Sequenced-based GWAS for linear classification traits in Belgian Blue beef cattle reveals new coding variants in genes regulating body size in mammals.

BACKGROUND: Cohorts of individuals that have been genotyped and phenotyped for genomic selection programs offer the opportunity to better understand genetic variation associated with complex traits. Here, we performed an association study for traits related to body size and muscular development in intensively selected beef cattle. We leveraged multiple trait information to refine and interpret the significant associations. RESULTS: After a multiple-step genotype imputation to the sequence-level for 14,762 Belgian Blue beef (BBB) cows, we performed a genome-wide association study (GWAS) for 11 traits related to muscular development and body size. The 37 identified genome-wide significant quantitative trait loci (QTL) could be condensed in 11 unique QTL regions based on their position. Evidence for pleiotropic effects was found in most of these regions (e.g., correlated association signals, overlap between credible sets (CS) of candidate variants). Thus, we applied a multiple-trait approach to combine information from different traits to refine the CS. In several QTL regions, we identified strong candidate genes known to be related to growth and height in other species such as LCORL-NCAPG or CCND2. For some of these genes, relevant candidate variants were identified in the CS, including three new missense variants in EZH2, PAPPA2 and ADAM12, possibly two additional coding variants in LCORL, and candidate regulatory variants linked to CCND2 and ARMC12. Strikingly, four other QTL regions associated with dimension or muscular development traits were related to five (recessive) deleterious coding variants previously identified. CONCLUSIONS: Our study further supports that a set of common genes controls body size across mammalian species. In particular, we added new genes to the list of those associated with height in both humans and cattle. We also identified new strong candidate causal variants in some of these genes, strengthening the evidence of their causality. Several breed-specific recessive deleterious variants were identified in our QTL regions, probably as a result of the extreme selection for muscular development in BBB cattle.

MiRNAs differentially expressed in skeletal muscle of animals with divergent estimated breeding values for beef tenderness.

BACKGROUND: MicroRNAs (miRNAs) are small noncoding RNAs of approximately 22 nucleotides, highly conserved among species, which modulate gene expression by cleaving messenger RNA target or inhibiting translation. MiRNAs are involved in the regulation of many processes including cell proliferation, differentiation, neurogenesis, angiogenesis, and apoptosis. Beef tenderness is an organoleptic characteristic of great influence in the acceptance of meat by consumers. Previous studies have shown that collagen level, marbling, apoptosis and proteolysis are among the many factors that affect beef tenderness. Considering that miRNAs can modulate gene expression, this study was designed to identify differentially expressed miRNAs that could be modulating biological processes involved with beef tenderness. RESULTS: Deep sequence analysis of miRNA libraries from longissimus thoracis muscle allowed the identification of 42 novel and 308 known miRNAs. Among the known miRNAs, seven were specifically expressed in skeletal muscle. Differential expression analysis between animals with high (H) and low (L) estimated breeding values for shear force (EBVSF) revealed bta-mir-182 and bta-mir-183 are up-regulated (q value < 0.05) in animals with L EBVSF, and bta-mir-338 is up-regulated in animals with H EBVSF. The number of bovine predicted targets for bta-mir-182, bta-mir-183 and bta-mir-338 were 811, 281 and 222, respectively, which correspond to 1204 unique target genes. Among these, four of them, MEF2C, MAP3K2, MTDH and TNRC6B were common targets of the three differentially expressed miRNAs. The functional analysis identified important pathways related to tenderness such as apoptosis and the calpain-calpastatin system. CONCLUSION: The results obtained indicate the importance of miRNAs in the regulatory mechanisms that influence muscle proteolysis and meat tenderness and contribute to our better understanding of the role of miRNAs in biological processes associated with beef tenderness.

Identification of putative regulatory regions and transcription factors associated with intramuscular fat content traits.

