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Premutation in the Fragile X Mental Retardation 1 (FMR1) Gene Affects Maternal Zn-milk and Perinatal Brain Bioenergetics and Scaffolding.

Napoli E, Ross-Inta C, Song G, Wong S, Hagerman R, Gane LW, Smilowitz JT, Tassone F, Giulivi C - Front Neurosci (2016)

Bottom Line: Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background.A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced.Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase).

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, School of Veterinary Medicine Davis, CA, USA.

ABSTRACT
Fragile X premutation alleles have 55-200 CGG repeats in the 5' UTR of the FMR1 gene. Altered zinc (Zn) homeostasis has been reported in fibroblasts from >60 years old premutation carriers, in which Zn supplementation significantly restored Zn-dependent mitochondrial protein import/processing and function. Given that mitochondria play a critical role in synaptic transmission, brain function, and cognition, we tested FMRP protein expression, brain bioenergetics, and expression of the Zn-dependent synaptic scaffolding protein SH3 and multiple ankyrin repeat domains 3 (Shank3) in a knock-in (KI) premutation mouse model with 180 CGG repeats. Mitochondrial outcomes correlated with FMRP protein expression (but not FMR1 gene expression) in KI mice and human fibroblasts from carriers of the pre- and full-mutation. Significant deficits in brain bioenergetics, Zn levels, and Shank3 protein expression were observed in the Zn-rich regions KI hippocampus and cerebellum at PND21, with some of these effects lasting into adulthood (PND210). A strong genotype × age interaction was observed for most of the outcomes tested in hippocampus and cerebellum, whereas in cortex, age played a major role. Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background. A higher gene expression of ZnT4 and ZnT6, Zn transporters abundant in brain and lactating mammary glands, was observed in the latter tissue of KI dams. A cross-fostering experiment allowed improving cortex bioenergetics in KI pups nursing on WT milk. Conversely, WT pups nursing on KI milk showed deficits in hippocampus and cerebellum bioenergetics. A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced. Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase). In premutation carriers, altered Zn homeostasis, brain bioenergetics and Shank3 levels could be compounded by Zn-deficient milk, increasing the risk of developing emotional and neurological/cognitive problems and/or FXTAS later in life.

No MeSH data available.


Related in: MedlinePlus

Brain bioenergetics of KI mice during neurodevelopment and adulthood. Mitochondria were isolated from cortex, cerebellum, and hippocampus of WT and KI pups as described in the Methods section. Activities of NADH oxidase, succinate oxidase, and cytochrome c oxidase (CCO), and respiratory control ratio (RCR) were evaluated at PND0 (cerebellum and cortex only, due to the scarcity of hippocampal tissue), PND7, PND21, and PND210. Data are reported as mean ± SEM, n = 3–7 per genotype per time point. Statistical analysis was performed by Two-way ANOVA. Post-hoc analysis performed by Tukey's HSD test revealed significant differences between WT and KI indicated by asterisks as follows NADH oxidase: *p = 0.0468; Succinate oxidase: *p = 0.0466 at PND21, *p = 0.0423 at PND210; Cytochrome c oxidase: *p = 0.0484; Coupling: **p = 0.001, ***p = 0.0004, *p = 0.0496. Letters indicate statistically significant differences amongst time points as follows. NADH oxidase: p = 0.0291 (a), p = 0.0033 (b), p = 0.0166 (c), p = 0.0320 (d), p = 0.0036 (e), p = 0.0001 (f), p = 0.0113 (g), p = 0.0032 (h), p = 0.0004 (i); Succinate oxidase: p < 0.0001 (a), p = 0.0002 (b), p = 0.0240 (c), p = 0.0434 (d), p = 0.0283 (e), p = 0.0009 (f), p = 0.0250 (g), p = 0.0013 (h); Cytochrome c oxidase: p = 0.0053 (a), p = 0.0035 (b). Coupling: p = 0.0003 (a), p = 0.0010 (b). Further statistical details on the genotype, age, and genotype × age effect can be found in the legend of Table 3.
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Figure 2: Brain bioenergetics of KI mice during neurodevelopment and adulthood. Mitochondria were isolated from cortex, cerebellum, and hippocampus of WT and KI pups as described in the Methods section. Activities of NADH oxidase, succinate oxidase, and cytochrome c oxidase (CCO), and respiratory control ratio (RCR) were evaluated at PND0 (cerebellum and cortex only, due to the scarcity of hippocampal tissue), PND7, PND21, and PND210. Data are reported as mean ± SEM, n = 3–7 per genotype per time point. Statistical analysis was performed by Two-way ANOVA. Post-hoc analysis performed by Tukey's HSD test revealed significant differences between WT and KI indicated by asterisks as follows NADH oxidase: *p = 0.0468; Succinate oxidase: *p = 0.0466 at PND21, *p = 0.0423 at PND210; Cytochrome c oxidase: *p = 0.0484; Coupling: **p = 0.001, ***p = 0.0004, *p = 0.0496. Letters indicate statistically significant differences amongst time points as follows. NADH oxidase: p = 0.0291 (a), p = 0.0033 (b), p = 0.0166 (c), p = 0.0320 (d), p = 0.0036 (e), p = 0.0001 (f), p = 0.0113 (g), p = 0.0032 (h), p = 0.0004 (i); Succinate oxidase: p < 0.0001 (a), p = 0.0002 (b), p = 0.0240 (c), p = 0.0434 (d), p = 0.0283 (e), p = 0.0009 (f), p = 0.0250 (g), p = 0.0013 (h); Cytochrome c oxidase: p = 0.0053 (a), p = 0.0035 (b). Coupling: p = 0.0003 (a), p = 0.0010 (b). Further statistical details on the genotype, age, and genotype × age effect can be found in the legend of Table 3.

