The piglet as a model for B cell and immune system development.

The ability to identify factors responsible for disease in all species depends on the ability to separate those factors which are environmental from those that are intrinsic. This is particularly important for studies on the development of the adaptive immune response of neonates. Studies on laboratory rodents or primates have been ambiguous because neither the effect of environmental nor maternal factors on the newborn can be controlled in mammals that: (i) transmit potential maternal immunoregulatory factors in utero and (ii) are altricial and cannot be reared after birth without their mothers. Employing the newborn piglet model can address each of these concerns. However, it comes at the price of having first to characterize the immune system of swine and its development. This review focuses on the porcine B cell system, especially on the methods used for its characterization in fetal studies and neonatal piglets. Understanding these procedures is important in the interpretation of the data obtained. Studies on neonatal piglets have (a) provided valuable information on the development of the adaptive immune system, (b) lead to important advances in evolutionary biology, (c) aided our understanding of passive immunity and (d) provided opportunities to use swine to address specific issues in veterinary and biomedical research and immunotherapy. This review summarizes the history of the development of the piglet as a model for antibody repertoire development, thus providing a framework to guide future investigators.

Identification and characterization of a hapten-modifiable TEPC 15 cross-reactive idiotype in swine.

Rabbits and swine immunized with TEPC 15 IgA, goats immunized with T15-positive IgM and swine immunized with affinity-pure swine anti-phosphorylcholine (PC) all produce antibodies which recognize a hapten-inhibitable idiotypic determinant on swine anti-PC. The similarity in reactivity and order of inhibitability with various PC analogs of the heterologous (swine anti-TEPC 15) and isologous (swine anti-swine anti-PC) reagents indicates that they recognize a related idiotype and suggest it may be the predominant idiotype expressed on swine anti-PC antibodies. The heterologous and isologous anti-idiotypic reagents generated in this study recognize swine and mouse anti-PC but not normal swine IgM, IgG or MOPC 460. Only reactions with swine anti-PC and mouse T15-positive anti-PC proteins are hapten-inhibitable. The greater inhibitory capacity of trimethylammonium and acetylcholine than PC suggests that the idiotope(s) recognized on swine anti-PC by the anti-idiotypic reagents is integral rather than peripheral to the anti-PC binding site. The nearly exclusive IgM anti-PC response of swine to Streptococcus pneumoniae R36A and PC-Brucella have so far hindered attempts to study the isotypic distribution of the idiotype.

The differential localization of IgA, IgM and IgG in the gut of suckled neonatal piglets.

The localization of immunoglobulin IgG, IgM and IgA in tissue sections prepared from the ileum of neonatal and adult swine were compared. Eighty percent of the immunoglobulin-containing lymphoid cells in the lamina propria of conventional adult German Landrasse swine were IgA-positive with lower numbers of IgM cells and occasionally an IgG cell. Anti-mu and alpha-chain reagents also stained the cytoplasm of the crypt epithelial cells. By comparison to these adult control tissues, the ileum of unsuckled neonates contained no immunoglobulins although after the ingestion of colostrum, the entire cytoplasm of the villus epithelial cells stained intensely when tested for IgG with only faint staining for IgM and IgA. On the other hand, IgA and IgM were readily localized on what appears to be only the apical border of the crypt epithelial cells but in contrast to the adult, the cytoplasm of these cells was unlabelled. IgG was absent from the crypt region. We interprete these findings to indicate an important, selective role for the villus epithelium in the absorption into the neonatal circulation of colostral IgG and probably IgA and IgM, and a specialized role for the crypt epithelium in adsorbing colostral IgA and IgM; possibly by complexing with mucin-bound secretory component.

Maternal-neonatal immunoregulation in swine. II. Influence of multiparity on de novo immunoglobulin synthesis by piglets.

