A targeted sequencing panel identifies rare damaging variants in multiple genes in the cranial neural tube defect, anencephaly.

Neural tube defects (NTDs) affecting the brain (anencephaly) are lethal before or at birth, whereas lower spinal defects (spina bifida) may lead to lifelong neurological handicap. Collectively, NTDs rank among the most common birth defects worldwide. This study focuses on anencephaly, which despite having a similar frequency to spina bifida and being the most common type of NTD observed in mouse models, has had more limited inclusion in genetic studies. A genetic influence is strongly implicated in determining risk of NTDs and a molecular diagnosis is of fundamental importance to families both in terms of understanding the origin of the condition and for managing future pregnancies. Here we used a custom panel of 191 NTD candidate genes to screen 90 patients with cranial NTDs (n = 85 anencephaly and n = 5 craniorachischisis) with a targeted exome sequencing platform. After filtering and comparing to our in-house control exome database (N = 509), we identified 397 rare variants (minor allele frequency, MAF < 1%), 21 of which were previously unreported and predicted damaging. This included 1 frameshift (PDGFRA), 2 stop-gained (MAT1A; NOS2) and 18 missense variations. Together with evidence for oligogenic inheritance, this study provides new information on the possible genetic causation of anencephaly.

In vitro anti-LPS dose determination of ketorolac tromethamine and in vivo safety of repeated dosing in healthy horses.

Flunixin meglumine (FM) is a commonly used Nonsteroidal anti-inflammatory drug (NSAID) in horses, but clinical efficacy is often unsatisfactory. Ketorolac tromethamine (KT) demonstrates superior efficacy compared to other NSAIDs in humans, but its anti-inflammatory effects have not been investigated in the horse. Safety of repeated dosing of KT has not been evaluated. The first objective was to conduct a dose determination study to verify that a previously described dosage of KT would inhibit Lipopolysaccharide (LPS)-induced eicosanoid production in vitro, and to compare KT effects of this inhibition to those of FM. Then, a randomized crossover study was performed using nine healthy horses to evaluate plasma concentrations of KT and FM following IV administration. Administered dosages of KT and FM were 0.5 mg/kg and 1.1 mg/kg, respectively. Safety following six repeated doses of KT was assessed. Ketorolac tromethamine and FM suppressed LPS-induced Thromboxane B2 (TXB2 ) and Prostaglandin E2 (PGE2 ) production in vitro for up to 12 hr. Intravenous administration produced plasma concentrations of KT and FM similar to previous reports. No adverse effects were observed. A KT dosage of 0.5 mg/kg IV inhibited LPS-induced eicosanoids in vitro, and repeated dosing for up to 3 days appears safe in healthy horses. Investigation of in vivo anti-inflammatory and analgesic effects of KT is warranted.

Impact of rare variants in ARHGAP29 to the etiology of oral clefts: role of loss-of-function vs missense variants.

Non-syndromic cleft lip with or without cleft palate (NSCL/P) is a prevalent, complex congenital malformation. Genome-wide association studies (GWAS) on NSCL/P have consistently identified association for the 1p22 region, in which ARHGAP29 has emerged as the main candidate gene. ARHGAP29 re-sequencing studies in NSCL/P patients have identified rare variants; however, their clinical impact is still unclear. In this study we identified 10 rare variants in ARHGAP29, including five missense, one in-frame deletion, and four loss-of-function (LoF) variants, in a cohort of 188 familial NSCL/P cases. A significant mutational burden was found for LoF (Sequence Kernel Association Test, p = 0.0005) but not for missense variants in ARHGAP29, suggesting that only LoF variants contribute to the etiology of NSCL/P. Penetrance was estimated as 59%, indicating that heterozygous LoF variants in ARHGAP29 confer a moderate risk to NSCL/P. The GWAS hits in IRF6 (rs642961) and 1p22 (rs560426 and rs4147811) do not seem to contribute to the penetrance of the phenotype, based on co-segregation analysis. Our data show that rare variants leading to haploinsufficiency of ARHGAP29 represent an important etiological clefting mechanism, and genetic testing for this gene might be taken into consideration in genetic counseling of familial cases.

