The effect of immunocastration on adipose tissue deposition and composition in pigs.

Immunocastrated pigs (IC) exhibit intensive fat deposition after immunisation, but the underlying mechanisms of intensified fat metabolism and deposition are not yet fully understood. Moreover, there is also a lack of comparative studies performed on IC, entire males (EM) and surgical castrates (SC). The main objective of our research was, therefore, to characterise the adipose tissue from the quantitative, histo-morphological and biochemical perspectives in IC 5 weeks after their immunisation in comparison to EM and SC. Immunocastrated pigs had an intermediate position in carcass fatness traits between EM (the leanest) and SC (the fattest). The histo-morphological traits of the subcutaneous adipose tissue of IC were similar to those of SC and differed from those of EM; i.e., they exhibited larger adipocytes in the outer backfat and a larger lobulus surface area in both backfat layers than EM. Intensive fat tissue development in IC was corroborated with higher activities of lipogenic enzymes (i.e., fatty acid synthase, malic enzyme, glucose 6-phosphate dehydrogenase, citrate cleavage enzyme), which was especially pronounced in the subcutaneous adipose tissue of IC (1.5- to 2.7-fold higher activity than in EM or SC). The fatty acid composition of the backfat in IC was similar to that in EM pigs. Both IC and EM exhibited less saturated and more polyunsaturated fatty acids than SC. In contrast, the fatty acid composition of the intramuscular fat of longissimus dorsi muscle in IC pigs was more similar to SC than to EM (higher monounsaturated and lower polyunsaturated fatty acid content in IC and SC than EM). In this study, it was demonstrated that immunocastration notably influenced lipid metabolism. This was shown by increased quantity of lipid depots and with changes in adipose tissue cellularity compared to EM, with changes in the fatty acid composition of the intramuscular fat and enhanced lipogenic activity compared to both EM and SC. These results provide new insights into the specificity of adipose tissue development and deposition in IC compared to EM and SC.

Dynamics of myosin heavy chain isoform transition in the longissimus muscle of domestic and wild pigs during growth: a comparative study.

Dynamics of myofiber differentiation/maturation in porcine skeletal muscle is associated with domestication, breeding and rearing conditions. This study was aimed to comparatively elucidate the age-dependent myosin heavy chain (MyHC) isoform expression and transition pattern in domestic and wild pig (WP) skeletal muscle from birth until adulthood. Domestic pigs (DPs) of Large White breed raised in conventional production system were compared with WPs reared in a large hunting enclosure. Muscle samples for immuno/enzyme histochemistry were taken from the longissimus dorsi muscle within 24 h postmortem at 24 to 48 h, 21 to 23 days, 7 months and ~2 years postpartum. Based on the antibody reactivity to MyHCs (NCL-MHCs, A4.74, BF-F3) and succinate dehydrogenase activity, myofibers were classified into I, I/IIa, IIa, IIx and IIb types. In addition, foetal MyHC expression was determined with the use of F158.4C10 antibody. Maturation of the longissimus dorsi muscle in the WP was characterized by an accelerated transformation of the fast to slow MyHC during the first hours postpartum, followed by differentiation towards oxidative myofibers in which type I, IIa and IIx MyHCs predominated. In the DP, the transformation shifted towards glycolytic myofibers that expressed MyHC-IIb. The expression of foetal MyHC was higher in the DP than in the WP at 1 day of age, and the decline in the foetal MyHC during the first 3 weeks was more rapid in the WP than in the DP denoting an accelerated early postnatal muscle maturation in WP than DP piglets. All foetal MyHC-positive myofibers co-expressed IIa isoform, but not vice versa. The intense myofiber hypertrophy was evident from 3 weeks until 7 months of age. In this period, the myofiber cross-sectional area increased up to 10- and 20-fold in the WP and the DP, respectively. In the DP, the hypertrophy of all myofiber types was more pronounced than in the WP, particularly the hypertrophy of IIx and IIb myofibers. To summarize, the comparison between growing DP with wild ancestors showed that genetic selection and rearing conditions lead to substantial changes in the direction and intensity of postnatal MyHC transformation as evidenced by different proportion of individual myofiber types and differences in their hypertrophic potential.

Corrosion cast study of the canine hepatic veins.

