Abnormal folic асid homocysteine metabolism as maternal risk factors for Down syndrome in Japan Текст научной статьи по специальности «Клиническая медицина»

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КЛИНИЧЕСКАЯ МЕДИЦИНА

ABNORMAL FOLIC ACm HOMOCYSTEINE METABOLISM AS MATERNAL RISK FACTORS FOR DOWN SYNDROME IN JAPAN

Noboru Takamura, Tatsuro Kondoh, Syohei Ohgi, Kokichi Arisawa, Mariko Mine,

Shunichi Yamashita, and Kiyoshi Aoyagi

Department of Public Health, Department of Pediatrics, Department of Molecular Epidemiology, Biostatistics Section and Department of Molecular Medicine,Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan

Background: Japan has been considered as «a folate sufficient area», since traditional Japanese food contains an adequate amount of folic acid. However, the recent westernized food style of young Japanese mothers may affect the intake of folic acid among them. This food style may contribute to the occurrence of Down syndrome, which has proved to be linked to abnormal folate and homocysteine metabolisms.

Aim of the study: To preliminary evaluate the levels of folic acid, homocysteine and other relevant factors which are associated with folate metabolism, among Japanese women who had pregnancies affected by Down syndrome.

Methods: Blood samples from 31 women who had pregnancies affected by Down syndrome (DS) were obtained. 60 age-matched control blood samples were also obtained from mothers who had not experienced miscarriages or abnormal pregnancies (CONT). Plasma homocysteine and serum folic acid, vitamin B12, and B6 were measured and compared between DS and CONT. Furthermore, the frequency of MTHFR polymorphism (C677T) was also investigated.

Results: Plasma levels of homocysteine were significantly increased in DS mothers (p=0.004). In contrast, serum levels of folic acid were significantly decreased in DS mothers (p=0.0001). There were no significant differences in the vitamin B12 and B6 levels between DS and CONT. Also, the frequency of 5-10 methylenetetrahydrofolate reductase gene (MTHFR) homozygous polymorphism showed no differences between DS and CONT.

Conclusion: Different levels of serum folic acid and plasma homocysteine between both groups may suggest the difference of food style may contribute the occurrence of Down syndrome even in Japan. Although there was no significant difference in the frequency of MTHFR polymorphism between the groups, probably because of inadequate number of samples, further studies may contribute to the understanding of the occurrence of Down syndrome in Japan.

Key words: folic acid, Down syndrome, homocysteine, 5-10 methylenetetrahydrofolate reductase gene (MTHFR)

НАРУШЕНИЯ МЕТАБОЛИЗМА ГОМОЦИСТЕИНА, ЗАВИСИМОГО ОТ ФОЛИЕВОЙ КИСЛОТЫ КАК РЕАЛЬНЫЙ ФАКТОР РИСКА СИНДРОМА ДАУНА В ЯПОНИИ

Нобору Такамура, Тацуро Кондо, Сиохи Ооги, Кокиси Арисава, Марико Мине,

Суници Ямасита, Киоси Аояги

Кафедра общественного здравоохранения, кафедра педиатрии, кафедра молекулярной эпидемиологии, отдел биостатистики, кафедра молекулярной

медицины,

Высшая школа биомедицины, Нагасаки, Япония

Обоснование: Япония считается «регионом с достатком фолатов», поскольку традиционная японская пища содержит достаточное количество фолиевой кислоты. Однако наметившаяся в последнее время «европеизация» образа питания молодых японских матерей способна повлиять на их потребление фолиевой кислоты. Это может вызвать увеличение частоты синдрома Дауна, поскольку доказано, что подобное питание связано с патологией метаболизма фолатов и гомоцистеина.

Целью настоящего исследования является первоначальная оценка уровня фолиевой кислоты, гомоцистеина и других факторов, связанных с фолатовым метаболизмом у японских женщин, чья беременность была отягощена синдромом Дауна.

Методы исследования: Были получены образцы крови от 31 женщины, чья беременность была отягощена синдромом Дауна (СД). Также были получены 60 образцов крови от женщин соответствующего возраста, чья беременность не закончилась выкидышем или патологией беременности. Было проведено измерение и сравнение гомоцистеина плазмы и сывороточной фолиевой кислоты, витамина Bi2 и Вб также была изучена частота полиморфизма MTHFR (С677Т).

Результаты: Уровень гомоцистеина плазмы был значительно увеличен у матерей из группы СД (р=0.004). Напротив, уровень фолиевой кислоты сыворотки в данной группе матерей был существенно снижен (р=0.0001). Значимой разницы в количестве витаминов Bi2 и В6 между двумя группами выявлено не было. Частота гомозиготного полиморфизма гена редуктазы 5-10 метилентетрагидрофолата не выявила различий между двумя группами.

