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Serum lipoproteins of patients with glycogen storage disease
99
Clinica Chimica Acta, 102 (1980) 99-104 @ EIsevier~North-HoIland Biomedical Press
CCA 1287
SERUM L~OPROTEINS DISEASE
OF PATIENTS WITH GLYCOGEN
E.L. ROSENFELD, I.V. CHIBISOV, I.M. KARMANSKY, andA.F.LEONTIEV
STORAGE
V.A. TABOLIN, L.V. CHISTOVA
~~s#itute of Riologi~al and Medical chemists USSR, Academy of medical Sciences and 2th Medical Institute, Institute of Pediatrics, USSR Academy of Medical Sciences, Moscow (USSR) (Received September 5th, 1979)
Seventeen patients with different types of glycogen storage disease (GSD) were under observation. The type of the disease was defined from glucaemic and lactotaemic curves obtained in glucose, galactose and adrenaline tolerance tests and by biochemical analysis of liver biopsy specimens. Seven patients were found to have Type I; five patients, Type III; one patient, Type VI; and four patients, the Type IX (or X) of GSD. The serum lipoprotein (LP) content was determined in all patients using analytical ultracentrifugation. Hyperlipoproteinaemia (HLP) was found in virtually all patients. Patients with Type I of GSD were found to have Types 2b and 4 of HLP; and patients with Type III of GSD, 2b Type of HLP. 2a Type of HLP was diagnosed in patients with GSD of VI and IX (X) Types. Patients with Type III GSD, in contrast to those with GSD of other types, had enhanced levels of Si 12-20 LP. The levels of Si 100-400 and Si 20-100 LP were greatly increased only in patients with Type I GSD.
Introduction GSD is a hereditary disease caused by different disturbances of glycogen metabolism leading to glycogen accumulation in affected tissues. According to the clinical m~~es~tions and enzymic defect more than 10 types of GSD are differentiated. The hepatic form of the disease is characterized by hepatomegaly, growth retardation, fasting hypoglycaemia and acidosis. * Correspondence should be addressed to: Prof. E.L. Rosenfeld, Institute of Biological and Medical Chemistry USSR, Academy of Medical Sciences, Pogodin Street 10, Moscow 9,119121, USSR.
100
Administration of galactose, glucose, adrenaline and glucagon leads to different changes in the glycaemic and lactotaemic curves of patients with the different types of GSD. Disturbances in lipid metabolism [l] and increased cholesterol and triglyceride in the serum of patients with GSD have been described [ 2,3]. This paper reports data on the low and very low density LP content (LDL and VLDL, respectively) in serum of patients with GSD. Materials and methods The substrates used were: rabbit liver glycogen; cp-limitdextrin, obtained from rabbit liver glycogen; glucose-6-phosphate dipotassium salt and glucose-lphosphate dipotassium salt; AMP sodium salt; horse-radish peroxidase (Reanal); glucose-oxidase from Penicillium vitale (L’ vov); amyloglucosidase (Serva). Determination of glucose and lactate in glucose, galactose and adrenaline tolerance tests were carried out as described [ 41. LDL and VLDL were isolated from the serum of patients by preparative ultracentrifugation after adjusting the serum density to 1.063 g/ml [ 51. Floating LP were collected and subjected to analytical ultracentrifugation by the method of De Lalla et al. [6] to determine the concentration of LP with floatation coefficients of 100-400, 20-100, 12-20 and O-12 Sf. Liver tissue (15-20 mg) obtained by needle biopsy under general anaesthesia was immediately frozen to -78°C and stored at this temperature for 20-44 h. Glycogen concentration in liver tissue was determined by the micromethod of Huijing [ 71, amylo-1,6-glucosidase and glucose-6-phosphatase activities were determined by the micromethod of Hers [8], phosphorylase activity (in the presence and absence of AMP and NaZS04) was determined by the method of Stalman and Hers [9], phosphoglucomutase activity was estimated by the method of Najjar [lo] and phosphohexoisomerase activity by the method of Slain [ 111. The phosphates formed were estimated using FeSO, [ 71. Results Seven patients were diagnosed as having Type I GSD. In all these patients glucose administration resulted in the lowering of lactate levels in the whole blood, whereas the glycaemic curves were diabetic-like. The blood glucose failed to rise in response to adrenaline injections and the blood lactate level, which was high even before the adrenaline administration, increased. Table I presents data on the biochemical analysis of liver tissue of five patients with Type I glycogenosis. In all patients the liver glycogen content was sharply enhanced. In two patients (D.Yu. and Sh.A.) glucose-6-phosphatase activity was virtually absent. In three other patients, M.A., S.G. and L.M., the activity of this enzyme was within normal limits, although the data obtained from tolerance tests were characteristic of Type I GSD. It is known that conversion of glucose-g-phosphate to glucose in hepatocytes includes two stages, namely: (1) transfer of glucose-g-phosphate from the cytosol to the inner surface of the cytoplasmic reticulum which is mediated by a specific permease, and (2) its hydrolysis by glucose-6-phosphatase [12]. The lack of each of them leads to GSD Type I.
