Microalbuminuria in diabetes mellitus: more on urine storage and accuracy of colorimetric assays.
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Biomedical subjects
Publications and source records attributed to O Giampietro.
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We have developed a radioimmunoassay method (RIA) to measure urinary albumin excretion. We determined the albumin excretion rate (AER) (micrograms/min) of 122 healthy subjects and 145 diabetic patients (115 type I, 30 type II). The results indicate that the RIA is sensitive (0.39 +/- 0.08 mg/L), precise (CV 5-8%), and gives reliable results on previously frozen urine samples. The distribution of the AER values in healthy subjects and diabetic patients was not normal. It was normalized by log or square-root transformation of the data. Seventy-three percent of diabetic patients lay within the normal range (0.6-10.6 micrograms/min). Twenty percent could be considered "at risk" to develop overt diabetic nephropathy because their albuminuria exceeded a threshold level of 15 micrograms/min chosen previously as the cutoff value for microalbuminuria. We found no correlation between AER and glycated hemoglobin, and only a weak correlation between AER and diabetes duration in type I diabetic patients.
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Excretion of digoxin-like immunoreactivity (DLIS) was measured by RIA in timed overnight urine collections from 91 normotensive nondiabetic subjects and 104 normotensive insulin-dependent diabetic (IDDM) patients. The mean +/- SD DLIS excretion rate for the diabetic patients significantly exceeded that for the controls (73 +/- 41 vs 63 +/- 36 pg/min, P = 0.024). In both groups, the mean DLIS excretion rates for men were significantly higher (P = 0.0014, P = 0.006) than for women. In the controls, the DLIS excretion rate significantly correlated with the urinary excretion rate of creatinine (P less than 0.01), Na+ (P less than 0.05), and K+(P less than 0.05), and with the subjects' body weight (P less than 0.01), body mass index (P less than 0.05), and systolic blood pressure (P less than 0.05). In the diabetics, the DLIS excretion rate was significantly correlated with body weight (P less than 0.05) and with urinary excretion rates for albumin (P less than 0.01), creatinine (P less than 0.01), Na+ (P less than 0.05), and K+(P less than 0.05). Our data indicate that: (a) increased amounts of a cardiac glycoside-like substance (or a group of substances) are excreted in the urine of IDDM patients; (b) the urinary excretion of DLIS seems to depend on glomerular filtration rate and physiocochemical properties of glomerular membrane, as well as on subjects' body mass; and (c) because cardiac glycoside-like substances may increase peripheral vascular resistance, increased urinary excretion of DLIS by IDDM patients may indicate a tendency to develop hypertension.
While proteinuria detectable by dip-sticks is the hallmark of overt diabetic nephropathy, urinary albumin excretion (UAE) above normal but Albustix-negative (so-called microalbuminuria) is the main characteristic of the preproteinuric phase of the diabetic renal disease. Reliable measurement of UAE requires very sensitive and accurate methods, such as radioimmunoassay (RIA), which, however, are not suitable for routine UAE analysis. The Coomassie Brilliant Blue dye binding method has been suggested to be a simple and economical way of indirectly measuring albuminuria in diabetic patients. In the present paper, we compared the chemico-clinical characteristics of a RIA method with those of the Coomassie dye binding method, in order to verify if it is really possible to calculate albuminuria from proteinuria (and vice versa) by a simple linear regression equation, as previously suggested. The RIA has shown a better sensitivity and accuracy in comparison to the dye binding method. Our study suggests that there is not a linear relationship between proteinuria and albuminuria in diabetic patients. Indeed, the ratio between the total proteinuria, as measured with the dye method, and the albuminuria, as measured by a specific and sensitive RIA, varies greatly in diabetic patients with or without glomerular nephropathy. While the dye binding method appears the best procedure for the assay of total microproteinuria, since it is precise, cheap and feasible, the RIA, due to its high sensitivity and specificity, is more suitable for early and accurate detection of microalbuminuria, as well for the close follow-up of subjects at risk of developing overt diabetic nephropathy.
To verify whether there is a variation in the 24-h urinary excretion of digoxin-like immunoreactivity (DLIS) in humans, we studied 18 normal adults, who collected their urines for 24-h in several portions. We then measured DLIS (by means of a sensitive RIA method), creatinine, sodium, and potassium concentrations in the urine samples. The mean urinary excretion rate for DLIS in the complete 24-h collection was 84.8 (SD 31.3) pg/min. The mean DLIS urinary excretion rate calculated for overnight collections was significantly lower than those of afternoon collections (P less than 0.01) and the 24-h collection (P less than 0.05). Significant positive correlations were found between urinary DLIS and excretion rates for creatinine (r = 0.347, P = 0.0016), Na+ (r = 0.232, P = 0.038), and K+ (r = 0.323, P = 0.003), respectively. Our data suggest that urinary excretion of DLIS is higher during "active" hours of the day, especially in the afternoon, than at rest, during the night.
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We randomly administered luteinizing hormone-releasing hormone (LHRH) or thyrotropin releasing hormone (TRH) (25 micrograms and 200 micrograms, respectively, as a bolus), to 16 diabetic male subjects (9 type I, 7 type II) and to 9 healthy male controls in two different mornings. While GH in the basal state was similar in type I, type II, and normal subjects, LHRH administration surprisingly evoked a significant GH release in 7 (5 type 1, 2 type II) diabetic patients. GH-responders had higher glycated hemoglobin than non-responders (11 +/- 1 nu 8.3 +/- 0.5%) but superimposable fasting and intratest average glucose levels. Only one patient among the GH-responders to LHRH showed a GH release also after TRH. These data support the hypothesis that GH secretion in diabetes, especially when poorly controlled, is abnormal.