What's New in Old Horses? Postmortem Diagnoses in Mature and Aged Equids.

Postmortem findings in 241 equids admitted to a teaching hospital that were at least 15 years old at autopsy were reviewed (1) to determine disease prevalence, (2) to compare the cause of death (or euthanasia) in equids 15 to 19 years of age (n = 116) with that in equids ≥20 years of age (n = 125), and (3) to catalog coexisting lesions in equids with pituitary pars intermedia dysfunction (PPID). Breed and sex were evenly distributed between the age groups. Death or euthanasia was attributed to disease of the digestive system (41.5%), pituitary gland (12.9%), locomotor system (10.0%), nervous system (7.9%), cardiovascular system (4.6%), urinary system (4.6%), reproductive system (4.2%), respiratory system (4.2%), integumentary system (4.2%), lymphoid system (2.5%), liver (2.5%), or systemic neoplasia (1.2%). Nervous system disease was more common in the 15- to 19-year group; urinary tract disease was more common in the ≥20-year group. Neoplastic disease, regardless of systemic location, was the basis for death or euthanasia in 18.7% of all equids. Squamous cell carcinoma, lymphoma, and melanoma were the most common malignant neoplasms. PPID was the most common specific diagnosis, based on the postmortem presence of hyperplasia or adenoma, and was the reason for euthanasia in 47.7% of 65 equids with PPID. The most common nonpituitary causes for death or euthanasia in equids with PPID were colic, lameness, cancer, and spinal cord disease. Coexisting conditions in equids with PPID that were not considered the basis for euthanasia included neoplasms, infections, lameness, and recurrent airway obstruction.

Genome-wide methylation analysis in Silver-Russell syndrome patients.

Silver-Russell syndrome (SRS) is a clinically heterogeneous disorder characterised by severe in utero growth restriction and poor postnatal growth, body asymmetry, irregular craniofacial features and several additional minor malformations. The aetiology of SRS is complex and current evidence strongly implicates imprinted genes. Approximately, half of all patients exhibit DNA hypomethylation at the H19/IGF2 imprinted domain, and around 10% have maternal uniparental disomy of chromosome 7. We measured DNA methylation in 18 SRS patients at >485,000 CpG sites using DNA methylation microarrays. Using a novel bioinformatics methodology specifically designed to identify subsets of patients with a shared epimutation, we analysed methylation changes genome-wide as well as at known imprinted regions to identify SRS-associated epimutations. Our analysis identifies epimutations at the previously characterised domains of H19/IGF2 and at imprinted regions on chromosome 7, providing proof of principle that our methodology can detect DNA methylation changes at imprinted loci. In addition, we discovered two novel epimutations associated with SRS and located at imprinted loci previously linked to relevant mouse and human phenotypes. We identify RB1 as an additional imprinted locus associated with SRS, with a region near the RB1 differentially methylated region hypermethylated in 13/18 (~70%) patients. We also report 6/18 (~33%) patients were hypermethylated at a CpG island near the ANKRD11 gene. We do not observe consistent co-occurrence of epimutations at multiple imprinted loci in single SRS individuals. SRS is clinically heterogeneous and the absence of multiple imprinted loci epimutations reflects the heterogeneity at the molecular level. Further stratification of SRS patients by molecular phenotypes might aid the identification of disease causes.

Complications and putative risk factors for cecal or colonic surgery in dogs: 79 cases (2002-2015).

OBJECTIVES: To evaluate the complication rate, mortality rate and putative risk factors for cecal or colonic surgery in dogs. MATERIALS AND METHODS: A multi-institutional retrospective study including dogs that had undergone surgery that involved the cecum or the colon. Medical records from three referral hospitals were reviewed for patient demographics and clinical data. The association between putative risk factors and survival to discharge or complications was assessed using univariable and multivariable analysis. RESULTS: Seventy-nine dogs met the criteria to be included in this study. Fifty-five dogs had full thickness incision surgeries, while 24 dogs had partial thickness surgeries. The complication and mortality rates for full thickness and partial thickness cecal/colonic surgeries were not statistically different. The dehiscence rate of colonic anastomosis in this study was four of 47 (8.5%). On univariate analysis, performing full thickness procedures out of hours had an association with increased complications and mortality. On multivariable analysis, no factors were associated with survival to discharge or complications. There was no association of board-certified surgeon presence in surgery with complications or mortality. CLINICAL SIGNIFICANCE: The performance of full thickness cecal/colonic surgery is not associated with a statistically significant increased risk for complications or mortality compared to partial thickness procedures, with a possible increased risk of complications and mortality in full thickness procedures out of hours.

