Effect of arteriovenous flow reversal on blood flow and metabolism in a skin flap

Twelve pig buttock island flaps (10 X 10 cm) were studied for 6 hours after arteriovenous flow reversal at the level of the pedicle. Follow-up was 48 hours. Blood pressure, Po2, pH, and lactate were measured in flap arteries and veins. Oxygen consumption was calculated. Data indicated true flow reversal. Blood pressure and Po2 in flap veins increased to systemic arterial levels. Outflow was provided by the arterial system, demonstrating venous pressure and Po2 values. Lactate increased significantly (1.8 +/- 0.5 to 4.0 +/- 2.3 mmol/liter), while pH dropped from 7.43 +/- 0.03 to 7.11 +/- 0.02. Oxygen consumption remained below baseline. In four flaps thrombosis occurred within 6 hours; no flap survived 48 hours. The results of this study do not encourage clinical application of the concept of flow reversal.     

Linkage relationships and gene order around the locus for X-linked retinoschisis.

X-linked recessive retinoschisis (RS) is a hereditary disorder with variable clinical features. The main symptoms are poor sight; radial, cystic macula degeneration; and peripheral superficial retinal detachment. The disease is quite common in Finland, where at least 300 hemizygous males have been diagnosed. We used nine polymorphic DNA markers to study the localization of RS on the short arm of the X chromosome in 31 families comprising 88 affected persons. Two-point linkage results confirmed close linkage of the RS gene to the marker loci DXS43, DXS16, DXS207, and DXS41 and also revealed close linkage to the marker loci DXS197 and DXS9. Only one recombination was observed between DXS43 and RS in 59 informative meioses, giving a maximum lod score of 13.87 at the recombination fraction .02. No recombinations were observed between the RS locus and DXS9 and DXS197 (lods between 3 and 4), but at neither locus was the number of informative meioses sufficient to provide reliable estimates of recombination fractions. The most likely gene order on the basis of multilocus analysis was Xpter-DXS85-(DXS207,DXS43)-RS-DXS41-DXS 164-Xcen. Because multilocus linkage analysis indicated that the most probable location of RS is proximal to DXS207 and DXS43 and distal to DXS41, these three flanking markers are the closest and most informative markers currently available for carrier detection.     

Dielectric properties of human blood and erythrocytes at radio frequencies (0.2–10 MHz); dependence on cell volume fraction and medium composition

The dielectric properties of human erythrocytes (red blood cells) suspended in whole blood and in isotonic media at various volume fractions (haematocrits) have been studied in the frequency range 0.2–10 MHz, in which the so-called-dispersion due to the Maxwell-Wagner effect is known to occur. The capacitance and conductance at 25 °C were measured by an instrument interfaced to a computer. The rectangular sample cavity (1 ml volume) contained four pure gold electrode pins, and the sample could be circulated by a roller pump. The frequency-dependence of the permittivity and conductivity were fitted by non-linear least squares regression. Corrections were applied for non-linearity in the dielectric increment at high haematocrit, and for electrode polarisation when diluting the blood in saline. Data were interpreted in terms of a simple equivalent resistor-capacitor circuit. From the measured haematological values the specific membrane capacitance (C_m) and the conductivities internal and external to the cells (_i and _o respectively) were estimated. The conductivities behaved in a predictable manner with a mean of 0.458 S · m^–1 (s.d. ± 0.044) for _i, whereas the value of C_m (and indeed the actual capacitance of the suspension) was dependent on the amount of plasma present. Hence, in stationary normal (anticoagulated) whole blood samples, C_m was as high as 2.98 F · cm^–2 (s.d. ± 0.40), in contrast to about 0.9 F · cm^–2 in blood diluted more than two-fold (to less than 20% hct) in isotonic media. The high value remained when the diluent was plasma. The C_m value returned to a high value when washed erythrocytes were reconstituted with plasma, provided that this was present at above a critical or threshold concentration of about 30 vol % in the medium, irrespective of the haematocrit in the range studied (15–44%). The C_m remained low in serum. When added to washed cells in saline, purified fibrinogen had no effect. However, high C_m values were obtained by fibrinogen supplementation to serum and diluted plasma. Applying moderate flow to whole blood approximately halved its high C_m value in an exponential manner with flow rate, whilst the C_m of washed cells (31–67% hct) slightly increased, and converged to the value for whole blood under flow. We interpret the highapparent C_m value in stationary samples to be a result of rapid cell aggregation in the presence of plasma, where rouleaux formation takes place before visible sedimentation sets in.     

Phylogeny of theCyclanthaceae

The affinities ofCyclanthaceae are discussed, and it is concluded that the sister-group to this family is most probablyPandanaceae. A hypothesis of generic relationships inCyclanthaceae, based on cladistic methods, is presented. Bootstrap analysis and Bremer support (decay index) have been used to test the strength of individual clades, and the result is compared with previously made phylogenetic analyses. TheSphaeradenia group (Chorigyne, Stelestylis, Sphaeradenia, andLudovia) is supported as monophyletic and acceptably resolved, while theAsplundia group (remaining genera inCarludovicoideae) may be paraphyletic, with largely uncertain relationships. A formal recognition of these groups is therefore not justified. The probable character evolution inCarludovicoideae is discussed.     

