Heterotopic Renal Autotransplantation in a Porcine Model: A Step-by-Step Protocol

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality of life when compared to dialysis. Although the last few decades have seen major improvements in patient outcomes following kidney transplantation, the increasing shortage of available organs represents a severe problem worldwide. To expand the donor pool, marginal kidney grafts recovered from extended criteria donors (ECD) or donated after circulatory death (DCD) are now accepted for transplantation. To further improve the postoperative outcome of these marginal grafts, research must focus on new therapeutic approaches such as alternative preservation techniques, immunomodulation, gene transfer, and stem cell administration.Experimental studies in animal models are the final step before newly developed techniques can be translated into clinical practice. Porcine kidney transplantation is an excellent model of human transplantation and allows investigation of novel approaches. The major advantage of the porcine model is its anatomical and physiological similarity to the human body, which facilitates the rapid translation of new findings to clinical trials. This article offers a surgical step-by-step protocol for an autotransplantation model and highlights key factors to ensure experimental success. Adequate pre- and postoperative housing, attentive anesthesia, and consistent surgical techniques result in favorable postoperative outcomes. Resection of the contralateral native kidney provides the opportunity to assess post-transplant graft function. The placement of venous and urinary catheters and the use of metabolic cages allow further detailed evaluation. For long-term follow-up studies and investigation of alternative graft preservation techniques, autotransplantation models are superior to allotransplantation models, as they avoid the confounding bias posed by rejection and immunosuppressive medication.     

Inhibition of the Arp2/3 complex impairs early embryo development of porcine parthenotes

The Arp2/3 complex, which nucleates actin filaments, comprises a stable assembly of seven-protein subunits including two actin-related proteins (Arp2 and Arp3). Previous work showed that Arp2/3 binds to the sides of actin filaments and is concentrated at the leading edges of motile cells. In the present study, we show that the Arp2/3 complex is critical for cytokinesis during early embryonic development in porcine parthenotes. The Arp2/3 complex is concentrated at the cortex of each cell at the 1-, 2-, and 4-cell stages, and at the periphery at the morula stage. The amount of Arp2/3 significantly decreased at the blastocyst stage in parthenogenetically activated porcine embryos. Inhibition of the Arp2/3 complex in the pig embryos by the Arp2/3-specific inhibitor CK666 resulted in abnormal cell division, a decrease in developmental rate and total cell numbers, and an increase in the ratio of trophectoderm cell number to inner cell mass number in blastocyst-stage embryos. In addition, 4-cell stage embryos subjected to CK666 treatment exhibited significantly decreased expression of ZGA genes (Pou5f1, Sox2, and Nanog), suggesting that the Arp2/3 complex plays an important role in early porcine embryo development. Thus, our data demonstrate that the Arp2/3 complex is required for early embryonic development in pigs and appears to regulate the expression of pluripotency genes.     

A molecularly imprinted polymer based chemiluminescence array sensor for one‐step determination of phenothiazines and benzodiazepines in pig urine

The residues of phenothiazines and benzodiazepines in foods of animal origin are dangerous to consumers. For inspection of their abuses, this study for the first time reported on the use of a chemiluminescence array sensor for the simultaneous determination of four phenothiazines and five benzodiazepines in pig urine. Two molecularly imprinted polymers were coated in different wells of a conventional 96‐well microtiter plate as the recognition reagents. After sample loading, the absorbed analytes were initiated directly by using an imidazole enhanced bis(2,4,6‐trichlorophenyl)oxalate–hydrogen peroxide system to emit light. The assay process consisted of only one sample‐loading step prior to data acquisition, so one test was finished within 10 min. The limits of detection for the nine drugs in the pig urine were in a range of 0.1 to 0.6 pg/mL, and the recoveries from the fortified blank urine samples were in a range of 80.3 to 95%. Furthermore, the sensor could be reused six times. Therefore, this sensor could be used as a simple, rapid, sensitive and reusable tool for routine screening for residues of phenothiazines and benzodiazepines in pig urine.     