In the first-litter sows lower serum levels were found for all three Ig classes as compared to multiparous sows. The same was true for IgA in lacteal secretions and in piglet serum during the first days of life, while no differences were found for IgG levels. In contrast to these findings, IgM levels were found to be higher in lacteal secretions of first-litter sows and in piglet serum during the first days of life as compared to their counterparts. From three weeks after birth Igs found in piglet serum mainly originate from de novo synthesis. In this period piglets of first litter sows showed a higher IgA level up to the 6th week of life and higher IgG and IgM levels up to the end of the investigation period. Results are discussed in terms of maternal-neonatal immune regulation, focussing on the apparent suppressive role of maternally-derived IgG on total de novo Ig synthesis by suckling piglets.

Changes in the concentrations of serum IgG, IgA and IgM of sows throughout the reproductive cycle.

The concentrations of IgG, IgM and IgA in sera collected from 3855 sows (3208 pregnant and 647 lactating) at a single time point were determined. This experimental design allowed changes in serum immunoglobulin over the reproductive cycle to be studied without bias from seasonal influence. The concentrations of the three immunoglobulins changed independently during the reproductive cycle. Serum levels of IgM and IgG began a progressive postpartal decline during the 14th-17th week of gestation. At the onset of lactation serum IgG levels progressively increased while IgM levels continued to decline, the latter reaching their lowest level during the third week of lactation. In contrast to IgM and IgG, serum IgA levels increased 35% during weeks 14-17 of gestation and continued to increase throughout lactation, reaching their highest serum levels in the third week of lactation; the serum IgA concentration at this time was twice that observed during the first 13 weeks of gestation. Results of these studies allowed the reproductive cycle to be classified into four phases on the basis of serum immunoglobulin concentrations: (1) weeks 1-4 of gestation; (2) weeks 5-13 of gestation; (3) weeks 14-17 of gestation and (4) lactation.

The influence of age and breed on the concentrations of serum IgG, IgA and IgM in sows throughout the reproductive cycle.

The influence of age and breed on the concentrations of IgG, IgA and IgM in the sera of sows throughout the reproductive cycle was investigated in 4137 sows which had had 0-20 gestations and representing three breeds: Swedish Landrace, German Landrace and L-12 (= Swedish Landrace x Large White). Data revealed an increase in total immunoglobulins (Ig), IgM and IgG serum levels with increasing gestation number; the latter contributed greater than 80% to total Ig levels. IgG was significantly increased up to the fourth gestation, whereas IgM showed a significant increase only to the third gestation. IgA showed only minor differences. Age-dependent increases in serum IgM were ascribed to the increased probability of antigenic exposure during suckling, while failure to observe this change in serum IgA was ascribed to its role as a local Ig. Breed differences were observed to be significant for all three Ig's. It is concluded that establishment of group norms for serum Ig's should consider age and breed difference as well as stage of gestation or lactation.

Composition of sow milk during lactation.

The composition of sow colostrum and milk was quantitated in 25 sows at 14 time points throughout lactation. All animals belonged to the same experimental herd of German Landrace, farrowed within 4 d, and were of various lactation numbers and various litter sizes. In the first 6 h of lactation colostrum total solids (TS) and protein contents were higher, while fat and lactose contents were lower than in mature milk. Decreased total protein and whey protein contents and concomitantly increased fat and lactose content, with nearly unchanged TS levels, indicate transition from colostrum to mature milk. The high protein content of colostrum was largely due to immunoglobulin (Ig). During the first 6 h, IgG accounts for nearly all the protein in colostrum but plays a decreasing role in sow milk as lactation proceeds. After 2 wk, IgA levels begin to increase and at the end of lactation, IgA constitutes 40% of the total whey protein. No influences of lactation number and litter size on milk composition could be ascertained in this study.

Comparison of freezing and lyophilizing for preservation of colostrum as a source of immunoglobulins for calves.

Lyophilizing was compared to freezing as a method of colostrum storage. Eight lots of colostrum from the first milking were divided into two equal parts; one was frozen, and the other was lyophilized. Twenty-two newborn calves were divided into two groups and fed either 2 L of frozen and thawed colostrum or 2 L of reconstituted lyophilized colostrum. The calves were bled at 12, 18, 24, and 72 h after feeding, and levels of the immunoglobulins IgG1, IgG2, IgM, and IgA were determined with a radial immunodiffusion assay, in colostrum and sera. The mean concentration of individual immunoglobulin isotypes in the sera of calves fed either frozen or lyophilized colostrum did not differ significantly. Calves fed from the same lots of colostrum had similar immunoglobulin concentrations in their sera, irrespective of the method of storage. All immunoglobulin isotypes were absorbed with equal efficiency from frozen and lyophilized colostrum as determined by calculation of the absorption coefficient.

