Aust N Z J Med (1979), 9, pp 4Y 53 ~~~
Haemoglobin A, in Diabetes Mellitus P. C. Bartley* and T. E. Hamblingt From t h e R e p a t r i a t i o n G e n e r a l Hospital, G r e e n s l o p e s , Q u e e n s l a n d
SUmmary:
Haemoglobin A, in diabetes mellitus. P. C. Bartley a n d T: E. Harnbling, Aust. N.Z. J . Med., 1979: 9, p p . 4S.53.
HbA, was measured by a rapid method in a routine hospital laboratory. The normal range of 5.0-8.5% compares favourably with other studies. Patients with uncontrolled diabetes had levels of HbA, of up to 20.5% while well controlled patients had levels of HbA, in the normal range. The level of HbA, is relatively stable in normal subjects and does not correlate with age, sex or weight. There was a correlation (p 90% negative tests in three months. Fair = 50-90% negative tests in three months. Poor = i 50% negative tests in three months.
Two patients were studied for three months from their initial presentation with diabetic ketoacidosis. The first patient (Fig. 5 ) presented with a blood sugar of 55 mmo1:I at which time the HbA, was 13.8>”. Within five days, the blood
sugar was less tlyan 10 mmol/l and the HbA, reached normal ( < 8.5’3;,) within one month. The second patient (Fig. 6) prcsentcd with a blood sugar of 60mmolil and the HbA, was 1 9 . 1 ~ Twelve ~. days later the H b h , was unchanged despite a large reduction in his blood sugar and one week later it had decreased by 50‘),,, but it was not until three months after onset that the HbA, was returned t o within the normal range ( < 8 . So/;,). Over the last two months fourtimes-a-day urine tests with “Clinitest” were negative. The blood sugar taken at the diabetic clinic at this hospital is taken 1 - 2 hours after breakfast and insulin injection. When the sugar level obtained at that time was correlated with the ”.;, HbA,, there was a highly significant correlation (Fig. 7) P < 0.001.
52
12. I
Q
0 8 I
-9.
VOL.
HAKTI.1.Y A h l ) HAMHLING
-
s
10
20
30 40 50 G l u c o s e m m o l /l
FIGURE 7. Correlation of % HbA, with blood glucose as taken at the diabetic clinic.
Discussion
HbAI, was isolated in 195812and shown by ionexchange chromatography to cotnprise 5-79,:, of the Hb found in normal human erythrocytes. In 1968 an unusual haemoglobin was reported in patients with diabetes mellitus.'3 Later it was reported that this unusual Hb rcsemblcd HbA,, and was elevated two-fold in patients with diabetes mellitus.'" Studies have revealed that an amino-terminal sugar was bound to the beta chains of HbA,, in diabetic subjects.15 In diabetic patients HbAI, and HbAlh which comprisc 1-2(vn of the total Hb in normal subjects, increased to 2-37;. But HbAI, which was 3.3-3.5% in normals was increased LO 6 10% in patients with diabetes inellitus." This study did not find any relationship between HbA, and age or duration of. or type of, therapy of diabetes or presence of renal impairment. but in patients with pneumonia or ketoacidosis the percentage of HbAI, was significantly higher than in other diabetic patients. Later it was shown that HbAI, was reduced towards normal after controlling the diabetic state.5 Also, any change downwards in the blood sugar preceded the decrease in HbA1, by three to four weeks. This is compatible with the fact that the glycosylation of the H b is a post-translational one and therefore continuous formation
YO.
