N-Acetylcysteine and α-cyano-4-hydroxycinnamic acid alter protein kinase C (PKC)-δ and PKC-ζ and diminish dysmorphogenesis in rat embryos cultured with high glucose in vitro

Abstract

Malformations and growth disturbances are two-to threefold more common in infants of diabetic mothers than in offspring of non-diabetic pregnancy. Several suggestions have emerged to explain the reasons for diabetic embryopathy, including enhanced mitochondrial production of reactive oxygen species leading to altered activation of protein kinase C. This study aimed to evaluate the effect of a-cyano-4-hydroxycinnamic acid (CHC) and N-acetylcysteine (NAC) addition on morphology and activity of protein kinase C-d and protein kinase C-z in rat embryos exposed to a high glucose concentration in vitro. Day 9 embryos from normal rats were cultured in 10 or 30 mM glucose concentrations with or without supplementation of CHC, NAC, or protein kinase C inhibitors specific for protein kinase C-d and protein kinase C-z. Embryos were evaluated for malformations, crown rump length, and somite number. Protein kinase C-d and protein kinase C-z activities were estimated by western blot by separating membranous and cytosolic fractions of the embryo. We found increased malformations and growth retardation in embryos cultured in high versus low glucose concentrations. These abnormalities were diminished when CHC and NAC or specific protein kinase C-inhibitors were added to the culture medium. The activities of embryonic protein kinase C-d and protein kinase C-z were increased in the high glucose environment after 24-h culture, but were normalized by the addition of CHC and NAC as well as respective inhibitor to the culture medium. These findings suggest that mitochondrial overproduction of reactive oxygen species is involved in diabetic embryopathy. Furthermore, such overproduction may affect embryonic development, at least partly, by enhancing the activities of protein kinase C-d and protein kinase C-z.

The Monocarboxylate Transporter Inhibitor α-Cyano-4-Hydroxycinnamic Acid Disrupts Rat Lung Branching

“…This was described by Halestrap and co workers in 1974 [45], however in this study they use isolated mitochondria. On one hand, there are no studies reporting the entry of CHC into cells and, on the other hand, there is evidence showing that CHC acts only outside the cell [45,46,47,48]. Thus, we believe that effect of CHC in our model is mediated by inhibition of MCT activity at the plasma membrane.…”

Hyperglycemia-mediated activation of the hexosamine biosynthetic pathway results in myocardial apoptosis

“…We also administered 250 M ␣-cyano-4-hydroxycinnamic acid (4-OHCA) as an antioxidant as employed by other researchers (12,22). 4-OHCA is a mitochondrial pyruvate transport inhibitor, thereby attenuating the increased ROS production as a result of pyruvate-mediated overloading of the electron transport chain (13). In addition, we also assessed the effects of phosphorylation on O-GlcNAc levels and apoptosis by including experiments promoting phosphorylation and used 225 nM okadaic acid (25) to explore the "yin-yang'" relationship between phosphorylation and O-GlcNAcylation.…”

Maternal diet modulates the risk for neural tube defects in a mouse model of diabetic pregnancy

“…This is a spectrum of birth defects that include heart defects, neural tube defects, and caudal growth defects; all these serious morphogenetic defects show increased frequency in diabetic pregnancies [5–7]. The mechanisms by which these defects arise in the presence of high maternal blood glucose are being unraveled, and are thought to involve crucial changes of gene expression in the embryo [8–10] as well as the yolk sac [11], with alterations in the Wnt signaling pathway [12], protein Kinase C signaling [13, 14], Pax3-dependent [15] and p53-dependent processes [16], and the hypoxia and oxidative stress response systems [17–20]. …”