BACKGROUND: Integration of high throughput DNA genotyping and RNA-sequencing data allows for the identification of genomic regions that control gene expression, known as expression quantitative trait loci (eQTL), on a whole genome scale. Intramuscular fat (IMF) content and carcass composition play important roles in metabolic and physiological processes in mammals because they influence insulin sensitivity and consequently prevalence of metabolic diseases such as obesity and type 2 diabetes. However, limited information is available on the genetic variants and mechanisms associated with IMF deposition in mammals. Thus, our hypothesis was that eQTL analyses could identify putative regulatory regions and transcription factors (TFs) associated with intramuscular fat (IMF) content traits. RESULTS: We performed an integrative eQTL study in skeletal muscle to identify putative regulatory regions and factors associated with intramuscular fat content traits. Data obtained from skeletal muscle samples of 192 animals was used for association analysis between 461,466 SNPs and the transcription level of 11,808 genes. This yielded 1268 cis- and 10,334 trans-eQTLs, among which we identified nine hotspot regions that each affected the expression of > 119 genes. These putative regulatory regions overlapped with previously identified QTLs for IMF content. Three of the hotspots respectively harbored the transcription factors USF1, EGR4 and RUNX1T1, which are known to play important roles in lipid metabolism. From co-expression network analysis, we further identified modules significantly correlated with IMF content and associated with relevant processes such as fatty acid metabolism, carbohydrate metabolism and lipid metabolism. CONCLUSION: This study provides novel insights into the link between genotype and IMF content as evident from the expression level. It thereby identifies genomic regions of particular importance and associated regulatory factors. These new findings provide new knowledge about the biological processes associated with genetic variants and mechanisms associated with IMF deposition in mammals.

Lower follicular fluid vitamin D concentration is related to a higher number of large ovarian follicles.

Vitamin D receptor-knockout mice fail to produce mature oocytes, indicating vitamin D is crucial for folliculogenesis in mice. However, the actions of vitamin D during folliculogenesis remain unknown. This prospective study aimed to assess whether follicular fluid (FF) vitamin D (25OHD3) concentrations are related to specific responses to ovarian stimulation. Women undergoing ovarian stimulation for IVF participated in the study. FF 25OHD3 concentrations were assessed in the first follicle aspirate on oocyte retrieval day. Oestradiol and progesterone concentrations were assessed on the trigger day. K-means grouping analysis showed that 25OHD3 FF concentrations clustered into a higher and lower group (mean ± SEM 17.4 ± 6.61 ng/ml and 35.5 ± 7.17 ng/ml, respectively, P < 0.001). The clusters were analysed according to the oestradiol and progesterone concentrations, follicle number and size and resulting oocyte number and maturity. The FF 25OHD3 concentrations were no different among the infertility diagnoses. The lower 25OHD3 group had more follicles (≥16.0 mm, P = 0.009) and higher serum oestradiol concentrations (P < 0.03) on the day of HCG administration. In this study, lower follicular 25OHD3 concentrations predicted a better response to ovarian stimulation shown by a greater production of larger follicles and higher serum oestradiol concentrations.

Agronomic viability of New Zealand spinach and kale intercropping

ABSTRACT The intercropping is a production system that aims to provide increased yield with less environmental impact, due to greater efficiency in the use of natural resources and inputs involved in the production process. An experiment was carried out to evaluate the agronomic viability of kale and New Zealand spinach intercropping as a function of the spinach transplanting time. (0, 14, 28, 42, 56, 70, 84 and 98 days after transplanting of the kale). The total yield (TY) and yield per harvest (YH) of the kale in intercropping did not differ from those obtained in monoculture. The spinach TY was influenced by the transplanting time, the earlier the transplanting, the higher the TY. The spinach YH was not influenced by the transplanting time, but rather by the cultivation system. In intercropping, the spinach YH was 13.5% lower than in monoculture. The intercropping was agronomically feasible, since the land use efficiency index, which was not influenced by the transplanting time, had an average value of 1.71, indicating that the intercropping produced 71% more kale and spinach than the same area in monoculture. Competitiveness coefficient, aggressiveness and yield loss values showed that kale is the dominating species and spinach is the dominated.

Agronomic viability of New Zealand spinach and kale intercropping.

The intercropping is a production system that aims to provide increased yield with less environmental impact, due to greater efficiency in the use of natural resources and inputs involved in the production process. An experiment was carried out to evaluate the agronomic viability of kale and New Zealand spinach intercropping as a function of the spinach transplanting time. (0, 14, 28, 42, 56, 70, 84 and 98 days after transplanting of the kale). The total yield (TY) and yield per harvest (YH) of the kale in intercropping did not differ from those obtained in monoculture. The spinach TY was influenced by the transplanting time, the earlier the transplanting, the higher the TY. The spinach YH was not influenced by the transplanting time, but rather by the cultivation system. In intercropping, the spinach YH was 13.5% lower than in monoculture. The intercropping was agronomically feasible, since the land use efficiency index, which was not influenced by the transplanting time, had an average value of 1.71, indicating that the intercropping produced 71% more kale and spinach than the same area in monoculture. Competitiveness coefficient, aggressiveness and yield loss values showed that kale is the dominating species and spinach is the dominated.

