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Salivary secretion of albumin in type 1 (insulin-dependent) diabetes
Diabetes Research and Clinical Practice, 11 (1991) 117-120
117
Elsevier DIABET
00468
Brief Report
Salivary secretion of albumin in type 1 (insulin-dependent) diabetes B.M. Fisher I, P.-J. Lamey3, D. Sweeney 3, J.A. Beeley3, R.J. Spooner2 ‘Diabetic Department
and ‘Department
of Pathological Biochemistry,
and B.M. Frier ’
Western InJirmary and Gartnavel General Hospital and
3Department of Oral Medicine and Pathology, Glasgow Dental Hospital and School, Glasgow, U.K.
(Received 26 August 1990) (Accepted 5 October 1990)
Summary The concentration of albumin in saliva is low in healthy humans. To determine whether alterations in capillary permeability in diabetes affects the salivary glands, the concentration of albumin in parotid saliva was measured in 26 Type 1 (insulin-dependent) diabetic patients, and compared to 32 non-diabetic control subjects. The diabetic patients were subdivided into 3 groups on the basis of the urinary excretion of albumin in timed overnight collections of urine: (1) normal albumin excretion (< 30 pg/min) n = 13 ; (2) microalbuminuria (30-300 pg/min) y1= 7, and (3) macroalbuminuria (> 300 pg/min) n = 6. Saliva was collected for one minute following stimulation with 1 ml 10% citric acid, and the concentration of albumin was measured by a sensitive ELBA method. No significant difference in salivary albumin concentration was found between the control group and any of the diabetic groups. Thus, although urinary albumin excretion was increased, suggesting altered capillary permeability, simultaneous leakage of albumin into saliva was not observed. Measurement of salivary albumin concentration does not, therefore, provide a marker of occult microvascular disease in diabetes. Key words: Saliva; Albumin;
Insulin-dependent
diabetes;
Introduction In healthy, non-diabetic humans albumin is excreted in small amounts in the urine and saliva. The composition of saliva is altered in patients with diabetes, with increased secretion of calcium and immunoglobulin [ 11, and it has been sugCorrespondence to: B.M. Fisher, Wards 415, Royal Infirmary, Glasgow, G4 OSF, U.K.
0168-8227/91/$03.50
0 1991 Elsevier Science Publishers
Proteinuria;
Microalbuminuria
gested that this indicates an increase in microvascular permeability [ I]. If this is so, other salivary constituents such as albumin which has a lower molecular weight than immunoglobulins, and which is not secreted by salivary acinar cells but enters saliva following leakage from surrounding blood vessels, should be present in increased concentration in the saliva. The aim of the present study was to examine the concentration of albumin in the saliva of Type 1 (insulin-depen-
B.V. (Biomedical
Division)
118
dent) diabetic patients, and to examine whether any relationship exists with urinary albumin excretion, which might provide a marker for generalised microangiopathy in diabetes.
Materials and Methods Approval for this study was given by the local medical ethical advisory committee and informed consent was obtained from all subjects. Twentysix Type 1 diabetic patients, age range 23 to 57 years, were studied. Patients who had a long duration of diabetes (range 5 to 39 years, mean 18 years), with varying degrees of glycaemic control (HbAl mean 9.8x, normal < 8%) and who had established complications of diabetes were recruited. Eleven patients had background retinopathy, assessed by direct ophthalmoscopy, and 9 patients had received photocoagulation for proliferative retinopathy. Four patients had clinical evidence of peripheral neuropathy with diminished reflexes and absent vibration sense in the feet. A series of cardiovascular reflexes were performed to assess autonomic function [2], and seventeen patients were found to have normal autonomic function (score O-2), six to have definite autonomic neuropathy (score 3-5), and three to have severe autonomic neuropathy (score 6-10). However, because of the small numbers in each group it was not possible to evaluate salivary albumin excretion depending on the severity of autonomic neuropathy. Patients were subdivided into 3 groups on the basis of the urinary excretion of albumin measured in two, timed overnight collections of urine as follows: (a) normal urinary albumin excretion ( < 30 pg/min) it = 13, (b) microalbuminuria (30-300 pg/min) n = 7, and (c) macroalbuminuria (> 300 pg/min) n = 6. Comparisons were made with 32 healthy age- and sex-matched nondiabetic control subjects. Stimulated parotid salivary flow was performed at 09.00 hours in all subjects. A Carlsson-Crittenden cup was placed over each parotid orifice and 1 ml of a 10% solution of citric acid applied to the
Control
Group
1
Group
2
Group
3
Groups
Fig. 1. Salivary albumin concentrations in control subjects, and in diabetic patients with no albuminuria (Group l), microalbuminuria (Group 2) and macroalbuminuria (Group 3). No significant differences in salivary albumin concentrations existed between the control group and any of the diabetic groups.