Factors affecting piglet mortality during the first 24 h after the onset of parturition in large litters: effects of farrowing housing on behaviour of postpartum sows

The present study aimed to identify the factors that affect immediate (within 24 h after farrowing onset) postnatal piglet mortality in litters with hyperprolific sows, and investigate their associations with behaviour of postpartum sows in two different farrowing housing systems. A total of 30 sows were housed in: (1) CRATE (n=15): the farrowing crate closed (0.80×2.20 m) within a pen (2.50×1.70 m), and (2) OPEN (n=15): the farrowing crate open (0.80×2.20×1.80 m) within a pen (2.50×2.40 m) with a provision of 20 ls of hay in a rack. A total of 518 live born piglets, produced from the 30 sows, were used for data analyses during the first 24 h after the onset of parturition (T24). Behavioural observations of the sows were assessed via video analyses during T24. Total and crushed piglet mortality rates were higher in OPEN compared with CRATE (P<0.01, for both). During T24, the OPEN sows tended to show higher frequency of postural changes (P=0.07) and duration of standing (P=0.10), and showed higher frequencies of bar-biting (P<0.05) and piglet trapping (P<0.01), when compared with the CRATE sows. During T24, the mortality rates caused by crushing were correlated with the piglet trapping event (r=0.93, P<0.0001), postural changes (r=0.37, P<0.01), duration of standing (r=0.32, P<0.01) and frequency of bar-biting behaviour (r=0.51, P<0.01) of the sows (n=30). In conclusion, immediate postnatal piglet mortality, mainly due to crushing, may be associated with potential increases in frequency of postural changes, duration of standing and incidence of piglet trapping in postpartum sows in the open crate system with large litters.     

Economic weights of maternal and direct traits of pigs calculated by applying gene flow methods

Multiple trait selection indexes in pig breeding programmes should take into account the population structure and time delay between parent selection and expressions of traits in all production levels next to the trait impacts on economic efficiency of production systems. Gene flow procedures could be used for the correct evaluation of maternal and direct traits of pig breeds involved in breeding or crossbreeding systems. Therefore, the aim of this study was to expand a previously developed bioeconomic model and computer program to calculate the marginal economic values by including a gene flow procedure to calculate the economic weights for maternal and direct traits in pig breeds. The new program was then applied to the three-way crossbreeding system of the Czech National Programme for Pig Breeding. Using this program, the marginal economic values of traits for dam breeds Czech Large White in the dam position and Czech Landrace in the sire position, and for the sire breed Pietrain were weighted by the number of discounted gene expressions of selected parents of each breed summarised within all links of the crossbreeding system during the 8-year investment period. Economic weights calculated in this way were compared with the approximate economic weights calculated previously without a gene flow procedure. Taking into account the time delay between parent selection and trait expression (using discounting with half-year discount rates of 2% or 5%) and including more than one generation of parent progeny had little impact on the relative economic importance of maternal and direct traits of breeds involved in the evaluated three-way crossbreeding system. These results indicated that this gene-flow method could be foregone when estimating the relative economic weights of traits in pig crossbreeding systems applying artificial insemination at all production levels.     

‘Would it sell more pork?’ Pig farmers’ perceptions of Real Welfare,the welfare outcome component of their farm assurance scheme

