PubMed Health⌕ Search

Biomedical subjects

Kathleen K Sulik

Publications and source records attributed to Kathleen K Sulik.

11 recordsLinked to original sources
PubMed2007

Reprogramming of genetic networks during initiation of the Fetal Alcohol Syndrome.

Fetal Alcohol Spectrum Disorders (FASD) are birth defects that result from maternal alcohol use. We used a non a priori approach to prioritize candidate pathways during alcohol-induced teratogenicity in early mouse embryos. Two C57BL/6 substrains (B6J, B6N) served as the basis for study. Dosing pregnant dams with alcohol (2x 2.9 g/kg ethanol spaced 4 hr on day 8) induced FASD in B6J at a higher incidence than B6N embryos. Counter-exposure to PK11195 (4 mg/kg) significantly protected B6J embryos but slightly promoted FASD in B6N embryos. Microarray transcript profiling was performed on the embryonic headfold 3 hr after the first maternal alcohol injection (GEO data series accession GSE1074). This analysis revealed metabolic and cellular reprogramming that was substrain-specific and/or PK11195-dependent. Mapping ethanol-responsive KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways revealed down-regulation of ribosomal proteins and proteasome, and up-regulation of glycolysis and pentose phosphate pathway in B6N embryos; and significant up-regulation of tight junction, focal adhesion, adherens junction, and regulation of the actin cytoskeleton (and near-significant up-regulation of Wnt signaling and apoptosis) pathways in both substrains. Expression networks constructed computationally from these altered genes identified entry points for EtOH at several hubs (MAPK1, ALDH3A2, CD14, PFKM, TNFRSF1A, RPS6, IGF1, EGFR, PTEN) and for PK11195 at AKT1. Our findings are consistent with the growing view that developmental exposure to alcohol alters common signaling pathways linking receptor activation to cytoskeletal reorganization. The programmatic shift in cell motility and metabolic capacity further implies cell signals and responses that are integrated by the mitochondrial recognition site for PK11195.

Analysis of Variance↗
PubMed2006

Novel role for the orphan nuclear receptor Dax1 in embryogenesis, different from steroidogenesis.

Cytomegalic adrenal hypoplasia congenita (AHC) is an X-linked disease caused by mutations in DAX1-encoding gene NR0B1, previously thought to function primarily in steroidogenesis. We sought to determine the expression pattern for Dax1 along with known network partners in early embryogenesis and to determine a steroidogenic capacity for the embryo prior to the establishment of the urogenital ridge at embryonic day 9 (E9). Here, we report that murine Dax1 is a unique marker in early embryonic development, distinguishing the extraembryonic (proximal) endoderm from the remainder of the developing embryo. We showed that Wilms tumor 1, steroidogenic factor 1, and estrogen receptor beta were expressed throughout the embryo, but the progesterone, estrogen alpha and androgen receptors, cytochrome P450 (Cyp11a1) and Nur77 were not observed in any of the embryonic layers. Lack of Cyp11A1 expression at this stage confirmed an absence of inherent steroidogenic capacity for the early embryo. The role of Nr0b1 in embryonic stem (ES) cells was investigated using siRNA knockdown, resulting in differentiation toward endoderm-like fate. Nr0b1 conditional knockout in ES cells led to differentiation, confirming our knockdown results. Our investigations suggest that Nr0b1 functions in a novel role in the maintenance of a relatively undifferentiated state. Our results further suggest that the failure of conventional murine Nr0b1 knockout attempts may be due to disregulated differentiation.

Animals↗
PubMed2006

Maternal oral intake mouse model for fetal alcohol spectrum disorders: ocular defects as a measure of effect.

