Fluctuating facial asymmetry and visual perceptive background during a tissue diagnostic histopathological

Background. Fluctuating facial asymmetry (FFA) is accentuated throughout life and has perceptual psychological implications; tissue diagnosis shows interindividual differences at first glance, for example, in the number of fixations, but no reports are available regarding the visual perceptual background in relation to individuals with less or more FFA during the tissue diagnostic task. Materials and methods. In medical students, including 13 men (SD = 19.4 years) and 8 women (SD = 18.1 years), FFA was determined as follows: n = 9 FFA. The entire population performed tissue diagnostic analysis of normal skin and skin with squamous cell carcinoma pathology from digital images to establish the duration and number of fixations and the total time taken for diagnosis. Results. Individuals with > FFA show significant differences in the visual perceptual background during diagnostic analysis of normal and pathological skin, which are magnified by the fixation duration and the number of fixations when the tissue diagnosis is pathological. Conclusion. Compared to those with lower FFA, medical students with greater FFA performing tissue diagnosis of pathological tissue have visual perceptual backgrounds characterized by less time spent in each fixation but with more fixations

Forward vs. reverse genetics: a bovine perspective based on visible and hidden phenotypes of inherited disorders

In modern cattle production, we have seen a negative trend for decades in reproduction while productivity and performance have improved. Although considered genetically complex, part of these fecundity, fertility, and rearing success issues are caused by Mendelian monogenic disorders. Traditionally, such disorders are investigated opportunistically based on their sporadic occurrence and through subsequent targeted analysis of affected individuals. This approach is called the forward genetic approach (FGA). Modern genomic technologies, such as single nucleotide polymorphism (SNP) array genotyping and whole-genome sequencing (WGS), allow for straightforward locus mapping and the identification of candidate causal variants in affected individuals or families. Nevertheless, a major drawback is the arbitrary sampling and availability of well-phenotyped individuals for research, especially for mostly invisible defects affecting fecundity, early embryonic death, and abortions. Therefore, the reverse genetic approach (RGA) is applied to screen for underlying recessive lethal or sub-lethal variants. This approach requires the availability of massive population-wide genomic data. By applying a haplotype screen for a significant deviation of the Hardy-Weinberg equilibrium, genomic regions potentially harboring candidate causal variants are identified. The subsequent generation of WGS data of haplotype carriers allows for the mining for pathogenic variants potentially causing a reduction in homozygosity. In the first part of my thesis, I present 18 successful, 1 inconclusive example, and 1 example addressing co-dominant effects of a known disorder. These FGA analyzes include heritable skin (n=7), bone (n=7), neuromuscular (n=1), eye (n=2), as well as syndromic disorders (n=3) in various European cattle breeds. Missense and frameshift variants in the IL17RA, DSP, and FA2H genes were described in three recessive genodermatoses: immunodeficiency with psoriasis-like skin alterations, syndromic ichthyosis, and ichthyosis congenita, respectively. Hypohidrotic ectodermal dysplasia was described as X-linked disorder that is associated with a gross deletion in the EDA gene. In dominant genodermatoses, a missense variant in COL5A2 was shown to lead to classical Ehlers-Danlos syndrome, an in-frame deletion in KRT5 was shown to cause epidermolysis bullosa simplex, and results of a study using an individual case of juvenile angiomatosis remained inconclusive. A recessive disorder described as hemifacial macrosomia was associated with a missense variant in LAMB1. Chondrodysplasia in a single family was shown to be caused by a de novo mutation in the bull leading to a stop-loss of the gene FGFR3. De novo mutations (missense and large deletions) in the COL2A1 and COL1A1 genes were associated with achondrogenesis type II (bulldog calf syndrome), and osteogenesis imperfecta type II, respectively. Another mutation that we found to affect bone morphology was a trisomy in chromosome 29 leading to proportional dwarfism with facial dysplasia. Congenital neuromuscular channelopathy was for the first time associated with a missense variant in KCNG1. Furthermore, a de novo missense variant in ADAMTSL4 and a recessive missense variant in CNGB3 were shown to cause congenital cataract and achromatopsia, respectively. Additionally, cases of pulmonary hypoplasia and anasarca syndrome were analyzed and shown to be caused by trisomy 20 in two unrelated calves and a recessively inherited missense variant in ADAMTS3. Moreover, the fatal syndromic disorder skeletal-cardo-enteric dysplasia was described to be caused by a de novo missense variant in MAP2K2. Finally, I investigated the effects on blood cholesterol and triglyceride levels of heterozygous carriers of the previously described APOB-related cholesterol deficiency. In the second part of my thesis, I present the outcome of the RGA in four main Swiss populations, that was validated with the SWISScow custom array. In the Brown Swiss dairy population, 72 haplotype regions showed significant depletions in homozygosity. Four of these haplotypes (BH6, BH14, BH24, and BH34) were associated with missense and nonsense variants in different genes (MARS2, MRPL55, CPT1C, and ACSL5, respectively). In the Original Braunvieh population, eight haplotype regions were identified. Candidate causal variants included a missense variant in TUBGCP5 gene associated with haplotype OH2, and a splice site frameshift variant in LIG3 gene associated with haplotype OH4. In the Holstein population, 24 haplotype regions were identified with a significant reduction of homozygosity. Subsequently, four novel candidate variants were proposed: a nonsense variant in KIR2DS1 for haplotype HH13, in-frame deletion in the genes NOTCH3 for HH21 haplotype, and RIOX1 for HH25 haplotype, and finally, a missense variant in PCDH15 for HH35 haplotype. In the Simmental population, eleven haplotype regions were detected. The haplotype SH5 was associated with a frameshift variant in DIS3 gene and the haplotypes SH8 and SH9 with missense variants in the CYP2B6 and NUBPL genes, respectively. For the breeds Brown Swiss, Original Braunvieh, and Holstein, association studies were carried out including traits describing fertility, birth, growth, and survival. Thereby most of the described mentioned haplotypes show additive effects. Regardless of the approach, all the described candidate causal variants can be used as a tool of precision diagnostics and represent a step forward towards personalized medicine in cattle. Furthermore, these variants can be easily genotyped and allow for targeted breeding to reduce the number of risk matings, which would lead to a reduction of affected animals and significant improvement in animal health and welfare