dorsum of the tongue. Saliva was collected for 1 min, placed in plastic vials, and stored at - 25 ‘C. Samples were assayed in quadruplicate for albumin using a double antibody sandwich ELISA technique [ 31. Comparisons of salivary flow between the control and each of the diabetic groups were assessed by the Mann-Whitney U test. The results are expressed as mean + SEM.
Results The salivary concentrations of albumin are shown in Fig. 1. No significant difference was found between the control group and any of the diabetic sub-groups.
Discussion Many functional similarities between the kidneys and salivary glands exist in humans. In both organs, a fluid is produced which passes through a collecting system, where its concentration and constituents are altered by both active and pas-
119
sive mechanisms. A small amount of protein normally enters the urine via the glomerulus of the kidney. The urine passes through the loop of Henle and collecting tubules where there is reabsorption of water, and exchange of electrolytes. By contrast, in the salivary gland, water, electrolytes and protein are actively secreted by the acinar cells, resulting in an isotonic fluid. During passage through the collecting ducts there is active resorption of electrolytes, and to a lesser extent absorption of water, producing a hypotonic saliva. Increased urinary excretion of albumin in diabetic patients indicates the development of early diabetic nephropathy, with increased capillary permeability allowing leakage of albumin into the urine [4]. In the present study, the albumin concentration was not increased in the parotid saliva of diabetic patients, including patients with diabetic nephropathy, and measurement of salivary albumin excretion in diabetic patients does not appear to be of value in indicating the presence of microvascular disease. Although the lack of an increase of parotid salivary albumin in diabetic patients who had either microalbuminuria or macroproteinuria could indicate selective organ damage, it is likely that the differences in structure and physiology underlay this apparent discrepancy. It is unlikely that examination of other salivary glands would have modified the present findings, as the saliva produced by other glands contains more mucin and less protein than
the parotid salivary secretion. An increase in the transcapillary escape rate of albumin has been demonstrated in patients with microalbuminuria [5], suggesting the presence of a generalised microvascular dysfunction. A similar result was observed when transfer of smaller protein molecules such as transferrin was estimated [6]. It is possible that capillary leakage of some smaller proteins may occur into the saliva of diabetic patients, and this may be worthy of further investigation.
References Marder, M., Abelson, D.C. and Mandel, I.D. (1975) Salivary alterations in diabetes mellitus. J. Periodontol. 46, 567-569. Fisher, B.M. and Frier, B.M. (1989) Diagnostic yield of tests used to screen asymptomatic patients for diabetic autonomic neuropathy. Diabetes Res. Clin. Pratt. 6, 157-160. Sweeney, D. and Beeley, J.A. (1990) An enzyme linked immunoassay for human salivary albumin. Arch. Oral Biol., in press. Viberti, G. and Keen, H. (1984) The pattern of proteinuria in diabetes mellitus. Diabetes 33, 686-692. Bent-Hansen, L., Feldt-Rasmussen, B., Kverneland, A. and Deckert, T. (1987) Transcapillary escape rate and relative metabolic clearance of glycated and non-glycated albumin in Type 1 (insulin-dependent) diabetes mellitus. Diabetologia 30, 2-4. Bell, D., Collier, A., Nicoll, J.J., Jackson, M., Millar, A.M., Clarke, B.F. and Muir, A.L. (1988) Reduced venous compliance and increased transcapillary escape of protein in insulin-dependent diabetic patients. Diabetic Med. 5, 454-458.