Positive end-expiratory pressure and variable ventilation in lung-healthy rats under general anesthesia.

OBJECTIVES: Variable ventilation (VV) seems to improve respiratory function in acute lung injury and may be combined with positive end-expiratory pressure (PEEP) in order to protect the lungs even in healthy subjects. We hypothesized that VV in combination with moderate levels of PEEP reduce the deterioration of pulmonary function related to general anesthesia. Hence, we aimed at evaluating the alveolar stability and lung protection of the combination of VV at different PEEP levels. DESIGN: Randomized experimental study. SETTING: Animal research facility. SUBJECTS: Forty-nine male Wistar rats (200-270 g). INTERVENTIONS: Animals were ventilated during 2 hours with protective low tidal volume (VT) in volume control ventilation (VCV) or VV and PEEP adjusted at the level of minimum respiratory system elastance (Ers), obtained during a decremental PEEP trial subsequent to a recruitment maneuver, and 2 cmH2O above or below of this level. MEASUREMENTS AND MAIN RESULTS: Ers, gas exchange and hemodynamic variables were measured. Cytokines were determined in lung homogenate and plasma samples and left lung was used for histologic analysis and diffuse alveolar damage scoring. A progressive time-dependent increase in Ers was observed independent on ventilatory mode or PEEP level. Despite of that, the rate of increase of Ers and lung tissue IL-1 beta concentration were significantly lower in VV than in VCV at the level of the PEEP of minimum Ers. A significant increase in lung tissue cytokines (IL-6, IL-1 beta, CINC-1 and TNF-alpha) as well as a ventral to dorsal and cranial to caudal reduction in aeration was observed in all ventilated rats with no significant differences among groups. CONCLUSIONS: VV combined with PEEP adjusted at the level of the PEEP of minimal Ers seemed to better prevent anesthesia-induced atelectasis and might improve lung protection throughout general anesthesia.

Asymmetrical attenuation of vibration sensation in unilateral diabetic Charcot foot neuroarthropathy.

AIMS: To further characterize the distal sensory neuropathy in subjects with unilateral diabetic Charcot foot neuroarthropathy. METHODS: A retrospective cohort study to assess the level to which the sensory modalities of pinprick, light touch, vibration, joint position and temperature were attenuated in the affected and unaffected limbs in subjects with unilateral Charcot. The level to which the sensory modality was attenuated in each limb was assigned a score. The Wilcoxon signed rank test was used to compare the scores in the affected and unaffected limbs and also to compare the scores of the different sensory modalities in the affected and unaffected limbs. RESULTS: Fifty subjects with unilateral Charcot foot neuroarthropathy were assessed. Mean age was 45 ± SD 6 years for the 17 subjects with Type 1 diabetes and 62 ± 10 years for the 33 subjects with Type 2 diabetes. Duration of diabetes was 21 ± 13 years, HbA(1c) was 70 ± 19 mmol/mol [8.6 ± 1.8 %] and 15 subjects (30%) required renal replacement therapy. The level of attenuation of vibration sensation was more proximal in the affected compared with the unaffected limbs (P = 0.002). Pinprick, light touch, joint position and temperature sensations were not different. Joint position sensation was less attenuated bilaterally than the other sensory modalities. CONCLUSIONS: Asymmetrical attenuation of vibration sensation may predict the side that will develop a Charcot joint and may suggest a more important role for vibration sense loss than loss of other sensory modalities in the pathophysiology of Charcot.

Matching the numerator with an appropriate denominator to demonstrate low amputation incidence associated with a London hospital multidisciplinary diabetic foot clinic.