1
and accumulation occurs within the erythrocyte during its life span.'7. '* In this study, we have used a method which is cheap. takes three hours and is precise. In normal subjects the range of 5-8.5'5b for the total glycosylated H b compares favourably with earlier reports (Table 1). We have confirmed the finding of Trivelli et d.",that there is no correlation of HbA, with weight and have extended that to show the absence of a coi-relation with agc or sex. The range for our diabetic subjects of 5.5-20.5:,, overlapped with that of our normal subjects. When the patients with diabetes treated with insulin were separated on the basis of their previous thrcc months' urine tests (performed four timesiclay), it was apparent that the level of HbA, decreased with better diabetic control and in some patients, returned to normal levels. This change was shown best in the study of the two ketoacidotic patients. In one patient (P.D.). the HbA, remained unchanged over the first 12 days and then in a further seven days decreased by 501;; and returned to normal in thrcc months. The blood sugar taken over the last two months was always less than 10 nimolil. We are unable to explain the sudden drop in HbA, which occurred between the twelfth and twentieth days. The second patient (R.) exhibited a fall 01- 2.5% in the HbA, in the first fourteen days and had returned to normal in six weeks. Therc is clearly a lag of about six weeks between return of the blood sugar to acceptable levels and the return of HbA, to normal. TABLE 1
Book~liitiand Gallop" HbAI, 5-7 Rahbar c r a/ l4 HbAI, 4 6
Trivelli er a1 ''
7 . 5 - 10.6 -- 3 6 -10 KetnacidoGs 1 4 3 i 3 . 6 L;iicontrolled Y 8 Controlled 5 8 1 h A l 7.45 12.45 HbAb, -1, 2 - 1 3-2 HhAi.. 4.Y 10.0
HbA1, . h I 2 HhA,, 3.3-3.5
2
This study
Oral hypoglycacinics I 1-09 t 2 - 0 9 (SD) Diet alone 10.2 1 0 . 6 2 (SD)
When patients presenting with diabetes mellitus, which was previously untreated, were
FEBRUARY
1979
53
HAEMOGLOBIN A, lh 1)IABbTbS MELLITUS
subdivided on the basis of their therapy (decided upon without any knowledge of the HbA,), it can be seen that there exists a difference. Patients who were subsequently trea tcd with insulin had the highest levels of HbA, and those treated with diet the lowest. What the difference in the mean HbA, level indicates is unknown. It is possible that it is only an indicator of the height of the blood sugar which rises more rapidly in insulin deficiency but it is also probable that the patients not treated with insulin had hyperglycaemia, albeit milder, for a longer period of time. What combination of duration of hyperglycaemia, rate of rise of blood sugar and height of blood sugar results in the level of HbA, found at presentation is as yet unknown. The blood sugar level taken at the diabetic clinic at this hospital is drawn about two hours after the breakfast and insulin. Thc blood sugar at this time in insulin treated patients correlated highly with the level of HbA,. When the two patients with ketoacidosis were included, the correlation coefficient was reduced but still highly significant. It is possible that this correlation is a chance finding, but we believe that taking a blood sugar in the above manner represents one of the higher levels of blood glucose which is achieved during the day. This study has shown that measurement of HbA, can be performed in a routine hospital laboratory. It has also confirmed that HbA, alters with the state of diabetic control and that it can be returned to normal levels. The fact that HbA, reflects the blood sugar over the previous six weeks is an invaluable adjunct to the
""