X-linked CHARGE-like Abruzzo-Erickson syndrome and classic cleft palate with ankyloglossia result from TBX22 splicing mutations.

X-linked cleft palate (CPX) is caused by mutations in the gene encoding the TBX22 transcription factor and is known to exhibit phenotypic variability, usually involving either a complete, partial or submucous cleft palate, with or without ankyloglossia. This study hypothesized a possible involvement of TBX22 in a family with X-linked, CHARGE-like Abruzzo-Erickson syndrome, of unknown etiology. The phenotype extends to additional features including sensorineural deafness and coloboma, which are suggested by the Tbx22 developmental expression pattern but not previously associated in CPX patients. A novel TBX22 splice acceptor mutation (c.593-5T>A) was identified that tracked with the phenotype in this family. A novel splice donor variant (c.767+5G>A) and a known canonical splice donor mutation (c.767+1G>A) affecting the same exon were identified in patients with classic CPX phenotypes and were comparatively analyzed using both in silico and in vitro splicing studies. All three variants were predicted to abolish normal mRNA splicing and an in vitro assay indicated that use of alternative splice sites was a likely outcome. Collectively, the data showed the functional effect of several novel intronic splice site variants but most importantly confirms that TBX22 is the gene underlying Abruzzo-Erickson syndrome, expanding the phenotypic spectrum of TBX22 mutations.

Immunohistochemical and ultrastructural features of congenital melanocytic naevus cells support a stem-cell phenotype.

BACKGROUND: Multiple congenital melanocytic naevi (CMN) in one individual are caused by somatic mosaicism for NRAS mutations; however, the lineage of the mutated cells remains uncertain. OBJECTIVES: To test the hypothesis that CMN may be derived from cutaneous stem cells. METHODS: Sixty-six CMN samples from 44 patients were stained for immunohistochemical (IHC) markers of melanocytic differentiation (TYR, TRP1, TRP2, LEF1, MITF, cKit), pluripotency (nestin, fascin, CD133, CD20, CD34), monocyte/macrophage lineage (CD68, CD163, CD14), proliferation (Ki67) and MTOR/Wnt-signalling pathway activation (pS6, ß-catenin). Semiquantitative scoring compared samples with naevus cell nesting (group 1) with those with only diffuse dermal infiltration (group 2). Transmission electron microscopy (TEM) was performed on 10 samples. RESULTS: A normal melanocyte population was seen overlying many dermal CMN. Group 1 samples were significantly more likely to express melanocytic differentiation markers than group 2, and expression decreased significantly with depth. Expression of these markers was correlated with each other, and with nestin and fascin. CD20 staining was positive in a substantial proportion and was stronger superficially. Expression of ß-catenin and pS6 was almost universal. Some samples expressed monocyte/macrophage markers. TEM revealed variable naevus cell morphology, striking macromelanosomes, double cilia and microvilli. CONCLUSIONS: Congenital melanocytic naevi development frequently coexists with normal overlying melanocyte development, leading us to hypothesize that in these cases CMN are likely to develop from a cell present in the skin independent of, or remaining after, normal melanocytic migration. IHC and TEM findings are compatible with CMN cells being of cutaneous stem-cell origin, capable of some degree of melanocytic differentiation superficially.

The T-box transcription factor gene TBX22 is mutated in X-linked cleft palate and ankyloglossia.