Absolute and relative concentrations of immunoglobulins G, M, and A, and albumin in the lacteal secretion of sows of different lactation numbers.

The concentrations of immunoglobulin (Ig)G, IgM, IgA, and albumin in the lacteal secretion of 80 sows were measured not more than 14 times during lactation to determine whether the age of the sow, the anatomic location of the mammae, or the farrowing barn housing of the sow influenced Ig concentrations. The combined data indicated that there were lactation stage-dependent changes in the total Ig concentration, as well as changes in the relative importance of the 3 classes of Ig studied. Mean values of any one time point showed enormous variation in absolute values, with coefficients of variation ranging up to 70% The largest variations were observed between colostral samples obtained on the first day. The IgM showed the least variation on day 1. Such variations hindered attempts to statistically identify their source. Normalization of Ig concentrations in relation to albumin concentrations decreased some of the apparent variation. Normalized data accentuated the changeover from colostrum to milk which occurred after the first 3 days. There was also a 3-fold increase in IgA concentrations which occurred during the last 4 weeks of lactation. Data showed that older sows tended to have higher concentrations of all Ig. Lactation number-dependent increases in IgM and IgG were restricted to colostrum in lactation numbers higher than 4. Immunoglobulin A increased in a similar pattern and, in addition, increased in mature milk in sows with more than 2 lactations. The enormous variations observed seemed to be individual animal variations. This allowed only trends, rather than statistically significant differences, to be recognized.(ABSTRACT TRUNCATED AT 250 WORDS)

Maternal-neonatal immunoregulation: suppression of de novo synthesis of IgG and IgA, but not IgM, in neonatal pigs by bovine colostrum, is lost upon storage.

Fifty-four neonatal pigs were allotted to 4 groups and reared in an electrically controlled automatic feeding device (autosow). Each group was reared on a different pool of bovine colostrum: fresh, stored 1 month, stored 6 months, and stored 8 years. Bovine and porcine immunoglobulins in the sera of these pigs, and in a group of conventionally reared pigs, were measured periodically during the first 42 days after birth. The maximal concentration of absorbed bovine immunoglobulin was reached between 12 and 18 hours and equaled or exceeded the amount of porcine immunoglobulin absorbed by the conventionally reared pigs. Large differences in the concentrations of the bovine immunoglobulin isotypes among the various pools of colostrum were positively correlated with concentration of these isotypes in the sera of the neonatal pigs fed these pools. Relative to their concentrations in colostrum, approximately 41% of the IgG1, 55% of the IgG2, 29% of the IgM, and 67% of the IgA was absorbed. The IgA was absorbed the best and IgM was least absorbed. Significant trends or differences in absorption were not observed among groups. Neonatal pigs given fresh colostrum, which had a higher fat content, had significantly more weight gain (P less than 0.05). This occurred, despite the fact that the fresh colostrum had the lowest concentration of bovine immunoglobulin. Serum half-lives for bovine IgG1 and IgG2 were significantly less than for porcine IgG (P less than 0.05), whereas the half-lives for bovine and porcine IgM and IgA were similar. De novo-synthesized immunoglobulins were detectable in serum after 6 days; IgM concentrations reached a maximum at 15 days in neonatal pigs given stored, but not fresh, colostrum.(ABSTRACT TRUNCATED AT 250 WORDS)

[ACTH stimulation test for the determination of salivary cortisol and of cortisol responses as markers of the training status/fitness of warm-blooded sports horses]. / ACTH Stimulationstest und Bestimmung von Cortisol im Blut und Speichel zur Bewertung des Trainingszustands/der Kondition beim Warmblutpferd.