AIMS: To establish a method to assess amputation incidence that addresses the problems matching a numerator with an appropriate denominator in London and to demonstrate low amputation incidence associated with the activity of our multidisciplinary diabetic foot clinic. METHODS: Hospital-coded inpatient data was examined to derive the numerator: the number of non-traumatic amputations performed on subjects with diabetes each financial year where the Primary Care Trust commissioner code was our main local Primary Care Trust. Denominators were derived from the main local Primary Care Trust's Quality and Outcomes Framework data sets. Not all Primary Care Trust subjects with diabetes receive inpatient care at our hospital, so that the denominators were corrected for the hospital's percentage market share for the provision of inpatient diabetes care for the Primary Care Trust each financial year, derived from the Dr Foster database. RESULTS: Between April 2004 and April 2009, 44 Primary Care Trust subjects with diabetes underwent 34 minor and 10 major amputations at the hospital. Although the Primary Care Trust populations with and without diabetes increased, the hospital's Primary Care Trust percentage market share decreased, so that overall denominators decreased. The mean annual incidence of minor, major and total amputations over the five financial years was 14.7, 4.2 and 18.9 per 10 000 subjects with diabetes,respectively, and 3.9, 1.1 and 5.0 per 100 000 of the general population, respectively. CONCLUSIONS: We report for the first time amputation incidence in a London population. Acknowledging the limitations of accurately defining incidence in London, we demonstrate low amputation incidence associated with our multidisciplinary diabetic foot clinic.

Optic chiasm enhancement associated with giant aneurysm and yttrium treated pituitary adenoma.

A patient with a history of pituitary tumour treated with yttrium 29 years before presented with an asymmetrical chiasmal neuropathy. Magnetic resonance imaging showed a partially thrombosed giant aneurysm of the right internal carotid artery, with enhancement of the chiasm and right optic tract adjacent to the aneurysm. It was thought that, in addition to the effects of compression, a peri-aneurysmal inflammatory reaction had developed, causing breakdown of the blood-brain barrier and consequent inflammatory changes in the optic chiasm. High dose steroid treatment led to significant improvement in vision within two weeks. Steroids may have a role in the acute preservation of vision in similar cases, as well as in cases of deterioration following coiling or embolisation of aneurysms where thrombosis within the aneurysm has been induced.

Immunological study of hereditary motor and sensory neuropathy type 1a (HMSN1a).

OBJECTIVES: Fifty three patients were studied to investigate whether autoimmune or inflammatory mechanisms could explain the phenotypic heterogeneity of patients with hereditary motor and sensory neuropathy type 1a (HMSN1a). METHODS: Serum samples were examined for antibodies to peripheral nerve myelin protein 22 (PMP22), ganglioside GM1 and cauda equina homogenate, and interleukin-6 (IL-6) and soluble tumour necrosis factor receptor 1 (sTNF R1) concentrations. Serological results were compared with those from patients with other neuropathies (ONPs, n=30) and with normal subjects (n=51). RESULTS: In the group as a whole, no relation emerged between clinical severity and any immune parameters. Immunohistochemical examination of four sural nerve biopsies did not show significant inflammatory infiltration. In a subset of 12 patients who experienced stepwise progression of disease, there was a trend towards a higher proportion having anti-PMP22 antibodies (33% v 15% of those with gradual disease progression, 3% ONPs, and no normal controls) and complement fixing antibodies to human cauda equina (25% v 5% with gradual progression, 8.6% ONPs, 3.9% normal controls, p=0.07). CONCLUSIONS: Patients with HMSN1a and a stepwise disease progression may have an inflammatory, autoimmune component superimposed on the genetic condition.

Prospective study of the usefulness of sural nerve biopsy.

OBJECTIVE: This study aimed to determine the usefulness of sural nerve biopsy in neurological practice. METHODS: The first prospective study of sural nerve biopsy in 50 consecutive patients was undertaken. The investigating neurologist declared the prebiopsy diagnosis and management plan and after 3 months an independent neurologist evaluated the contribution of the biopsy to diagnosis and management. An independent audit officer sought information from the patient about the adverse effects and value of the biopsy after 6 weeks and 6 months. RESULTS: In seven cases the nerve biopsy changed the diagnosis, in 35 cases the biopsy confirmed the suspected diagnosis, and in eight cases the biopsy was non-contributory. The biopsy either changed or was helpful in guiding patient management in 60%, especially those with demyelinating neuropathy and multiple mononeuropathy. Seven patients reported having had infection and 10 reported increased pain at the biopsy site 6 months later. CONCLUSION: In a consecutive series of 50 cases, sural nerve biopsy altered the diagnosis in 14%, affected management in 60%, and caused persistent increased pain at the biopsy site in 33%.

Anti-PMP22 antibodies in patients with inflammatory neuropathy.

