Oxidative stress and cellular and tissue damage in organogenic outbred mouse embryos after moderate perigestational alcohol intake.

Perigestational alcohol consumption by CF-1 mouse, from before mating up to the period of embryo organogenesis, leads to retarded early embryo development and neural tube defects. Here, we addressed if perigestational alcohol ingestion up to Day 10 of pregnancy induces oxidative stress and changes in macromolecules and organ tissues of early organogenic embryos. Adult CF-1 female mice were administered 10% ethanol in their drinking water for 17 days prior to mating and until Day 10 of gestation, whereas control females were administered ethanol-free water. Our results demonstrated significantly reduced Catalase abundance and activity and increased glutathione content in the embryos of ethanol-treated females. The nitrite level was significantly reduced, but TBARS (thiobarbituric acid reactive substances) content, an index of lipid peroxidation, did not change. Embryos derived from ethanol-treated females also showed higher abundance of 3-nitrotyrosine (3-NT)-containing proteins in all tissues, compared to the control group. Apoptosis was significantly increased in the ectoderm and mesoderm, but not in the heart-although this organ did contain more cleaved Caspase-3-positive cardiomyocytes per area of ventricular myocardium than controls. In sum, moderate perigestational alcohol ingestion up to Day 10 of gestation in mice induces oxidative stress by altering radical nitrogen species and antioxidant enzymatic and non-enzymatic mechanisms in embryos. Further, generalized protein nitration, due to unbalanced nitric oxide levels associated with tissue-specific apoptosis, was detected in embryos, suggesting that oxidative mechanisms may play an important role in the perigestational alcohol-induced malformation of organogenic embryos exposed to ethanol.

Matrix metalloproteinase expression and activity in trophoblast-decidual tissues at organogenesis in CF-1 mouse.

During early placentation, matrix metalloproteinases (MMPs) play important roles in decidualization, trophoblast migration, invasion, angiogenesis, vascularization and extracellular matrix (ECM) remodeling of the endometrium. The aim of our study was to analyze the localization, distribution and differential expression of MMP-2 and -9 in the organogenic implantation site and to evaluate in vivo and in vitro decidual MMP-2 and -9 activities on day 10 of gestation in CF-1 mouse. Whole extracts for Western blotting of organogenic E10-decidua expressed MMP-2 and -9 isoforms. MMP-2 immunoreactivity was found in a granular and discrete pattern in ECM of mesometrial decidua (MD) near maternal blood vessels and slightly in non-decidualized endometrium (NDE). Immunoexpression of MMP-9 was also detected in NDE, in cytoplasm of decidual cells and ECM of vascular MD, in trophoblastic area and in growing antimesometrial deciduum. Gelatin zymography showed that MMP-9 activity was significantly lower in CM compared to the active form of direct (not cultured) and cultured decidua. The decidual active MMP-9 was significantly higher than the active MMP-2. These results show differential localization, protein expression and enzymatic activation of MMPs, suggesting specific roles for MMP-2 and MMP-9 in decidual and trophoblast tissues related to organogenic ECM remodeling and vascularization during early establishment of mouse placentation.

Embryo developmental disruption during organogenesis produced by CF-1 murine periconceptional alcohol consumption.

The aim was to study the control females (CF)-1 mouse embryo differentiation, growth, morphology on embryonic E- and N-cadherin expression at midgestation after periconceptional moderate alcohol ingestion. Adult female mice were exposed to 10% ethanol in drinking water for 17 days previous to and up to day 10 of gestation (ethanol-exposed females, EF) and were compared with nonexposed CF. EF presented reduced quantities of E10 to E10.5 embryos, greater percentage of embryos at stages less than E7.5, reduced implantation site numbers/female, and increased resorptions compared with CF. EF-embryo growth was significantly affected as evidenced by reduced cephalic and body sizes of E10 and E10.5 embryos (scanning electron microscopy) and decreased protein content of E10.5 embryos vs. CF embryos. A significantly higher percentage of EF-E10-10.5 embryos presented abnormal neural tube (NT) closure vs. the percentage of CF. E10 embryos from EF presented elevated tissue disorganization, pyknosis and nuclear condensation in somites, mesenchymal and neuroepithelial tissue. Immunohistochemical E- and N-cadherin distribution patterns were similar in organic structures of E10 embryos between groups. However, western blot revealed that E- and N-cadherin expression levels were significantly increased in EF-derived embryos vs. controls. Perigestational ethanol consumption by CF-1 mice induced significant damage in the organogenic embryogenesis by producing delayed differentiation, growth deficiencies, and increasing the frequency of NT defects. Ethanol exposure may disrupt cell-cell adhesion leading to upregulation of E- and N-cadherin expression suggesting that deregulation of cell adhesion molecules could be involved in the disruption of embryo development at organogenesis in CF-1 mouse.