Gastrulation-stage alcohol exposure induces similar rates of craniofacial malformations in male and female C57BL/6J mice.

BACKGROUND: Prenatal alcohol exposure during gastrulation (embryonic day [E] 7 in mice, ~3rd week of human pregnancy) impairs eye, facial, and cortical development, recapitulating birth defects characteristic of Fetal Alcohol Syndrome (FAS). However, it is not known whether the prevalence or severity of craniofacial features associated with FAS is affected by biological sex. METHODS: The current study administered either alcohol (2.9 g/kg, two i.p. doses, 4 hr apart) or vehicle to pregnant C57BL/6J females on E7, prior to gonadal sex differentiation, and assessed fetal morphology at E17. RESULTS: Whereas sex did not affect fetal size in controls, alcohol-exposed females were smaller than both control females and alcohol-treated males. Alcohol exposure increased the incidence of eye defects to a similar degree in males and females. Together, these data suggest that females might be more sensitive to the general developmental effects of alcohol, but not effects specific to the craniofacies. Whole transcriptomic analysis of untreated E7 embryos found 214 differentially expressed genes in females vs. males, including those in pathways related to cilia and mitochondria, histone demethylase activity, and pluripotency. CONCLUSION: Gastrulation-stage alcohol induces craniofacial malformations in male and female mouse fetuses at similar rates and severity, though growth deficits are more prevalent females. These findings support the investigation of biological sex as a contributing factor in prenatal alcohol studies.

The pro-apoptotic Bax gene modifies susceptibility to craniofacial dysmorphology following gastrulation-stage alcohol exposure.

BACKGROUND: During early development, alcohol exposure causes apoptotic cell death in discrete regions of the embryo which are associated with distinctive patterns of later-life abnormalities. In gastrulation, which occurs during the third week of human pregnancy, alcohol targets the ectoderm, the precursor of the eyes, face, and brain. This midline tissue loss leads to the craniofacial dysmorphologies, such as microphthalmia and a smooth philtrum, which define fetal alcohol syndrome (FAS). An important regulator of alcohol-induced cell death is the pro-apoptotic protein Bax. The current study determines if mice lacking the Bax gene are less susceptible to the pathogenic effects of gastrulation-stage alcohol exposure. METHODS: Male and female Bax+/- mice mated to produce embryos with full (-/- ) or partial (+/- ) Bax deletions, or Bax+/+ wild-type controls. On Gestational Day 7 (GD 7), embryos received two alcohol (2.9 g/kg, 4 hr apart), or control exposures. A subset of embryos was collected 12 hr later and examined for the presence of apoptotic cell death, while others were examined on GD 17 for the presence of FAS-like facial features. RESULTS: Full Bax deletion reduced embryonic apoptotic cell death and the incidence of fetal eye and face malformations, indicating that Bax normally facilitates the development of alcohol-induced defects. An RNA-seq analysis of GD 7 Bax+/+ and Bax-/- embryos revealed 63 differentially expressed genes, some of which may interact with the Bax deletion to further protect against apoptosis. CONCLUSIONS: Overall, these experiments identify that Bax is a primary teratogenic mechanism of gastrulation-stage alcohol exposure.

Differences in [18F]FDG uptake in BAT of UCP1 -/- and UCP1 +/+ during adrenergic stimulation of non-shivering thermogenesis.

BACKGROUND: Brown adipose tissue (BAT) is a fat tissue found in most mammals that helps regulate energy balance and core body temperature through a sympathetic process known as non-shivering thermogenesis. BAT activity is commonly detected and quantified in [18F]FDG positron emission tomography/computed tomography (PET/CT) scans, and radiotracer uptake in BAT during adrenergic stimulation is often used as a surrogate measure for identifying thermogenic activity in the tissue. BAT thermogenesis is believed to be contingent upon the expression of the protein UCP1, but conflicting results have been reported in the literature concerning [18F]FDG uptake within BAT of mice with and without UCP1. Differences in animal handling techniques such as feeding status, type of anesthetic, type of BAT stimulation, and estrogen levels were identified as possible confounding variables for [18F]FDG uptake. In this study, we aimed to assess differences in BAT [18F]FDG uptake between wild-type and UCP1-knockout mice using a protocol that minimizes possible variations in BAT stimulation caused by different stress responses to mouse handling. RESULTS: [18F]FDG PET/CT scans were run on mice that were anesthetized with pentobarbital after stimulation of non-shivering thermogenesis by norepinephrine. While in wild-type mice [18F]FDG uptake in BAT increased significantly with norepinephrine stimulation of BAT, there was no consistent change in [18F]FDG uptake in BAT of mice lacking UCP1. CONCLUSIONS: [18F]FDG uptake within adrenergically stimulated BAT of wild-type and UCP1-knockout mice can significantly vary such that an [18F]FDG uptake threshold cannot be used to differentiate wild-type from UCP1-knockout mice. However, while an increase in BAT [18F]FDG uptake during adrenergic stimulation is consistently observed in wild-type mice, in UCP1-knockout mice [18F]FDG uptake in BAT seems to be independent of ß3-adrenergic stimulation of non-shivering thermogenesis.

Computed Tomographic Measures of Funnel-Shaped Lumbar Vertebral Canal and Articular Process Dysplasia Malformations Differ Between German Shepherd and Belgian Malinois Military Working Dogs.

Researchers who study the selection and breeding program criteria for military working dogs aim to help maximize the years of active duty service. Computed tomographic (CT) quantitative phenotyping has been previously described as a method for supporting these research studies. Funnel-shaped lumbar vertebral foramen malformations have been previously described in Labrador retriever military working dogs and proposed to be risk factors for impaired arterial perfusion of nerve tissues during exercise. Articular process dysplasia malformations have been previously described in varying dog breeds and proposed to be risk factors for articular process degenerative joint disease and vertebral foramen stenosis. Aims of this retrospective, cross-sectional study were to describe quantitative CT phenotyping methods for characterizing funnel-shaped lumbar vertebral foramina and articular process dysplasia malformations and to apply these methods in a comparison between groups of German shepherd and Belgian Malinois military working dogs. A military working dog hospital's database was searched for German shepherd and Belgian Malinois dogs aged <6 years that had CT scans of the lumbosacral region during the period of 2008-2016. Observers unaware of CT findings recorded available clinical data for each of the dogs. An observer unaware of clinical data recorded CT measures of funnel-shaped lumbar vertebral foramina and articular process dysplasia malformations for each of dogs and each of the lumbar vertebrae that were available in the scans. A total of 59 dogs were sampled: 41 German shepherd and 18 Belgian Malinois. Articular process dysplasia and funnel-shaped vertebral foramen phenotypic traits were present in both breeds in this sample, with the frequency and quantitative measure of these traits being greater in German shepherd dogs and heavier dogs. Lower weight dogs had a lesser degree of a funnel-shaped foramen at all sampled vertebral locations. A consistent relationship between articular process dysplasia measures and body weight was not seen. Computed tomography measures of funnel shaped vertebral foramina were greater in German shepherd vs. Belgian Malinois dogs at the L7 vertebra (P < 0.01). The CT measures of cranial articular process dysplasia were greater in German shepherd vs. Belgian Malinois dogs at the L4 (P < 0.01) and L5 (P < 0.05) vertebrae.