Formation of the secondary palate is a complex step during craniofacial development. Disturbance of the events affecting palatogenesis results in a failure of the palate to close. As a consequence of deformity, an affected child will have problems with feeding, speech, hearing, dentition and psychological development. Cleft palate occurs frequently, affecting approximately 1 in 1,500 births; it is usually considered a sporadic occurrence resulting from an interaction between genetic and environmental factors. Although several susceptibility loci have been implicated, attempts to link genetic variation to functional effects have met with little success. Cleft palate with ankyloglossia (CPX; MIM 303400) is inherited as a semidominant X-linked disorder previously described in several large families of different ethnic origins and has been the subject of several studies that localized the causative gene to Xq21 (refs. 10-13). Here we show that CPX is caused by mutations in the gene encoding the recently described T-box transcription factor TBX22 (ref. 14). Members of the T-box gene family are known to play essential roles in early vertebrate development, especially in mesoderm specification. We demonstrate that TBX22 is a major gene determinant crucial to human palatogenesis. The spectrum of nonsense, splice-site, frameshift and missense mutations we have identified in this study indicates that the cleft phenotype results from a complete loss of TBX22 function.

Adaptation of land treadmill scoring system for underwater treadmill in dogs with thoracolumbar intervertebral disc extrusion.

The underwater treadmill (UWTM) is utilized in dogs recovering from thoracolumbar intervertebral disc extrusion (TL-IVDE). Gait scoring is validated for dogs with TL-IVDE walking on the land treadmill (LT) but has not been reported for the UWTM. Our objective was to investigate if LT gait analysis could be applied to the UWTM and if non-ambulatory dogs walking unassisted on the UWTM, at a standardized water level, would be more likely to generate gait scores compared to on the LT. This was a prospective, observational study in dogs with TL-IVDE managed surigcally. At 0, 2, 4, 8 and 12 weeks post-operatively, paired video footage of dogs walking on the LT and UWTM (water level at the greater trochanter) was used to generate 0-100 stepping (SS) and coordination (regularity index, RI) scores. Scores were compared between treadmill type and over time. Twenty dogs were enrolled and seventy-eight paired recordings were available for review. Median gait scores increased over time but did not differ by treadmill type (P = 0.262 for SS, P = 0.533 for RI). Combining SS and RI, more recordings received scores of 0 for the LT (n = 58/156; 37.2 %) compared to the UWTM (n = 44/156; 28.2 %; P = 0.043). Scores of 0, at visits when there was at least movement present at multiple joints, was more common on the LT (n = 11/108; 10.2 %) compared to the UWTM (n = 2/108, 1.9 %; P = 0.026). In dogs recovering from TL-IVDE, LT-based gait scoring was feasible in dogs walking on the UWTM and might complement other gait analysis methods, especially for non-ambulatory dogs.

Epigenotype-phenotype correlations in Silver-Russell syndrome.

BACKGROUND: Silver-Russell syndrome (SRS) is characterised by intrauterine growth restriction, poor postnatal growth, relative macrocephaly, triangular face and asymmetry. Maternal uniparental disomy (mUPD) of chromosome 7 and hypomethylation of the imprinting control region (ICR) 1 on chromosome 11p15 are found in 5-10% and up to 60% of patients with SRS, respectively. As many features are non-specific, diagnosis of SRS remains difficult. Studies of patients in whom the molecular diagnosis is confirmed therefore provide valuable clinical information on the condition. METHODS: A detailed, prospective study of 64 patients with mUPD7 (n=20) or ICR1 hypomethylation (n=44) was undertaken. RESULTS AND CONCLUSIONS: The considerable overlap in clinical phenotype makes it difficult to distinguish these two molecular subgroups reliably. ICR1 hypomethylation was more likely to be scored as 'classical' SRS. Asymmetry, fifth finger clinodactyly and congenital anomalies were more commonly seen with ICR1 hypomethylation, whereas learning difficulties and referral for speech therapy were more likely with mUPD7. Myoclonus-dystonia has been reported previously in one mUPD7 patient. The authors report mild movement disorders in three further cases. No correlation was found between clinical severity and level of ICR1 hypomethylation. Use of assisted reproductive technology in association with ICR1 hypomethylation seems increased compared with the general population. ICR1 hypomethylation was also observed in affected siblings, although recurrence risk remains low in the majority of cases. Overall, a wide range of severity was observed, particularly with ICR1 hypomethylation. A low threshold for investigation of patients with features suggestive, but not typical, of SRS is therefore recommended.

