Abstracts

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Maeroplwga+womal mzyme rhcalhtion and inactivation by oxidized LDL and pmtective effects of antioxidants Wei Li, Xi Ming Yuan, Am&m G. Olsson, and U!f T. Bnmk Departments of Internal Medicine, Pathology II, and Cl&xl Research Center University of Liokoping, Sweden

The cytotoxicity of oxidized low density lipoprotein (oxLDL) to several types of artery wall cells might contribute to atherosclerosis by causing cell death, presumable by apop- tosis. After its uptake., by receptor-mediated endocytosis, into macrophage lysosomes, OXLDL is poorly degraded resulting in foamcell formation. In order to identify the influ- ence of OXLDL on lyy~somal enzymatic activity and particular, on membrane stability, and its modulation by high density lipoprotein (HDL) and vitamin E J-774 cells and human monocyte-deriued macmphages (HMDMS) were incubated for 24 to 48 b under normal cultore conditions with either OXLDL alone or with OXLDL mixed with HDL or vitamin E. Tbc lysosomal marker enzymes, catbepsin-L and N-ace@-5-glucosaminidasc @&!3Gase), were assayed after cell fractionation while cathcpsin-D was demonstrated immonocybxbemically. A test for lysosomal integrity, estimating tbe uptake of the lyso- somotmpic weak base acridine orange, was applied as well. We found that endocytosed oxLDL not only partially inactivated the lysosomal marker enzymes, but also destabilized the acidic vacuolar compartment and caused relocalization of lytic enzymes to tbe cytosol. It was fort&r found that HDL and vitamin E, when mixed with OXLDL, significantly diminished the cytotoxicity of tbe latter compound. Cells exposed to ultraviolet-oxidized LDL enlarged and acquired a foam-cell morphology. Compared to normal control cells, tbe latter cells showed an enhanced amount of cytosolic cathepsin-D. The results indicate that lysosomal membt;mes and contents were damaged by oxLDL-exposure, resulting in partial relocalization of lysosomal enzymes. Such subcellular damage could be dimin- ished by HDL and vitamin E.

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Serum antioxidant status in type 1 diabetic young patients D. WZlems*, H. Dorchy’* D. Lhfillsne* Department of Clinical Chemistry*, Bmgmann University Hospital and Diabetology Unit**, Children’s Umversity Hospital Queen Fabiola, Brussels, Belgium.

In some stadies, a decreased antioxidant activity has been shown in the semm of diabet- ic patients. The aim of the present study is to investigate parameters involved in cell defence against oxygen radicals ( vitamin .4 and E ), total antioxidant status ( TAS Randox ), autoantibodies against oxidized-LDL-lipoproteins ( o-LAB-Elisa Biomedica ) in rela- tionship with diabetes duration, level of metabolic control ( glycated hemoglobin-HbAlc ), lipids abnormalities ( total cholesterol, HDL-c, LDL-c, triglycerides ) and subclinical complications ( retinopathy, neoropatby, nepbropatby). The shldy includes 110 young type 1 diabetic patients. Tbe median age is 15 years ( 2 to 36 years ) and the median dura- don of diabetes is 5 years ( 6 months to 30 years )~ There is a positive correlation between age and vitamin A ( I = 0.5; p < 0.001 ), TAS ( r = 0.23; p < 0.01 ) and a negative correlation with o-LAB ( r = -0.28; p < 0.005 ). The same relationships an: found with diabetes duration. Serum vitamin E is independent of age or diabetes duration, is positively related to TAS ( I = 0.2, p < 0.05 ) and negatively related to o-LAB ( I= -0.22; p < 0.05 ). HbAlc levels are not related to o-LAB, vitamin A, vitamin E, or TAS. The patients with bigb serum lipids levels have tbe highest levels of serum vitamin A and E.Wben the patients are classified in fimction of tbe presence of subclinical complications, no significant differences are noted for o-LAB, vitamin E and TAS. Yet, tbe patients with nepbmpathy or neuropatby present higher values of vitamin A than tbe patients w&out complications. It is to be noted tbat vitamin A, vitamin E and TAS patients values are within the normal range but 33 patients present o-LAB values above the normal range. Conclusion: Antioxidant status ( TAS and vitamin A ) of young type 1 diabetic patients change with age and diabetes duration, independently of tbe level of metabolic control. A&antibodies against oxidized-LDL lipoproteins decrease with age, as reported in healthy non diabetic patieots.Tbis decrease with age may be associated with tbe onset of atherosclerotic processes if we consider that o-LAB are an accompanying immunological expression of lipid peroxidation

