Börje W. Karlsson

Diseases and environmental contaminants in seals from the Baltic and the Swedish west coast

Investigations have shown that Baltic grey seal (Halichoerus grypus) and ringed seal (Phoca hispida) suffer from a disease complex described as a primary lesion in the adrenals causing secondary reactions in various other organs. Studies on historical Baltic grey seal skull bone material show that the prevalence of affected animals started to increase after World War II. The disease complex explains the dramatic decrease in the Baltic grey and ringed seal population during the 1960s and 1970s and is believed to be caused by environmental pollutants. In 1988, about 60% of the harbor seal population (Phoca vitulina) along the Swedish west coast and in the southwestern part of the Baltic died in the PDV epizootic (Phocine Distemper Virus). Whether the course of the epizootic was altered by environmental pollutants is still an open question. Studies on historical harbor seal skull bone material from both the Baltic and the Swedish west coast show that the incidence of skull bone lesions has also increased in these populations since World War II, indicating the presence of unnatural stress factors. After the epizootic, the harbor seal populations both in the Baltic and along the Swedish west coast have increased in number. Chemical analysis of tissues has been performed on the three seal species collected in various areas of the Baltic and the Swedish west coast. The concentrations of 17 metals and non-metal elements, sDDT and PCBs, DDE and PCB methylsulfones, toxaphene, chlordanes, polybrominated diphenyl ethers, PCDDs and PCDFs have been determined in selected groups of seals in order to determine spatial, species and age variations in concentrations. Furthermore, healthy animals have been compared to diseased animals. Spatial variation was found mostly within the group of organohalogenated compounds, a group of contaminants where a strong covariation between the various compounds was also found. On the basis of the analytical results as well as the pathological findings on Baltic seals, the group of DDE and PCB methyl sulfones is tentatively suggested to be more important in explaining the disease complex than coplanar structures including dioxins.

Intestinal Transmission of Macromolecules (BSA and FITC-Dextran) in the Neonatal Pig: Enhancing Effect of Colostrum, Proteins and Proteinase Inhibitors

The effects of colostrum and constituents/factors in colostrum which may influence intestinal macromolecular transmission in the newborn preclosure pig were investigated. Unsuckled piglets were given, by use of a stomach tube, bovine serum albumin (BSA) and fluorescein-isothiocyanate (FITC)-labelled dextran 70,000 (FITC-D) as markers together with colostrum or the factors under study. The serum levels of BSA and FITC-D 4 h after feeding were then determined as a measure of the transfer. It was found that the two colostrums tested, bovine and especially porcine, markedly enhanced the transmission of both BSA and FITC-D. Furthermore, increasing amounts of the model proteins, BSA and bovine IgG (50-200 mg/ml), significantly increased the transfer of FITC-D, whereas unlabelled dextran 70,000 given in similar amounts did not. Proteinase inhibitors obtained from sow colostrum or soy bean also enhanced the transmission of both BSA and FITC-D while the inactive inhibitors, given as trypsin-inhibitor complexes, had no effect. On the other hand, addition of a proteinase, porcine trypsin, significantly decreased the transmission of FITC-D. These findings indicate that the intestinal transmission of macromolecules in the preclosure piglet is governed by the amount of protein available in the intestine. Therefore, feeding colostrum with a high protein content and proteinase inhibitors is likely to favour efficient intestinal transmission, although other colostrum factors may also be of importance.

Development of exocrine pancreas function in chronically cannulated pigs during 1-13 weeks of postnatal life

The development of exocrine pancreas function was studied in Swedish Landrace pigs surgically fitted with a chronic pancreatic duct catheter and a duodenal re-entrant cannula. The juice secretion and output of total protein and trypsin activity were followed before (basal secretion) and after feeding (postprandial secretion) during the first 1–13 weeks of life. The results showed that throughout the suckling period, up to 4–5 weeks of age, the basal pancreas function remained low and the secretory response to feeding, i.e., nursing sow milk, was also low. After weaning, the pancreatic juice secretion as well as the output of protein and trypsin activity markedly increased with respect to both basal and postprandial levels. Furthermore, the enzyme composition of the pancreatic juice changed qualitatively during this period. During the first 2 weeks of life, the intravenous administration of cholecystokinin (CCK) and secretin did not stimulate exocrine function, but a significant effect was achieved from 3–4 weeks of age. These results showed that there was both an increase in exocrine pancreas function and a qualitative change in the hydrolytic enzyme pattern during porcine postnatal ontogeny, apparently correlated with the changes in diet around weaning. An increase in the response of the pancreas to hormonal stimulation was also observed during the suckling period.