Previous work (Marc et al., 2000) suggested that plasma cortisol responses to treadmill exercise or ACTH injection are a reliable marker for performance evaluation in warmblood horses. For practical purposes blood sample collections and treadmill exercise tests are somewhat troublesome and time consuming. The goal of this study was thus to evaluate the use of saliva for cortisol determination (by direct EIA) as a marker for performance and to investigate the reliability and repeatability of plasma cortisol responses to a single i.v. injection of ACTH (50 micrograms or 250 micrograms). Furthermore, the effect of training horses for 8 weeks 3 times per week covering the same distance (increasing from 3.5 km during the first week to 8 km during the last week) either by trotting (approximately 240 m/min) or by cantering (375 m/min) was investigated. For this purpose initially ten four-year-old Hannovarian geldings, all reared in the same State stud, were used. Mean overall correlation between salivary cortisol and plasma cortisol concentrations was 0.64 when samples of various points of time were used. However, in spite of attempts to standardize saliva sample collection, correlation between salivary cortisol levels and plasma cortisol levels at distinct points of time in different tests were low and significant (r = 0.85, p < 0.02) only in one test. Thus, salivary cortisol measurements for diagnostic purposes are not reliable or useful. The repeatability of plasma cortisol responses to ACTH for untrained and trained horses were r = 0.86 and r = 0.8 respectively (p < or = 0.01 and p < or = 0.05 respectively). Training horses either by trotting or cantering did not affect the cortisol response either to treadmill exercise or to stimulation by ACTH. It is concluded that the relationship between salivary cortisol levels and plasma cortisol levels is not close enough to allow the use of salivary cortisol determination as marker of the training status/fitness of horses. The repeatability of the cortisol response to ACTH is similar to the cortisol response to treadmill exercise. Based on plasma cortisol responses to ACTH or treadmill exercise training horses by cantering at low speed is not superior to training by trotting for the fitness of horses.

[Variations in the concentrations of the immunoglobulins IgG1, IgG2, IgM and IgA in sheep. 1. Changes in the blood serum and in the milk of sheep of different breeds and cross-breeds during lactation]. / Variationen in den Konzentrationen der Immunglobuline IgG1, IgG2, IgM und IgA beim Schaf. 1. Mitteilung: Veränderungen im Blutserum und in der Milch der Schafe verschiedener Rassen und Rassekreuzungen im Verlauf der Laktation.

A total of 103 ewes from the breeds Black mutt., Texel, Finn. L., Heidschnucke and the crossbreeds Texel x Finn. L. and Black. mutt. x Finn. L. was studied. Blood samples were drawn at days 1, 7, 21 and 42 and milk samples at the 4th, 12th, 24th and 72nd hour after the onset of lactation and subsequently on days 7, 21 and 42. The concentrations of the immunoglobulins IgG1, IgG2, IgM and IgA were assayed in serum and milk. The following results were obtained: 1. The total immunoglobulin contents in the serum was not significantly different between breeds. 2. All ewes showed a rise in serum immunoglobulin concentrations by about one third over the first six weeks of lactation. Between 50-60% of this increase were on the account of IgG1. 3. The serum concentration of IgG1 and IgG1 rose as of the third day, those of IgM as of day 21 after lambing. 4. The rise in serum immunoglobulin concentration continued after the weaning of the lambs. 5. The ratio of IgG1 to IgG1 in ewe serum was 2:1. 6. The immunoglobulin concentration in milk dropped sharply on the first day of lactation, followed by a continuous, more gradual decrease over the entire course of lactation. A terminal rise, as observed in sows, could not be detected. 7. The ratio of IgG1:IgG2 : IgM : IgA in the whey changed from 85 : 1 : 12 : 2 on day one to 70 : 7 : 12 : 11 on the last day of lactation. 8. While characteristic trends in immunoglobulin patterns in the sera of ewes over the course of lactation are clearly discernible, it is not possible to denote "normal" values.

