THE PPARGC1A - GLY482SER POLYMORPHISM (RS8192678) AND THE METABOLIC SYNDROME IN A CENTRAL ROMANIAN POPULATION.

BACKGROUND: The peroxisome proliferator-activated receptor-γ co-activator 1-α (PPARGC1A), a key transcription factor involved in the control of metabolism and energy homeostasis, is an important biological and positional candidate of the metabolic syndrome. Association studies of its polymorphisms, however, yielded inconsistent sometimes conflicting results, pointing to important ethnic differences, which call for replication in various populations. OBJECTIVE: In order to study its most common - potentially functional - polymorphism Gly482Ser (rs8192678), we carried out a case-control study in a central Romanian population. MATERIAL AND METHODS: Two hundred and ninety six patients affected by the metabolic syndrome diagnosed according to the International Diabetes Federation proposed criteria and 166 middle-aged control subjects have been investigated. Genotyping was done by PCR-RFLP, using the restriction enzyme MspI. RESULTS: While the G(Gly)/A(Ser) allele frequencies (66.89/33.11 vs. 71.68/28.31 %) and GG/GA/AA genotype distribution (45.27-43.24-11.48 vs. 54.21-34.93-10.84 %) differed in the metabolic syndrome and control group, the risk of developing the metabolic syndrome did not reach the limit of statistical significance (OR=1.43; p=0.06, CI 95%: 0.97-2.09). Metabolic parameters in the two study groups did not show significant differences according to the genotype (p>0.05). CONCLUSION: rs8192678 could be a functional polymorphism contributing to the development of the metabolic syndrome, but probably its effect is minor, and might depend on gene-gene and gene-environment interactions. Clarification of very small effects would require larger sample sizes.

Ultrastructural localization of Hsp-72 examined with a new polyclonal antibody raised against the truncated variable domain of the heat shock protein.

In spite of the intensive search for the determination of the continuously widening physiological and pathological roles of different stress proteins, their ultrastructural localization at the electron microscopic (EM) level has hardly been examined. As it becomes increasingly evident that the function and physiological effectiveness of stress proteins are highly dependent on their spatial location and their associations with diverse regulator proteins, the demand for morphological studies which can identify their detailed distribution within the cells is evident. The reason for the practical lack of studies carried out at the EM level, lies in the shortage of reagents with suitable specificity and avidity necessary for this type of examination. To create such a reagent, a polyclonal antibody was raised using a recombinant truncated form of the inducible Hsp-72 protein. The antibody was extensively characterized, using different immunochemical methods to determine and verify its specificity, and then it was tried in ultrastructural examinations. Using the new antibody, it was possible to analyze the intracellular distribution of Hsp-72 with the immunogold technique. The localization of Hsp-72 was demonstrated directly at the ultrastructural level in the cytoplasm (especially at the cisterns of the RER), in the nucleus (mainly around the heterochromatic regions) and at both sides of the nuclear envelope close to the membrane pores. Apart from these localizations, Hsp-72 was found in several membrane bordered intracellular structures, which mainly belong to the endosomal-lysosomal system. We provide the first morphological verification of the appearance of Hsp-72 on the surface of the cells. Also novel is the indication, that the stress protein may recycle from the cell surface using a common route which includes coated pits and the endosomal system.

Polymorphism of 5' of the catalase gene in Hungarian acatalasemia and hypocatalasemia.

The amplified fragment length polymorphism of Hinf1 on the promoter region of the catalase gene in Hungarian acatalasemic and hypocatalasemic patients yielded three different patterns with five bands in total. The sequence analyses revealed A-to-T, C-to-A, and C-to-T mutations at positions -21, -20, and -18 upstream of the translational initiation site. The -21 A-to-T mutations were more frequent in acatalasemic and hypocatalasemic patients (36/2) than in controls (18/14). This mutation had been detected in Japanese acatalasemic patients while the other two are novel mutations. Two extra bands in the Hinf1 pattern are due to star-like activity that cleaved a G/ATTT sequence at position -4 to 0 upstream of the initiation site.

Reference ranges of normal blood catalase activity and levels in familial hypocatalasemia in Hungary.

In 1756 healthy individuals the mean and S.D. values of blood catalase activity were 111.3 +/- 16.5 MU/l with lower blood catalase for females (107.7 +/- 14.4 MU/l, n = 880) than for males (117.9 +/- 16.8 MU/l, n = 876) while the ratios of blood catalase activity to blood hemoglobin concentration were not different (0.841 +/- 0.107 MU/g versus 0.849 +/- 0.119 MU/g). The decrease of blood catalase with age was greater in males (b = -0.084 MU/l year) than in females (b = -0.016 MU/l year). The screening of 3300 healthy citizens for hypocatalasemia yielded six families (0.18%), and three families were identified out of 1630 clinic patients. These nine families revealed 37 hypocatalasemic patients with 57.5 +/- 11.7 MU/l mean and S.D. of blood catalase activity. Similarly to the Japanese and the Hungarian actalasemic patients, the electrophoretic mobilities of catalase in erythrocytes of hypocatalasemic patients were indistinguishable from that of healthy controls.

Hungarian hereditary acatalasemia and hypocatalasemia are not associated with chronic hemolysis.

Two Hungarian acatalasemic and eight hypocatalasemic patients revealed normal erythropoesis. Contrary to their decreased defence system against the toxic hydrogen peroxide, the biochemical tests (serum catalase, serum hemoglobin, serum lactate dehydrogenase (LDH) ratio of LDH1 and LDH2 isoenzymes and serum haptoglobin) excluded hemolysis. The normal activity of glutathione peroxidase and the decreased catalase activity could prevent the lysis of the erythrocytes. In the presence of extremely high levels of hydrogen peroxide acute hemolysis may not be excluded; therefore, follow-up of these patients is required.