Uterine histotroph and conceptus development: III. Adrenomedullin stimulates proliferation, migration and adhesion of porcine trophectoderm cells via AKT-TSC2-MTOR cell signaling pathway.

Adrenomedullin (ADM) as a highly conserved peptide hormone has been reported to increase significantly in the uterine lumen during the peri-implantation period of pregnancy in pigs, but its functional roles in growth and development of porcine conceptus (embryonic/fetus and its extra-embryonic membranes) as well as underlying mechanisms remain largely unknown. Therefore, we conducted in vitro experiments using our established porcine trophectoderm cell line (pTr2) isolated from Day-12 porcine conceptuses to test the hypothesis that porcine ADM stimulates cell proliferation, migration and adhesion via activation of mechanistic target of rapamycin (MTOR) cell signaling pathway in pTr2 cells. Porcine ADM at 10-7 M stimulated (P < 0.05) pTr2 cell proliferation, migration and adhesion by 1.4-, 1.5- and 1.2-folds, respectively. These ADM-induced effects were abrogated (P < 0.05) by siRNA-mediated knockdown of ADM (siADM) and its shared receptor component calcitonin-receptor-like receptor (CALCRL; siCALCRL), as well as by rapamycin, the inhibitor of MTOR. Using siRNA-mediated knockdown of CALCRL coupled with Western blot analyses, ADM signaling transduction was determined in which ADM binds to CALCRL to increase phosphorylation of MTOR, its downstream effectors (4EBP1, P70S6K, and S6), and upstream regulators (AKT and TSC2). Collectively, these results suggest that porcine ADM in histotroph acts on its receptor component CALCRL to activate AKT-TSC2-MTOR, particularly MTORC1 signaling cascade, leading to elongation, migration and attachment of conceptuses.

Evaluation of a novel ELISA test using synthetic mycolic acid antigens for serodiagnosis of non-tuberculous mycobacterial (NTM) infections.

The diagnosis of non-tuberculous mycobacteria (NTM) is a particular challenge in people with cystic fibrosis. Current standard diagnostic approaches rely on serial sputum culture, which is resource demanding, dependent on patient expectoration and may be compromised by excessive decontamination, conventional bacterial overgrowth and masking by concomitant oral and nebulised antibiotics. An alternative rapid, reliable and inexpensive diagnostic method is therefore urgently needed. Serum of patients with Mycobacterium abscessus infection and chronic suppurative lung disease without NTM infection was tested against an array of novel synthetic mycolic acids, identical or similar to natural components of mycobacterial cell walls, and glycopeptidolipid (GPL)-core antigen, which has previously been investigated in Mycobacterium avium pulmonary infection. Diagnostic accuracy of individual antigens and combination of various antigens were calculated. An ELISA using individual trehalose dimycolates and GPL-core antigen was able to effectively distinguish serum from infected and non-infected individuals with a specificity of 88% and a sensitivity of up to 88%, which increased to 88% sensitivity and 93% specificity by combining several antigens in the test. These results suggest synthetic mycolic acid antigens, used individually or in combination with GPL-core antigen could be successfully used to distinguish patients with M. abscessus infection from disease controls.

Litter-of-origin traits and their association with lifetime productivity in sows and boars.

Litter-of-origin variables refer to unique characteristics of piglets between birth and weaning. They also are associated with reproductive organ development, so it seems plausible that they should influence lifetime productivity. Birthweight exhibited positive relationships with total sperm per ejaculate and total pigs produced over three parities for boars and sows, respectively. There were minimum birthweights below which the likelihood of future reproductive success was low so its use as an early selection criterion to exclude replacements with poor lifetime potential merits consideration. Colostrum is critical for normal adult reproductive physiology, and its intake can be estimated by measuring immunocrit levels after birth. Immunocrit had positive linear relationships with total pigs born alive over four parities in sows and total sperm produced per ejaculate in boars. Consequently, management strategies that enhance both the quality and quantity of colostrum consumed hold considerable potential for improving lifetime productivity. Results from both retrospective and prospective studies demonstrated that preweaning growth and weaning weight were associated positively with the development of reproductive organs, adult reproductive processes, and lifetime productivity. These should be, perhaps, the easiest to improve from a management perspective through the implementation of strategic cross-fostering, split-nursing, or supplemental feeding techniques.