Effects of a large-scale, natural sediment deposition event on plant cover in a Massachusetts salt marsh.

In mid-winter 2018, an unprecedented sediment deposition event occurred throughout portions of the Great Marsh in Massachusetts. Evaluation of this event in distinct marsh areas spanning three towns (Essex, Ipswich, and Newbury) revealed deposition covering 29.2 hectares with an average thickness of 30.1±2.1 mm measured shortly after deposition. While sediment deposition helps marshes survive sea level rise by building elevation, effects of such a large-scale deposition on New England marshes are unknown. This natural event provided an opportunity to study effects of large-scale sediment addition on plant cover and soil chemistry, with implications for marsh resilience. Sediment thickness did not differ significantly between winter and summer, indicating sediment is not eroding or compacting. The deposited sediment at each site had similar characteristics to that of the adjacent mudflat (e.g., texture, bivalve shells), suggesting that deposited materials resulted from ice rafting from adjacent flats, a natural phenomenon noted by other authors. Vegetative cover was significantly lower in plots with rafted sediment (75.6±2.3%) than sediment-free controls (93.1±1.6%) after one growing season. When sorted by sediment thickness categories, the low thickness level (1-19 mm) had significantly greater percent cover than medium (20-39 mm) and high (40-90 mm) categories. Given that sediment accretion in the Great Marsh was found to average 2.7 mm per year, the sediment thickness documented herein represents ~11 years of sediment accretion with only a 25% reduction in plant cover, suggesting this natural sediment event will likely increase long-term marsh resilience to sea level rise.

A functional haplotype variant in the TBX22 promoter is associated with cleft palate and ankyloglossia.

BACKGROUND: Mutations in the T-box transcription factor gene TBX22 are found in patients with X-linked cleft palate and ankyloglossia (CPX), and are reported in approximately 5% of all non-syndromic cleft palate patients. Clinical variability in CPX ranges from a mild or occult submucous cleft palate to a severe, complete cleft of the secondary palate. AIMS: To explore the possibility that mutations lying outside of the TBX22 coding region might contribute to the phenotype, a non-coding upstream exon and its upstream regulatory region were investigated. METHODS AND RESULTS: We sequenced 137 patients with cleft palate without coding region mutations and 295 controls. While no unique mutations were identified, seven single nucleotide polymorphisms (SNPs) were noted. These variants segregate into four distinct haplotypes. Individually, two of the SNPs associate significantly with cleft palate, as does the haplotype containing the rare allele of both SNPs. Analysis of the patient cohorts stratified for the presence of ankyloglossia significantly increases these associations. Reporter assays were used to analyse each of these haplotypes and the impact of individual SNPs. An important functional role for rs41307258 results in a decreased promoter activity of up to 50%. CONCLUSIONS: CPX-like patients harbouring this promoter haplotype are therefore associated with decreased TBX22 transcriptional activity. The risk haplotype, in concert with additional genetic and/or environmental factors, may contribute to the phenotypic variation observed and provide a novel causative mechanism for cleft palate, especially in patients with ankyloglossia.

Effect of tissue processing on assessment of endoscopic intestinal biopsies in dogs and cats.

BACKGROUND: Prior studies failed to detect significant association between hypoalbuminemia and small intestinal lesions. HYPOTHESIS: Use of pictorial templates will enhance consistency of interpathologist interpretation and identification of intestinal lesions associated with hypoalbuminemia. ANIMALS: Tissues from 62 dogs and 25 cats examined as clinical cases at 7 referral veterinary practices in 4 countries. METHODS: Retrospective, observational study. Histopathology slides from sequential cases undergoing endoscopic biopsy were examined by 4 pathologists by pictorial templates. Changes for 9 microscopic features were recorded as normal, mild, moderate or severe, and 2- and 4-point scales were tested for consistency of interpretation. Logistic regression models determined odds ratios (OR) of histologic lesions being associated with hypoalbuminemia while kappa statistics determined agreement between pathologists on histologic lesions. RESULTS: There was poor agreement (kappa = -0.013 to 0.3) between pathologists, and institution of origin of slides had effect (kappa = 1.0 for 3 of 4 lesions on slides from Institution 5) on agreement between pathologists on selected histologic features. Using 2 point as opposed to 4-point grading scale increased agreement between pathologists (maximum kappa = 0.69 using 4-point scale versus maximum kappa = 1.0 using 2-point scale). Significant association (P = .019- .04; 95% OR = 3.14-10.84) between lacteal dilation and hypoalbuminemia was found by 3 pathologists. CONCLUSIONS AND CLINICAL IMPORTANCE: Substantial inconsistency between pathologists remains despite use of pictorial template because of differences in slide processing. Distinguishing between mild and moderate lesions might be important source of the disagreement among pathologists.