Ribonucleic acid labelling perfused livers of protein-fed and protein-deprived rats

Several observations suggest an increased RNA synthesis in livers of protein-deprived rats, though the RNA/DNA ratio is decreased. A number of hormones may be involved in these changes. Therefore, we studied in RNA metabolism in isolated perfused livers taken from protein-fed and protein-deprived rats. (3H)-orotic acid was given to the rats 2 h before liver explantation, and (14C)-orotic acid was added to the perfusate. Other rats, called controls in vivo, whose livers were not transplanted were also given (3H)-orotic acid followed by (14C)-orotic acid. The livers of these rats, which were not hormone supplemented, were labelled for the same length of times as the livers in vitro. The ratio specific RNA radioactivity/specific nucleotide radioactivity x RNA/DNA was determined and taken as a measure of the RNA synthesis per liver cell. In the controls in vivo, this ratio was significantly higher for protein-deprived than for protein-fed rats. In livers from the protein-fed rats, labelling in vitro increased significantly when growth hormone, hydrocortisone, insulin and tri-iodothyronine were added to the perfusate. Labelling was also significantly higher in these livers than in the controls in vivo. In livers from protein-deprived rats, the ratio in question was the same whether the hormones were added to the perfusate or not, and was significantly lower than in the controls in vivo. Differences in RNA labelling are thus obtained in our in vitro system. Gel electrophoresis of RNA demonstrated normal RNA labelling, showing that the system is suitable for studying liver RNA synthesis. Further refinement can be made by studying the labelling of UTP and CTP. The results might suggest that the liver from a protein-fed rat, explanted in vitro, may increase its RNA synthesis under the influence of the four hormones in question, and that the RNA synthesis of the liver of a protein-deprived rat is high in-vivo and that it might decrease, when it is explanted to in vitro conditions.     

Methylation changes in the human IGF2 p3 promoter parallel IGF2 expression in the primary tumor, established cell line, and xenograft of a human hepatoblastoma

Hepatoblastoma (HB) is a rare malignant embryonal liver tumor. Its pathogenesis has been associated with altered regulation of the IGF2 and H19 genes, and previous studies have suggested a correlation between abnormal methylation and altered expression of these genes in hepatoblastoma. Upregulation of the activity of the IGF2 promoter P3 has previously been shown to be tightly correlated with demethylation in hepatoblastoma. Here, we have used bisulfite genomic sequencing to characterize the methylation pattern of the IGF2 promoter P3 in the hepatoblastoma-derived cell line Hep T1, in the original tumor from which Hep T1 is derived, and in nude mouse xenografts of the Hep T1 cell line. The results show a clear difference in methylation pattern of the most proximal region of the IGF2 P3 promoter between the primary tumor, the cell line, and the xenografts. RNase protection and mRNA in situ hybridization revealed that variations in methylation patterns was paralleled by the levels of IGF2 P3 mRNA, which was detectable in the primary tumor and xenografts, but not in the cell line. Furthermore, it was demonstrated that H19 was reactivated and demethylated in the HepT1 cell line by 5-azaCytidine, in contrast to IGF2 P3, which was not demethylated or reactivated. We suggest that methylation of the proximal IGF2 P3 is important for its regulation.     

Species pools in cultural landscapes – niche construction,ecological opportunity and niche shifts

This paper discusses the ecology of species that were favoured by the development of the cultural landscape in central and NW Europe beginning in the Neolithic and the Bronze Age, with a focus on mechanisms behind species responses to this landscape transformation. A fraction of species may have maintained their realized niches from the pre‐ agricultural landscape and utilized similar niches created by the landscape transformation. However, I suggest that many species responded by altering their niche relationships, and a conceptual model is proposed for this response, based on niche construction, ecological opportunity and niche shifts. Human‐mediated niche construction, associated with clearing of forests and creation of pastures and fields promoted niche shifts towards open habitats, and species exploited the ecological opportunity provided by these created environments. This process was initially purely ecological, i.e. the new habitats must have been included in the original fundamental niche of the species. Two other features of human‐mediated niche construction, increased interconnectivity and increased spatial stability of open habitats, resulted in species accumulating in the habitats of the constructed landscape. As a consequence, selection processes were initiated favouring traits promoting fitness in the constructed landscape. This process implied a feed‐back to niche shifts, but now also including evolutionary changes in fundamental niches. I briefly discuss whether this model can be applied also to present‐day anthropogenic impact on landscapes. A general conclusion is that ecological and evolutionary changes in species niches should be more explicitly considered in modeling and predictions of species response to present‐day landscape and land‐use changes.     

Hepatocellular uptake of 3H-dihydromicrocystin-LR, a cyclic peptide toxin

The cellular uptake of microcystin-LR, a cyclic heptapeptide hepatotoxin from the cyanobacterium Microcystis aeruginosa, was studied by means of a radiolabelled derivative of the toxin. 3H-dihydromicrocystin-LR. The uptake of 3H-dihydromicrocystin-LR was shown to be specific for freshly isolated rat hepatocytes whereas the uptake in the human hepatocarcinoma cell line Hep G2 as well as the mouse fibroblast cell line NIH-3T3, and the human neuroblastoma cell line SH-SY5Y, was negligible. By means of a surface barostat technique it was shown that the membrane penetrating capacity (surface activity) of microcystin-LR was low, indicating that the toxin requires an active uptake mechanism. The hepatocellular uptake of microcystin-LR could be inhibited in the presence of bile acid transport inhibitors such as antamanide (5 microM), sulfobromophthalein (50 microM) and rifampicin (30 microM). The uptake was also reduced in a concentration dependent manner when the hepatocytes were incubated in the presence the bile salts cholate and taurocholate. A complete inhibition of the hepatocellular uptake was achieved by 100 microM of either bile salt. The overall results indicate that the uptake of microcystin-LR is through the multispecific transport system for bile acids. This mechanism of cell entry would explain the previously observed cell specificity and organotropism of microcystin-LR.