Levels of immunoreactive insulin, neurotensin, and bombesin in porcine colostrum and milk.

High concentrations of insulin (411 ± 214 μU/ml), neurotensin-like (265 ± 72 pg/ml), and bombesin-like immunoreactivities (1995 ± 288 pg/ml) were detected in porcine colostrum using radioimmunoassay, as compared to the levels found in sow blood serum at farrowing (5 μU/ml, <12 pg/ml, and 17 pg/ml, respectively). After 72 h of lactation, the levels of insulin and neurotensin-like immunoreactivities had decreased to 28 ± 17 μU/ml and 89 ± 23 pg/ml, respectively, while the bombesin-like activity remained constant. Characterization with reversed-phase high performance liquid chromatography showed that the insulin immunoreactivity eluted at the same position as the insulin standard, while the elution patterns of the neurotensin-like and bombesin-like immunoreactivities (eluted in three separate peaks) did not correspond to that of their respective standards. The biological function of the peptide hormones in colostrum/milk may be as triggers of the developmental changes taking place in the nursing neonate, especially in the gastrointestinal tract.

Enzymoblotting: A method for localizing proteinases and their zymogens using para-nitroanilide substrates after agarose gel electrophoresis and transfer to nitrocellulose

A method--enzymoblotting--was developed for localizing various enzymes after electrophoretic separation, transfer to nitrocellulose, and incubation with specific substrates. As an application, the proteinases porcine trypsin (EC 3.4.21.4), bovine chymotrypsin (EC 3.4.21.1), porcine elastase (EC 3.4.22.11), and their zymogen forms from porcine pancreas homogenate were analyzed utilizing specific p-nitroanilide substrates. After agarose gel electrophoresis, transfer of the separated proteinases to a nitrocellulose membrane was performed by capillary diffusion for 30 min. After air-drying of the nitrocellulose membrane, it was incubated in the appropriate substrate solution for 60 min. N-alpha-Benzoyl-DL-arginine-para-nitroanilide HCl was used as a substrate for trypsin, N-benzoyl-L-tyrosine-para-nitroanilide and succinyl-L-phenylalanine-para-nitroanilide for chymotrypsin, and N-succinyl-L-alanyl-L-alanyl-L-alanine-para-nitroanilide for elastase. p-Nitroaniline, the product thus obtained, was diazotized with N-(1-naphthyl)ethylenediamine to a red azo dye, visible at the site of the proteinases on the nitrocellulose membrane. The results could be preserved at -18 degrees C. Zymogen forms of the pancreas proteinases were detected in a similar manner. They were converted to active proteinases in situ on the nitrocellulose membrane after preincubating the nitrocellulose membrane in the activation enzymes enteropeptidase or trypsin.

Induction of Exocrine Pancreas Maturation at Weaning in Young Developing Pigs

The influence of weaning, at either 4 or 6 weeks of age, on the maturation of the exocrine pancreas was studied in naturally reared Swedish Landrace pigs that had no access to solid food. The pigs were surgically fitted with chronic catheters at 3 weeks of age, permitting periodic sampling of pancreatic juice and blood in conscious animals ≤4 weeks after weaning. During the suckling period, pancreatic fluid and enzyme secretion remained low, both before (preprandial) and after (postprandial) milk ingestion. After weaning at 4 or 6 weeks of age, juice secretion, output of total protein, and levels of different hydrolases (amylase, trypsin, lipase, and carboxylester lipase) and the cofactor colipase all increased markedly postprandially. Moreover, after weaning, the plasma insulin level increased postprandially. This did not happen before weaning, although blood glucose levels always rose after feeding. The data showed a relationship between the time of weaning and the induction of exocrine pancreatic maturation in pigs. This finding implies that postnatal development of pancreatic function is triggered by the dietary change from sow milk to dry solid food. In contrast, the age of the pig appears to be of minor importance, since weaning at either 4 or 6 weeks of age gave a similar result.