117
Elsevier DIABET
00468
Brief Report
Salivary secretion of albumin in type 1 (insulin-dependent) diabetes B.M. Fisher I, P.-J. Lamey3, D. Sweeney 3, J.A. Beeley3, R.J. Spooner2 ‘Diabetic Department
and ‘Department
of Pathological Biochemistry,
and B.M. Frier ’
Western InJirmary and Gartnavel General Hospital and
3Department of Oral Medicine and Pathology, Glasgow Dental Hospital and School, Glasgow, U.K.
(Received 26 August 1990) (Accepted 5 October 1990)
Summary The concentration of albumin in saliva is low in healthy humans. To determine whether alterations in capillary permeability in diabetes affects the salivary glands, the concentration of albumin in parotid saliva was measured in 26 Type 1 (insulin-dependent) diabetic patients, and compared to 32 non-diabetic control subjects. The diabetic patients were subdivided into 3 groups on the basis of the urinary excretion of albumin in timed overnight collections of urine: (1) normal albumin excretion (< 30 pg/min) n = 13 ; (2) microalbuminuria (30-300 pg/min) y1= 7, and (3) macroalbuminuria (> 300 pg/min) n = 6. Saliva was collected for one minute following stimulation with 1 ml 10% citric acid, and the concentration of albumin was measured by a sensitive ELBA method. No significant difference in salivary albumin concentration was found between the control group and any of the diabetic groups. Thus, although urinary albumin excretion was increased, suggesting altered capillary permeability, simultaneous leakage of albumin into saliva was not observed. Measurement of salivary albumin concentration does not, therefore, provide a marker of occult microvascular disease in diabetes. Key words: Saliva; Albumin;
Insulin-dependent
diabetes;
Introduction In healthy, non-diabetic humans albumin is excreted in small amounts in the urine and saliva. The composition of saliva is altered in patients with diabetes, with increased secretion of calcium and immunoglobulin [ 11, and it has been sugCorrespondence to: B.M. Fisher, Wards 415, Royal Infirmary, Glasgow, G4 OSF, U.K.
0168-8227/91/$03.50
0 1991 Elsevier Science Publishers
Proteinuria;
Microalbuminuria
gested that this indicates an increase in microvascular permeability [ I]. If this is so, other salivary constituents such as albumin which has a lower molecular weight than immunoglobulins, and which is not secreted by salivary acinar cells but enters saliva following leakage from surrounding blood vessels, should be present in increased concentration in the saliva. The aim of the present study was to examine the concentration of albumin in the saliva of Type 1 (insulin-depen-
B.V. (Biomedical
Division)
118
dent) diabetic patients, and to examine whether any relationship exists with urinary albumin excretion, which might provide a marker for generalised microangiopathy in diabetes.
Materials and Methods Approval for this study was given by the local medical ethical advisory committee and informed consent was obtained from all subjects. Twentysix Type 1 diabetic patients, age range 23 to 57 years, were studied. Patients who had a long duration of diabetes (range 5 to 39 years, mean 18 years), with varying degrees of glycaemic control (HbAl mean 9.8x, normal < 8%) and who had established complications of diabetes were recruited. Eleven patients had background retinopathy, assessed by direct ophthalmoscopy, and 9 patients had received photocoagulation for proliferative retinopathy. Four patients had clinical evidence of peripheral neuropathy with diminished reflexes and absent vibration sense in the feet. A series of cardiovascular reflexes were performed to assess autonomic function [2], and seventeen patients were found to have normal autonomic function (score O-2), six to have definite autonomic neuropathy (score 3-5), and three to have severe autonomic neuropathy (score 6-10). However, because of the small numbers in each group it was not possible to evaluate salivary albumin excretion depending on the severity of autonomic neuropathy. Patients were subdivided into 3 groups on the basis of the urinary excretion of albumin measured in two, timed overnight collections of urine as follows: (a) normal urinary albumin excretion ( < 30 pg/min) it = 13, (b) microalbuminuria (30-300 pg/min) n = 7, and (c) macroalbuminuria (> 300 pg/min) n = 6. Comparisons were made with 32 healthy age- and sex-matched nondiabetic control subjects. Stimulated parotid salivary flow was performed at 09.00 hours in all subjects. A Carlsson-Crittenden cup was placed over each parotid orifice and 1 ml of a 10% solution of citric acid applied to the
Control
Group
1
Group
2
Group
3
Groups
Fig. 1. Salivary albumin concentrations in control subjects, and in diabetic patients with no albuminuria (Group l), microalbuminuria (Group 2) and macroalbuminuria (Group 3). No significant differences in salivary albumin concentrations existed between the control group and any of the diabetic groups.
