Developmental expression of paraoxonase 2

Abstract

Paraoxonase 2 (PON2) is a member of the paraoxonase gene family also comprising PON1 and PON3. PON2 functions as a lactonase and exhibits anti-bacterial as well as antioxidant properties. At the cellular level, PON2 localizes to the mitochondrial and endoplasmic reticulum membranes where it scavenges reactive oxygen species. PON2 is of particular interest as it is the only paraoxonase expressed in brain tissue and appears to play a critical role in mitigating oxidative stress in the brain. The aim of this study was to investigate the expression of PON2 at the protein and mRNA level in the brain and liver of mice through development to identify potential age windows of susceptibility to oxidative stress, as well as to compare expression of hepatic PON2 to expression of PON1 and PON3. Overall, PON2 expression in the brain was lower in neonatal mice and increased with age up to postnatal day (PND) 21, with a significant decrease observed at PND 30 and 60. In contrast, the liver showed continuously increasing levels of PON2 with age, similar to the patterns of PON1 and PON3. PON2 protein levels were also investigated in brain samples from non-human primates, with PON2 increasing with age up to the infant stage and decreasing at the juvenile stage, mirroring the results observed in the mouse brain. These variable expression levels of PON2 suggest that neonatal and young adult animals may be more susceptible to neurological insult by oxidants due to lower levels of PON2 in the brain.

Keywords: Brain; Development; Liver; Oxidative stress; Paraoxonase 1; Paraoxonase 2; Paraoxonase 3.

Figure 1. PON2 protein levels in whole brain of mice

A. Quantification of PON2 protein in whole brain of female and male PND 21 mice after normalization to β-actin. Results show the means (± SE) of 3 animals/sex. Significantly different from male, *** p < 0.001. (Student’s t-test). B. Western blots of the same brain samples. PON2 is seen at ~43kDa.

Figure 2. PON2 protein and mRNA levels in brain of female mice during postnatal development

A. Quantification of PON2 protein after normalization to total protein. Results are expressed as percentage of PND 1 optical density (mean ± SE, with n = 4). a, b, c: significantly different from PND 1, PND 30, and PND 60, respectively (all p < 0.01). B. Levels of PON2 mRNA normalized for β-actin. Results represent the mean (± SE) of 3–4 mice. Significantly different from PND 1, * p < 0.05.

Figure 3. PON2 protein levels in brain of non-human primates

A. Quantification of PON2 protein in brain (ventral caudate) of African green monkeys, after normalization to β-actin. Results represent the mean (± SE) with n = 5 (midgestation, late-gestation, infant and aged) 8 (juvenile) and 11 (young adult). Samples are mixed gender. Infant and juvenile stage significantly different, ** p < 0.01 B. Quantification of PON2 protein in brain (ventral caudate) of male and female young-adult monkeys after normalization to β-actin. Results show the mean (± SE) of 4–6 samples for each group. Significantly different from male, * p < 0.05 (Student t-test). C. Representative Western blot of monkey brain samples. PON2 is seen at ~43kDa.

Figure 4. Developmental expression of PON2 protein and mRNA levels in female mouse liver

A. Quantification of PON2 protein after normalization to β-actin. Results are expressed as percentage of PND 1 optical density, and represent the mean (± SE) of 5 mice/group. Significantly different from PND 1, * p < 0.05. B. Levels of PON2 mRNA in liver from female mice of different ages, corrected for β-actin. Results are the mean (± SE) of 3 animals/group.

Figure 5. Developmental expression of PON1 protein and mRNA levels in female mouse liver

A. Quantification of PON1 protein after normalization to β-actin. Results are expressed as percentage of PND 1 optical density, and represent the mean (± SE) of 5 mice/group. B. Levels of PON1 mRNA in liver from female mice of different ages, corrected for β-actin. Results are the mean (± SE) of 3–4 animals/group except for PND 21 in which n = 2 and error bar denotes range. a, b, c: significantly different from PND 1, PND 7, and PND 14 (all, p < 0.05).

Figure 6. Developmental expression of PON3 protein and mRNA levels in female mouse liver

A. Quantification of PON3 protein after normalization to β-actin. Results are expressed as percentage of PND 1 optical density, and represent the mean (± SE) of 4 mice/group. Significantly different from PND 1,* p < 0.05. B. Levels of PON3 mRNA in liver from female mice of different ages, corrected for β-actin. Results are the mean (± SE) of 4 animals/group except for PND 21 in which n = 2 and error bar denotes range. a, b: significantly different from PND 7 (p < 0.01), and PND 14 (p < 0.05), respectively.