4-l
1.01 1.22 1.04 1.14
0.3-0.5
K.D. Sb.M. J.Zh. S.M.
* mmol phosphate . miu-1 . g-I tissue. ** mmol fructose-6-phosphate - mine1 + g-1 tissue. *** mm01 giucose ’ n~in-~ . g-l tissue.
6
6.7 6.4 3.7 8.3
1.40
VI
IX IX IX IX
G.S.
NOllXld
2.0 2.6 4.0 3.1 0
1.27 1.16 1.40 1.16 0.97
III III III III III
M.L. V.Yu. S.S. L.O. Sh.L.
1.8
0 0‘8 3.8 6.5 7.6
0.79 1.13 0.77 1.17 0.93
la la lb lb lb
D.Yu. Sh.a. M.a. s.g. L.M.
(W (X) 00 W)
Glucose-6phosphatase *
Glycogen
(g/v
OF LIVER TISSUE
Type of GSD
ANALYSIS
Patients
BIOCHEMICAL
TABLE I
15-56
5 8.8 4.5 6.6
3
3.5 29 9 14.5 0
27 23 22 54 20.5
Phosphorylase a*
15-W
16.6 28.6 11.8 22
4
18 12
6.6 27
33 23 17 50 20.6
Phosphorylase b*
25-100
56 40 27 135
40
64 27 22 36
44 65 80 173 -
Phosphoglucomutsse *
25-100
12.5 15 -
70
44 23 48 8
33 71 28 54 37
Phosphexoisomerase * *
0.4-1.1
0.81 0.44 0.4 0.6
0.39
0 0.1 0 0.23 0
1.22 0.36 0.43 0.50 0.74
Amylo-1,6glucosidase * * *
m+f
N0ma.l
* The number of controls.
3.0-15.0
0.5 9.5 15.0 5.5 1.5 3.0 23.0 9.0 3.5 7.0 4.0 5.0 4.0 2.0 4.5 4.0 2.0
f f f m f f m f f m f f m m m m m
D.Yu. Sh.a Yu.g M.a. s.g. L.M. D.V. M.L. V.Yu. S.S. L.O. Sh.L. G.Z. K.D. Sh.m. J.Zh. S.M.