Contribution of variant alleles of ABCB11 to susceptibility to intrahepatic cholestasis of pregnancy.

BACKGROUND: Intrahepatic cholestasis of pregnancy (ICP) has a complex aetiology with a significant genetic component. ABCB11 encodes the bile salt export pump (BSEP); mutations cause a spectrum of cholestatic disease, and are implicated in the aetiology of ICP. METHODS: ABCB11 variation in ICP was investigated by screening for five mutant alleles (E297G, D482G, N591S, D676Y and G855R) and the V444A polymorphism (c.1331T>C, rs2287622) in two ICP cohorts (n = 333 UK, n = 158 continental Europe), and controls (n = 261) for V444A. PCR primers were used to amplify and sequence patient and control DNA. The molecular basis for the observed phenotypes was investigated in silico by analysing the equivalent residues in the structure of the homologous bacterial transporter Sav1866. RESULTS: E297G was observed four times and D482G once. N591S was present in two patients; D676Y and G855R were not observed. The V444A polymorphism was associated with ICP (allelic analysis for C vs T: OR 1.7 (95% CI 1.4 to 2.1, p<0.001)). In addition, CC homozygotes were more likely to have ICP than TT homozygotes: OR 2.8 (95% CI 1.7 to 4.4 p<0.0001). Structural analyses suggest that E297G and D482G destabilize the protein fold of BSEP. The molecular basis of V444A and N591S was not apparent from the Sav1866 structure. CONCLUSIONS: Heterozygosity for the common ABCB11 mutations accounts for 1% of European ICP cases; these two mutants probably reduce the folding efficiency of BSEP. N591S is a recurrent mutation; however, the mechanism may be independent of protein stability or function. The V444A polymorphism is a significant risk factor for ICP in this population.

Lymphocytoid lines from persons with sex chromosome anomalies.

Permanent lymphocytoid cell lines have been established from patients with the XYY and XXY chromosome constitutions. Large amounts of these and other cell lines with genetic defects can be provided for various studies.

The genetic aetiology of Silver-Russell syndrome.

Silver-Russell syndrome (SRS MIM180860) is a disorder characterised by intrauterine and/or postnatal growth restriction and typical facies. However, the clinical picture is extremely diverse due to numerous diagnostic features reflecting a heterogeneous genetic disorder. The mode of inheritance is variable with sporadic cases also being described. Maternal uniparental disomy (mUPD) of chromosome 7 accounts for 10% of SRS cases and many candidate imprinted genes on 7 have been investigated. Chromosome 11 has moved to the forefront as the key chromosome in the aetiology, with reports of methylation defects in the H19 imprinted domain associated with the phenotype in 35-65% of SRS patients. Methylation aberrations have been described in a number of other imprinted growth related disorders such as Beckwith-Wiedmann syndrome. This review discusses these recent developments as well as the previous work on chromosome 7. Other candidate genes/chromosomal regions previously investigated are tabled.

Disruption of FOXF2 as a Likely Cause of Absent Uvula in an Egyptian Family.

