Only amyloidogenic intermediates of transthyretin induce apoptosis

In diseases like Alzheimer's disease and familial amyloidotic polyneuropathy (FAP) amyloid deposits co-localize with areas of neurodegeneration. FAP is associated with mutations of the plasma protein transthyretin (TTR). We can here show an apoptotic effect of amyloidogenic mutants of TTR on a human neuroblastoma cell line. Toxicity could be blocked by catalase indicating a free oxygen radical dependent mechanism. The toxic effect was dependent on the state of aggregation and unexpectedly mature fibrils from FAP-patients who failed to exert an apoptotic response. Morphological studies revealed a correlation between toxicity and the presence of immature amyloid. Thus, we can show that toxicity is associated with early stages of fibril formation and propose that mature full-length fibrils represent an inert end stage, which might serve as a rescue mechanism.     

Macromolecule absorption and cortisol secretion in newborn calves derived from in vitro produced embryos

Earlier reports indicate that calves derived from in vitro produced (IVP) embryos are more susceptible to neonatal disease than calves produced after artificial insemination (AI) or natural mating. The aims of the present study were to investigate whether calves born after IVP embryos show an altered macromolecule absorption (immunoglobulin G (IgG) and porcine serum albumin (PSA)) compared with AI calves and whether the macromolecule absorption could be related to the degree of acidosis or to the cortisol secretion around birth. Hence, IgG and PSA absorption in control AI calves (n=7) was compared with that in two groups of IVP calves (IVP-defined: SOFaa embryo culture with polyvinyl alcohol, n=6; IVP-serum: SOFaa embryo culture with serum and co-culture, n=8). The calves were fed colostrum (40ml/kg) at 2, 6 and 12h after birth. At 24h after birth, both AI and IVP calves had achieved a level of plasma IgG sufficient to provide passive immunization (>15mg/ml). When the values were adjusted for the varying colostral IgG contents and the degree of acidosis, the IVP-defined calves had significantly lower peak plasma IgG concentrations than the AI calves at 18-24h after birth (P<0.04). However, when the macromolecule marker (PSA), was fed to all calves at 2 and 12h after birth the resulting plasma PSA levels were significantly lower in the AI calves compared with the IVP calves during the whole observation period (P<0.0001). Calves with a moderate neonatal acidosis (mean pH<7.2 during the first 30min after birth) had reduced peak plasma IgG concentration at 18-24h after birth (P<0.02) compared to calves without acidosis. The basal and ACTH-stimulated cortisol levels were lower in the newborn IVP-defined calves than in the AI calves (P<0.05) and the IVP-serum calves (P<0.002). Cortisol levels shortly after birth correlated positively with birth weight (r=0.60, P<0.0001) and with gestation length (r=0.34, P<0.04). Since, the IVP calves absorbed sufficient amounts of IgG from colostrum to acquire sufficient passive immunity, we conclude that the lower viability described in IVP offspring probably is not caused by an impaired passive immunization. IVP-defined calves had significantly lower absorption efficiency of IgG compared with AI calves, whereas absorption of a non-Ig macromolecule (PSA) was higher for IVP than AI calves. This might indicate a more selective absorption in AI calves in favor of IgG. Acidosis around birth affected immunoglobulin absorption negatively. IVP-defined calves had significantly lower cortisol levels the first 3h after birth and during an ACTH-challenge and a lower IgG absorption efficiency, which might indicate a mild degree of organ dysmaturity in these calves.     

Physical activity, cardiorespiratory fitness, and the metabolic syndrome in youth.

The metabolic syndrome is defined as the coexistence of multiple cardiovascular and diabetes risk factors, the prevalence of which has increased dramatically in adult populations in the last decades. More recently, the same cluster of metabolic risk factors has also been recognized in children and adolescents. Epidemiological evidence suggests that high levels of cardiorespiratory fitness (CRF) and physical activity are associated with a favorable metabolic risk profile in adults. However, in youth the role of these factors is less clear. Therefore, the purpose of this mini-review is to examine the recent evidence between objectively measured habitual physical activity and CRF with clustered metabolic risk in youth. In general, it appears that both physical activity and CRF are separately and independently associated with metabolic risk factors in youth, possibly through different causal pathways. Further research is necessary to quantify how much physical activity is needed to prevent the metabolic syndrome and the diseases with which it is associated. Public health approaches that encourage increased physical activity and reduce sedentary behaviors may prove useful in reducing the population burden associated with metabolic risk.     

A comparative analysis of the rainbow trout genome with 2 other species of fish (Arctic charr and Atlantic salmon) within the tetraploid derivative Salmonidae family (subfamily: Salmoninae).

