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Frataxin levels in peripheral tissue in Friedreich ataxia.

Lazaropoulos M, Dong Y, Clark E, Greeley NR, Seyer LA, Brigatti KW, Christie C, Perlman SL, Wilmot GR, Gomez CM, Mathews KD, Yoon G, Zesiewicz T, Hoyle C, Subramony SH, Brocht AF, Farmer JM, Wilson RB, Deutsch EC, Lynch DR - Ann Clin Transl Neurol (2015)

Bottom Line: Such mutations, usually expanded guanine-adenine-adenine (GAA) repeats, give rise to decreased levels of frataxin protein in both affected and unaffected tissues.Site-directed mutant frataxin was also transfected into human embryonic kidney cells to model results from specific point mutations.The G130V mutation led to decreased levels of frataxin in vitro as well as in vivo, while the R165C mutation produced normal immunoreactive levels of frataxin both in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Departments of Pediatrics and Neurology, Children's Hospital of Philadelphia, Perelman School of Medicine, University of Pennsylvania Philadelphia, Pennsylvania, 19104.

ABSTRACT

Objective: Friedreich ataxia (FRDA) is an autosomal recessive ataxia resulting from mutations in the frataxin gene (FXN). Such mutations, usually expanded guanine-adenine-adenine (GAA) repeats, give rise to decreased levels of frataxin protein in both affected and unaffected tissues. The goal was to understand the relationship of frataxin levels in peripheral tissues to disease status.

Methods: Frataxin levels were measured in buccal cells and blood, and analyzed in relation to disease features. Site-directed mutant frataxin was also transfected into human embryonic kidney cells to model results from specific point mutations.

Results: There was no evidence for change in frataxin levels over time with repeated measures analysis, although linear regression analysis of cross-sectional data predicted a small increase over decades. GAA repeat length predicted frataxin levels in both tissues, and frataxin levels themselves predicted neurological ratings (accounting for age). Compound heterozygous patients for a GAA expansion and a point mutation in FXN generally had lower levels of frataxin than those homozygous for the presence of two GAA repeat expansions, though levels varied dramatically between tissues in some compound heterozygotes for point mutations. The G130V mutation led to decreased levels of frataxin in vitro as well as in vivo, while the R165C mutation produced normal immunoreactive levels of frataxin both in vitro and in vivo. Start codon mutations led to low levels of frataxin in buccal cells but preserved immunoreactive frataxin levels in blood.

Interpretation: The present data show that peripheral frataxin levels reflect disease features in FRDA, but emphasize the need for interpretation of such levels in the context of specific mutations.

No MeSH data available.


Related in: MedlinePlus

Repeated-measures analysis of frataxin in blood and buccal cells. The change in frataxin levels between samples (ordinate, expressed as a percentage of mean control value from unaffected individuals) was evaluated as a function of time between samples (abscissa; A and B) and initial sample value (abscissa, expressed as a percentage of mean control value from unaffected individuals; C and D) in buccal cells (A and C) and whole blood (B and D). There was essentially no relationship between the difference between the first and second measurements of frataxin levels and the time difference between those two samples. Although the relationship was statistically significant in buccal cells, the R2 value was very low and the slope of the association revealed a change in frataxin value of less than 1% per year. In contrast there was an inverse relationship between the initial frataxin value and the change in value, consistent with regression to the mean as the reason for the mean decrease in frataxin level with serial measurement in both blood and buccal swabs. N = 143 for buccal swabs, n = 137 for blood.
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fig01: Repeated-measures analysis of frataxin in blood and buccal cells. The change in frataxin levels between samples (ordinate, expressed as a percentage of mean control value from unaffected individuals) was evaluated as a function of time between samples (abscissa; A and B) and initial sample value (abscissa, expressed as a percentage of mean control value from unaffected individuals; C and D) in buccal cells (A and C) and whole blood (B and D). There was essentially no relationship between the difference between the first and second measurements of frataxin levels and the time difference between those two samples. Although the relationship was statistically significant in buccal cells, the R2 value was very low and the slope of the association revealed a change in frataxin value of less than 1% per year. In contrast there was an inverse relationship between the initial frataxin value and the change in value, consistent with regression to the mean as the reason for the mean decrease in frataxin level with serial measurement in both blood and buccal swabs. N = 143 for buccal swabs, n = 137 for blood.

