Blood oxidative stress and metallothionein expression in Rett syndrome: Probing for markers.

TLDR: As the haematic level of MT-1A directly correlates with the phenotype severity, this metallothionein can represent a marker for RTT severity, and the attempt to link the level of blood oxidative stress with MECP2 mutation and specific clinical features led us to draw some interesting conclusions.

Abstract: Objectives: Oxidative stress seems to be involved in Rett syndrome (RTT). The aim of this study was to assess the antioxidant status in RTT children with MECP2 gene mutations with respect to healthy controls, and to explore novel blood antioxidant markers for RTT severity. Methods: In erythrocytes from RTT females aged 2–14 years (n = 27) and age-matched controls (n = 27), we measured the levels of malonaldehyde and the activity of two antioxidant enzymes, Cu/Zn-superoxide dismutase and catalase, by spectrophotometric assays. In leukocytes, the expression of metallothioneins, the main non-enzymatic antioxidants, was assessed by real-time RT-PCR. In nine selected RTT children, methylome analysis was also performed. Results: Blood of RTT patients showed increased lipid peroxidation and a dysregulated pattern of MT expression, while enzymatic activities did not change significantly with respect to controls. Moreover, we observed no epigenetic dysregulation in CpG-enriched promoter regions of the anal...

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Supplementary data
Table 1: Clinical and Genetic data of patients
Patient MT-2A MT-1A MT-1E TBARS CAT
Age
(years)
Clinical
phenotype
Protein mutation Gene mutation
Group of
mutation
1 2,72 65,45 5,87 0,06 143,52 9 Co p-Val331GlyfsX34 c.1052_1188del137 LT
2 11,75 9,68 1,45 0,11 157,41 9 C p.Arg168X c 502 C>T ET
3 2,54 69,61 0,52 0,13 226,39 5 Co p.Arg270Glufs288X c.808delC IT
4 0,80 14,46 0,86 11 C 3-4 Exon deletion ET
5 1,32 26,22 3,00 0,17 219,32 6 C p.Arg255X c.763C>T IT
6 0,93 0,35 4,92 0,15 298,92 3 C p.Lys135Asn c.405A>C M
7 1,06 19,69 1,79 0,11 182,26 14 C p.Arg294X c.880C>T LT
8 1,43 54,76 2,55 0,21 226,09 3 C p.Arg168X c 502 C>T ET
9 0,51 44,69 0,53 0,18 146,44 11 C p.Leu386His fs390X C.1156_1197del41 LT
10 1,39 1,16 6,29 0,13 271,26 6 C p.Leu386His fs390X C.1156_1197del41 LT
11 0,95 62,63 0,72 0,09 334,08 10 PSV p.P387fsX9 c.1160_1166del7 LT
12 2,70 3,43 2,20 0,15 213,49 12 C p.Arg306Cys c 916 C>T M
13 3,20 0,00 2,00 0,16 273,67 7 C p.P388fsX c.1163del26 LT
14 0,88 64,88 0,45 0,20 260,78 7 C p.Lys144Arg fs2X c.431delA ET
15 6,00 4,65 8,04 0,20 256,19 6 C p.Asp156Glu c.468 C>G M
16 2,87 60,06 4,96 0,17 252,34 4 C p.Arg306Cys c 916 C>T M
17 4,51 50,04 6,91 4 C p.Pro225Arg c 674 C>G M
18 1,98 46,09 3,66 4 C p.Arg255X c.763C>T IT
19 1,25 0,58 2,42 0,08 190,78 4 C p.Arg255X c.763C>T IT
20 0,45 9,59 1,28 0,20 301,89 3 C p.Glu37Xfs c.108-111 del ET
21 3,96 52,37 2,88 0,14 238,25 6 C p. Arg168X c 502 C>T ET
22 0,71 0,16 0,09 0,15 251,42 7 C p.Arg306Cys c 916 C>T M
23 1,50 24,91 0,01 0,21 270,07 3 Co p. Thr158Met c 473 C>T M
24 1,42 8,57 3,21 0,10 178,93 12 PSV p.Arg294X c.880C>T LT
25 3,21 11,81 9,94 0,14 161,66 6 Co p.Thr158Met c 473 C>T M
26 1,78 6,08 4,46 0,13 206,41 4 Co p.Thr158Met c 473 C>T M
27 2,59 8,41 6,35 0,11 190,19 8 PSV p. P389X c 1164_1207del44 LT