(14 *I
Age (Yl.1
OF LIPOPROTEINS
Patients
THE CONTENT
TABLE II
la la 1 lb lb lb 1 III III III Hi III VI IX IX IX IX
(K) (K) (K) (K)
Type of glycogenosis
0.03 f 0.01
8.6 8.0 8.3 5.2 16.2 4.1 0.1 0.7 0.1 0.3 0.4 0.2 0.03 0.03 0.03 0.03 0.03
100400
Liproproteins Sf
IN BLOOD SERUM (ED) AND GLUCOSE
0.09 f 0.05
0.03 0.9 0.4 0.2 0.03 0.03 0.03 7.0 0.8 0.7 3.0 1.5 0.03 0.03 0.03 0.4 0.03
4.4 3.4 6.4 9.6 7.9 5.4 0.6 1.6 0.8 0.8 0.1 1.3 0.03 0.8 0.1 0.4 0.03 0.2 * 0.06
1 Z-20
CONTENT
20-100
AND LACTATE
3.6-5.3
1.84 4 0.2
<2.8
10.0 13.1 7.3 11.1 11.5 9.3 7.1 1.4 2.2 2.1 6.4 1.8 2.2 2.7 2.9 1.8 4.2
(mmol/l)
(mmoI!H
0.4 2.2 2.5 2.1 2.3 1 .I 3.1 1.9 1.4 2.3 1.7 2.4 1.3 3-3 3.7 2.9 2.4
Lactate
GIUCOSe
6.1 6.3 2.7 5.2 4.1 3.6 2.9 7.9 7.3 4.6 7.5 5.3 4.6 3.2 3.4 4.0 5.1
O-l 2
--
IN THE WHOLE BLOOD
103
The method employed to estimate the ability of liver homogenates to hydrolyse glucose-6-phosphate only reveals the absence of glucose-6-phosphatase. In cases of GSD Type I in which the activity of glucose-6-phosphatase in liver tissue homogenates was within normal limits, the permease activity was probably absent (GSD Type Ib) [ 13,141. Five of the patients (M.L., S.S., L.O., Sh.L., V.Yu.) were found to have GSD Type III. Amylo-1,6-glucosidase activity was lacking in the liver tissue of these patients. In two of them glucose-6-phosphatase activity was markedly reduced. However, the results obtained by glucose and adrenaline tolerance tests suggested that the lowering of glucose-6-phosphatase activity in these patients was secondary. Low phosphorylase activity was also found in the same patients. Such secondary changes in the glucose-6-phosphatase and phosphorylase activities are often observed in patients with GSD Type III [15]. Absorption spectra of the iodine complexes of glycogens isolated from the liver tissue of these patients were abnormal [ 4,16,17]. In patient G.S. with a considerable decrease in liver phosphorylases a and b activities, glucose-6-phosphatase activity was reduced; however, the tolerance test data also indicated in this case that the reduction of glucose-6-phosphatase activity was of a secondary nature thus establishing the GSD as Type VI. In patients K.D., Sh.M., I.Zh. and S.M. the activity of phosphorylase was normal in the presence, but decreased in the absence, of AMP and Na$04 suggesting a normal level of phosphorylase b activity and a deficiency either of phosphorylase b kinase (GSD Type IX) or of proteinkinase (GSD Type X). Table II presents data on the LDL and VLDL content in serum of 17 patients with GSD. As will be seen from these data, in six patients with GSD Type I, the content of VLDL with Sf 100-400 and 20-100, as well as the content of LDL with Sf O-12, was sharply increased. However, negligible or no changes were observed in the content of LDL with Sf 12-20 of these patients. According to the HLP classification proposed by the WHO [18] five patients with GSD Type I had HLP Type 2b, and one patient HLP Type 4. A different picture was observed in GSD of other types. For instance, GSD Type III was characterized by increased content of all LP fractions, especially of fractions with Sf O-12 and 12-20. All patients with this type of the disease had HLP 2b. In phosphorylase deficiency (Types VI, IX, X) only the content of LDL with Sf O-12 was increased (HLP 2a type). Thus, according to our data different changes in LP spectra are observed in different types of GSD. Discussion Quantitative determination of serum lipoproteins by analytical ultracentrifugation showed that the patients with Type I GSD had HLP Types 2b and 4, those with Type III had HLP Type 2b, and in those with Types VI and IX (or X) there was HLP Type 2a. Some patients with Types I and III disease had HLP of the same Type (2b). However, none of the patients with Type III had such a high VLDL level as those with Type I. On the other hand, none of the patients with GSD Type I had such a high level of LP with Sf 12-20 as those with GSD Type III.