Mentions: The short-term reproducibility of the frataxin assay previously was excellent for technical replicates, but had larger variability in repeated sampling in buccal and whole blood samples.7,14 In addition, previous regression analyses suggested that peripheral frataxin levels changed minimally over the course of a lifetime. To revisit these issues, the variability in frataxin measurements was assessed in repeated samplings over several years by examining the relationship between the difference in frataxin level between the first and second samplings and the interval in time between samplings. There was no relationship between the difference in frataxin levels and the interval between measurements in buccal cells (R2 = 0.013, P = 0.17) and whole blood (R2 = 0.0004, P = 0.81) (Fig.1A and B). There was also little-to-no correlation of the difference in frataxin levels between measurements with GAA repeat length (buccal: R2 = 0.0405, P = 0.016; blood: R2 = 0.0003, P = 0.858) or age (buccal: R2 = 0.047, P = 0.009; blood: R2 = 0.0048, P = 0.422) in either tissue. Although these models were statistically significant in buccal cells, the low R2 values demonstrated a minimal relationship identified only due to the large size of the cohort. In contrast, the difference between the first and second frataxin samplings was predicted by the first frataxin measurement from buccal cells (R2 = 0.47, P < 0.001) and whole blood samples (R2 = 0.23, P < 0.001) (Fig.1C and D). The second measurement significantly proceeded toward the mean frataxin level for the cohort, showing that regression to the mean is the dominant identifiable feature of sampling variability with no evidence for biological changes in frataxin levels over time.


Frataxin levels in peripheral tissue in Friedreich ataxia.

Lazaropoulos M, Dong Y, Clark E, Greeley NR, Seyer LA, Brigatti KW, Christie C, Perlman SL, Wilmot GR, Gomez CM, Mathews KD, Yoon G, Zesiewicz T, Hoyle C, Subramony SH, Brocht AF, Farmer JM, Wilson RB, Deutsch EC, Lynch DR - Ann Clin Transl Neurol (2015)

Repeated-measures analysis of frataxin in blood and buccal cells. The change in frataxin levels between samples (ordinate, expressed as a percentage of mean control value from unaffected individuals) was evaluated as a function of time between samples (abscissa; A and B) and initial sample value (abscissa, expressed as a percentage of mean control value from unaffected individuals; C and D) in buccal cells (A and C) and whole blood (B and D). There was essentially no relationship between the difference between the first and second measurements of frataxin levels and the time difference between those two samples. Although the relationship was statistically significant in buccal cells, the R2 value was very low and the slope of the association revealed a change in frataxin value of less than 1% per year. In contrast there was an inverse relationship between the initial frataxin value and the change in value, consistent with regression to the mean as the reason for the mean decrease in frataxin level with serial measurement in both blood and buccal swabs. N = 143 for buccal swabs, n = 137 for blood.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4554444&req=5