management of diabetes inellitus. The advantage of measurement of HbA, over a random blood sugar is immense. Whether the decrease in the percentage of glycosylated H b with "good" control means that other proteins can also be "deglycosylated" will only be answered in time. References 1.i BoSI)Y. P. (1971): Relation of diabetic cantrol to H o s i n . P. 1'. K. K.and and I'F.LIG. I ' t i i i i . P. 11971): Rclattm :\fed. Clin .2V..411wr, 55. 889.897. dc\ielopment complications. dc\elopment ofofrascirlar ~ n ~ c i i cornplic,itions. l~ir l f i d ('im 2.3 RYAS. J. R., BALOUIMOS. M. C., 8. I.. Rtnn. FI. I:.. MAKRI.I..A.. ny,\\. J R , H,m1jriios. M. c ,CIUZ.A\, C I I A ~ A \ ,H. I.. ni WIIITX. P. and Jiai.ix. A. P. (1970): Quarter century victory nicdal lor W t i n k . I' and J i ~iz. i 9 P. (1970) Quartrr w n diabetes: tu twenty ycars later. Murabohi 19. illnhetGs .4A follow-up Iblloa-upof otpatients pat,cnt>onc o w 10 I*C111> ycars Ia1cr. 493. 493,501. 501 3.7 CAHII.I., G. F., JxR., I:lzwIi.I:~. D. I ) . FRli~snii.,X. (1976): "Conlrol" C'.\HIII . Ci I J\n., I l l w i i I'R, D 11.and hiid I ~ I . 1 ~ 6 1 and diabetcs, NW h g l . J. Med 294, 1004 .I005 4. WIKKO. A. and li:I.lG, 1'. (1Y7X): Elfwts of leg exercise on insulin absorption in diabetic palients. Xeir h g l . J. h f d 298. 79-43. 5 . KOFNIG.K.Ci., PFTIIRSOX. C.Y.,los~s,R.I... SACDEK, C.. I.ERKMAS, M.and CERAMI. A. (1976): Cornlation ofglucox regulationnand Alein a n dhaemoglobin haei;,oglnhinAiCiii diabetes mcllitus. N r u R i g / . .I. M v d 295, 417.420. 6. KOEMG. R.I.. P E ~ E ~ OC. X M., . Kiln. C..CI:RAMI.A. and WII.I.IAMMIS~ and Wit I IAVSOS. J.J K. K (19%): Aicas an indicator ofthc degrcccol'glucusc (19%))-Hacmoglobin Ha~mo_eli.knAiuncsnlndlcati d'plucoszinlulcrance iritdcrancc inIIIdidhelcs. I)iuhucv 25. 230-232. diahctch. l)tuhk,\25. 230 232 7. PE~IXWJN. C. %l.,JIISI:%. R. I... K o l w i t ; . R. J.. Mei.vi~.. E. T. and LIEIRMAV. M. L. (1977): Reversible hacmatologic rcquclac or diabetcs mellitus. .4nn. intern. hled. 86. 425. 429. 8. GABBAY. K.It., tinsru. K..BW~I.OW,J. I... 1:i.i.ism. K.C..131:zs. H. I:.and GAI.I.oP.P. Xj. 1 1977): Glywsylatcd haemoglobins and long-term blood gluwse control in diabetes mellitus. J. rliii. Eridurr. 44, 8SY..8M. 9. Ci)sEs, B., KLnmsTias. A. I
Haemoglobin A, in Diabetes Mellitus P. C. Bartley* and T. E. Hamblingt From t h e R e p a t r i a t i o n G e n e r a l Hospital, G r e e n s l o p e s , Q u e e n s l a n d
SUmmary:
Haemoglobin A, in diabetes mellitus. P. C. Bartley a n d T: E. Harnbling, Aust. N.Z. J . Med., 1979: 9, p p . 4S.53.
HbA, was measured by a rapid method in a routine hospital laboratory. The normal range of 5.0-8.5% compares favourably with other studies. Patients with uncontrolled diabetes had levels of HbA, of up to 20.5% while well controlled patients had levels of HbA, in the normal range. The level of HbA, is relatively stable in normal subjects and does not correlate with age, sex or weight. There was a correlation (p 90% negative tests in three months. Fair = 50-90% negative tests in three months. Poor = i 50% negative tests in three months.
Two patients were studied for three months from their initial presentation with diabetic ketoacidosis. The first patient (Fig. 5 ) presented with a blood sugar of 55 mmo1:I at which time the HbA, was 13.8>”. Within five days, the blood
sugar was less tlyan 10 mmol/l and the HbA, reached normal ( < 8.5’3;,) within one month. The second patient (Fig. 6) prcsentcd with a blood sugar of 60mmolil and the HbA, was 1 9 . 1 ~ Twelve ~. days later the H b h , was unchanged despite a large reduction in his blood sugar and one week later it had decreased by 50‘),,, but it was not until three months after onset that the HbA, was returned t o within the normal range ( < 8 . So/;,). Over the last two months fourtimes-a-day urine tests with “Clinitest” were negative. The blood sugar taken at the diabetic clinic at this hospital is taken 1 - 2 hours after breakfast and insulin injection. When the sugar level obtained at that time was correlated with the ”.;, HbA,, there was a highly significant correlation (Fig. 7) P < 0.001.
52
12. I
Q
0 8 I
-9.
VOL.