Evidence for recombination variability in purebred swine populations.

This study aimed to investigate interpopulation variation due to sex, breed and age, and the intrapopulation variation in the form of genetic variance for recombination in swine. Genome-wide recombination rate and recombination occurrences (RO) were traits studied in Landrace (LR) and Large White (LW) male and female populations. Differences were found for sex, breed, sex-breed interaction, and age effects for genome-wide recombination rate and RO at one or more chromosomes. Dams were found to have a higher genome-wide recombination rate and RO at all chromosomes than sires. LW animals had higher genome-wide recombination rate and RO at seven chromosomes but lower at two chromosomes than LR individuals. The sex-breed interaction effect did not show any pattern not already observable by sex. Recombination increased with increasing parity in females, while in males no effect of age was observed. We estimated heritabilities and repeatabilities for both investigated traits and obtained the genetic correlation between male and female genome-wide recombination rate within each of the two breeds studied. Estimates of heritability and repeatability were low (h2  = 0.01-0.26; r = 0.18-0.42) for both traits in all populations. Genetic correlations were high and positive, with estimates of 0.98 and 0.94 for the LR and LW breeds, respectively. We performed a GWAS for genome-wide recombination rate independently in the four sex/breed populations. The results of the GWAS were inconsistent across the four populations with different significant genomic regions identified. The results of this study provide evidence of variability for recombination in purebred swine populations.

Food for Thought: Dietary Intervention in a Rare Cause of Severe Ventilatory Failure.

Plastic bronchitis is a rare condition characterised by endobronchial cast formation. We report the case of a 53-year-old women who deteriorated following an elective bronchoscopy procedure. She developed refractory ventilatory failure and required repeated bronchoscopy, which identified thick tenacious casts as the cause of her airway compromise. She did not respond to conventional therapies including endoscopic clearance, mucolytic therapy and nebulised tissue plasminogen activator (TPA). Total parenteral nutrition and a fat-free enteral diet were instituted while the patient was on extracorporeal membrane oxygenation (ECMO), which led to substantial improvement in her condition and demonstrated the importance of dietary strategies in this case. LEARNING POINTS: Plastic bronchitis can rarely present in adults with acute ventilatory failure and life-threatening airway obstruction.Although there are no established guidelines on management, dietary intervention (e.g., a fat-free diet) should be strongly considered as a therapeutic option.Extracorporeal membrane oxygenation (ECMO) is feasible and should be considered in plastic bronchitis with airway compromise.

Transcriptome analysis identifies genes and co-expression networks underlying heat tolerance in pigs.

BACKGROUND: Heat stress adversely affects pig growth and reproduction performance by reducing feed intake, weight gain, farrowing rate, and litter size. Heat tolerance is an important characteristic in pigs, allowing them to mitigate the negative effects of heat stress on their physiological activities. Yet, genetic variation and signaling pathways associated with the biological processes of heat-tolerant pigs are currently not fully understood. This study examined differentially expressed genes and constructed gene co-expression networks on mRNAs of pigs under different heat-stress conditions using whole transcriptomic RNA-seq analyses. Semen parameters, including total sperm number per ejaculate, motility, normal morphology rate, droplets, and rejected ejaculate rate, were measured weekly on 12 boars for two time periods: thermoneutral (January to May), and heat stress (July to October). Boars were classified into heat-tolerant (n = 6) and heat-susceptible (n = 6) groups based on the variation of their ejaculate parameters across the two periods. RNA was isolated from the blood samples collected from the thermoneutral and heat stress periods for gene expression analysis. RESULTS: Under heat stress, a total of 66 differentially expressed genes (25 down-regulated, 41 up-regulated) were identified in heat-tolerant pigs compared to themselves during the thermoneutral period. A total of 1041 differentially expressed genes (282 down-regulated, 759 up-regulated) were identified in the comparison between heat-tolerant pigs and heat-susceptible pigs under heat stress. Weighted gene co-expression network analysis detected 4 and 7 modules with genes highly associated (r > 0.50, p < 0.05) with semen quality parameters in heat-tolerant and heat-susceptible pigs under the effects of heat stress, respectively. CONCLUSION: This study utilized the sensitivity of semen to heat stress to discriminate the heat-tolerance ability of pigs. The gene expression profiles under the thermoneutral and heat stress conditions were documented in heat-tolerant and heat-susceptible boars. Findings contribute to the understanding of genes and biological mechanisms related to heat stress response in pigs and provide potential biomarkers for future investigations on the reproductive performance of pigs.