BACKGROUND: This work was conducted in an effort to establish an oral intake model system in which the effects of ethanol insult that occur during early stages of embryogenesis can be easily examined and in which agents that may modulate ethanol's teratogenicity can be readily tested in vivo. The model system described utilizes the alcohol deprivation effect to obtain teratogenic levels of maternal ethanol intake on days 7 and 8 of pregnancy in C57Bl/6J mice. Ocular defects including microphthalmia and uveal coloboma, which have previously been shown to result from ethanol administered by gavage or via intraperitoneal injection on these days, served as the developmental end point for this study. The ocular defects are readily identifiable and their degree of severity is expected to correlate with concurrently developing defects of the central nervous system (CNS). METHODS: Female C57Bl/6J mice were maintained on an ethanol-containing (4.8% v/v) liquid diet for 14 days and then mated during a subsequent abstinence period. Mice were then reexposed to ethanol on days 7 and 8 of pregnancy only. Control as well as ethanol-exposed dams were killed on their 14th day of pregnancy. Fetuses were then weighed, measured for crown rump length, photographed, and analyzed for ocular abnormalities. Globe size, palpebral fissure length, and pupil size and shape were noted for both the right and left eyes of all fetuses and informative comparisons were made. RESULTS: This exposure paradigm resulted in peak maternal blood alcohol concentrations that ranged from 170 to 220 mg/dL on gestational day (GD) 8. Compared with the GD 14 fetuses from the normal control group, the pair-fed, acquisition controls, as well as the ethanol-exposed fetuses, were developmentally delayed and had reduced weights. Confirming previous studies, comparison of similarly staged control and treated GD 8 embryos illustrated reductions in the size of the forebrain in the latter. Subsequent ocular malformations were noted in 33% of the right eyes and 25% of the left eyes of the 103 GD 14 ethanol-exposed fetuses examined. This incidence of defects is twice that observed in the control groups. Additionally, it was found that the palpebral fissure length is directly correlated with globe size. CONCLUSIONS: The high incidence of readily identifiable ocular malformations produced by oral ethanol intake in this model and their relevance to human fetal alcohol spectrum disorders (FASD) makes this an excellent system for utilization in experiments involving factors administered to the embryo that might alter ethanol's teratogenic effects. Additionally, the fact that early ethanol insult yields ocular and forebrain abnormalities that are developmentally associated allows efficient specimen selection for subsequent detailed analyses of CNS effects in this in vivo mammalian FASD model.

Abnormalities, Drug-Induced↗
PubMed2005

Genesis of alcohol-induced craniofacial dysmorphism.

The initial diagnosis of fetal alcohol syndrome (FAS) in the United States was made because of the facial features common to the first cohort of patients. This article reviews the development of an FAS mouse model whose craniofacial features are remarkably similar to those of affected humans. The model is based on short-term maternal treatment with a high dosage of ethanol at stages of pregnancy that are equivalent to Weeks 3 and 4 of human gestation. At these early stages of development, alcohol's insult to the developing face is concurrent with that to the brain, eyes, and inner ear. That facial and central nervous system defects consistent with FAS can be induced by more "realistic" alcohol dosages as illustrated with data from an oral alcohol intake mouse model in which maternal blood alcohol levels do not exceed 200 mg/dl. The ethanol-induced pathogenesis involves apoptosis that occurs within 12 hrs of alcohol exposure in selected cell populations of Day 7, 8, and 9 mouse embryos. Experimental evidence from other species also shows that apoptosis underlies ethanol-induced malformations. With knowledge of sensitive and resistant cell populations at specific developmental stages, studies designed to identify the basis for these differing cellular responses and, therefore, to determine the primary mechanisms of ethanol's teratogenesis are possible. For example, microarray comparisons of sensitive and resistant embryonic cell populations have been made, as have in situ studies of gene expression patterns in the populations of interest. Studies that illustrate agents that are effective in diminishing or exacerbating ethanol's teratogenesis have also been helpful in determining mechanisms. Among these agents are antioxidants, sonic hedgehog protein, retinoids, and the peptides SAL and NAP.

Alcohol Drinking↗
PubMed2004

Protection from ethanol-induced limb malformations by the superoxide dismutase/catalase mimetic, EUK-134.

Based on previous in vitro studies that have illustrated prevention of ethanol-induced cell death by antioxidants, using an in vivo model, we have tested the anti-teratogenic potential of a potent synthetic superoxide dismutase plus catalase mimetic, EUK-134. The developing limb of C57BL/6J mice, which is sensitive to ethanol-induced reduction defects, served as the model system. On their ninth day of pregnancy, C57BL/6J mice were administered ethanol (two intraperitoneal doses of 2.9 g/kg given 4 h apart) alone or in combination with EUK-134 (two doses of 10 mg/kg). Pregnant control mice were similarly treated with either vehicle or EUK-134, alone. Within 15 h of the initial ethanol exposure, excessive apoptotic cell death was observed in the apical ectodermal ridge (AER) of the newly forming forelimb buds. Forelimb defects, including postaxial ectrodactyly, metacarpal, and ulnar deficiencies, occurred in 67.3% of the ethanol-exposed fetuses that were examined at 18 days of gestation. The right forelimbs were preferentially affected. No limb malformations were observed in control fetuses. Cell death in the AER of embryos concurrently exposed to ethanol and EUK-134 was notably reduced compared with that in embryos from ethanol-treated dams. Additionally, the antioxidant treatment reduced the incidence of forelimb malformations to 35.9%. This work illustrates that antioxidants can significantly improve the adverse developmental outcome that results from ethanol exposure in utero, diminishing the incidence and severity of major malformations that result from exposure to this important human teratogen.