HAKTI.1.Y A h l ) HAMHLING
-
s
10
20
30 40 50 G l u c o s e m m o l /l
FIGURE 7. Correlation of % HbA, with blood glucose as taken at the diabetic clinic.
Discussion
HbAI, was isolated in 195812and shown by ionexchange chromatography to cotnprise 5-79,:, of the Hb found in normal human erythrocytes. In 1968 an unusual haemoglobin was reported in patients with diabetes mellitus.'3 Later it was reported that this unusual Hb rcsemblcd HbA,, and was elevated two-fold in patients with diabetes mellitus.'" Studies have revealed that an amino-terminal sugar was bound to the beta chains of HbA,, in diabetic subjects.15 In diabetic patients HbAI, and HbAlh which comprisc 1-2(vn of the total Hb in normal subjects, increased to 2-37;. But HbAI, which was 3.3-3.5% in normals was increased LO 6 10% in patients with diabetes inellitus." This study did not find any relationship between HbA, and age or duration of. or type of, therapy of diabetes or presence of renal impairment. but in patients with pneumonia or ketoacidosis the percentage of HbAI, was significantly higher than in other diabetic patients. Later it was shown that HbAI, was reduced towards normal after controlling the diabetic state.5 Also, any change downwards in the blood sugar preceded the decrease in HbA1, by three to four weeks. This is compatible with the fact that the glycosylation of the H b is a post-translational one and therefore continuous formation
YO.
1
and accumulation occurs within the erythrocyte during its life span.'7. '* In this study, we have used a method which is cheap. takes three hours and is precise. In normal subjects the range of 5-8.5'5b for the total glycosylated H b compares favourably with earlier reports (Table 1). We have confirmed the finding of Trivelli et d.",that there is no correlation of HbA, with weight and have extended that to show the absence of a coi-relation with agc or sex. The range for our diabetic subjects of 5.5-20.5:,, overlapped with that of our normal subjects. When the patients with diabetes treated with insulin were separated on the basis of their previous thrcc months' urine tests (performed four timesiclay), it was apparent that the level of HbA, decreased with better diabetic control and in some patients, returned to normal levels. This change was shown best in the study of the two ketoacidotic patients. In one patient (P.D.). the HbA, remained unchanged over the first 12 days and then in a further seven days decreased by 501;; and returned to normal in thrcc months. The blood sugar taken over the last two months was always less than 10 nimolil. We are unable to explain the sudden drop in HbA, which occurred between the twelfth and twentieth days. The second patient (R.) exhibited a fall 01- 2.5% in the HbA, in the first fourteen days and had returned to normal in six weeks. Therc is clearly a lag of about six weeks between return of the blood sugar to acceptable levels and the return of HbA, to normal. TABLE 1
Book~liitiand Gallop" HbAI, 5-7 Rahbar c r a/ l4 HbAI, 4 6
Trivelli er a1 ''
7 . 5 - 10.6 -- 3 6 -10 KetnacidoGs 1 4 3 i 3 . 6 L;iicontrolled Y 8 Controlled 5 8 1 h A l 7.45 12.45 HbAb, -1, 2 - 1 3-2 HhAi.. 4.Y 10.0
HbA1, . h I 2 HhA,, 3.3-3.5
2
This study
Oral hypoglycacinics I 1-09 t 2 - 0 9 (SD) Diet alone 10.2 1 0 . 6 2 (SD)
When patients presenting with diabetes mellitus, which was previously untreated, were
FEBRUARY
1979
53
HAEMOGLOBIN A, lh 1)IABbTbS MELLITUS
