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Table 3 GAA and Cr synthesis experiments in rat astrocytes

From: In vitro study of uptake and synthesis of creatine and its precursors by cerebellar granule cells and astrocytes suggests some hypotheses on the physiopathology of the inherited disorders of creatine metabolism

Concentration of [15N2]Arg and [13C2,15N]Gly

in the medium (mmol/L)

[15N2]Arg in the cells

(nmol/mg protein)

Arg in the cells

(nmol/mg protein)

[15N2]Arg/Arg

0

1.8 ± 1.1

172.0 ± 78.2

0.01 ± 0.00

5

137.9 ± 71.6 NS

145.5 ± 73.4

1.12 ± 0.52

10

504.0 ± 390.0 *

178.6 ± 137.9

3.43 ± 1.66

15

957.4 ± 500.45 **

215.5 ± 132.5

6.74 ± 4.72

 

[13C2,15N]Gly in the cells

(nmol/mg protein)

Gly in the cells

(nmol/mg protein)

[13C2,15N]Gly/Gly

0

2.8 ± 0.9

365.7 ± 43.2

0.01 ± 0.00

5

1003.8 ± 1083.3 NS

696.3 ± 604.2

1.28 ± 0.49

10

2205.5 ± 2382.0 *

893.8 ± 958.4

2.53 ± 0.22

15

3213.9 ± 2482.9 *

744.8 ± 512.7

4.43 ± 0.85

 

[13C215N3]GAA in the cells

(nmol/mg protein)

GAA in the cells

(nmol/mg protein)

[13C215N3]GAA/GAA

0

0.25 ± 0.09

1.46 ± 1.46

0.29 ± 0.14

5

10.34 ± 7.90 *

2.37 ± 2.23

4.47 ± 3.97

10

17.18 ± 13.36 *

1.88 ± 1.87

13.30 ± 14.32

15

21.59 ± 15.00 **

1.64 ± 1.11

18.18 ± 17.96

 

[13C215N3]Cr in the cells

(nmol/mg protein)

Cr in the cells

(nmol/mg protein)

[13C215N3]Cr/Cr

0

0.10 ± 0.05

100.4 ± 118.7

0.00 ± 0.00

5

12.13 ± 12.64 *

86.53 ± 71.65

0.12 ± 0.04

10

10.88 ± 8.48 **

59.48 ± 23.72

0.17 ± 0.08

15

11.00 ± 6.85 **

48.54 ± 10.58

0.21 ± 0.09

  1. Intracellular concentrations of Gly, [13C2,15N]Gly, Arg, [15N2]Arg, GAA, [1,2-13C2,15N3]GAA, Cr and [1,2-13C2,15N3]Cr after 24 h in vitro incubation with [15N2]Arg and [13C2,15N]Gly under different concentrations of labeled precursors in the medium: no labeled precursors (0), 5, 10 and 15 mmol/L. The statistical significance of the differences between intracellular concentrations of labeled and unlabeled compounds was assessed by paired Student's t-test: NS not significant, * p < 0.05, ** p < 0.01. The concentrations of [13C2,15N]Gly and [15N2]Arg were significantly higher than those of Gly and Arg (except for cells incubated with [15N2]Arg mmol/L), showing that the labeled precursors are highly available into the cells for Cr synthesis. [13C2,15N3]GAA concentrations were also significantly higher than those of GAA, whereas [13C2,15N3]Cr levels remained significantly lower than those of Cr. The ratios express the relative intracellular availability of labeled and unlabeled forms (right column).