[Variations in the concentrations of the immunoglobulins IgG1, IgG2, IgM and IgA in sheep. 2. Changes in the blood of lambs of different breeds and crossbreeds during the course of the rearing period]. / Variationen in den Konzentrationen der Immunglobuline IgG1, IgG2, IgM und IgA beim Schaf. 2. Mitteilung: Veränderungen im Blut der Lämmer verschiedener Rassen und Rassekreuzungen im Verlauf der Aufzuchtperiode.

The sera of 188 lambs from seven breed groups were analyzed for the concentrations of IgG1, IgG2, IgM and IgA by radial immunodiffusion using monospecific antibodies. From each lamb, 14 blood samples were drawn before and 5 samples after weaning. The following results were obtained: 1. Immunoglobulins could not be detected in sera drawn before the first intake of colostrum. 2. In normally suckling lambs, the peak concentrations of maternal immunoglobulins are attained at 0-18 hrs after birth. They can be assessed in a single blood sample drawn between 18 and 24 hrs. 3. The half-life times of maternal immunoglobulin in lamb sera are 11 days for IgG1, 7 days for IgG2, 6 days for IgM and 18 hours for IgA. 4. The absolute peak heights relate to the amounts of colostrum ingested before 12-18 hrs after birth. 5. The decline of maternal immunoglobulins in lamb sera over-laps with the onset of lamb immunoglobulin synthesis. Renewed rises of concentrations are observed for IgG2 after week 2, for IgM after week 3 and for IgG1 after week 7. The concentrations of IgA remain at the low levels characteristic for the serum of grown sheep. 6. The role of immunoglobulin synthesis in suckling lambs is only briefly and to a small extent reduced after weaning.

[The quantitative passage of immunoglobulins from the intestine into the blood vessels in newborn lambs]. / Untersuchungen zum quantitativen Ubergang von Immunoglobulinen aus dem Darm in die Blutbahn neugeborener Lämmer.

A trial with 69 lambs examined the effect of the feeding regime on the transfer of colostral immunoglobulins from the intestinal lumen to the circulating blood. The lambs received 125 ml of bovine colostral milk per kg birth weight at each feeding in the first 24 hours after birth. There were four experimental groups. Feeding intervals were either 2 or 6 hours and the milk was administered with or without doubling the fluid volume by addition of tap water. Serum concentrations of IgG1, IgG2, IgM and IgA were measured during the experimental period and the transport efficiency index for each isotype was calculated. The transport efficiency index represents the relationship between the amount of immunoglobulin administered in colostral milk and the concentration measured in the blood. Feeding in 2 hour intervals produced maximum immunoglobulin concentrations at 30 hours, while 6 hour feeding intervals achieved the same maximal levels at 24 hours. The relative proportions of the four immunoglobulin isotypes measured in the blood were unaffected by the feeding interval. Doubling the fluid volume administered while holding the absolute amount of colostral milk constant, resulted in a disproportionate increase in blood IgG1 concentration as compared to the other isotypes. Concentrations for IgG2, IgM and IgA remained essentially the same as those achieved with undiluted colostral milk. The feeding regimes tested did not affect endogenous immunoglobulin synthesis, growth rate or the health of the lambs.

[The "waiting period" for follow-up administration of bovine colostrum to newborn lambs]. / Untersuchungen über "Wartezeiten" mit anschliessender Verabreichung von Rinderkolostralmilch bei neugeborenen Lämmern.

Extent and duration of the permeability of the small intestine of newborn lambs for intact proteins from cow's colostrum was analyzed in five groups of animals. Cow's colostrum in quantities of 200 ml/kg birth weight, was offered in 3-hr intervals for 24 hrs, beginning either immediately after birth or after fasting, for 12 and 24 hrs respectively. In two groups each either 5% glucose solution or saline was offered to the fasting animals. The following results were obtained: 1. All lambs fed immediately after birth drank eagerly. The intake in all fasting groups dropped between 12 and 24 hrs of the feeding period. 2. While the duration of the permeation of the small intestine for intact proteins remained unaffected by the various regimens, fasting with or without supplementation decreased the maxima. 3. The decrease in protein permeation was class-depending as demonstrated by the lower levels of bovine IgM in the sera of fasting lambs. 4. The permeability of the small intestine for proteins was substantially reduced by 5% glucose. The difference between fasting lambs or lambs with saline substitution was minimal. 5. All lambs fed immediately after birth survived. The losses in the other groups amounted to 29% after 12 hrs of starving, 33% after 24 hrs of starving, 30% after starving with glucose supplementation and 50% after saline solution. 6. The surviving animals showed no differences in weight gain.