In the UK, the pig industry is leading the way in the adoption of welfare outcome measures as part of their farm assurance scheme. The welfare outcome assessment (WOA), known as Real Welfare, is conducted by the farmers’ own veterinary surgeon. For the first time, this has allowed the pig industry to evaluate welfare by directly assessing the animal itself and to document the welfare of the UK pig industry as a whole. Farmer perspectives of the addition of a welfare outcome assessment to their farm assurance scheme have yet to be explored. Here, we investigate how the introduction of the Real Welfare protocol has been perceived by the farmers involved, what value it has (if any), whether any practical changes on farm have been a direct consequence of Real Welfare and ultimately whether they consider that the welfare of their pigs has been improved by the introduction of the Real Welfare protocol. Semi-structured interviews with 15 English pig farmers were conducted to explore their perceptions and experiences of the Real Welfare process. Our findings fall into three key areas: the lived experience of Real Welfare, on-farm changes resulting from Real Welfare and suggested improvements to the Real Welfare process as it currently stands. In all the three areas, the value farmers placed on the addition of WOA appeared to reflect their veterinary surgeon’s attitude towards the Real Welfare protocol. If the vet was engaged in the process and actively included the farmer, for example through discussion of their findings, the farmers interviewed had a greater appreciation of the benefits of Real Welfare themselves. It is recommended that future similar schemes should work with veterinary surgeons to ensure their understanding and engagement with the process, as well as identifying and promoting how the scheme will practically benefit individual farmers rather than assuming that they will be motivated to engage for the good of the industry alone. Retailers should be encouraged to use Real Welfare as a marketing tool for pig products to enhance the perceived commercial value of this protocol to farmers.     

Nurse sow strategies in the domestic pig: II. Consequences for piglet growth,suckling behaviour and sow nursing behaviour

Nurse sow strategies are used to manage large litters on commercial pig farms. However, new-born piglets transferred to nurse sows in late lactation might be compromised in terms of growth and survival. We investigated the effects of two nurse sow strategies on piglet growth, suckling behaviour and sow nursing behaviour. At 1-day post-farrowing, the four heaviest piglets from large litters were transferred to a nurse sow either 21 (1STEP21, n=9 litters) or 7 (2STEP7, n=10 litters) days into lactation. The remainder of the litter remained with their mother and was either kept intact (remain intact (RI), n=10 litters) or had some piglets cross-fostered to equalise birth weights (remain equalised (RE), n=9 litters). The 7-day-old piglets from 2STEP7 were transferred onto a sow 21 days into lactation (2STEP21, n=10 litters). The growth of new-born piglets on 1STEP21 and 2STEP7 nurse sows was initially lower than in RI litters (F_3,33.8=4.61; P<0.01), but weaning weights did not significantly differ (F_4,32.7=0.78; P>0.5). After the 1^st week of lactation, the weights and growth rates did not differ between treatments. Fighting behaviour during nursing bouts decreased over time. The frequency of fights was higher in 1STEP21 and 2STEP21 litters compared with RI litters (t₁₂₂=3.06 and t₁₂₃=3.00, respectively, P<0.05). The 2STEP21 litters had shorter nursing bouts than RI and 1STEP21 litters (t₁₀₇=−2.81 and t_81.7=2.8, respectively, P<0.05), which were more frequently terminated by 2STEP21 than RI sows (t₅₉₅=2.93; P<0.05). Transferring heaviest piglets from RI and RE litters to nurse sows reduced the percentage of teat changes during nursing bouts (RI: F_1,275=16.61; RE: F_1,308=43.59; P<0.001). In conclusion, nurse sow strategies do not appear to compromise piglet growth. However, new-born piglets transferred onto sows in late lactation experienced more competition at the udder, suggesting that the sows’ stage of lactation is of importance to how achievable nurse sow strategies are. Thus, the two-step nurse sow strategy is likely the best option (in relation to growth and suckling behaviour), as it minimises the difference between piglet age and sow stage of lactation.     

Reduced growth performance in gilt progeny is not improved by segregation from sow progeny in the grower–finisher phase