New perspectives of genetic disorders in cattle

In the last decades a negative trend in inbreeding has accompanied the evident improvement in productivity and performance of bovine domestic population, predisposing to the occurrence of recessively inherited disorders. The objectives of this thesis were: a) the study of genetic diseases applying a “forward genetic approach” (FGA); b) the estimation of the prevalence of deleterious alleles responsible for eight recessive disorders in different breeds; c) the collection of well-characterized materials in a Biobank for Bovine Genetic Disorders. The FGA allowed the identification of seven new recessive deleterious variants (Paunch calf syndrome - KDM2B; Congenital cholesterol deficiency - APOB; Ichthyosis congenita - FA2H; Hypotrichosis - KRT71; Hypotrichosis - HEPHL1; Achromatopsia - CNGB3; Hemifacial microsomia – LAMB1) and of seven new de novo dominant deleterious variants (Achondrogenesis type II - two variants in COL2A1; Osteogenesis imperfecta - COL1A1; Skeletal-cardio-enteric dysplasia - MAP2K2; Congenital neuromuscular channelopathy - KGNG1; Epidermolysis bullosa simplex - KRT5; Classical Ehlers-Danlos syndrome - COL5A2) in different breeds, associated with a large spectrum of phenotypes affecting different systems. The FGA was based on the sequence of a clinical, genealogical, gross- and/or histopathological and genomic study. In particular, a WGS trio-approach (patient, dam and sire) was applied. The prevalence of deleterious alleles was calculated for the Pseudomyotonia congenita, Paunch calf syndrome, Hemifacial microsomia, Congenital bilateral cataract, Ichthyosis congenita, Ichthyosis fetalis, Achromatopsia and Hypotrichosis. A particular concern resulted the allelic frequency of 12% for the Paunch calf syndrome in Romagnola cattle. In respect to the Biobank for Bovine Genetic Diseases, biological materials of clinical cases and their available relatives as well as controls used for the allelic frequency estimations were stored at -20 °C. Altogether, around 16.000 samples were added to the biobank

Shape Directional Asymmetry in Hindlimb Pairs among Calves (Bos Taurus)

The aim of this study was to determine paired asymmetries (right–left) in the autopods of bovine hindlimbs using geometric morphometry (GM). A total of 28 hindlimb right–left matched autopods belonging to healthy Brown Pyrenean calves were assessed. Dorsoplantar radiographs were obtained for each autopod. The bone shape was compared on right and left pairs by means of GM techniques, using a set of 15 landmarks. The results suggest that right and left distal limbs are, despite a perceived resemblance of symmetry, differently directionally developed in shape, with right hindlimbs tending to supinate (rotate outwards) and left hindlimbs tending to pronate (rotate inwards). This unevenness is probably related to the mediolateral forces’ contribution of each limb in carrying out the tasks of propulsion and control during walking, and/or a consequence of a laterality associated with a lateralized grazing posture. Our findings prompt a new reassessment of the function of each bovine hindlimb during standing and locomotion