Animals↗
PubMed2004

Ethanol antagonist peptides: structural specificity without stereospecificity.

Increasing evidence suggests that ethanol damages the developing nervous system partly by disrupting the L1 cell adhesion molecule. Ethanol inhibits L1-mediated cell adhesion, and compounds that antagonize this action also prevent ethanol-induced embryotoxicity. Two such compounds are the small peptides NAPVSIPQ (NAP) and SALLRSIPA (SAL). We showed previously that NAP and SAL antagonize ethanol inhibition of L1 adhesion at femtomolar to picomolar concentrations. Here we demonstrate that, despite this extraordinary potency, both NAP and SAL lack stereospecificity. d-NAP, a peptide composed entirely of d-amino acids, was an effective ethanol antagonist in NIH/3T3 cells transfected with human L1 and in the NG108-15 neural cell line. Interestingly, Ala-substituted derivatives of d-NAP demonstrate the same structure-activity relation as the corresponding derivatives of l-NAP. The Ser-Ile-Pro motif was important for the ethanol antagonist activity of d-NAP, l-NAP, and l-SAL, with Ile being the most critical element in all three. Like l-NAP, d-NAP effectively reduced ethanol-induced growth retardation in mouse whole embryo culture. The potential resistance of d-peptides to proteases makes d-NAP a potentially attractive agent for the prevention of fetal alcohol syndrome.

3T3 Cells↗
PubMed2003

Differential effects of ethanol antagonism and neuroprotection in peptide fragment NAPVSIPQ prevention of ethanol-induced developmental toxicity.

NAPVSIPQ (NAP), an active fragment of the glial-derived activity-dependent neuroprotective protein, is protective at femtomolar concentrations against a wide array of neural insults and prevents ethanol-induced fetal wastage and growth retardation in mice. NAP also antagonizes ethanol inhibition of L1-mediated cell adhesion (ethanol antagonism). We performed an Ala scanning substitution of NAP to determine the role of ethanol antagonism and neuroprotection in NAP prevention of ethanol embryotoxicity. The Ser-Ile-Pro region of NAP was crucial for both ethanol antagonism and protection of cortical neurons from tetrodotoxin toxicity (neuroprotection). Ala replacement of either Ser-5 or Pro-7 (P7A-NAP) abolished NAP neuroprotection but minimally changed the efficacy of NAP ethanol antagonism. In contrast, Ala replacement of Ile-6 (I6A-NAP) caused a decrease in potency (>2 logarithmic orders) with only a small reduction (<10%) in the efficacy of NAP neuroprotection but markedly reduced the efficacy (50%) and the potency (5 logarithmic orders) of NAP ethanol antagonism. Ethanol significantly reduced the number of paired somites in mouse whole-embryo culture; this effect was prevented significantly by 100 pM NAP or by 100 pM P7A-NAP, but not by 100 pM I6A-NAP. The structure-activity relation for NAP prevention of ethanol embryotoxicity was similar to that for NAP ethanol antagonism and different from that for NAP neuroprotection. These findings support the hypothesis that NAP antagonism of ethanol inhibition of L1 adhesion plays a central role in NAP prevention of ethanol embryotoxicity and highlight the potential importance of ethanol effects on L1 in the pathophysiology of fetal alcohol syndrome.

Abnormalities, Drug-Induced↗
PubMed2002

Hindbrain and cranial nerve dysmorphogenesis result from acute maternal ethanol administration.

