Stroke research in GenomEUtwin.

Stroke is one of the leading causes of severe disability and death in the world. In the present article we outline possibilities and limitations for future stroke research within the GenomEUtwin. The combined sample of twins born before 1958 from Denmark, Finland, and Sweden, and available for follow-up into the second millennium for non-fatal and fatal stroke events through national inpatient and death registers exceeds 70,000 twin pairs. This sample size will enable the study of genetic influences on stroke and major stroke subtypes. Large samples of twins in GenomEUtwin have been followed up repeatedly through interviews and questionnaires concerning a variety of exposures and potential risk factors for stroke. We briefly outline how this information can be combined with the health register information for epidemiologic and genetic epidemiologic studies of stroke. We also present the number of twin pairs concordant and discordant for stroke in Denmark, Finland and Sweden, and time lags between events for twins concordant for stroke. This information illustrates that the number of affected sib pairs for linkage studies is relatively limited, but the sample sizes are promising for association studies.     

Hydrocortisone regulation of hyaluronan metabolism in human skin organ culture

We studied the influence of hydrocortisone (HC) on hyaluronan (HA) metabolism in explants of human skin, a model retaining normal three-dimensional architecture of dermal connective tissue and dynamic growth and stratification of epidermal keratinocytes. The synthesis of hyaluronan and proteoglycans (PGs), and DNA, were determined with ³H-glucosamine and ³H-thymidine labelings, respectively. The total content and histological distribution of hyaluronan was studied utilizing a biotinylated aggrecan-link protein complex. A low concentration of HC (10^?9 M) stimulated the incorporation of ³H-glucosamine into hyaluronan in epidermis by 23% and reduced the disappearance rate of hyaluronan by 25% in chase experiments, resulting in a 74% increase in total hyaluronan (per epidermal dry weight) after a 5-day culture in 10^?9 M HC. On the other hand, a high concentration of HC (10^?5 M) reduced both synthesis (-42%) and degradation (-46%) of epidermal hyaluronan during 24 h labeling and chase periods. The cumulative effect of a 5-day treatment was a 24% decrease of total epidermal hyaluronan. The high dose (10^?5 M) also reduced keratinocyte DNA synthesis and epidermal thickness. In dermis, only the high (10^?5 M) concentration of HC was effective, inhibiting the incorporation of ³H-glucosamine into hyaluronan by 28%. No significant influences on total hyaluronan content or the disappearance rate of hyaluronan in dermal tissue was found. All HC concentrations lacked significant effects on newly synthesized PGs in epidermal and dermal tissues, but reduced the labeled PGs diffusing into culture medium. A low physiological concentration of HC thus maintains active synthesis and high concentration of hyaluronan in epidermal tissue, while high pharmacological doses of HC slows hyaluronan turnover and reduces its content in epidermis, an effect correlated with enhanced terminal differentiation, reduced proliferation rate and reduced number of vital keratinocyte layers. © 1995 Wiley-Liss, Inc.     

Determination of cis,cis-methylene interrupted polyunsaturated fatty acids in aqueous solutions by lipoxygenase chemiluminescence

The chemiluminescent reaction of luminol during lipoxygenase-catalyzed oxygenations was studied with the purpose of developing a specific luminometric assay for cis,cis-1,4-pentadiene fatty acids directly in aqueous solutions. The addition of picomole levels of either linoleic or arachidonic acids to reaction systems containing 0.04 mM luminol and 40 micrograms/ml of purified soybean lipoxygenase-1 gave light emission curves with a single sharp maximum. Under these conditions the peak heights were linearly dependent on the fatty acid concentration and the detection limit for both of the fatty acids was 2 pmol with a signal to noise ratio of 2. For maximum reproducibility of the assays a procedure for the proper quantitation of the enzyme was developed. The fact that the assay proved to be relatively interference-free was ascribed to the high molar enzyme/substrate ratio (above 1).     

Effect of prolonged ethanol ingestion on hepatic lipogenesis and related enzyme activities

1. Hepatic lipogenesis in vivo and the activities of enzymes associated with fatty acid synthesis in the liver were studied in rats fed for 21 days on liquid diets containing ethanol. 2. The ethanol-fed rats developed a moderate hepatic triacylglycerol accumulation during this period. When carbohydrate was replaced by ethanol in the diet, the rate of fatty acid synthesis was slower in the ethanol-fed rats on low-, medium- and high-fat diets than in the appropriate controls. However, when the fat/carbohydrate ratio was kept the same in the ethanol-fed and control rats, ethanol had no influence on the rate of fatty acid synthesis. 3. Glucose 6-phosphate dehydrogenase activity was lower in the ethanol-fed group. ;Malic' enzyme activity did not change during the ethanol treatment when the fat/carbohydrate ratio was kept unchanged. 4. The ATP citrate lyase activity was lower in the ethanol-fed rats on all diets, whereas acetyl-CoA synthetase activity was independent of the composition of the control diet, but was lower in the ethanol-fed rats, in which the concentration of the active form of pyruvate dehydrogenase was also lower. 5. It is concluded that hepatic fatty acid synthesis does not play any major role in ethanol-induced triacylglycerol accumulation. Careful design of the diets is necessary to reveal the specific effects of ethanol on the enzymes associated with lipogenesis.     