We updated the genetic map of rainbow trout (Oncorhynchus mykiss) for 2 outcrossed mapping panels, and used this map to assess the putative chromosome structure and recombination rate differences among linkage groups. We then used the rainbow trout sex-specific maps to make comparisons with 2 other ancestrally polyploid species of salmonid fishes, Arctic charr (Salvelinus alpinus) and Atlantic salmon (Salmo salar) to identify homeologous chromosome affinities within each species and ascertain homologous chromosome relationships among the species. Salmonid fishes exhibit a wide range of sex-specific differences in recombination rate, with some species having the largest differences for any vertebrate species studied to date. Our current estimate of female:male recombination rates in rainbow trout is 4.31:1. Chromosome structure and (or) size is associated with recombination rate differences between the sexes in rainbow trout. Linkage groups derived from presumptive acrocentric type chromosomes were observed to have much lower sex-specific differences in recombination rate than metacentric type linkage groups. Arctic charr is karyotypically the least derived species (i.e., possessing a high number of acrocentric chromosomes) and Atlantic salmon is the most derived (i.e., possessing a number of whole-arm fusions). Atlantic salmon have the largest female:male recombination ratio difference (i.e., 16.81:1) compared with rainbow trout, and Arctic charr (1.69:1). Comparisons of recombination rates between homologous segments of linkage groups among species indicated that when significant experiment-wise differences were detected (7/24 tests), recombination rates were generally higher in the species with a less-derived chromosome structure (6/7 significant comparisons). Greater similarity in linkage group syntenies were observed between Atlantic salmon and rainbow trout, suggesting their closer phylogenetic affinities, and most interspecific linkage group comparisons support a model that suggests whole chromosome arm translocations have occurred in the evolution of this group. However, some possible exceptions were detected and these findings are discussed in relation to their influence on segregation distortion patterns. We also report unusual meiotic segregation patterns in a female parent involving the duplicated (homeologous) linkage group pair 12/16 and discuss several models that may account for these patterns.     

Effects of microgravity on organ development of the neonatal rat.

We analyzed various organs in the same rats to study effects of gravitational condition on organ development of the neonatal rat in this study. Eight-day old and 14-day old Sprague-Dawley rats were flown for 16 days on the Space Shuttle Columbia (April 17-May 3, 1998). The organs were weighed and the ratio of the organ weight to the body weight (organ weight ratio; OBR) was calculated. Tissues were analyzed using anatomical, immunohistochemical and molecular biological technique. Six animals of the 8-day old group were reared on the ground for 30 more days after landing. The differences between flight and control rats in 8-day group were drastic. The lung, heart, kidney and adrenal glands in flight rats were significantly larger than that of control rats in OBR comparison. However, only the lung and kidney were still larger after 30 more days on ground. The kidney in flight rats performed pelvis expansion with down-regulation of aquaporin-2 expression confirmed by immunohistochemistry. The thymus, spleen, mesentery and pancreas were smaller in OBR. But the thymus in flight rats was heavier after 30 more days. The organs in flight rats which had no differences in OBR showed normal characteristics in histological analysis. We also found that the number of unmyelinated fibers of the aortic nerve in flight rats of 8-day group was smaller than that in control rats. In flight rats of the 14-day group, only the kidney was heavier and the ovary was lighter as compared to the controls. These results implied the second week of life was important for development during spaceflight. And the sensitivity and the critical period on neonatal development under microgravity might differ in each organ.     

Full-term development of nuclear transfer calves produced from open-pulled straw (OPS) vitrified cytoplasts: work in progress

Cryopreservation of cytoplasts would help to resolve the logistics of matching the availability of oocytes with embryo donors in nuclear transfer. Therefore, the developmental potential of nuclear transfer bovine embryos reconstructed using vitrified cytoplasts was investigated. In vitro matured oocytes were denuded, enucleated, activated with calcium ionophore (10 microM, 5 min) and cycloheximide (10 microg/mL, 6 h) and then vitrified by the open pulled straw (OPS) method. After immediate warming, the nuclear transfer embryos were reconstructed using blastomeres from nonvitrified,in vitro-produced embryo donors. Compared with control nuclear transfer embryos that were reconstructed using nonvitrified cytoplasts, fusion rates (% +/- SEM) were not affected (83.7+/-9.2 vs. 79.8+/-4.6; P>0.05), but cleavage (55.7+/-2.9 vs. 92.8+/-3.9; P = 0.0002) and blastocyst rates (7.2+/-5.0 vs. 32.6+/-7.8; P = 0.0025, vitrified vs. nonvitrified cytoplasts, respectively) per successful fusion were reduced. One nuclear transfer blastocyst reconstructed from a vitrified cytoplast was transferred to a synchronized recipient. After a normal length gestation (265 d), twin calves (21 and 26 kg) were delivered. Microsatellite analysis confirmed that the calves were homozygotic (the embryo split in utero), and were derived from the in vitro-produced embryo donor. The twins were dead at birth, but post-mortem analysis of the calves indicated no abnormalities or infections, suggesting that their death was related to the twin pregnancy and the known fragility of nuclear transfer calves. These data demonstrate that open pulled straw-vitrified cytoplasts are capable of supporting full-term development of nuclear transfer embryos.     