This study presents a detailed description of the distribution, diameters and drainage patterns of hepatic veins on the basis of the corrosion cast analysis in 18 dogs. We classified the hepatic veins in three main groups: the right hepatic veins of the caudate process and right lateral liver lobe, the middle hepatic veins of the right medial and quadrate lobes and the left hepatic veins of both left liver lobes and the papillary process. The corrosion cast study showed that the number of the veins in the Nomina Anatomica Veterinaria and most anatomical textbooks is underestimated. The number of various-sized hepatic veins of the right liver division ranged from 3 to 5 and included 1 to 4 veins from the caudate process and 2 to 4 veins from the right lateral liver lobe. Generally, in all corrosion casts, one middle-sized vein from the right part of the right medial lobe, which emptied separately in the caudal vena cava, was established. The other vein was a large-sized vein from the remainder of the central division, which frequently joined the common left hepatic vein from the left liver lobes. The common left hepatic vein was the largest of all the aforementioned hepatic veins.

Variations in pectoral girdle muscles in dogs.

Muscle variations take the form of additional muscle bundle or belly, unusual muscle origin or termination, as well as complete muscle absence. Knowledge of such variations not only has clinical importance for guiding surgery, but also can help reveal phylogenetic relationships. To improve our understanding of muscle variations in dogs, 57 medium-sized, cross-breed male and female adult animals were dissected as part of a gross anatomy course between 2005 and 2011. Variations in pectoral girdle muscles were observed in 7 (12.3%) dogs and took the form of an additional muscle bundle in the brachiocephalicus muscle (n = 2), in the omotransversarius muscle (n = 2), in the cervical part of the rhomboideus muscle (n = 2) and in the cervical part of the serratus ventralis muscle (n = 1). Muscle variation was bilateral in only one dog; it involved the omotransversarius muscle. The variations did not seem to be sex dependent. Such variations can appear regularly in dogs and should be taken into consideration during anatomical dissection.

Steroid hormones, boar taint compounds, and reproductive organs in pigs according to the delay between immunocastration and slaughter.

The producer of vaccine against GnRH recommends that immunocastrated pigs are to be slaughtered within 4 to 6 weeks after the second vaccination (V2). The objective of the study was to examine the effect of shorter or longer delay on steroid hormones, boar taint compounds, and morphologic and histologic traits of reproductive organs. Forty male pigs (individually housed and fed a commercial diet) were assigned within litter to four treatment groups, 10 pigs were left entire (EM27) and the others were vaccinated against GnRH (Improvac, Pfizer Animal Health) at the age of 12 and 19 weeks. Pigs were slaughtered at 21 (IC21), 24 (IC24), and 27 (IC27 and EM27) weeks of age. Two EM27 pigs died during the experiment, one IC21 pig was excluded because of illness, one IC27 pig was a nonresponder, and two pigs (IC24 and IC27) were hermaphrodites. To assess the effect on steroid hormones, blood was taken at 12, 15, 19, 21, and 24 weeks of age. Subcutaneous fat and reproductive organs were sampled after slaughter for determination of androstenone, skatole, morphologic, and histologic measurements. Immmunocastration interrupted the rise of estrogen and caused a substantial fall of testosterone in IC21, IC24, and IC27 pigs. As a result, androstenone and skatole levels were successfully reduced regardless of the time elapsed from V2. The weight of the reproductive organs was also drastically reduced, the shrinkage being proportional to the length of the interval between V2 and slaughter and was the most evident for vesicular glands, followed by bulbourethral glands, and testes. Corresponding changes were observed also on a histologic level with a progressive decrease in the size and number of Leydig cells, a diminishing immunoreactivity of 3ß-hydroxysteroid dehydrogenase/Δ-5-4 isomerase, and luteinizing hormone receptor, along with a shrinkage of tubuli seminiferi, atrophy of seminiferous epithelium, and a loss of germ cells, indicating a disruption in testicular spermatogenetic function. Regression of the glandular tissue with a decreasing amount of secreta was also observed for bulbourethral and vesicular glands. The investigated physiologic, morphologic, and histologic traits were progressive with the increasing delay to slaughter (clearly seen already 2 weeks after V2), though no signs of functional or morphological restoration was observed within 8 weeks after V2.

Gross anatomy of the portal vein and hepatic artery ramifications in dogs: corrosion cast study.

The anatomical variations of the portal vein and the hepatic artery ramifications were analysed on liver corrosion casts in 20 dogs as a possible aid in the surgical management of the organ. The portal vein ramified similarly in all dogs. It divided into the smaller right portal branch from which vessels for the caudate process and both right lobes arose and the substantial left portal branch, which supplied the remaining liver portions and in 12 cases also the dorsal part of the right lateral lobe. Right lateral, right medial and left branches are the major arteries originating from the hepatic artery; however, their origin and course varied among individual animals. In 10 livers, the right lateral and the left branches originated from the hepatic artery, while the right medial branch arose from the left branch and usually supplied the right medial lobe solely. In nine livers, the right medial branch arose directly from the hepatic artery and supplied quadrate lobe and gallbladder as well, while in one liver the common artery, which subsequently divided into lobar branches, branched away from the hepatic artery. An additional branch for the caudate process, originating directly from the hepatic artery, was observed in 10 livers. Certain liver portions received the arterial blood from two major branches, which was particularly characteristic for the right medial lobe (six livers) and caudate process (10 livers). The course of the major arterial branches was also variable, although they proceeded in close anatomical relationship with the portal vein branches. The left arterial branch accompanied the left portal branch on its dorsal aspect (15 cases) or crossed it from the caudal aspect (five cases). The right lateral branch crossed the initial parts of the left and right portal branches either from cranial (12 cases) or caudal aspects (eight cases), while the right medial branch always crossed the left portal branch from its caudal aspect.

