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Laboratory of Molecular Structure and Function

In our laboratory, we aim at the establishment of tailor-made-medicine for hypertriglyceridemia. For this purpose, we have developed the methods for measuring activities and masses of lipoprotein lipase (LPL) and hepatic triglyceride lipase (HTGL) and have accumulated LPL gene mutations in Japanese, and analyzed gene environment interaction on the expression of hypertriglyceridemia.

1. Tailor-made medicine for hypertriglyceridemia through developing an early diagnostic method for identifying genetic mutations

Recently, hypertriglyceridemia attracts attention as one of the defined components of metabolic syndrome. A prolonged mild hypertriglyceridemia changes the properties of low-density lipoprotein to small-dense LDL, which is thought to elevate the risk for cardiovascular disease.

We established the method of sandwich-enzyme immunoassay for the quantification of LPL mass (J Lipid Res 31: 1911-1924, 1990) and SIIA (Selective Immuno Inactivation Assay) (Biochim Biophys Acta 1003: 254-269, 1989)for the quantification of LPL activity in human postheparin plasma (Fig.1). By primary screening of LPL mass in postheparin plasma, we have succeeded in identifying the LPL gene mutations from patients with hypertriglyceridemia.

We elucidated that mild hypertriglyceridemia results from superimposition of triglyceride-synthesis-stimulating factors, such as high alcohol intake and a hyperinsulinemic state, on a genetic background of a heterozygous mutation in the LPL gene (Fig. 2).

So far, in Japanese, 30 LPL gene mutations resulting in non-functional LPL molecule have been reported, in which 21 LPL gene mutations have been collected in our laboratory (J Clin Invest 89: 581-591, 1992., Bioch Biophys Acta 1502: 433-446, 2000., Nuc Acids Res 32: e141, 2004.). Now, we are aiming to apply the established early diagnostic system of LPL gene mutations to tailor-made medicine for hypertriglyceridemia.

2. Studies on physiological roles of hepatic triglyceride lipase and its contribution to atherogenic lipoprotein production

Hyperlipoproteinemia is one of risk factors for atherosclerosis. Lipoprotein lipase (LPL), hepatic triglyceride lipase (HTGL), cholesterol ester transfer protein (CETP), and lecithin cholesterol acyltransferase (LCAT) are involved in metabolism of plasma lipoproteins. However, the physiological role of HTGL remains to be elucidated. To clarify it, we have purified HTGL from human postheparin plasma and raised monoclonal and polyclonal antibodies against human HTGL. By using them, we have developed a sandwich-enzyme immunoassay (J Immunol Methods 235: 41-51, 2000.) and selective immunoincativation assay (Biochim Biophys Acta 1003: 254-269, 1989)for quantification of HTGL mass and activity in human postheparin plasma, respectively. These systems lead to identify a patient with HTGL deficiency in the first time in Japanese. By the analysis of the patient's lipoprotein profile, we have demonstrated that HTGL hydrolyzes triglycerides in smaller lipoprotein like low, intermediate and high density lipoproteins.

It is known that a prolonged hypertriglyceridemia changes the properties of low density lipoprotein (LDL) to small dense LDL (sLDL), which is thought to elevate the risk for cardiovascular heart disease. To know role of HTGL for sLDL formation, we systematically investigated the mechanism of sLDL formation in hypertriglyceridemia due to accumulation of triglyceride (TG)-rich lipoprotein (TGRL). Our investigation lead to the conclusion that high density lipoprotein (HDL) particles associated with CETP and LCAT played a central role for the production of sLDL. Our proposed mechanism is explained as follows: in the first step, HDL particles are modified to TG-rich/cholesteryl ester (CE)-poor HDL by CETP-mediated lipid transfer between HDL and TGRL, which is elevated in hypertriglyceridemia; (ii) in the next step, the modified TG-rich/CE-poor HDL remodels LDL to TG-rich/PL-, FC- and CE-poor LDL; (iii) in the final step, TG particles in the modified LDL is hydrolyzed by HTGL, and LDL size and density result in small and dense (Ikeda Y, Takagi A et al.: Physiological role of human hepatic triglyceride lipase (HTGL): HTGL regulates the multiple lipoprotein metabolism of IDL, LDL, and HDL2. In: Multiple risk factors in cardiovascular disease, Ed. Yamamoto A (Churchill Livingstone, Tokyo) pp.181-186, 1994)

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