This study investigated the genetic basis of an unusual autosomal dominant phenotype characterized by familial absent uvula, with a short posterior border of the soft palate, abnormal tonsillar pillars, and velopharyngeal insufficiency. Cytogenetic analysis and single-nucleotide polymorphism-based linkage analysis were investigated in a 4-generation family with 8 affected individuals. Whole exome sequencing data were overlaid, and segregation analysis identified a single missense variant, p.Q433P in the FOXF2 transcription factor, that fully segregated with the phenotype. This was found to be in linkage disequilibrium with a small 6p25.3 tandem duplication affecting FOXC1 and GMDS. Notably, the copy number imbalances of this region are commonly associated with pathologies that are not present in this family. Bioinformatic predictions with luciferase reporter studies of the FOXF2 missense variant indicated a negative impact, affecting both protein stability and transcriptional activation. Foxf 2 is expressed in the posterior mouse palate, and knockout animals develop an overt cleft palate. Since mice naturally lack the structural equivalent of the uvula, we demonstrated FOXF2 expression in the developing human uvula. Decipher also records 2 individuals with hypoplastic or bifid uvulae with copy number variants affecting FOXF2. Nevertheless, given cosegregation with the 6p25.3 duplications, we cannot rule out a combined effect of these gains and the missense variant on FOXF2 function, which may account for the rare palate phenotype observed.

Effect of clinical signs, endocrinopathies, timing of surgery, hyperlipidemia, and hyperbilirubinemia on outcome in dogs with gallbladder mucocele.

Gallbladder mucocele (GBM) is a common extra-hepatic biliary syndrome in dogs with death rates ranging from 7 to 45%. Therefore, the aim of this study was to identify the association of survival with variables that could be utilized to improve clinical decisions. A total of 1194 dogs with a gross and histopathological diagnosis of GBM were included from 41 veterinary referral hospitals in this retrospective study. Dogs with GBM that demonstrated abnormal clinical signs had significantly greater odds of death than subclinical dogs in a univariable analysis (OR, 4.2; 95% CI, 2.14-8.23; P<0.001). The multivariable model indicated that categorical variables including owner recognition of jaundice (OR, 2.12; 95% CI, 1.19-3.77; P=0.011), concurrent hyperadrenocorticism (OR 1.94; 95% CI, 1.08-3.47; P=0.026), and Pomeranian breed (OR, 2.46; 95% CI 1.10-5.50; P=0.029) were associated with increased odds of death, and vomiting was associated with decreased odds of death (OR, 0.48; 95% CI, 0.30-0.72; P=0.001). Continuous variables in the multivariable model, total serum/plasma bilirubin concentration (OR, 1.03; 95% CI, 1.01-1.04; P<0.001) and age (OR, 1.17; 95% CI, 1.08-1.26; P<0.001), were associated with increased odds of death. The clinical utility of total serum/plasma bilirubin concentration as a biomarker to predict death was poor with a sensitivity of 0.61 (95% CI, 0.54-0.69) and a specificity of 0.63 (95% CI, 0.59-0.66). This study identified several prognostic variables in dogs with GBM including total serum/plasma bilirubin concentration, age, clinical signs, concurrent hyperadrenocorticism, and the Pomeranian breed. The presence of hypothyroidism or diabetes mellitus did not impact outcome in this study.

Elevated placental expression of the imprinted PHLDA2 gene is associated with low birth weight.

The identification of genes that regulate fetal growth will help establish the reasons for intrauterine growth restriction. Most autosomal genes are expressed biallelically, but some are imprinted, expressed only from one parental allele. Imprinted genes are associated with fetal growth and development. The growth of the fetus in utero relies on effective nutrient transfer from the mother to the fetus via the placenta. Some current research on the genetic control of fetal growth has focused on genes that display imprinted expression in utero. The expression levels of four imprinted genes, the paternally expressed insulin growth factor 2 (IGF2), the mesoderm-specific transcript isoform 1 (MEST); the maternally expressed pleckstrin homology-like domain, family A, member 2 (PHLDA2); and the polymorphically imprinted insulin-like growth factor 2 (IGF2R) gene are all known to have roles in fetal growth and were studied in the placentae of 200 white European, normal term babies. The quantitative expression analysis with real-time PCR showed the maternally expressing PHLDA2 but not the paternally expressing IGF2 and MEST, nor the polymorphic maternally expressing IGF2R placental levels to have a statistically significant effect on birth weight. PHLDA2 expression levels are negatively correlated with size at birth. These data implicate PHLDA2 as an imprinted gene important in fetal growth and also as a potential marker of fetal growth.