fig01: Repeated-measures analysis of frataxin in blood and buccal cells. The change in frataxin levels between samples (ordinate, expressed as a percentage of mean control value from unaffected individuals) was evaluated as a function of time between samples (abscissa; A and B) and initial sample value (abscissa, expressed as a percentage of mean control value from unaffected individuals; C and D) in buccal cells (A and C) and whole blood (B and D). There was essentially no relationship between the difference between the first and second measurements of frataxin levels and the time difference between those two samples. Although the relationship was statistically significant in buccal cells, the R2 value was very low and the slope of the association revealed a change in frataxin value of less than 1% per year. In contrast there was an inverse relationship between the initial frataxin value and the change in value, consistent with regression to the mean as the reason for the mean decrease in frataxin level with serial measurement in both blood and buccal swabs. N = 143 for buccal swabs, n = 137 for blood.
Mentions: The short-term reproducibility of the frataxin assay previously was excellent for technical replicates, but had larger variability in repeated sampling in buccal and whole blood samples.7,14 In addition, previous regression analyses suggested that peripheral frataxin levels changed minimally over the course of a lifetime. To revisit these issues, the variability in frataxin measurements was assessed in repeated samplings over several years by examining the relationship between the difference in frataxin level between the first and second samplings and the interval in time between samplings. There was no relationship between the difference in frataxin levels and the interval between measurements in buccal cells (R2 = 0.013, P = 0.17) and whole blood (R2 = 0.0004, P = 0.81) (Fig.1A and B). There was also little-to-no correlation of the difference in frataxin levels between measurements with GAA repeat length (buccal: R2 = 0.0405, P = 0.016; blood: R2 = 0.0003, P = 0.858) or age (buccal: R2 = 0.047, P = 0.009; blood: R2 = 0.0048, P = 0.422) in either tissue. Although these models were statistically significant in buccal cells, the low R2 values demonstrated a minimal relationship identified only due to the large size of the cohort. In contrast, the difference between the first and second frataxin samplings was predicted by the first frataxin measurement from buccal cells (R2 = 0.47, P < 0.001) and whole blood samples (R2 = 0.23, P < 0.001) (Fig.1C and D). The second measurement significantly proceeded toward the mean frataxin level for the cohort, showing that regression to the mean is the dominant identifiable feature of sampling variability with no evidence for biological changes in frataxin levels over time.

Bottom Line: Such mutations, usually expanded guanine-adenine-adenine (GAA) repeats, give rise to decreased levels of frataxin protein in both affected and unaffected tissues.Site-directed mutant frataxin was also transfected into human embryonic kidney cells to model results from specific point mutations.The G130V mutation led to decreased levels of frataxin in vitro as well as in vivo, while the R165C mutation produced normal immunoreactive levels of frataxin both in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Departments of Pediatrics and Neurology, Children's Hospital of Philadelphia, Perelman School of Medicine, University of Pennsylvania Philadelphia, Pennsylvania, 19104.

ABSTRACT

Objective: Friedreich ataxia (FRDA) is an autosomal recessive ataxia resulting from mutations in the frataxin gene (FXN). Such mutations, usually expanded guanine-adenine-adenine (GAA) repeats, give rise to decreased levels of frataxin protein in both affected and unaffected tissues. The goal was to understand the relationship of frataxin levels in peripheral tissues to disease status.

Methods: Frataxin levels were measured in buccal cells and blood, and analyzed in relation to disease features. Site-directed mutant frataxin was also transfected into human embryonic kidney cells to model results from specific point mutations.

Results: There was no evidence for change in frataxin levels over time with repeated measures analysis, although linear regression analysis of cross-sectional data predicted a small increase over decades. GAA repeat length predicted frataxin levels in both tissues, and frataxin levels themselves predicted neurological ratings (accounting for age). Compound heterozygous patients for a GAA expansion and a point mutation in FXN generally had lower levels of frataxin than those homozygous for the presence of two GAA repeat expansions, though levels varied dramatically between tissues in some compound heterozygotes for point mutations. The G130V mutation led to decreased levels of frataxin in vitro as well as in vivo, while the R165C mutation produced normal immunoreactive levels of frataxin both in vitro and in vivo. Start codon mutations led to low levels of frataxin in buccal cells but preserved immunoreactive frataxin levels in blood.

Interpretation: The present data show that peripheral frataxin levels reflect disease features in FRDA, but emphasize the need for interpretation of such levels in the context of specific mutations.

No MeSH data available.


Related in: MedlinePlus