Identification of myosin heavy chain I, IIa and IIx in canine skeletal muscles by an electrophoretic and immunoblotting study.

To determine which myosin heavy chain (MHC) isoforms are expressed in canine skeletal muscles, different muscle samples of five mixed-breed dogs were analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The separated MHC isoforms were identified by immunoblotting technique using a set of specific monoclonal antibodies. To compare the results of the electrophoretic and immunoblotting study, the pattern of MHC isoform expression and histochemical profiles of canine fibres were additionally demonstrated on serial muscle sections by immunohistochemistry and myofibrillar adenosine triphosphatase (mATPase) histochemistry. Not more than three MHC isoforms were demonstrated by SDS-PAGE in the analysed canine muscles. By the immunoblotting technique, the fastest migrating MHC band was identified as slow or MHC-I, the intermediate one as MHC-IIx and the slowest migrating band as MHC-IIa isoform. Since none of the three MHC bands and none of the analysed fibres were recognized by the antibody specific to MHC-IIb of rats, we concluded that MHC-IIb is not expressed in large skeletal muscles of dogs. Similarly, only three major fibre types, i.e. I, IIA and IIX, were revealed according to the pattern of MHC immunohistochemistry and mATPase reaction. Type IIA fibres were more alkali- and acid-stable than type IIX fibres after mATPase histochemistry; hence, the latter corresponded to type IIDog fibres. However, beside the three major fibre types, scarce hybrid fibres co-expressing two MHC isoforms (I/IIA and IIA/IIX) were demonstrated by immunohistochemistry.

Pattern of myosin heavy chain isoforms in different fibre types of canine trunk and limb skeletal muscles.

The aim of this study was to determine the pattern of myosin heavy chain (MHC) isoform expressions within the muscle fibres of functionally diverse trunk and limb dog muscles using monoclonal antibodies that are specific to MHC isoforms. We found that three MHC isoforms are expressed in dog skeletal muscles. The pattern of their expressions determined the existence of 'pure' fibres, i.e. I and IIa, both expressing only one MHC isoform, and 'hybrid' fibres, i.e. I/IIa and IIa/x, that co-expressed two MHC isoforms. While the MHCI, MHCIIa and MHCI/IIa fibres corresponded to the myofibrillar ATPase type fibres I, IIA and IIC, respectively, the hybrid MHCIIa/x fibres mostly behaved like the IIDog fibre type in myofibrillar ATPase reaction as described by Latorre et al. No pure MHCIIx fibres were found. Though MHCIIa/x fibres were quite numerous, their presence varied not only within different muscles but within the same muscle of different animals as well. We suggest that the discrepancies in the classification of fibre types according to their myofibrillar ATPase activity between different studies of dog skeletal muscles are probably a consequence of the variable content of the MHCIIa and MHCIIx isoforms in the MHCIIa/x hybrid fibres. Estimating the histochemical metabolic profile of fibres we found that in all fast fibres oxidative-glycolytic metabolism prevailed, whereas in slow fibres oxidative metabolism was more pronounced.

Apoptosis and proliferation during seasonal testis regression in the brown hare (Lepus europaeus L.).

Testes samples of 52 brown hares (Lepus europaeus L.), sacrificed between July and January, were subjected to immuno histochemical analysis. The terminal deoxynucleotidyl transferase-mediated d'UTP nick end labelling (TUNEL) method was applied to detect apoptosis; and antibodies to proliferating cell nuclear antigen (PCNA) were used to evaluate cell proliferation in the testes. In the seminiferous epithelium, the apoptotic processes were evident from August to early November with maximal values in September. Cell death in germ cells occurs predominantly during the prophase of the first meiotic division. In July, and from mid-November onwards, only the occasional TUNEL-positive cells can be seen. The proliferation of germ cells continues during the testis regression phase. The average number of PCNA-positive cells decreases slightly from September onwards and rises again in mid-November.

Giant muscle fibres in pigs with different Ryr1 genotype.