Mentions: At PND0-7, no statistically significant differences were observed in mitochondrial outcomes between WT and KI pups in any of the brain regions tested (Figure 2). At PND21, hippocampus was the most affected brain area in KI pups, with significant decreases in NADH oxidase, succinate oxidase, and CCO activities, as well as increased uncoupling between ATP production and electron transfer, relative to WT (Figure 2). At PND21, uncoupling was also evident in cerebellum of KI pups, but to a lesser extent than hippocampus (Figure 2), whereas in adult age (PND210) succinate oxidase, and coupling (hippocampus) were still significantly lower in KI hippocampus than age-matched WT mice. No difference was observed in any of the outcomes in cortex at any of the time points (Figure 2).


Premutation in the Fragile X Mental Retardation 1 (FMR1) Gene Affects Maternal Zn-milk and Perinatal Brain Bioenergetics and Scaffolding.

Napoli E, Ross-Inta C, Song G, Wong S, Hagerman R, Gane LW, Smilowitz JT, Tassone F, Giulivi C - Front Neurosci (2016)

Brain bioenergetics of KI mice during neurodevelopment and adulthood. Mitochondria were isolated from cortex, cerebellum, and hippocampus of WT and KI pups as described in the Methods section. Activities of NADH oxidase, succinate oxidase, and cytochrome c oxidase (CCO), and respiratory control ratio (RCR) were evaluated at PND0 (cerebellum and cortex only, due to the scarcity of hippocampal tissue), PND7, PND21, and PND210. Data are reported as mean ± SEM, n = 3–7 per genotype per time point. Statistical analysis was performed by Two-way ANOVA. Post-hoc analysis performed by Tukey's HSD test revealed significant differences between WT and KI indicated by asterisks as follows NADH oxidase: *p = 0.0468; Succinate oxidase: *p = 0.0466 at PND21, *p = 0.0423 at PND210; Cytochrome c oxidase: *p = 0.0484; Coupling: **p = 0.001, ***p = 0.0004, *p = 0.0496. Letters indicate statistically significant differences amongst time points as follows. NADH oxidase: p = 0.0291 (a), p = 0.0033 (b), p = 0.0166 (c), p = 0.0320 (d), p = 0.0036 (e), p = 0.0001 (f), p = 0.0113 (g), p = 0.0032 (h), p = 0.0004 (i); Succinate oxidase: p < 0.0001 (a), p = 0.0002 (b), p = 0.0240 (c), p = 0.0434 (d), p = 0.0283 (e), p = 0.0009 (f), p = 0.0250 (g), p = 0.0013 (h); Cytochrome c oxidase: p = 0.0053 (a), p = 0.0035 (b). Coupling: p = 0.0003 (a), p = 0.0010 (b). Further statistical details on the genotype, age, and genotype × age effect can be found in the legend of Table 3.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4835505&req=5