The passage of orally fed proteins from mother to foetus in the rat

Pregnant rats were orally fed with a mixture of bovine IgG, bovine albumin, ovalbumin and beta-lactoglobulin. Using immunoprecipitation methods, these proteins were detected in the maternal blood serum, urine and uterine fluids, and also in the foetal blood serum and the amniotic fluid. The results imply that a variety of dietary proteins, still immunoprecipitable, are able to cross the materno-foetal barriers to be detected in the foetal circulation, where they may influence the maturation of the immune system of the foetus.

Influence of oat saponins on intestinal permeability in vitro and in vivo in the rat

The aim of the present study was to investigate whether oat saponins (avenacosides A and B) have any effect on the permeability of the rat intestine to actively and passively transported markers in vitro and in vivo. Intestinal segments were mounted in modified Ussing chambers, and the passage of the different marker compounds from the mucosal to the serosal side was measured for 120 min. Avenacosides (1 mg/ml) gave a significantly higher passage of the macromolecule ovalbumin and there was a tendency to increased passage of [14C]D-mannitol and [51Cr]EDTA. On the other hand, the saponins did not affect the active transport of [3H]methyl glucose. When rats were given saponins (40 mg/kg body weight) together with markers by gastric intubation, the passage of [51Cr]EDTA into blood and urine was somewhat reduced. For the macromolecule bovine serum albumin, no evident effect on the passage was observed in the presence of saponins. Thus, in contrast to the in vitro results, the in vivo marker passage seemed to be unaffected or even reduced in the presence of avenacosides. The study shows that saponins can affect the permeability of the rat intestine. However, this effect needs further investigation in vivo, especially regarding proteins.

Insulin involvement in intestinal macromolecular transmission and closure in neonatal pigs.

Summary: The involvement of insulin in the intestinal transmission of macromolecules to blood and the cessation of this transport (intestinal closure) was studied in neonatal pigs by measuring the serum levels of the markers bovine serum albumin and fluorescein isothiocyanate labelled dextran 70,000 (FITC‐D) at 4 h after gavage feeding. In naturally suckled pigs, intestinal closure at 18 h was shown to be associated with an increase in serum immunoreactive insulin (IRI) levels. Similarly, intestinal closure obtained at ?? lactose/kg, was accompanied by an increase in serum IRI levels. Neither high serum IRI levels nor closure were observed in fasted pigs or in pigs gavage fed a total of 12 g lactose/kg. The effect of exogenous insulin on intestinal macromolecular transmission was studied by injecting 5 IU insulin/kg subcutaneously at 3 and 6 h, respectively, in newborn pigs gavage fed 10 ml sow colostrum/kg at 3 h intervals. This resulted in a reduction in the transmission of the markers when tested at 12 h, in comparison to littermates receiving the same amount of colostrum and littermates suckling the sow. It appears as if insulin, reflected as high serum levels over an extended period of time, is involved in the regulation of macromolecular trans?? ?? ulated that insulin may be involved in these processes by initiating the synthesis of membrane structural proteins in the enterocytes.

α-FOETOPROTEIN, ALBUMIN AND TOTAL PROTEIN IN SERUM FROM PRETERM AND TERM INFANTS AND SMALL FOR GESTATIONAL AGE INFANTS

Blood serum levels of fetoprotein albumin and total protein were measured for infants of various gestational ages and with various birth weights. Laboratory assays used electrophoresis in agarose gel containing specific antibodies. The alpha fetoprotein levels for boys were numerically higher than for girls; these differences did not show up for either albumin or total protein. Results of the study show alpha fetoprotein levels to be a good indicator of gestational age. This is true because the levels did not differ between groups of infants of the same gestational age but different body weights; the levels did differ for groups of infants with the same birth weight but different gestational ages. In this study alpha fetoprotein levels were higher in preterm than in term infants. Albumin and total protein levels varied according to weight rather than gestational age.