Заключение: Разный уровень сыворотки в фолиевой кислоте и плазме гомоцистеина в двух группах позволяет предположить, что изменения в образе питания может вызвать увеличение частоты синдрома Дауна даже в Японии. Хотя значимых различий в частоте полиморфизма MTHFR между двумя группами выявлено не было, возможно, ввиду недостаточного количества образцов, дальнейшие исследования могут внести ясность в понимание частоты синдрома Дауна в Японии.

Ключевые слова: фолиевая кислота, синдром Дауна, гомоцистеин, ген 5-10 метилентетрагидрофолата редуктазы (MTHFR).

Introduction

Down syndrome (DS [MIM 190685]) is the most common genetic cause of human mental retardation, with an incidence of approximately 1 in 600 to 1000 live births [1]. It is estimated that 1 in 150 conceptions have trisomy 21 and that 80% of these are lost during early pregnancy. It has long been recognized that the risk of having a child with trisomy 21 increases with maternal age [2]. For example, the risk of having a livebom with Down syndrome at a maternal age of 30 is 1 in 1000 and at a maternal age of 40 is 9 in 1000 [3, 4].

Down syndrome has also been linked to the abnormal metabolism of homocysteine, which is led by variants in a critical folate metabolizing enzyme [5]. A thermolabile variant of 5-10 methylenetetrahydrofolate reductase (MTHFR) has been described in which a cytosine-to-thymine nucleotide (C677T) occurs, causing relatively reduced enzyme activity [6].

Although several studies have been performed on the causal relationship between homocysteine metabolism and Down syndrome in Western countries, there is scant data on folate metabolism in Japan, since Japan has been considered to be a «folate sufficient area» [7]. However, recent westernized food choices among young mothers may affect the incidence of Down syndrome in Japan.

In this study, we focused on the levels of homocysteine, folic acid and other relevant factors which contribute to homocysteine metabolism among mothers of Down syndrome children in Japan.

Key words: Down Syndrome, Folik Acid, homocysteine, polymorphism, 5-10 methylenetetrahydrofolate reductase (MTHFR)

Materials and Methods Study participants Before the study, ethical approval was obtained from the special committee of the Nagasaki University School of Medicine (project registration no. 0207160007). Blood samples from 31 women who had a pregnancy affected by Down syndrome (DS) were obtained at Nagasaki University Hospital. Mean age was 42.9 years. Blood samples of control mothers (CONT), who had experienced no miscarriages or abnormal pregnancies, were obtained at several clinics in Nagasaki city (see Acknowledgement). In all cases, informed consents were obtained. There was no significant difference between each group's mean age (42.9±8.3 and 42.9±9.5 years old, respectively, p=0.74).

Hematochemical analyses Plasma homocysteine, serum folate and vitamins В12 and B6 were measured in mothers of children with Down syndrome and in control mothers. Plasma homocysteine and serum vitamin B6 were measured using high performance liquid chromatography (HPLC). Serum folic acid and vitamin В12 were measured using the chemiluminescent immunoassay radioimmunoassay (CLIA) method. Genotype analyses Genomic DNA was extracted from blood cells using a QIAamp DNA blood mini kit (Qiagen, Tokyo, Japan). The MTHFR was amplified by polymerase chain reaction using the following set of primers: sense: 5' — CTG GGA AGA ACT CAG CGAAC — 3' antisense: 5' — GGA AGG TGC AAG АТС AGA GC — 3'.

The presence of the C677T mutation within the MTHFR gene creates a Hinfl restriction site that is detected by the appearance of226 and 165 base pair fragments on 2% agarose gel. Statistical Analysis Results of continuous data (e.g., folic acid, homocysteine, and vitamins В12 and B6) are expressed as means ± SD. Comparisons between groups were evaluated with the Mann Whitney test. For

count data (e.g., the number of individuals with each genotype), comparisons of percentage between groups were evaluated with the y2 test.

Results and Discussion

Plasma levels of homocysteine were significantly increased in mothers of DS children compared with the control group (8.98±2.09 vs. 7.80±1.77, p=0.004). Serum levels of folic acid were significantly decreased in mothers of DS children (6.43±1.38 vs. 9.88±3.60, p=0.0001). In contrast, there were no significant differences in vitamin B12 and B6 levels between DS and CONT groups (data not shown). Furthermore, analysis of the frequency of MTHFR homozygous polymorphism showed no differences between DS and CONT (10.6% vs 7.1%, p=0.85), probably due to an inadequate number of samples.