Figure 2: Brain bioenergetics of KI mice during neurodevelopment and adulthood. Mitochondria were isolated from cortex, cerebellum, and hippocampus of WT and KI pups as described in the Methods section. Activities of NADH oxidase, succinate oxidase, and cytochrome c oxidase (CCO), and respiratory control ratio (RCR) were evaluated at PND0 (cerebellum and cortex only, due to the scarcity of hippocampal tissue), PND7, PND21, and PND210. Data are reported as mean ± SEM, n = 3–7 per genotype per time point. Statistical analysis was performed by Two-way ANOVA. Post-hoc analysis performed by Tukey's HSD test revealed significant differences between WT and KI indicated by asterisks as follows NADH oxidase: *p = 0.0468; Succinate oxidase: *p = 0.0466 at PND21, *p = 0.0423 at PND210; Cytochrome c oxidase: *p = 0.0484; Coupling: **p = 0.001, ***p = 0.0004, *p = 0.0496. Letters indicate statistically significant differences amongst time points as follows. NADH oxidase: p = 0.0291 (a), p = 0.0033 (b), p = 0.0166 (c), p = 0.0320 (d), p = 0.0036 (e), p = 0.0001 (f), p = 0.0113 (g), p = 0.0032 (h), p = 0.0004 (i); Succinate oxidase: p < 0.0001 (a), p = 0.0002 (b), p = 0.0240 (c), p = 0.0434 (d), p = 0.0283 (e), p = 0.0009 (f), p = 0.0250 (g), p = 0.0013 (h); Cytochrome c oxidase: p = 0.0053 (a), p = 0.0035 (b). Coupling: p = 0.0003 (a), p = 0.0010 (b). Further statistical details on the genotype, age, and genotype × age effect can be found in the legend of Table 3.
Mentions: At PND0-7, no statistically significant differences were observed in mitochondrial outcomes between WT and KI pups in any of the brain regions tested (Figure 2). At PND21, hippocampus was the most affected brain area in KI pups, with significant decreases in NADH oxidase, succinate oxidase, and CCO activities, as well as increased uncoupling between ATP production and electron transfer, relative to WT (Figure 2). At PND21, uncoupling was also evident in cerebellum of KI pups, but to a lesser extent than hippocampus (Figure 2), whereas in adult age (PND210) succinate oxidase, and coupling (hippocampus) were still significantly lower in KI hippocampus than age-matched WT mice. No difference was observed in any of the outcomes in cortex at any of the time points (Figure 2).

Bottom Line: Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background.A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced.Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase).

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, School of Veterinary Medicine Davis, CA, USA.

ABSTRACT
Fragile X premutation alleles have 55-200 CGG repeats in the 5' UTR of the FMR1 gene. Altered zinc (Zn) homeostasis has been reported in fibroblasts from >60 years old premutation carriers, in which Zn supplementation significantly restored Zn-dependent mitochondrial protein import/processing and function. Given that mitochondria play a critical role in synaptic transmission, brain function, and cognition, we tested FMRP protein expression, brain bioenergetics, and expression of the Zn-dependent synaptic scaffolding protein SH3 and multiple ankyrin repeat domains 3 (Shank3) in a knock-in (KI) premutation mouse model with 180 CGG repeats. Mitochondrial outcomes correlated with FMRP protein expression (but not FMR1 gene expression) in KI mice and human fibroblasts from carriers of the pre- and full-mutation. Significant deficits in brain bioenergetics, Zn levels, and Shank3 protein expression were observed in the Zn-rich regions KI hippocampus and cerebellum at PND21, with some of these effects lasting into adulthood (PND210). A strong genotype × age interaction was observed for most of the outcomes tested in hippocampus and cerebellum, whereas in cortex, age played a major role. Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background. A higher gene expression of ZnT4 and ZnT6, Zn transporters abundant in brain and lactating mammary glands, was observed in the latter tissue of KI dams. A cross-fostering experiment allowed improving cortex bioenergetics in KI pups nursing on WT milk. Conversely, WT pups nursing on KI milk showed deficits in hippocampus and cerebellum bioenergetics. A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced. Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase). In premutation carriers, altered Zn homeostasis, brain bioenergetics and Shank3 levels could be compounded by Zn-deficient milk, increasing the risk of developing emotional and neurological/cognitive problems and/or FXTAS later in life.

No MeSH data available.


Related in: MedlinePlus