Gilt progeny (GP) are born and weaned lighter than sow progeny (SP) and tend to have higher rates of mortality and morbidity. This study quantified the lifetime growth performance differences between GP and SP and, additionally, evaluated whether segregating GP and SP in the grower–finisher period compared to mixing them within common pens reduced this variation. It was hypothesised that GP would be lighter than SP at every stage and segregation would improve growth performance of both GP and SP. All piglets born to 61 gilts (parity 1) and 47 sows (parities 2 to 7; mean 3.5 ± 0.2) were allocated to four treatments at 10 weeks of age: (i) GP housed together (GG), (ii) GP mixed (M) with SP (GM), (iii) SP housed together (SS) and (iv) SP mixed with GP (SM). The GM and SM pigs were housed together in common pens after movement into the grower–finisher facility. Individual live weight of all progeny was recorded at birth, weaning (WWT), 10 weeks of age (10WT) and sale (SWT). Individual hot carcass weight (HCW), fat depth at the head of the last rib (P2) and dressing percentage were measured at slaughter. Gilt progeny were lighter at birth (P = 0.038), weaning (P < 0.001) and through to sale (P = 0.001) than SP. Nursery and grower–finisher performance differences in GP were highly attributable to their lower WWT compared to SP (P < 0.001 when fitted as a covariate). Segregation of GP and SP increased grower–finisher average daily gain (ADG) in SP but decreased ADG and SWT in GP (P < 0.10). Segregated SP had increased average daily feed intake but only in males (P = 0.007); HCW (P < 0.001) and P2 fat depth (P = 0.055) were higher in mixed female GP, but there was no difference (P > 0.10) in female SP, or in males. In conclusion, GP were lighter at every stage than SP and differences after weaning were highly related to the lighter WWT of GP. Under the conditions of this study, overall segregation of GP and SP showed no consistent advantages in growth performance for both groups and differed significantly between males and females.     

Genetic diversity of autochthonous pig breeds analyzed by microsatellite markers and mitochondrial DNA D-loop sequence polymorphism

The evaluation of the genetic structure of autochthonous pig breeds is very important for conservation of local pig breeds and preservation of diversity. In this study, 18 microsatellite loci were used to detect genetic relationship between autochthonous pig breeds [Black Slavonian (BS), Turopolje pig (TP), and Croatian wild boar] and to determine phylogenetic relationship among Croatian autochthonous pig breeds and certain Asian and European pigs using the mitochondrial DNA (mtDNA) D-loop sequence polymorphism. Relatively high degree of genetic variation was found between the observed populations. The analysis of mtDNA showed that haplotypes of the studied pig populations are different from the other European and Chinese haplotypes. BS pigs showed some similarities with Mangalitsa and Duroc breeds. The genetic distances of TP can be explained by high degree of inbreeding during the past century. Despite the European origin of Croatian pig breeds with some impact of Chinese breeds in the past, the results of present study show that genetic diversity is still pronounced within investigated breeds. Furthermore, the genetic diversity is even more pronounced between Croatian breeds and other European and Chinese pig breeds. Thus, conservation of Croatian pig breeds will contribute to overall genetic diversity preservation of pig breeds.     

Behaviour influences cholesterol plasma levels in a pig model

Little is known about the relationship between feed intake behaviour and cholesterol levels in humans. This can be attributed to the fact that feed intake behaviour in humans is difficult to assess. The relationships between feed intake, feed efficiency and feed intake behaviour, and cholesterol and triglyceride levels were investigated at an average age of 187 days, in a pig model consisting of 202 Duroc barrows. Feed intake and feed intake behaviour were recorded individually and daily by means of an electronic identification system. Animals with high levels of total cholesterol also had high levels of high-density lipoprotein (HDL), low-density lipoprotein (LDL) cholesterol and triglycerides. Animals with high levels of HDL also had high levels of LDL and triglycerides, and animals with high levels of LDL also had high levels of triglycerides. Animals with higher BW, higher backfat thickness, higher BW gain, higher gain of backfat deposition, higher feed intake, higher residual feed intake (RFI) and higher feed intake rate had higher levels of total, HDL and LDL plasma cholesterol. Results indicate that the relationship between feed intake and cholesterol levels is a long-term relationship, while the relationship between RFI and cholesterol levels is more of a short-term nature. The relationship between intake rate and cholesterol plasma levels disappeared after correction for the amount of feed consumed. Results indicate that feed intake independent of metabolic BW, growth and fatness, i.e. 'RFI', was positively correlated with cholesterol plasma levels. This suggests that eating food over and above the maintenance and growth requirements constitutes a health risk independent of the level of fatness.