dorsum of the tongue. Saliva was collected for 1 min, placed in plastic vials, and stored at - 25 ‘C. Samples were assayed in quadruplicate for albumin using a double antibody sandwich ELISA technique [ 31. Comparisons of salivary flow between the control and each of the diabetic groups were assessed by the Mann-Whitney U test. The results are expressed as mean + SEM.
Results The salivary concentrations of albumin are shown in Fig. 1. No significant difference was found between the control group and any of the diabetic sub-groups.
Discussion Many functional similarities between the kidneys and salivary glands exist in humans. In both organs, a fluid is produced which passes through a collecting system, where its concentration and constituents are altered by both active and pas-
119
sive mechanisms. A small amount of protein normally enters the urine via the glomerulus of the kidney. The urine passes through the loop of Henle and collecting tubules where there is reabsorption of water, and exchange of electrolytes. By contrast, in the salivary gland, water, electrolytes and protein are actively secreted by the acinar cells, resulting in an isotonic fluid. During passage through the collecting ducts there is active resorption of electrolytes, and to a lesser extent absorption of water, producing a hypotonic saliva. Increased urinary excretion of albumin in diabetic patients indicates the development of early diabetic nephropathy, with increased capillary permeability allowing leakage of albumin into the urine [4]. In the present study, the albumin concentration was not increased in the parotid saliva of diabetic patients, including patients with diabetic nephropathy, and measurement of salivary albumin excretion in diabetic patients does not appear to be of value in indicating the presence of microvascular disease. Although the lack of an increase of parotid salivary albumin in diabetic patients who had either microalbuminuria or macroproteinuria could indicate selective organ damage, it is likely that the differences in structure and physiology underlay this apparent discrepancy. It is unlikely that examination of other salivary glands would have modified the present findings, as the saliva produced by other glands contains more mucin and less protein than
the parotid salivary secretion. An increase in the transcapillary escape rate of albumin has been demonstrated in patients with microalbuminuria [5], suggesting the presence of a generalised microvascular dysfunction. A similar result was observed when transfer of smaller protein molecules such as transferrin was estimated [6]. It is possible that capillary leakage of some smaller proteins may occur into the saliva of diabetic patients, and this may be worthy of further investigation.
References Marder, M., Abelson, D.C. and Mandel, I.D. (1975) Salivary alterations in diabetes mellitus. J. Periodontol. 46, 567-569. Fisher, B.M. and Frier, B.M. (1989) Diagnostic yield of tests used to screen asymptomatic patients for diabetic autonomic neuropathy. Diabetes Res. Clin. Pratt. 6, 157-160. Sweeney, D. and Beeley, J.A. (1990) An enzyme linked immunoassay for human salivary albumin. Arch. Oral Biol., in press. Viberti, G. and Keen, H. (1984) The pattern of proteinuria in diabetes mellitus. Diabetes 33, 686-692. Bent-Hansen, L., Feldt-Rasmussen, B., Kverneland, A. and Deckert, T. (1987) Transcapillary escape rate and relative metabolic clearance of glycated and non-glycated albumin in Type 1 (insulin-dependent) diabetes mellitus. Diabetologia 30, 2-4. Bell, D., Collier, A., Nicoll, J.J., Jackson, M., Millar, A.M., Clarke, B.F. and Muir, A.L. (1988) Reduced venous compliance and increased transcapillary escape of protein in insulin-dependent diabetic patients. Diabetic Med. 5, 454-458.