Acute exposure of mouse embryos to ethanol during stages of hindbrain segmentation results in excessive cell death in specific cell populations. This study details the ethanol-induced cell loss and defines the subsequent effects of this early insult on rhombomere and cranial nerve development. Ethanol at a teratogenic dosage (2.9 g/kg) or a comparable volume of vehicle was administered in each of two intraperitoneal injections to pregnant C57BL/6J mice on gestational day (GD) 8, 8 h, and GD 8, 12 h (defined hereafter as GD 8.5). Ethanol-exposed GD 9 embryos, visualized in three dimensions using laser scanning confocal microscopy of LysoTracker Red fluorescence or Nile blue sulphate vital staining, displayed excessive apoptosis in the rostral hindbrain, specifically within rhombomeres 1-3, as well as in cranial neural crest cells and ectodermal placodes. Comparably treated embryos examined on GD 10.5-11 illustrated a disproportionate reduction in the length of the rostral hindbrain. Examination of plastic histological sections of GD 9 embryos and via scanning electron microscopy on GD 10 revealed deficiencies in the hindbrain, with a phenotype including abnormal rhombomere segmentation and an extremely small fourth ventricular roofplate. Whole-mount antineurofilament immunohistochemistry on GD 10.5 and GD 11 illustrated a variety of cranial nerve abnormalities ranging from fused or absent ganglia to ectopic or disorganized fibers. In addition, a delay in the development of the glossopharyngeal (IX) nerve/ganglia complex was observed. These hindbrain and cranial nerve abnormalities are discussed in the context of the genesis of human alcohol-related birth defects and neurodevelopmental disorder.

Alcohol-Induced Disorders, Nervous System↗
PubMed2002

Distinct requirements for extra-embryonic and embryonic bone morphogenetic protein 4 in the formation of the node and primitive streak and coordination of left-right asymmetry in the mouse.

In the mouse and chick embryo, the node plays a central role in generating left-right (LR) positional information. Using several different strategies, we provide evidence in the mouse that bone morphogenetic protein 4 (Bmp4) is required independently in two different sites for node morphogenesis and for LR patterning. Bmp4 expression in the trophoblast-derived extra-embryonic ectoderm is essential for the normal formation of the node and primitive streak. However, tetraploid chimera analysis demonstrates that Bmp4 made in epiblast-derived tissues is required for robust LR patterning, even when normal node morphology is restored. In the absence of embryonic Bmp4, the expression of left-side determinants such as Nodal and Lefty2 is absent in the left lateral plate mesoderm (LPM). Noggin-mediated inhibition of Bmp activity in cultured wild-type embryos results in suppression of Nodal expression in the LPM. Thus, unlike previous models proposed in the chick embryo in which Bmp4 suppresses left-sided gene expression, our results suggest that Bmp acts as a positive facilitator of the left-sided molecular cascade and is required for Nodal induction and maintenance in the left LPM.

Animals↗
PubMedDate not supplied

Peptide-mediated protection from ethanol-induced neural tube defects.

Ethanol inhibition of L1-mediated cell adhesion may contribute to the spectrum of neurological, behavioral and morphological abnormalities associated with prenatal ethanol exposure. We showed previously that the neuroprotective peptides NAPVSIPQ (NAP) and SALLRSIPA (SAL) antagonize ethanol inhibition of L1 adhesion and prevent ethanol-induced growth retardation in mouse whole embryo culture. Here we ask whether NAP and SAL also prevent ethanol-induced major malformations of the nervous system. Gestational day 8.0 (3-5 somites) C57BL/6J mouse embryos were grown for 6 h in control medium, 100 mM ethanol and 10(-10) M peptides and then maintained for an additional 20 h in control medium. At the end of the culture period, only embryos having 18-19 somite pairs were examined and compared for the degree of neural tube closure. Ethanol exposure resulted in neural tube defects (NTDs) consistent with total dysraphia and anencephaly. Co-incubation with ethanol and L-NAP (all L-amino acids), D-NAP (all D-amino acids) or SAL significantly increased the percentage of embryos that had begun to close their neural folds at the level of the forebrain/midbrain junction or that had progressed beyond this stage of closure. P7A-NAP (NAPVSIAQ), which lacks neuroprotective activity, but retains activity as an antagonist of ethanol inhibition of L1 adhesion, was effective in preventing ethanol-induced NTDs. In contrast, I6A-NAP (NAPVSAPQ), which shows reduced efficacy as an ethanol antagonist but retains its neuroprotective efficacy, did not significantly diminish the induction of NTDs by ethanol. These findings demonstrate the ability of NAP and SAL to prevent ethanol-induced NTDs and support the hypothesis that ethanol teratogenesis is caused in part by ethanol inhibition of L1-mediated cell adhesion.

Animals↗