subdivided on the basis of their therapy (decided upon without any knowledge of the HbA,), it can be seen that there exists a difference. Patients who were subsequently trea tcd with insulin had the highest levels of HbA, and those treated with diet the lowest. What the difference in the mean HbA, level indicates is unknown. It is possible that it is only an indicator of the height of the blood sugar which rises more rapidly in insulin deficiency but it is also probable that the patients not treated with insulin had hyperglycaemia, albeit milder, for a longer period of time. What combination of duration of hyperglycaemia, rate of rise of blood sugar and height of blood sugar results in the level of HbA, found at presentation is as yet unknown. The blood sugar level taken at the diabetic clinic at this hospital is drawn about two hours after the breakfast and insulin. Thc blood sugar at this time in insulin treated patients correlated highly with the level of HbA,. When the two patients with ketoacidosis were included, the correlation coefficient was reduced but still highly significant. It is possible that this correlation is a chance finding, but we believe that taking a blood sugar in the above manner represents one of the higher levels of blood glucose which is achieved during the day. This study has shown that measurement of HbA, can be performed in a routine hospital laboratory. It has also confirmed that HbA, alters with the state of diabetic control and that it can be returned to normal levels. The fact that HbA, reflects the blood sugar over the previous six weeks is an invaluable adjunct to the
""
management of diabetes inellitus. The advantage of measurement of HbA, over a random blood sugar is immense. Whether the decrease in the percentage of glycosylated H b with "good" control means that other proteins can also be "deglycosylated" will only be answered in time. References 1.i BoSI)Y. P. (1971): Relation of diabetic cantrol to H o s i n . P. 1'. K. K.and and I'F.LIG. I ' t i i i i . P. 11971): Rclattm :\fed. Clin .2V..411wr, 55. 889.897. dc\ielopment complications. dc\elopment ofofrascirlar ~ n ~ c i i cornplic,itions. l~ir l f i d ('im 2.3 RYAS. J. R., BALOUIMOS. M. C., 8. I.. Rtnn. FI. I:.. MAKRI.I..A.. ny,\\. J R , H,m1jriios. M. c ,CIUZ.A\, C I I A ~ A \ ,H. I.. ni WIIITX. P. and Jiai.ix. A. P. (1970): Quarter century victory nicdal lor W t i n k . I' and J i ~iz. i 9 P. (1970) Quartrr w n diabetes: tu twenty ycars later. Murabohi 19. illnhetGs .4A follow-up Iblloa-upof otpatients pat,cnt>onc o w 10 I*C111> ycars Ia1cr. 493. 493,501. 501 3.7 CAHII.I., G. F., JxR., I:lzwIi.I:~. D. I ) . FRli~snii.,X. (1976): "Conlrol" C'.\HIII . Ci I J\n., I l l w i i I'R, D 11.and hiid I ~ I . 1 ~ 6 1 and diabetcs, NW h g l . J. Med 294, 1004 .I005 4. WIKKO. A. and li:I.lG, 1'. (1Y7X): Elfwts of leg exercise on insulin absorption in diabetic palients. Xeir h g l . J. h f d 298. 79-43. 5 . KOFNIG.K.Ci., PFTIIRSOX. C.Y.,los~s,R.I... SACDEK, C.. I.ERKMAS, M.and CERAMI. A. (1976): Cornlation ofglucox regulationnand Alein a n dhaemoglobin haei;,oglnhinAiCiii diabetes mcllitus. N r u R i g / . .I. M v d 295, 417.420. 6. KOEMG. R.I.. P E ~ E ~ OC. X M., . Kiln. C..CI:RAMI.A. and WII.I.IAMMIS~ and Wit I IAVSOS. J.J K. K (19%): Aicas an indicator ofthc degrcccol'glucusc (19%))-Hacmoglobin Ha~mo_eli.knAiuncsnlndlcati d'plucoszinlulcrance iritdcrancc inIIIdidhelcs. I)iuhucv 25. 230-232. diahctch. l)tuhk,\25. 230 232 7. PE~IXWJN. C. %l.,JIISI:%. R. I... K o l w i t ; . R. J.. Mei.vi~.. E. T. and LIEIRMAV. M. L. (1977): Reversible hacmatologic rcquclac or diabetcs mellitus. .4nn. intern. hled. 86. 425. 429. 8. GABBAY. K.It., tinsru. K..BW~I.OW,J. I... 1:i.i.ism. K.C..131:zs. H. I:.and GAI.I.oP.P. Xj. 1 1977): Glywsylatcd haemoglobins and long-term blood gluwse control in diabetes mellitus. J. rliii. Eridurr. 44, 8SY..8M. 9. Ci)sEs, B., KLnmsTias. A. I