104
In patients with Type I GSD not only the VLDL content but also that of blood lactate were markedly elevated, which was not observed in other patients, including those with a secondary deficiency of glucose-6-phosphatase (patients M.L., Sh.L., G.S.). It is not yet clear whether there is any interrelation between blood lactate content and changes in LP spectra in GSD. The cause of HLP in glycogenosis is not yet established. It is possible that the occurrence of HLP is connected with increased lipid synthesis, induced by disturbances of glycogen metabolism [ 31. The nature of changes in LP spectra in various types of glycogenoses is even less understood. In accordance with the generally accepted fact that LDL with Sf 12-20 and O-12 are formed as a result of metabolic transformations of VLDL with Si 100-400 and 20-100, these transformations may be expected to occur mainly in Types III, VI and IX (X) GSD but not in Type I disease. It might also be that in Type I GSD the VLDL synthesis is intensified to a greater extent than in other types of GSD. Data showing the differences in serum LP spectra may be used as additional evidence in the diagnosis of the type of GSD. References 1 MC Adams, A.J., Hug, G. and Bow, K.E. (1974) Hum. Pathol. 5,463-487 2 R. Rodney Howell (1972) in: The Met,aboIic Basis of Inherited Disease (Stanbury, J.B., Wyngaarden, J.B. and Fredrickson, D.S.. eds.), pp. 149-173, McGraw Hill, New York 3 Forget, P.P.. Fernandes, J. and Begemann, P.H. (1974) Pediatr. Res. 8, 114-119 4 Rosenfeld, E.L., Popova, LA. and Chibisov. I.V. (1976) Clin. Chin Acta 67, 123-130 5 Iahnke, K. and Scholtan, W. (1960) in Die Bluteiweissk6r:rper in der Ultracentrifuge. PP. 118-124. G. Thieme Verlag. Stuttgart 6 De Lalla. O.F., Tandy, R.K. and Loeb, H.G. (1967) Clin. Chem. 13. 85-100 7 Huijing, F. (1974) in Clinical Biochemistry (Curtius, H.Ch.. and Rot, M.. eds.). PP. 1203-1235. Walter de Gruyter. Berlin. New York 8 Hers, H.G. (1964) in Advances in Metabolic Disorders (Levine, R. and Jabt, R.. eds.) Vol. 1, PP. l44. Academic Press, New York 9 StaIrnan. J.W. and Hers, H.G. (1975) Europ. J. Biochem. 54, 341-350 10 Najjar. V.A. (1955) in Methods in Enzymology (Colowick. S.P. and KaPlan, N.O., eds.) Vol. 1. PP. 294-298, Academic Press Inc., New York 11 Slain, M.W. (1955) in Methods in Enzymology (Colowick, S.P. and Kaplan, N.O., eds.) Vol. 1, PP. 299-305, Academic Press Inc., New York 12 Arion, J.W., Lange, A.J. and BaIlas. L.M. (1976) J. Biol. Chem. 251.4901-4907 13 Naresawa, K., Jagarashi, L., Otomo. H. and Tada, K. (1978) Biochem. Biophys. Res. Comm. 83, 1360-1364 14 Rosenfeld, E.L., Chibisov, I.V., Chistova, L.V., Leontjev. A.F. and Karmansky, I.M. (1978) CIin. Chim. Acta 86, 295-299 15 Rossignol, A.M., Bost. M.. Marchal, A., Frappart, P., Stoebner. P. and Beaudoin, S.A. (1975) Ann. Pediatr. 22, 717-725 16 Chibisov. I.V., Karmansky. I.M. Cheljapina, V.V.. Leontjev, A.F. and Rosenfeld. E.L. (1978) Vo~r. Med. Chem. 24, 555-559. (Russian) 17 Chibisov, I.V., Rosenfeld, E.L., Tabolin, V.A., Komeva, T.I. and Muchina, Yu.G. (1977) Pediatria 11, 46-51, (Russian) 18 Beaumont, J.L., Carlsson, L.A.. Cooper, G.R., Fejfar. Z., Fredrickson, D.S. and Strasser, T. (1970) Bull. W.H.O. 43. 891
Clinica Chimica Acta, 102 (1980) 99-104 @ EIsevier~North-HoIland Biomedical Press
CCA 1287
SERUM L~OPROTEINS DISEASE
OF PATIENTS WITH GLYCOGEN
E.L. ROSENFELD, I.V. CHIBISOV, I.M. KARMANSKY, andA.F.LEONTIEV
STORAGE
V.A. TABOLIN, L.V. CHISTOVA
~~s#itute of Riologi~al and Medical chemists USSR, Academy of medical Sciences and 2th Medical Institute, Institute of Pediatrics, USSR Academy of Medical Sciences, Moscow (USSR) (Received September 5th, 1979)