High bovine blastocyst development in a static in vitro production system using SOFaa medium supplemented with sodium citrate and myo-inositol with or without serum-proteins

We describe a bovine embryo culture system that supports repeatable high development in the presence of serum or BSA as well as under defined conditions in the absence of those components. In the first experiment, embryo development in SOF with amino acids (SOFaa), sodium citrate (SOFaac) and myo-inositol (SOFaaci) and with BSA or polyvinyl alcohol (PVA) was compared with that in a M199 granulosa cell co-culture (M199 co-culture). Subsequently, development and cell numbers of blastocysts cultured under defined conditions in SOFaaci with PVA (SOFaaci-PVA), or under undefined conditions in SOFaaci with 5% cow serum (SOFaaci-CS) or M199 co-culture were compared. The repeatability of culture results in SOFaaci-CS was checked by weekly replicates (n = 30) spread over 11 months. The viability of embryos developed in SOFaaci-PVA was estimated by transfer of morphologically good blastocysts (n = 10) to synchronized recipients. In the second experiment, the effect of omitting CS or BSA from IVM and IVM-IVF on subsequent embryo development in SOFaaci-PVA or in SOFaaci-CS was investigated. Blastocyst development in SOFaa-PVA, SOFaac-PVA, SOFaa-BSA and M199 was 16 +/- 3b, 23 +/- 2ab, 30 +/- 8a and 36 +/- 7a%, respectively (Pab < 0.05). Additional inclusion of myoinositol resulted in 42 +/- 1a% blastocysts in SOFaaci-PVA vs 19 +/- 3b% in SOFaac-PVA, 47 +/- 7a% in SOFaac-BSA, and 36 +/- 7a% in M199 co-culture, respectively (Pab < 0.01). In 30 replicates, the average cleavage and blastocyst rates of oocytes in SOFaaci-CS were 87 +/- 4 and 49 +/- 5%, respectively. Five normal calves were produced after transfer of 10 blastocysts developed in defined culture medium (i.e., SOFaaci-PVA). Defined IVM or IVM-IVF (i.e., in absence of CS and BSA) reduced cleavage rates (83 +/- 3 and 55 +/- 3% vs 90 +/- 1% in presence of CS; P < 0.01). Subsequent embryo development in SOFaaci-CS was not affected in either of these defined conditions. However, cleavage and blastocyst rates under completely defined IVP conditions were 54 +/- 7 and 19 +/- 4%, respectively. It was concluded that under defined culture conditions, addition of citrate and myo-inositol improved blastocyst development to rates comparable to those obtained with serum, BSA or co-culture and that the quality of blastocysts was not affected by the absence of serum or BSA. However, serum was essential during IVM/IVF for normal fertilization and subsequent high blastocyst development.     

Internal electron transfer between hemes and Cu(II) bound at cysteine beta93 promotes methemoglobin reduction by carbon monoxide

Previous studies showed that CO/H2O oxidation provides electrons to drive the reduction of oxidized hemoglobin (metHb). We report here that Cu(II) addition accelerates the rate of metHb beta chain reduction by CO by a factor of about 1000. A mechanism whereby electron transfer occurs via an internal pathway coupling CO/H2O oxidation to Fe(III) and Cu(II) reduction is suggested by the observation that the copper-induced rate enhancement is inhibited by blocking Cys-beta93 with N-ethylmaleimide. Furthermore, this internal electron-transfer pathway is more readily established at low Cu(II) concentrations in Hb Deer Lodge (beta2His --> Arg) and other species lacking His-beta2 than in Hb A0. This difference is consistent with preferential binding of Cu(II) in Hb A0 to a high affinity site involving His-beta2, which is ineffective in promoting electron exchange between Cu(II) and the beta heme iron. Effective electron transfer is thus affected by Hb type but is not governed by the R left arrow over right arrow T conformational equilibrium. The beta hemes in Cu(II)-metHb are reduced under CO at rates close to those observed for cytochrome c oxidase, where heme and copper are present together in the oxygen-binding site and where internal electron transfer also occurs.     

The Danish Twin Registry: 127 birth cohorts of twins.

The Danish Twin Registry is the oldest national twin register in the world, initiated in 1954 by ascertainment of twins born from 1870 to 1910. During a number of studies birth cohorts have been added to the register, and by the recent addition of birth cohorts from 1931 to 1952 the Registry now comprizes 127 birth cohorts of twins from 1870 to 1996, with a total of more than 65,000 twin pairs included. In all cohorts the ascertainment has been population-based and independent of the traits studied, although different procedures of ascertainment have been employed. In the oldest cohorts only twin pairs with both twins surviving to age 6 have been included while from 1931 all ascertained twins are included. The completeness of the ascertainment after adjustment for infant mortality is high, with approximately 90% ascertained up to 1968, and complete ascertainment of all liveborn twin pairs since 1968. The Danish Twin Registry is used as a source for large studies on genetic influence on aging and age-related health problems, normal variation in clinical parameters associated with the metabolic syndrome and cardiovascular diseases, and clinical studies of specific diseases. The combination of survey data with data obtained by linkage to national health related registers enables follow-up studies both of the general twin population and of twins from clinical studies.