Clozapine-induced agranulocytosis

Summary An epidemic of agranulocytosis and granulocytopenia occurred in 1975 in conjunction with clozapine treatment of mental patients in Finland. An attempt was made to assess the epidemiologic and genetic factors contributing to the adverse drug effect. The estimated incidence rate in Finland was 2.1/1000 patient-months. This figure could not be compared with rates from other countries because of the inexact nature of the figures reported so far. All 16 cases occurred in seven hospitals in southwestern Finland, whereas the overall hospital net use of the drug was geographically evenly distributed. The difference between the observed and the proportionally expected incidence of cases amongst the hospitals where clozapine was used was statistically significant. The average consumption of the drug did not differ between the hospitals where cases occurred and those where no definite cases could be diagnosed. Six-generation pedigree analyses failed to reveal significant parental consanguinity or genetic kinship between probands. Neither did the birth places of the ancestors of the probands disclose a typical isolate pattern. In conclusion, the cases appeared to be confined to a few hospitals in southwestern Finland. Although a genetic factor is not excluded, we found no evidence in support of a genetic mechanism.     

Practical identifiability of growth and substrate consumption models

The estimation of parameters in several dynamic models, which describe growth and substrate consumption, has been carried out using a modified Gauss-Newton-type method. The four models considered are Monod, Contois, linear specific growth rate, and an enzyme kinetic model. The initial values of the differential equations are included in the parameter vector which will be estimated. The efficiency of the method and the confidence limits of the parameters were studied using simulated measurement noise. The experimental results describe Trichoderma viride growing on glucose as the main carbon source.     

Correlations between hyaluronan and epidermal proliferation as studied by [H]glucosamine and [H]thymidine incorporations and staining of hyaluronan on mitotic keratinocytes

The rates of keratinocyte proliferation and synthesis of Hyaluronan (HA) were studied in human whole-skin organ culture by labeling with [6-³H]glucosamine and [³H]thymidine, respectively, to reveal possible correlations between the two functions of the cell. HA distribution in epidermis was examined by staining with a specific probe prepared front cartilage proteoglycan. The keratinocyte proliferation rate was low on the first 2 culture days, but showed a tenfold increase on the third and fourth days while the synthesis of HA proceeded at a relatively stable level throughout the same period. The most intensive staining of HA occurred in the uppermost spinous cell layer, whereas mitotic cells resided in the basal and suprabasal layers. The keratinocytes under various stages of mitosis were surrounded by a HA staining not more intense than that around nondividing basal cells, but a thick pad of HA appeared rapidly between the daughter cells. These findings suggest that newly synthesized HA is associated with the separation of keratinocytes following mitosis but the majority of the synthesis and content of HA in epidermis is involved in other keratinocyte activities such as maintenance of the extracellular space and cell-cell interactions during migration and differentiation.     

Calbindin D-28k-immunoreactivity in rat muscle spindles during postnatal maturation and after denervation

Summary Calbindin D-28k-immunoreactivity has been demonstrated in some of the intrafusal muscle fibres and in the capsule of adult rat muscle spindles. In this study, the immunocytochemical localization of calbindin D-28k in the muscle spindles of triceps surae muscle was studied during postnatal maturation and after denervation. In young rats calbindin D-28k-immunoreactivity was seen in a few intrafusal fibres, first at the age of 4 days. At the 7th day, three calbindin D-28k-immunoreactive fibres and one unlabelled fibre were seen in most muscle spindles, as in adult rats. The spindle capsule and perineurial sheath of nerves were first seen to exhibit calbindin D-28k immunoreactivity at the age of 14 days, and thereafter the localization of calbinding D-28k-like immunoreactivity was similar to that in adult rats. After denervation, calbindin D-28k-immunoreactivity remained in intrafusal muscle fibres and the spindle capsule for a long period. After two months of denervation, calbindin D-28k immunoreactivity could still be seen in the spindle capsule, but the intrafusal fibres were not labelled.The innervation is known to have trophic effects on the intrafusal fibres. The present findings suggest that the expression of calbindin D-28k-immunoreactivity in maturating muscle spindles may be induced by the developing innervation. The decrease of calbindin D-28k-immunoreactivity in intrafusal fibres after denervation may be due to the loss of trophic factors released by the nerves.