Seventeen patients with different types of glycogen storage disease (GSD) were under observation. The type of the disease was defined from glucaemic and lactotaemic curves obtained in glucose, galactose and adrenaline tolerance tests and by biochemical analysis of liver biopsy specimens. Seven patients were found to have Type I; five patients, Type III; one patient, Type VI; and four patients, the Type IX (or X) of GSD. The serum lipoprotein (LP) content was determined in all patients using analytical ultracentrifugation. Hyperlipoproteinaemia (HLP) was found in virtually all patients. Patients with Type I of GSD were found to have Types 2b and 4 of HLP; and patients with Type III of GSD, 2b Type of HLP. 2a Type of HLP was diagnosed in patients with GSD of VI and IX (X) Types. Patients with Type III GSD, in contrast to those with GSD of other types, had enhanced levels of Si 12-20 LP. The levels of Si 100-400 and Si 20-100 LP were greatly increased only in patients with Type I GSD.
Introduction GSD is a hereditary disease caused by different disturbances of glycogen metabolism leading to glycogen accumulation in affected tissues. According to the clinical m~~es~tions and enzymic defect more than 10 types of GSD are differentiated. The hepatic form of the disease is characterized by hepatomegaly, growth retardation, fasting hypoglycaemia and acidosis. * Correspondence should be addressed to: Prof. E.L. Rosenfeld, Institute of Biological and Medical Chemistry USSR, Academy of Medical Sciences, Pogodin Street 10, Moscow 9,119121, USSR.
100
Administration of galactose, glucose, adrenaline and glucagon leads to different changes in the glycaemic and lactotaemic curves of patients with the different types of GSD. Disturbances in lipid metabolism [l] and increased cholesterol and triglyceride in the serum of patients with GSD have been described [ 2,3]. This paper reports data on the low and very low density LP content (LDL and VLDL, respectively) in serum of patients with GSD. Materials and methods The substrates used were: rabbit liver glycogen; cp-limitdextrin, obtained from rabbit liver glycogen; glucose-6-phosphate dipotassium salt and glucose-lphosphate dipotassium salt; AMP sodium salt; horse-radish peroxidase (Reanal); glucose-oxidase from Penicillium vitale (L’ vov); amyloglucosidase (Serva). Determination of glucose and lactate in glucose, galactose and adrenaline tolerance tests were carried out as described [ 41. LDL and VLDL were isolated from the serum of patients by preparative ultracentrifugation after adjusting the serum density to 1.063 g/ml [ 51. Floating LP were collected and subjected to analytical ultracentrifugation by the method of De Lalla et al. [6] to determine the concentration of LP with floatation coefficients of 100-400, 20-100, 12-20 and O-12 Sf. Liver tissue (15-20 mg) obtained by needle biopsy under general anaesthesia was immediately frozen to -78°C and stored at this temperature for 20-44 h. Glycogen concentration in liver tissue was determined by the micromethod of Huijing [ 71, amylo-1,6-glucosidase and glucose-6-phosphatase activities were determined by the micromethod of Hers [8], phosphorylase activity (in the presence and absence of AMP and NaZS04) was determined by the method of Stalman and Hers [9], phosphoglucomutase activity was estimated by the method of Najjar [lo] and phosphohexoisomerase activity by the method of Slain [ 111. The phosphates formed were estimated using FeSO, [ 71. Results Seven patients were diagnosed as having Type I GSD. In all these patients glucose administration resulted in the lowering of lactate levels in the whole blood, whereas the glycaemic curves were diabetic-like. The blood glucose failed to rise in response to adrenaline injections and the blood lactate level, which was high even before the adrenaline administration, increased. Table I presents data on the biochemical analysis of liver tissue of five patients with Type I glycogenosis. In all patients the liver glycogen content was sharply enhanced. In two patients (D.Yu. and Sh.A.) glucose-6-phosphatase activity was virtually absent. In three other patients, M.A., S.G. and L.M., the activity of this enzyme was within normal limits, although the data obtained from tolerance tests were characteristic of Type I GSD. It is known that conversion of glucose-g-phosphate to glucose in hepatocytes includes two stages, namely: (1) transfer of glucose-g-phosphate from the cytosol to the inner surface of the cytoplasmic reticulum which is mediated by a specific permease, and (2) its hydrolysis by glucose-6-phosphatase [12]. The lack of each of them leads to GSD Type I.