On the definition of prematoro mrneal MW A.F FEndq L.B Day PF flutowski, J.C Jolleys. Royal Free Hospital School of Medicine London and Glentield Hospital NHS Trust, Leicester, UK

Comeal arcus is both common and age dependent, but premature anxs is listed in many guidelines as an indication for lipid screening. what is premature:, We have attempted to define this for a British population by assessing arcus expression in unselected patients consecutively attending a geneml practice, and in confirmed familial hypercbolestero- lacmia [FH] betemzygotes assessed at presentation before lipid therapy. Arcus was recorded by two experienced observers, on a previously established graded scale of O-4, nil to full ring arcus. Progression with age for males, females and ovetall for the normal and FH groups was expressed in three ways: as the mean age of patients with each arcus grade, as the cumulative proportion of cases with a specified grade or any grade of arcus, and as the proportion of patients within age steps of 5 years with a specified grade or any grade of arcus to avoid any bias through age at recruitment on tbe cumulative distribution. For tbe 275 male, 351 female total 626 unselected patients age range 16-76 years, 50% bad some degree of arcus of any grade by age 4145 years, and by 3 l-35 years for tbe 59 male, 41 female total 100 FH of similar age profile. Heavy grade 4 arcus was present in 50% of the FH group by age 50 years and in the practice group by age 68 years, of whom 5% were then affected by age 50 years. Arcus progression was advanced by average 3 years in males for both tbe unselected and FH groups, this difference reducing with age, consistent with the increased proportion of cholesterol as LDL in males. These observa- tions show that some degree of arcus is common in early middle age on assessment by experienced observers using good illumination, and confum that premature heavy arcus has a variable relationship with but can alert to hypercbolesterolaemia. This study also supports a broad definition of premature arcus for males and females combined as heavy full ring involvement by age 50 years.

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The discovery of leptin and the leptin receptor: Perspectives for the treatment of obesity and wasting disorders Rem Dews Roche Research Gent

The abihty to store large quanhties of energy-dense fuel in the form of adipose tissue allows survival during prolonged periods of food deprivation. To maintain such fix1 stores without undergoing continual alterations in body size and shape, a balance between ener- gy intake and expenditure needs to be achieved. Leptin, the product of the ob gene, is secreted exclusively by white fat adipocytes, and appears to be tbe long sought a&x sati- ety factor released in tbe periphery to regulate long-term body weight. In tbis lipstatic theory of body weight maintenance, as body fat increases more leptin is secreted, thereby bringing the fat mass back to a certain normal set point. Once released into tbe circula- tion, leptin lowers body weight by decreasing appetite and altering metabolic processes. While the exact mechanism whereby leptin enters the brain needs to be elucidated, the hypothalamus appears to be the major target tissue for this hormone, since leptin recep- tors are present within the appetite control centem of this region, and since injection of leptin into the ventricular system of the brain reduces food intake and increases energy expenditure. Although the ceniral mechanism of action has not yet been elucidated, leptin seems to alter energy expenditure by enhancing sympathetic outflow thmugb modulation of synaptic transmission in the arcoate nucleus of the hypothalamus rather than through gene regulation. NPY, a neumpeptide known to increase food intake and body weight when injected in the brain, is coexpressed with tbe leptin receptor in the anxate nucleus neurons and placed downstream in the biological response of @tin. Leptin is present at high concentration in the serum of obese individuals, which suggests a central resistance to leptin, and brings into question the value of recombinant leptin for therapeutic applications. While the molecular mechanism for this resistance might per- haps be explained at tbe level of a defective transport to the brain or a defective leptin receptor signaling and/or translocation, a more plausable explanation is to be sought in defects of other components regulating energy expenditure, leading to a satiration of lep- tin. Clinically, lcptin might however be effective for body weight mainwce after weight-loss through surgery (virtual fat). Antagonists of NPY or its hypothalamic recep- tor are currently under development for treatment of obesity. However, unlike for genetic animal models of obesity, there is no evidence for an increased central NPY level as a causative agent in human obesity. Recenly, genetic shldies in mice have identified anoth- er component of tbe system that mediates weight control, the melanocortin-4 receptor, which when antagonized lea& to obesity and lack of response to increased concentrations of leptin in tbe blood. Since this situation resembles the prominent leptin resitance observed in human obesity, this component will certainly receive much further attention for understanding and fighting tbis disease and all its complications. It has been demon- strated that inflammatory cytokines such as TNF, produces anorexia through the induction of leptin. This strongly advances the hypothesis that leptin may play a significant role in the anorexia and cachcxia of inflammatory diseases and cancer. We have developed a human leptin antagonist which induces a progressive increase in body weight in normal nuce. This antagonist could be of therapeutic use for wasting disorders.