinase. J Bioenerg
Biomembr 29, 339-343.

Miller, R.L., James-Kracke, M., Sun, G.Y., and Sun, A.Y. (2009). Oxidative and
inflammatory pathways in Parkinson's disease. Neurochem Res 34, 55-65.

Meira, D.D., Marinho-Carvalho, M.M., Teixeira, C.A., Veiga, V.F., Da Poian, A.T.,
Holandino, C., de Freitas, M.S., and Sola-Penna, M. (2005). Clotrimazole
decreases human breast cancer cells viability through alterations in
cytoskeleton-associated glycolytic enzymes. Mol Genet Metab 84, 354-362

Nakashima, R.A. (1989). Hexokinase-binding properties of the mitochondrial VDAC
protein: inhibition by DCCD and location of putative DCCD-binding sites. J Bioenerg
Biomembr 21, 461-470.

Nakashima, R.A., Mangan, P.S., Colombini, M., and Pedersen, P.L. (1986). Hexokinase
receptor complex in hepatoma mitochondria: evidence from
N,N'-dicyclohexylcarbodiimide-labeling studies for the involvement of the
pore-forming protein VDAC. Biochemistry 25, 1015-1021.

Nakashima, R.A. (1989). Hexokinase-binding properties of the mitochondrial VDAC
protein: inhibition by DCCD and location of putative DCCD-binding sites. J Bioenerg
Biomembr 21, 461-470.

Nabi, I.R., Watanabe, H., and Raz, A. (1992). Autocrine motility factor and its receptor:
role in cell locomotion and metastasis. Cancer Metastasis Rev 11, 5-20.

Nabi, I.R., Watanabe, H., and Raz, A. (1990). Identification of B16-F1 melanoma
autocrine motility-like factor receptor. Cancer Res 50, 409-414.

Nakashima, K., Miwa, S., Oda, S., Tanaka, T., and Imamura, K. (1974). Electrophoretic
and kinetic studies of mutant erythrocyte pyruvate kinases. Blood 43, 537-548.

Noguchi, T., Inoue, H., and Tanaka, T. (1986). The M1- and M2-type isozymes of rat
pyruvate kinase are produced from the same gene by alternative RNA splicing. J Biol
Chem 261, 13807-13812.

Polakis, P.G., and Wilson, J.E. (1985). An intact hydrophobic N-terminal sequence is
critical for binding of rat brain hexokinase to mitochondria. Arch Biochem Biophys
236, 328-337.

Qu, Y., Mao, M., Zhao, F., Zhang, L., and Mu, D. (2009). Proapoptotic Role of Human
Growth and Transformation-Dependent Protein in the Developing Rat Brain After
Hypoxia-Ischemia. Stroke.

Reisch, A.S., and Elpeleg, O. (2007). Biochemical assays for mitochondrial activity: assays
of TCA cycle enzymes and PDHc. Methods Cell Biol 80, 199-222.

Semenza, G.L., Jiang, B.H., Leung, S.W., Passantino, R., Concordet, J.P., Maire, P.,
and Giallongo, A. (1996). Hypoxia response elements in the aldolase A, enolase 1,
and lactate dehydrogenase A gene promoters contain essential binding sites for
hypoxia-inducible factor 1. J Biol Chem 271, 32529-32537.

Shoshan-Barmatz, V., Keinan, N., and Zaid, H. (2008). Uncovering the role of VDAC
in the regulation of cell life and death. J Bioenerg Biomembr 40, 183-191.

Sun, L., Shukair, S., Naik, T.J., Moazed, F., and Ardehali, H. (2008). Glucose
phosphorylation and mitochondrial binding are required for the protective effects of
hexokinases I and II. Mol Cell Biol 28, 1007-1017.

Takashi, M., Zhu, Y., Nakano, Y., Miyake, K., and Kato, K. (1992). Elevated levels of
serum aldolase A in patients with renal cell carcinoma. Urol Res 20, 307-311.

Takeuchi, H., Yanagida, T., Inden, M., Takata, K., Kitamura, Y., Yamakawa, K.,
Sawada, H., Izumi, Y., Yamamoto, N., Kihara, T., et al. (2009). Nicotinic receptor
stimulation protects nigral dopaminergic neurons in rotenone-induced Parkinson's
disease models. J Neurosci Res 87, 576-585.

Tsutsumi, S., Hogan, V., Nabi, I.R., and Raz, A. (2003). Overexpression of the autocrine
motility factor/phosphoglucose isomerase induces transformation and survival of
NIH-3T3 fibroblasts. Cancer Res 63, 242-249.

Xie, G.C., and Wilson, J.E. (1988). Rat brain hexokinase: the hydrophobic N-terminus of
the mitochondrially bound enzyme is inserted in the lipid bilayer. Arch Biochem
Biophys 267, 803-810.

Xu, X., Zur Hausen, A., Coy, J.F., and Lochelt, M. (2009). Transketolase-like protein
1 (TKTL1) is required for rapid cell growth and full viability of human tumor cells.
Int J Cancer 124, 1330-1337.

Vander Heiden, M.G., Cantley, L.C., and Thompson, C.B. (2009). Understanding the
Warburg effect: the metabolic requirements of cell proliferation. Science 324,1029-1033

Warburg, O. (1956). On the origin of cancer cells. Science 123, 309-314.

Wilson, J.E. (1995). Hexokinases. Rev Physiol Biochem Pharmacol 126, 65-198.

Wentzensen, N., and Klug, S.J. (2009). Cervical cancer control in the era of HPV
vaccination and novel biomarkers. Pathobiology 76, 82-89.

Wang, X., Figueroa, B.E., Stavrovskaya, I.G., Zhang, Y., Sirianni, A.C., Zhu, S.,
Day, A.L., Kristal, B.S., and Friedlander, R.M. (2009). Methazolamide and melatonin
inhibit mitochondrial cytochrome C release and are neuroprotective in experimental
models of ischemic injury. Stroke 40, 1877-1885.

Watanabe, H., Carmi, P., Hogan, V., Raz, T., Silletti, S., Nabi, I.R., and Raz, A. (1991).
Purification of human tumor cell autocrine motility factor and molecular cloning of its
receptor. J Biol Chem 266, 13442-13448.

Zundorf, G., Kahlert, S., Bunik, V.I., and Reiser, G. (2009). alpha-Ketoglutarate
dehydrogenase contributes to production of reactive oxygen species in
glutamate-stimulated hippocampal neurons in situ. Neuroscience 158, 610-61