This study examined the frequency, morphological and immunohistochemical characteristics of the giant fibres in the longissimus muscle of local Krsko polje pigs with different Ryr1 genotypes. Giant fibres were round-shaped and had significantly increased cross-sectional area compared with normal muscle fibres. Only fast-twitch glycolytic fibres were affected, usually showing enhanced succinate dehydrogenase activity. On the ultrastructural level, the dilation of the sarcoplasmic reticulum, swelling of mitochondria and destruction of myofilaments was observed. The incidence of giant fibres was the highest in Ryr1 dimutant pigs (Ryr1 nn), which also exhibited lower muscle pH1 than heterozygous (Ryr1 Nn) or pigs with the wild Ryr1 gene (Ryr1 NN). However, the giant fibres were also present in pigs free of Ryr1 gene mutation. Our results suggest that the giant fibre syndrome depends mostly upon the rate and intensity of early post-mortem glycolysis, which results in acidity of muscle tissue. We suppose that the giant fibre formation is a result of excessive intracellular lactate accumulation in some fast-twitch glycolytic fibres. This process could also explain the ultrastructural alterations and the consequent changes in the oxidative enzymes and myofibrillar ATPase staining pattern observed in our and some previous studies.

Growth dynamics of lipizzan horses and their comparison to other horse breeds.

A typical body format of Slovenian Lipizzan horse was investigated. The study included 6 foals (5 colts and 1 filly) at the Lipica stud farm. They were measured from birth to twenty-seven months and again at forty-four months of age. Measurements included body length, chest circumference, withers height and body mass. All those measurements were statistically evaluated and compared to some other horse breeds to determine the similarity of the growth dynamics of those horse breeds. It was concluded that the parameters of the head and neck reached their full growth at the age of 27 months and also exceeded the values of their parents. The same results were obtained with the length and heights of the body (body length, withers height, etc.) and with the parameters of the front and hind legs. However the girth width, hip width and some other parameters had not yet reached their full growth at 27 months. We observed different growth dynamics with the body weight. The parameter continued to grow after 27 months of age. The same growth dynamics also seem to apply to other investigated horse breeds although the actual measurements were slightly different.

Immunohistochemical study of normal and mange (S. scabiei var. rupicaprae) infested chamois (Rupicapra rupicapra L.) skin.

The immunohistochemical study of chamois (Rupicapra rupicapra L.) skin showed that a limited number of available monoclonal and polyclonal antibodies expressed reactivity with skin cell components. These included cytokeratins, vimentin, desmin, neuron-specific enolase and S-100 protein with almost the same distribution pattern as already described in the skin of humans and animals. Antibodies used for labelling skin-associated lymphoid tissues and other cells with the immunologic function in human skin failed to demonstrate these cells in the chamois skin with the exception of LCA and OKT6 antibodies. Epidermal Langerhans cells were reliably demonstrated only by the enzyme histochemical method for adenosine triphosphatase, while the majority of mononuclear cells in dermal infiltrates showed a strong immunoreaction with OKT6 antibody. The histologic and histochemical analysis showed that the dermal infiltrations in infested skin consisted of macrophages, lymphocytes, granulocytes, mastocytes and fibroblasts. The chamois skin affected with sarcoptes mange showed a significant loss of cytokeratins in the epidermis and its derivatives. Particular keratinocytes showing nonspecific staining with several antibodies were also described and discussed in this paper.

Vimentin- and desmin-positive cells in the moulting budgerigar (Melopsittacus undulatus) skin.

The distribution of vimentin- and desmin-positive cells in the budgerigar (Melopsittacus undulatus) dermis was investigated by means of immunohistochemical reactivity with the commercially available (Euro-Diagnostics) polyclonal antibodies. The staining pattern for vimentin in the paraffin sections was generally comparable to that in other animal species with regard to endothelial cells, vascular wall cells, muscle cells and fibroblasts. The modified Schwann cells in the inner core of the Herbst corpuscles reacted distinctly with anti-vimentin and anti-desmin. Some connective tissue cells in the superficial dermal layer, in the feather papilla and along the pulp core inside of the regenerating feathers were particularly well stained with anti-vimentin. Fibroblast-like cells of the regenerating feathers, particularly at the base of the pulp, also reacted strongly with anti-desmin. The findings were discussed with regard to references.

Combined histochemical and immunohistochemical determination of three muscle fibre types in a single section of porcine skeletal muscle.

The method of successive staining for SDH and immunohistochemical detection of fast contracting muscle fibres in pig muscle tissue has been used for demonstration of three muscle fibre types (slow oxidative, SO; fast metabolic intermediate, FI; fast glycolytic, FG). It gives basic information on contractile and metabolic muscle fibre properties in a single frozen section and can be well applied to the pale, soft and exudative (PSE) changes affecting pig muscles.