4-l
1.01 1.22 1.04 1.14
0.3-0.5
K.D. Sb.M. J.Zh. S.M.
* mmol phosphate . miu-1 . g-I tissue. ** mmol fructose-6-phosphate - mine1 + g-1 tissue. *** mm01 giucose ’ n~in-~ . g-l tissue.
6
6.7 6.4 3.7 8.3
1.40
VI
IX IX IX IX
G.S.
NOllXld
2.0 2.6 4.0 3.1 0
1.27 1.16 1.40 1.16 0.97
III III III III III
M.L. V.Yu. S.S. L.O. Sh.L.
1.8
0 0‘8 3.8 6.5 7.6
0.79 1.13 0.77 1.17 0.93
la la lb lb lb
D.Yu. Sh.a. M.a. s.g. L.M.
(W (X) 00 W)
Glucose-6phosphatase *
Glycogen
(g/v
OF LIVER TISSUE
Type of GSD
ANALYSIS
Patients
BIOCHEMICAL
TABLE I
15-56
5 8.8 4.5 6.6
3
3.5 29 9 14.5 0
27 23 22 54 20.5
Phosphorylase a*
15-W
16.6 28.6 11.8 22
4
18 12
6.6 27
33 23 17 50 20.6
Phosphorylase b*
25-100
56 40 27 135
40
64 27 22 36
44 65 80 173 -
Phosphoglucomutsse *
25-100
12.5 15 -
70
44 23 48 8
33 71 28 54 37
Phosphexoisomerase * *
0.4-1.1
0.81 0.44 0.4 0.6
0.39
0 0.1 0 0.23 0
1.22 0.36 0.43 0.50 0.74
Amylo-1,6glucosidase * * *
m+f
N0ma.l
* The number of controls.
3.0-15.0
0.5 9.5 15.0 5.5 1.5 3.0 23.0 9.0 3.5 7.0 4.0 5.0 4.0 2.0 4.5 4.0 2.0
f f f m f f m f f m f f m m m m m
D.Yu. Sh.a Yu.g M.a. s.g. L.M. D.V. M.L. V.Yu. S.S. L.O. Sh.L. G.Z. K.D. Sh.m. J.Zh. S.M.