Table 2: Patients Kerr
Score
Pt
A B C D E F G H I J K L M N O P Q R S T
TOT
1 0 2 1 0 1 2 2 2 1 2 0 2 1 2 2 0 1 1 2 1 25
2 1 0 2 2 2 1 1 1 1 2 0 1 1 2 2 2 1 1 0 0 23
3 2 2 2 1 1 1 2 1 2 1 0 2 2 2 2 2 2 1 1 2 31
4 0 1 1 1 1 1 1 0 1 1 0 1 1 2 2 1 1 1 0 0 17
5 0 1 2 2 2 1 1 1 1 1 0 1 0 2 2 2 1 1 1 1 23
6 0 0 2 1 1 0 0 0 1 1 0 0 1 1 1 0 1 1 0 0 11
7 1 2 0 0 1 1 1 1 2 2 1 0 1 2 2 1 1 1 0 0 19
8 0 1 1 1 1 1 0 0 2 2 0 2 1 2 2 1 1 1 1 1 21
9 1 0 2 2 1 1 1 1 1 2 0 0 1 2 1 2 1 1 2 1 23
10 0 1 1 2 1 1 1 1 0 1 0 1 0 1 1 1 1 1 1 1 17
11 1 1 1 0 0 1 1 1 0 1 0 1 0 1 1 1 0 1 1 1 14
12 1 1 2 1 1 1 0 1 1 2 0 1 2 1 2 1 0 1 2 1 22
13 1 1 1 0 1 1 1 1 1 1 0 1 0 1 1 0 0 1 0 0 13
14 2 0 1 2 0 1 0 0 1 2 0 1 2 2 2 0 1 1 0 1 19
15 1 0 1 0 0 1 1 1 1 2 1 1 1 1 2 1 2 1 0 0 18
16 0 1 1 0 1 1 1 1 1 2 0 1 0 1 0 0 1 1 0 0 14
17 1 0 2 1 1 1 0 0 1 2 0 2 0 1 2 0 1 1 0 0 16
18 0 1 2 0 1 2 0 0 2 2 0 1 1 2 2 1 1 0 0 1 19
19 0 1 0 0 0 1 0 0 2 2 0 1 1 1 2 1 0 0 0 1 13
20 2 1 1 0 0 2 0 0 1 1 0 1 1 1 1 0 0 0 0 0 12
21 0 0 0 0 0 0 1 0 1 2 0 1 1 2 2 0 0 0 0 1 11
22 0 0 0 1 0 2 0 0 1 1 0 1 0 2 1 0 0 0 1 0 10
23 0 2 1 1 1 2 0 0 2 0 1 2 2 2 2 2 2 0 1 2 25
24 0 0 0 0 2 0 1 0 1 1 0 1 0 1 1 1 0 0 2 1 12
25 1 1 2 0 0 2 1 0 2 1 0 1 0 2 2 0 0 1 0 0 16
26 1 1 1 0 0 2 1 2 2 1 0 2 1 2 2 2 1 0 0 1 22
27 0 0 0 0 0 0 1 0 0 1 1 0 0 1 1 1 0 1 1 0 8

Table 3: Patients Clinical Severity Score
Patient
age onset
regression
somatic
growth
head
growth
motor
sitting
ambulation
hand
use
scoliosis
language
non verbal
communication
respiratory
dysfunction
autonomic
symptoms
onset
stereotype
epilepsy
Total CSS
1 3 0 1 2 3 2 2 3 1 1 2 2 0 22
2 3 4 u 2 1 3 2 3 2 0 1 2 4 27
3 3 0 u 2 5 3 1 3 2 1 1 3 4 28
4 3 4 u 1 1 2 0 4 0 1 1 2 3 22
5 3 2 u 1 3 3 2 4 1 1 1 3 1 25
6 3 1 u 0 1 2 1 3 1 1 1 3 0 17
7 4 4 u 2 2 2 2 3 0 1 1 2 4 27
8 4 1 u 2 5 2 1 4 2 1 1 3 0 25
9 3 0 u 1 1 3 2 3 2 2 4 3 4 28
10 4 4 u 1 1 2 2 3 0 2 1 2 1 23
11 1 0 u 0 0 1 1 1 0 1 0 1 0 6
12 4 4 u 2 1 2 3 3 2 1 1 4 0 27
13 3 0 u 1 1 0 1 2 1 0 1 3 0 13
14 2 4 u 0 0 3 0 3 1 3 1 2 0 19
15 2 4 u 2 0 3 1 3 1 2 1 2 1 22
16 2 0 u 1 2 3 0 3 2 1 1 2 0 17
17 1 1 u 0 0 1 0 4 1 1 0 1 0 10
18 2 0 4 1 5 3 0 4 1 2 0 2 0 24
19 3 0 0 5 5 2 3 4 1 0 3 3 0 29
20 0 0 0 5 5 2 0 2 2 0 0 3 0 19
21 3 0 0 1 3 3 2 3 2 0 0 3 0 20
22 3 0 0 0 1 1 0 2 2 0 0 3 0 12
23 5 0 0 5 5 3 0 4 2 2 1 0 1 28
24 2 0 0 0 0 1 2 3 1 0 0 1 0 10
25 4 3 2 2 4 3 0 4 2 2 3 3 4 36

26 5 2 2 4 5 5 1 3 3 1 2 3 0 36
27 2 0 0 1 1 2 3 2 2 2 1 1 0 17