Shape Directional Asymmetry in Hindlimb Pairs among Calves (<i>Bos Taurus</i>)

The aim of this study was to determine paired asymmetries (right–left) in the autopods of bovine hindlimbs using geometric morphometry (GM). A total of 28 hindlimb right–left matched autopods belonging to healthy Brown Pyrenean calves were assessed. Dorsoplantar radiographs were obtained for each autopod. The bone shape was compared on right and left pairs by means of GM techniques, using a set of 15 landmarks. The results suggest that right and left distal limbs are, despite a perceived resemblance of symmetry, differently directionally developed in shape, with right hindlimbs tending to supinate (rotate outwards) and left hindlimbs tending to pronate (rotate inwards). This unevenness is probably related to the mediolateral forces’ contribution of each limb in carrying out the tasks of propulsion and control during walking, and/or a consequence of a laterality associated with a lateralized grazing posture. Our findings prompt a new reassessment of the function of each bovine hindlimb during standing and locomotion

Evaluation of the ingestive behaviour of the dairy cow under two systems of rotation with slope

The ingestive behaviour of grazing animals is modulated by the vegetation characteristics, topography and the type of stocking method. This research was carried out in 2019, at the Rumipamba CADER-UCE. It aimed to evaluate the impact of two contrasting stocking methods of dairy cows grazing a pasture with an average of slope >8.5%. Four dairy cows were set to graze a 0.4 ha paddock for 5 days for continuous stocking methods, while for the electric fence methods the dairy cows were restricted to 0.2 ha and the fence was moved uphill every 3 hours, repeating this process four times a day. Cow were equipped with activity sensors for 12 h per day. The whole procedure was repeated 2 times after realizing an equalization cuts and both paddocks, a rest time of 30 days and a random reassignment of paddocks to one of the treatments. The cows showed a difference in terms of the percentage of grazing P=0.0072, being higher with the electric fence (55% of the measurement time). From rising-plate-meter estimates of available biomass along the grazing periods, we calculated despite similar forage allowances (electric fence = 48.06 kg DM/cow/d and continuous = 48.21 DM/cow/d) a higher forage intake was obtained in the electric fence treatment (17.5 kg DM/cow/d) compared the continuous stocking (15.7 kg DM/cow/d) (P=0.006). In terms of milk production animals grazing under the differences electrical fence stocking method tended (P=0.0985) to produce more milk (17.39 kg/d) than those grazing in the continuous system (15.16 kg/d) due to the influence of the slope (P=0.05), while for milk quality the protein content was higher for the electric fence (33.7 g/l) than the continuous method (30.5 g/l) (P=0.039). None of the other milk properties differed between methods (P>0.05)

Dichotomic role of NAADP/two-pore channel 2/Ca2+ signaling in regulating neural differentiation of mouse embryonic stem cells

Poster Presentation - Stem Cells and Pluripotency: abstract no. 1866The mobilization of intracellular Ca2+stores is involved in diverse cellular functions, including cell proliferation and differentiation. At least three endogenous Ca2+mobilizing messengers have been identified, including inositol trisphosphate (IP3), cyclic adenosine diphosphoribose (cADPR), and nicotinic adenine acid dinucleotide phosphate (NAADP). Similar to IP3, NAADP can mobilize calcium release in a wide variety of cell types and species, from plants to animals. Moreover, it has been previously shown that NAADP but not IP3-mediated Ca2+increases can potently induce neuronal differentiation in PC12 cells. Recently, two pore channels (TPCs) have been identified as a novel family of NAADP-gated calcium release channels in endolysosome. Therefore, it is of great interest to examine the role of TPC2 in the neural differentiation of mouse ES cells. We found that the expression of TPC2 is markedly decreased during the initial ES cell entry into neural progenitors, and the levels of TPC2 gradually rebound during the late stages of neurogenesis. Correspondingly, perturbing the NAADP signaling by TPC2 knockdown accelerates mouse ES cell differentiation into neural progenitors but inhibits these neural progenitors from committing to the final neural lineage. Interestingly, TPC2 knockdown has no effect on the differentiation of astrocytes and oligodendrocytes of mouse ES cells. Overexpression of TPC2, on the other hand, inhibits mouse ES cell from entering the neural lineage. Taken together, our data indicate that the NAADP/TPC2-mediated Ca2+signaling pathway plays a temporal and dichotomic role in modulating the neural lineage entry of ES cells; in that NAADP signaling antagonizes ES cell entry to early neural progenitors, but promotes late neural differentiation.postprin