(14 *I
Age (Yl.1
OF LIPOPROTEINS
Patients
THE CONTENT
TABLE II
la la 1 lb lb lb 1 III III III Hi III VI IX IX IX IX
(K) (K) (K) (K)
Type of glycogenosis
0.03 f 0.01
8.6 8.0 8.3 5.2 16.2 4.1 0.1 0.7 0.1 0.3 0.4 0.2 0.03 0.03 0.03 0.03 0.03
100400
Liproproteins Sf
IN BLOOD SERUM (ED) AND GLUCOSE
0.09 f 0.05
0.03 0.9 0.4 0.2 0.03 0.03 0.03 7.0 0.8 0.7 3.0 1.5 0.03 0.03 0.03 0.4 0.03
4.4 3.4 6.4 9.6 7.9 5.4 0.6 1.6 0.8 0.8 0.1 1.3 0.03 0.8 0.1 0.4 0.03 0.2 * 0.06
1 Z-20
CONTENT
20-100
AND LACTATE
3.6-5.3
1.84 4 0.2
<2.8
10.0 13.1 7.3 11.1 11.5 9.3 7.1 1.4 2.2 2.1 6.4 1.8 2.2 2.7 2.9 1.8 4.2
(mmol/l)
(mmoI!H
0.4 2.2 2.5 2.1 2.3 1 .I 3.1 1.9 1.4 2.3 1.7 2.4 1.3 3-3 3.7 2.9 2.4
Lactate
GIUCOSe
6.1 6.3 2.7 5.2 4.1 3.6 2.9 7.9 7.3 4.6 7.5 5.3 4.6 3.2 3.4 4.0 5.1
O-l 2
--
IN THE WHOLE BLOOD
103
The method employed to estimate the ability of liver homogenates to hydrolyse glucose-6-phosphate only reveals the absence of glucose-6-phosphatase. In cases of GSD Type I in which the activity of glucose-6-phosphatase in liver tissue homogenates was within normal limits, the permease activity was probably absent (GSD Type Ib) [ 13,141. Five of the patients (M.L., S.S., L.O., Sh.L., V.Yu.) were found to have GSD Type III. Amylo-1,6-glucosidase activity was lacking in the liver tissue of these patients. In two of them glucose-6-phosphatase activity was markedly reduced. However, the results obtained by glucose and adrenaline tolerance tests suggested that the lowering of glucose-6-phosphatase activity in these patients was secondary. Low phosphorylase activity was also found in the same patients. Such secondary changes in the glucose-6-phosphatase and phosphorylase activities are often observed in patients with GSD Type III [15]. Absorption spectra of the iodine complexes of glycogens isolated from the liver tissue of these patients were abnormal [ 4,16,17]. In patient G.S. with a considerable decrease in liver phosphorylases a and b activities, glucose-6-phosphatase activity was reduced; however, the tolerance test data also indicated in this case that the reduction of glucose-6-phosphatase activity was of a secondary nature thus establishing the GSD as Type VI. In patients K.D., Sh.M., I.Zh. and S.M. the activity of phosphorylase was normal in the presence, but decreased in the absence, of AMP and Na$04 suggesting a normal level of phosphorylase b activity and a deficiency either of phosphorylase b kinase (GSD Type IX) or of proteinkinase (GSD Type X). Table II presents data on the LDL and VLDL content in serum of 17 patients with GSD. As will be seen from these data, in six patients with GSD Type I, the content of VLDL with Sf 100-400 and 20-100, as well as the content of LDL with Sf O-12, was sharply increased. However, negligible or no changes were observed in the content of LDL with Sf 12-20 of these patients. According to the HLP classification proposed by the WHO [18] five patients with GSD Type I had HLP Type 2b, and one patient HLP Type 4. A different picture was observed in GSD of other types. For instance, GSD Type III was characterized by increased content of all LP fractions, especially of fractions with Sf O-12 and 12-20. All patients with this type of the disease had HLP 2b. In phosphorylase deficiency (Types VI, IX, X) only the content of LDL with Sf O-12 was increased (HLP 2a type). Thus, according to our data different changes in LP spectra are observed in different types of GSD. Discussion Quantitative determination of serum lipoproteins by analytical ultracentrifugation showed that the patients with Type I GSD had HLP Types 2b and 4, those with Type III had HLP Type 2b, and in those with Types VI and IX (or X) there was HLP Type 2a. Some patients with Types I and III disease had HLP of the same Type (2b). However, none of the patients with Type III had such a high VLDL level as those with Type I. On the other hand, none of the patients with GSD Type I had such a high level of LP with Sf 12-20 as those with GSD Type III.