In this study, we demonstrated the possibility that a difference in mothers diets may contribute to the occurrence of Down syndrome in Japan. Less attention has been paid to the importance of folate intake during pregnancy in this country since Japan has been considered to be a «folate rich country» based upon its traditional food style that includes abundant vegetables and rice. On the other hand, it has been noted that food choice among young Japanese has been rapidly westernized [8]. Already in the United States, the Food and Drug Administration (FDA) recommended the addition of folic acid to enriched grain products in 1986 and made compliance mandatory by 1998 [9]. In the future, such an interventional policy to reduce the occurrence of Down syndrome may also be needed in Japan.

Our preliminary study suggests the importance of folate intake among Japanese reproductive-aged women. Further molecular epidemiological studies, as well as nutritional education to emphasize the importance of folate intake, may be beneficial for the effective prevention of Down syndrome in Japan.

Acknowledgement

We appreciate the study participants and the three clinics in Nagasaki (Yamashita clinic, Oshibuchi clinic and Takamura clinic) that collected control blood samples.

LITERATURE

1. Smith G, Berg J (1976). Down's anomaly. 2d ed. Churchill Livingstone, Edinburgh and New York.

2. Penrose L.S (1933). The relative effects of paternal and maternal age in mongolism. J. Genet 27: 219.

3. HookE.G (1982). Epidemiology of Down syndrome. In: Pueschel S.M; Rynders J.E (ed) Down Syndrome. Advances in Biomedicine and the Behavioral Sciences. Ware Press, Cambridge.

4. Hook E.B, Cross P.K, Schreinemachers D.M. (1983). Chromosomal abnormality rates at amniocentesis and in live-bom infants. JAMA 249: 2034—2038.

5. James S.J., Pogribna M., Pogribny I.P., Melnyk S., Hine R.J., Gibson J.B., Yi P., Tafoya D.L., Swenson D.H., Wilson V.L., Gaylor D.W. (1999).

Abnormal folate metabolism and mutation in the methylenetetrahydrofolate reductase gene may be maternal risk factors for Down syndrome. Am J Clin Nutr 70:495—501.

6. Botto L.D., Yang Q. (2000). 5,10-Methylenetetrahydrofolate reductase gene variants and congenital anomalies: a HuGE review. Am J Epidemiol. 151: 862—77.

7. The National Nutrition Survey in Japan (2001) Ministry of Health, Labour and Welfare, Japan, pp 1—244 (Japanese).

8. Couch S.C., Cross A.T., KidaK., RosE., Plaza I., Shea S., Deckelbaum R. (2000). Rapid westernization of children's blood cholesterol in 3 countries: evidence for nutrient-gene interactions? Am J Clin Nutr. 72: 12668—12748.

9. Honein M.A. Paulozzi L.J., Mathews T.J., Erickson J.D., WongL.Y. (2001). Impact of folic acid fortification of the US food supply on the occurrence of neural tube defects. JAMA 285:2981—6.

УДК 617-089: 616.089: 614.88

СИМУЛЬТАННЫЕ ОПЕРАЦИИ У БОЛЬНЫХ ОСТРЫМ ХОЛЕЦИСТИТОМ

Б. Б. Осипов

Гомельская городская клиническая больница скорой медицинской помощи

Обобщен опыт выполнения симультанных операций во время лапароскопической холецистэктомии у больных острым холециститом в Гомельской городской клинической больнице скорой медицинской помощи за период с 1995 по 2003 год. На основании анализа результатов 145 сочетанных вмешательств, изучения изменений индекса APACHE II в группах симультанных операций и лапароскопических холецистэктомий, а также предшествующих исследований по микробиологическим аспектам острого холецистита представлено обоснование возможности выполнения симультанных операций у больных острым холециститом.

Ключевые слова: симультанная операция, лапароскопическая холецистэктомия, острый холецистит.

SIMULTANEOUS OPERATIONS FOR PATIENTS WITH ACUTE

CHOLECYSTITIS

B.B. Osipov

The experience of performance simultaneous operations during laparoscopic cholecystectomy for patients with acute cholecystitis in Gomel emergency hospital between 1995 and 2003 is summarized. Grounded on analysis of outcomes of 145 operations, the analyses of changes of an index APACHE II in groups simultaneous operations and laparoscopic cholecystectomy precursor researches on microbiological aspects of an acute cholecystitis the indications and contraindications to performance of similar interferences Eire formulated.

Key words: simultaneous operation, laparoscopic cholecystectomy, acute cholecystitis.