[The absorption of colostral immunoglobulins in newborn piglets. II. Effect of water or glucose solutions on the permeability of the newborn intestine]. / Untersuchungen über die Absorption der kolostralen Immunoglobuline bei neugeborenen Ferkeln. II. Mitteilung: Einfluss von Wasser oder von Glukoselösung auf die Durchlässigkeit des neugeborenendarmes.

The extent and the duration of the passage of intact proteins from the intestinal lumen into the bloodstream of newborn piglets was investigated. A total of 48 piglets from 4 litters were used in groups of 6 animals per experiment. The piglets were removed from the sow immediately after birth and placed in individual boxes of an automatic feeding device. Liquids were offered in trays, access to which was controlled by electronically operated gates. Volumes of 25 ml were allotted in hourly intervals. Porcine colostrum (6 x 25 ml) or bovine colostrum (24 x 25 ml) were either offered immediately, or after a 24 h fasting period without or with access to tap water. Two groups received 24 hourly rations of a 13 % glucose solution prior to the onset of feeding with porcine and bovine colostrum, respectively. The plasma levels of immunoglobulins were assessed by radial diffusion in blood samples drawn from a subcutaneous abdominal vein. The passage of intact proteins ceases between 12-18 hrs after the onset of feeding. Both the time course of the passage and the maximal levels achieved are unaffected after previous fasting or after tap water allowances only for 24 hrs. Previously fed glucose solution--in contrast--closes the gut barrier to subsequently ingested proteins. There was no difference in the postexperimental weight developments between groups.

[The absorption of colostral immunoglobulins in newborn piglets. III. Influence of the duration of colostrum administration]. / Untersuchungen über die Absorption der kolostralen Immunglobuline bei neugeborenen Ferkeln. III. Mitteilung: Einfluss der Verabreichungsdauer der Kolostralmilch.

The gastrointestinal tract of newborn piglets is permeable for intact immunoglobulins ingested with the colostrum. The duration of this passage was investigated by administering hourly rations of 25 ml of either porcine or bovine colostrum for 6, 12, 18 or 24 hrs after birth. The plasma concentrations of the subclasses porcine IgG, IgM and IgA or bovine IgG1, IgG2, IgM and IgA were determined at 12, 18 and 24 hrs after birth and on days 3 and 6. Feeding periods of 6 hrs resulted in plasma Ig levels of the same order of magnitude as observed in natural rearing. These levels were not substantially increased after prolonged feeding. The 6% gain from 6 to 12 feedings seen with porcine colostrum as compared with a gain of 24% for bovine colostrum points at an earlier closure of the intestinal wall for the species-specific proteins. There was no further increase of Ig permeation after 12 hourly feedings. Growth performances and losses were identical in all groups.

[Necessary provision of newborn lambs with sheep colostrum]. / Erforderliche Versorgung neugeborener Lämmer mit Schafskolostralmilch. 1. Mitteilung: Verabreichungsintervall.

Twenty new-born lambs of each of the breeds Schwarzkopf, Texel and Finnish were fed with 600 ml of pooled colostrum (0-12 h) on the first day of life. The total allocation was fed in portions ranging from 25 to 150 ml at intervals of 1, 2, 3, or 6 hours. The effect of the different intervals and portion volumes on the transintestinal passage of immunoglobulins, the onset of endogenous synthesis, the growth performance of the lambs and the mortality rate was investigated. The concentrations of IgG1, IgG2, IgM and IgA in the sera were analyzed over the course of a nine-week period. 1. The extent of passive immunization by transintestinal passage remained essentially unaffected by different feeding schedules. 2. There was also no effect on the onset of endogenous Ig synthesis. 3. Growth performance and mortality rate likewise showed no discernible differences.