An experimental model of inflammatory radiculoneuropathy is induced by immunising rats with peripheral myelin protein 22 (PMP22). We have investigated whether PMP22 may be important in inducing human inflammatory neuropathy. We examined sera of patients with Guillain-Barré syndrome (GBS), chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), other neuropathies (ONP) and normal controls for IgM and IgG antibodies against PMP22 by ELISA (against synthetic PMP22 extracellular peptide fragments) and Western blot (against cauda equina). Antibodies were detected by both methods in 52% of patients with GBS, 35% with CIDP, 3% with ONP and no normal controls. We conclude that an immune response against PMP22 may play a role in the pathogenesis of the inflammatory neuropathies.

Vasculitic neuropathy in association with chronic graft-versus-host disease.

A 43-year-old woman is reported who developed acute and later chronic graft-versus-host disease following an unrelated donor bone marrow transplantation for chronic myeloid leukaemia. Four years later, she developed a sensory multiple mononeuropathy with biopsy features of chronic vasculitis. This is the first report of vasculitic neuropathy in association with graft-versus-host disease.

Induction of experimental autoimmune neuritis with peripheral myelin protein-22.

Two myelin proteins, P2 basic protein and P0 glycoprotein, can induce experimental autoimmune neuritis (EAN), a model of human inflammatory neuropathy. We investigated whether peripheral nerve myelin protein-22 (PMP22), the gene for which is duplicated in hereditary motor sensory neuropathy type la, can also induce EAN. PMP22 cDNA produced by the reverse transcriptase-polymerase chain reaction from rat sciatic nerve was expressed in Escherichia coli as a fusion protein with glutathione-S-transferase (GST). Ten Lewis rats were immunized with purified PMP22 fusion protein (50-100 microg) and eight controls with the same amount of GST. Two additional animals were immunized with each of two peptides (250 microg) of the human PMP22 extracellular sequences. Animals were examined daily until 20 days following immunization, when they underwent neurophysiological examination. A serum sample was then taken, prior to perfusion with glutaraldehyde and removal of the sciatic nerves and cauda equina. PMP22-immunized animals developed antibodies to the fusion protein and five out of 10 developed limp tails. No changes were observed in controls immunized with GST or in animals immunized with peptide. The mean compound motor action potentials elicited in the foot muscles by stimulation of the sciatic nerve at the sciatic notch and of the tibial nerve at the ankle were significantly reduced in the PMP22-immunized group (P < 0.05). Spinal roots from the group of animals immunized with PMP22 showed sparse infiltration of mononuclear cells, oedema and demyelination. PMP22 now deserves consideration as an autoantigen in human acute inflammatory demyelinating polyradiculoneuropathy.

Human immunoglobulin ameliorates rat experimental autoimmune neuritis.

Human immunoglobulin is an effective treatment for Guillain-Barré syndrome, although the mechanism of action is not understood. We have investigated the effect of human immunoglobulin in an animal model of Guillain-Barré syndrome, namely experimental autoimmune neuritis (EAN), induced in Lewis rats by immunization with bovine spinal root myelin. Human immunoglobulin administered intraperitoneally at the time of onset of disease accelerated the rate of recovery from EAN. This improvement was associated with a reduction in the titre of anti-rat myelin antibodies and may be due to earlier remyelination of demyelinated nerve fibres. This model may facilitate further investigation of the mechanism of therapeutic action of immunoglobulin in inflammatory neuropathy.

Treatment of inflammatory neuropathy.

Experimental autoimmune neuritis (EAN) provides an accurate model for understanding the mechanism of acute and chronic inflammatory demyelinating polyradiculoneuropathy (AIDP and CIDP). Treatments aimed at every stage of the immune process in EAN have been effective in inhibiting or treating the disease, including antibodies directed against cell adhesion molecules on the endothelium, inhibition of T cells, removal or blockade of antibodies, depletion of complement, and interference with the release or action of macrophage effector molecules. In human disease the only proven treatments are plasma exchange and intravenous immunoglobulin (IVIg) in AIDP, and either of these regimens and also corticosteroids in CIDP. However the outcome from AIDP and CIDP remains unsatisfactory with existing immunosuppressive regimens. This problem arises from the fact that while demyelination appears to be effectively and promptly repaired by remyelination, it may be accompanied by axonal degeneration which can cause severe persistent disability. In addition to limiting demyelination, it will also be important to develop strategies to protect axons from degeneration and to enhance regeneration.