Supplementation of Maternal Diets with Docosahexaenoic Acid and Methylating Vitamins Impacts Growth and Development of Fetuses from Malnourished Gilts.

BACKGROUND: Like many species, pregnant swine mobilize and repartition body nutrient stores during extreme malnutrition to support fetal development. OBJECTIVE: The objective of this study was to model chronic human maternal malnutrition and measure effects of methylating-vitamins (MVs, containing choline, folate, B-6, B-12, and riboflavin) and docosahexaenoic acid (DHA) supplementation on fetal growth and development. METHODS: Pregnant gilts (n = 24) were either fully nourished (2.0 kg/d) with a corn-plus-isolated-soy-protein basal diet (control) supplemented with MVs and DHA or nourishment was restricted throughout gestation. Basal diet fed to malnourished gilts was reduced progressively from 50% to 70% restriction (1.0 to 0.6 kg/d) and was supplemented following a 2 (±MVs) x 2 (±DHA) factorial design. Full-term c-sections were performed to assess impacts on low and normal birth weight (LBW/NBW) fetuses (n = 238). RESULTS: Body weight gain of malnourished gilts was 10% of full-fed control dams (P < 0.05), but offspring birth weight, length, girth, and percentage of LBW fetuses were not different between treatments. The number of pigs per litter was reduced by 30% in malnourished control dams. Fetal brain weights were reduced by 7% compared to positive controls (P < 0.05). Micronutrient supplementation to malnourished dams increased fetal brain weights back to full-fed control levels. Dams with DHA produced offspring with higher DHA concentrations in brain and liver (P < 0.05). Plasma choline concentration was 4-fold higher in fetuses from unsupplemented malnourished dams (P < 0.0001). Global DNA methylation status of fetuses from restricted dams was higher than in control fetuses, including brain, liver, heart, muscle, and placenta tissues (P < 0.05). Addition of DHA increased methylation in LBW fetal brains (P < 0.05). CONCLUSIONS: Despite the mobilization of maternal stores, malnourished litters displayed reduced brain development that was fully mitigated by micronutrient supplementation. Severe maternal malnutrition increased global DNA methylation in several fetal tissues that was unaltered by choline and B-vitamin supplementation.

High mobility group A2 (HMGA2) deficiency in pigs leads to dwarfism, abnormal fetal resource allocation, and cryptorchidism.

Expression of HMGA2 is strongly associated with body size and growth in mice and humans. In mice, inactivation of one or both alleles of Hmga2 results in body-size reductions of 20% and 60%, respectively. In humans, microdeletions involving the HMGA2 locus result in short stature, suggesting the function of the HMGA2 protein is conserved among mammals. To test this hypothesis, we generated HMGA2-deficient pigs via gene editing and somatic cell nuclear transfer (SCNT). Examination of growth parameters revealed that HMGA2-/+ male and female pigs were on average 20% lighter and smaller than HMGA2+/+ matched controls (P < 0.05). HMGA2-/- boars showed significant size reduction ranging from 35 to 85% of controls depending on age (P < 0.05), and organ weights were also affected (P < 0.05). HMGA2-/+ gilts and boars exhibited normal reproductive development and fertility, while HMGA2-/- boars were sterile due to undescended testes (cryptorchidism). Crossbreeding HMGA2-/+ boars and gilts produced litters lacking the HMGA2-/- genotype. However, analysis of day (D) D40 and D78 pregnancies indicated that HMGA2-/- fetuses were present at the expected Mendelian ratio, but placental abnormalities were seen in the D78 HMGA2-/- concepti. Additionally, HMGA2-/- embryos generated by gene editing and SCNT produced multiple pregnancies and viable offspring, indicating that lack of HMGA2 is not lethal per se. Overall, our results show that the effect of HMGA2 with respect to growth regulation is highly conserved among mammals and opens up the possibility of regulating body and organ size in a variety of mammalian species including food and companion animals.