104
In patients with Type I GSD not only the VLDL content but also that of blood lactate were markedly elevated, which was not observed in other patients, including those with a secondary deficiency of glucose-6-phosphatase (patients M.L., Sh.L., G.S.). It is not yet clear whether there is any interrelation between blood lactate content and changes in LP spectra in GSD. The cause of HLP in glycogenosis is not yet established. It is possible that the occurrence of HLP is connected with increased lipid synthesis, induced by disturbances of glycogen metabolism [ 31. The nature of changes in LP spectra in various types of glycogenoses is even less understood. In accordance with the generally accepted fact that LDL with Sf 12-20 and O-12 are formed as a result of metabolic transformations of VLDL with Si 100-400 and 20-100, these transformations may be expected to occur mainly in Types III, VI and IX (X) GSD but not in Type I disease. It might also be that in Type I GSD the VLDL synthesis is intensified to a greater extent than in other types of GSD. Data showing the differences in serum LP spectra may be used as additional evidence in the diagnosis of the type of GSD. References 1 MC Adams, A.J., Hug, G. and Bow, K.E. (1974) Hum. Pathol. 5,463-487 2 R. Rodney Howell (1972) in: The Met,aboIic Basis of Inherited Disease (Stanbury, J.B., Wyngaarden, J.B. and Fredrickson, D.S.. eds.), pp. 149-173, McGraw Hill, New York 3 Forget, P.P.. Fernandes, J. and Begemann, P.H. (1974) Pediatr. Res. 8, 114-119 4 Rosenfeld, E.L., Popova, LA. and Chibisov. I.V. (1976) Clin. Chin Acta 67, 123-130 5 Iahnke, K. and Scholtan, W. (1960) in Die Bluteiweissk6r:rper in der Ultracentrifuge. PP. 118-124. G. Thieme Verlag. Stuttgart 6 De Lalla. O.F., Tandy, R.K. and Loeb, H.G. (1967) Clin. Chem. 13. 85-100 7 Huijing, F. (1974) in Clinical Biochemistry (Curtius, H.Ch.. and Rot, M.. eds.). PP. 1203-1235. Walter de Gruyter. Berlin. New York 8 Hers, H.G. (1964) in Advances in Metabolic Disorders (Levine, R. and Jabt, R.. eds.) Vol. 1, PP. l44. Academic Press, New York 9 StaIrnan. J.W. and Hers, H.G. (1975) Europ. J. Biochem. 54, 341-350 10 Najjar. V.A. (1955) in Methods in Enzymology (Colowick. S.P. and KaPlan, N.O., eds.) Vol. 1. PP. 294-298, Academic Press Inc., New York 11 Slain, M.W. (1955) in Methods in Enzymology (Colowick, S.P. and Kaplan, N.O., eds.) Vol. 1, PP. 299-305, Academic Press Inc., New York 12 Arion, J.W., Lange, A.J. and BaIlas. L.M. (1976) J. Biol. Chem. 251.4901-4907 13 Naresawa, K., Jagarashi, L., Otomo. H. and Tada, K. (1978) Biochem. Biophys. Res. Comm. 83, 1360-1364 14 Rosenfeld, E.L., Chibisov, I.V., Chistova, L.V., Leontjev. A.F. and Karmansky, I.M. (1978) CIin. Chim. Acta 86, 295-299 15 Rossignol, A.M., Bost. M.. Marchal, A., Frappart, P., Stoebner. P. and Beaudoin, S.A. (1975) Ann. Pediatr. 22, 717-725 16 Chibisov. I.V., Karmansky. I.M. Cheljapina, V.V.. Leontjev, A.F. and Rosenfeld. E.L. (1978) Vo~r. Med. Chem. 24, 555-559. (Russian) 17 Chibisov, I.V., Rosenfeld, E.L., Tabolin, V.A., Komeva, T.I. and Muchina, Yu.G. (1977) Pediatria 11, 46-51, (Russian) 18 Beaumont, J.L., Carlsson, L.A.. Cooper, G.R., Fejfar. Z., Fredrickson, D.S. and Strasser, T. (1970) Bull. W.H.O. 43. 891