Blocking ligand occupancy of the αVß3 integrin inhibits the development of nephropathy in diabetic pigs.

Hyperglycemia stimulates secretion of αVß3 ligands from vascular cells, including endothelial cells, resulting in activation of the αVß3 integrin. This study determined whether blocking ligand occupancy of αVß3 would inhibit the development of diabetic nephropathy. Ten diabetic pigs received an F(ab)2 fragment of an antibody directed against the extracellular domain of the ß3-subunit, and 10 received a control IgG F(ab)2 for 18 weeks. Nondiabetic pigs excreted 115 ± 50 µg of protein/mg creatinine compared with control F(ab)2-treated diabetic animals (218 ± 57 µg/mg), whereas diabetic animals treated with the anti-ß3 F(ab)2 excreted 119 ± 55 µg/mg (P < .05). Mesangial volume/glomerular volume increased to 21 ± 2.4% in control-treated diabetic animals compared with 14 ± 2.8% (P < .01) in animals treated with active antibody. Diabetic animals treated with control F(ab)2 had significantly less glomerular podocin staining compared with nondiabetic animals, and this decrease was attenuated by treatment with anti-ß3 F(ab)2. Glomerular basement membrane thickness was increased in the control, F(ab)2-treated diabetic animals (212 ± 14 nm) compared with nondiabetic animals (170 ± 8.8 nm), but it was unchanged (159.9 ± 16.4 nm) in animals receiving anti-ß3 F(ab)2. Podocyte foot process width was greater in control, F(ab)2-treated, animals (502 ± 34 nm) compared with animals treated with the anti-ß3 F(ab)2 (357 ± 47 nm, P < .05). Renal ß3 tyrosine phosphorylation decreased from 13 934 ± 6437 to 6730 ± 1524 (P < .01) scanning units in the anti-ß3-treated group. We conclude that administration of an antibody that inhibits activation of the ß3-subunit of αVß3 that is induced by hyperglycemia attenuates proteinuria and early histologic changes of diabetic nephropathy, suggesting that it may have utility in preventing the progression of this disease complication.

Genetic and phenotypic variation in reproductive traits of AI boars.

The primary objective of this paper is to review our current understanding of phenotypic variation in reproductive traits of AI boars. The proportion of boars that cannot be trained for collection in commercial studs is low and differences among genetic lines are small. In contrast, there is a considerable variation in sperm production and significant differences are present among genotypes. The general pattern is for sperm numbers to increase rapidly between 9 and 13 months of age and then gradually reach a plateau. This initial period of enhanced production occurs over a longer period in some genetic lines, resulting in differences of 30 x 10(9) sperm cells or more per ejaculate. There also are genetic lines of boars that seem to have a high "heat tolerance". Decreases in sperm production during periods of high environmental temperatures average 5-7% in these lines, compared with 15-20% in others. Finally, there are boars currently being used in the industry that are capable of producing exceptional fertility results with low numbers of sperm. Unfortunately, several breeding practices common to swine AI make their routine identification difficult. Based on the phenotypic variation observed in modern terminal sire lines of AI boars, current prospects for influencing sperm production, boar fertility, and mounting behaviours through genetic selection are viewed as being good, moderate to low, and poor, respectively.