We administered growth-hormone releasing hormone (GHRH), clonidine or thyrotropin-releasing hormone (TRH) as intravenous boli each in three different randomized mornings to nine well-controlled Type 1 diabetic men and to six age-matched healthy men who served as controls. GHRH and clonidine evoked a prompt and brisk GH release both in diabetic and in control subjects with no significant difference being evident between the two groups. Only one diabetic subject showed a paradoxical GH release after TRH when he was under long-term poor metabolic control. These results indicate that in insulin-dependent patients with good control of the metabolic disease the response of somatotropes to pituitary- or central nervous system-directed stimuli is normal. These data are supportive of the idea that altered GH secretion in Type 1 diabetes rather than reflecting a primary hypothalamic and/or pituitary alteration may be a state-dependent phenomenon related to the metabolic state of the disease.
In this paper we studied the effects of a low-protein, low-phosphorus supplemented diet in 8 type I diabetics with 'overt' diabetic nephropathy and mild or severe renal insufficiency. We examined the following parameters: the rate of decline of creatinine clearance, the urinary protein loss, the total serum protein, the daily insulin requirement, the serum fasting glucose, the pattern of serum lipids (serum total cholesterol, HDL cholesterol and serum triglycerides), the mean blood pressure and body weight. The rate of decline of creatinine clearance decreased monthly from 1.48 +/- 0.20 ml/min during a previous 15.6-month period of unrestricted protein diet (UPD), to 0.13 +/- 0.3 ml/min during the 11.4 months on the supplemented diet (SD). The mean blood pressure did not differ during UPD (130.9 +/- 7.0 mmHg) and during SD (128.1 +/- 1.6 mmHg). Urinary protein loss significantly decreased on SD, and total serum protein increased. The daily insulin requirement and the serum fasting glucose levels significantly decreased on SD. Serum cholesterol was lower during SD than during UPD, while serum HDL cholesterol and serum triglycerides were not significantly modified. In some patients the body weight decreased on SD as a consequence of the disappearance of edema. In conclusion, on the basis of these preliminary observations, the SD slows the progression of renal failure and seems to exert several beneficial and no unwanted side-effects in renal failure of type I diabetics.
Plasma lipid levels have been proposed as probable risk factors of diabetic retinopathy. To clarify this question, we evaluated the apolipoprotein levels in 68 type I diabetic patients (39 +/- SD 14 years; duration of diabetes 13 +/- SD 8.4 years). By the analysis of fluorescein angiography we have classified diabetic retinopathy as follows: absent retinopathy (AR, n = 23), minimal retinopathy (MR, n = 16), exudative retinopathy (ER, n = 15), proliferative retinopathy (PR, n = 14). For all patients we measured: total, LDL- and HDL-cholesterol (T-CH, LDL-CH, HDL-CH), apolipoproteins A and B (Apo A and B), fasting plasma glucose (FPG), glycosylated hemoglobin (HbA1). The mean age of the patients with ER (47 +/- SD 14 years) was greater than those with MR (36 +/- SD 15 years) or AR (33 +/- SD 13 years) (p less than 0.05). Significant differences within groups were not found for relative body weight, daily insulin dose, FPG, HbA1% (analysis of variance, ANOVA). The groups differed between each other with regard to T-CH, LDL-CH and HDL-CH/LDL-CH ratio (T-cholesterol: AR 186 +/- SD 34, MR 191 +/- SD 32, ER 212 +/- SD 52, PR 215 +/- SD 41 mg/dl; LDL-cholesterol: AR 124 +/- SD 28, MR 122 +/- SD 18, ER 148 +/- SD 38, PR 145 +/- SD 33 mg/dl; HDL/LDL-cholesterol ratio: AR 0.38 +/- SD 0.1, MR 0.35 +/- SD 0.1, ER 0.30 +/- SD 0.1, PR 0.29 +/- SD 0.1). The total-CH levels increased and HDL-CH/LDL-CH ratio decreased along with the severity of the retinopathy.(ABSTRACT TRUNCATED AT 250 WORDS)
The diagnosis of sexual dysfunctions (SD) in diabetic male patients is usually based on interviews about the sexual behavior of the patients themselves. The absence of a standardized procedure may explain the discrepancies in the prevalence reported in previous papers. In this work, we show a rational procedure to assess the presence and the type of SD in a group of 128 diabetic men. By means of an appropriate questionnaire, SD were pointed out in 40% of insulin-dependent diabetics (IDD) and 52% of non-insulin-dependent diabetic patients (NIDD). Among the patients with SD, 28% had organic impotence, 11% psychogenic impotence and 8% reduction of libido. IDD with SD were older (p less than 0.01) and with a longer duration of diabetes (p less than 0.02) than IDD without SD. NIDD with and without SD had similar ages and known duration of diabetes. Both in IDD and in NIDD, the prevalence of SD was not affected by the quality of short-term metabolic control. In the patients in whom it was studied by a different technique, the predominant cause of organic impotence (76%) was a damage of parasympathetic nervous system, as evaluated by non-invasive tests. In 12% of the cases with organic impotence, a reduced penile flow alone was observed, while hormonal parameters were similar in patients with and without organic impotence.
We developed a simple isocratic high performance liquid chromatography (HPLC) method for the quantitative determination of 5-hydroxymethyl-2-furfuraldehyde (5-HMF) liberated by mild hydrolysis of small amounts of glycosyl proteins. The absorbance of hydrolysate components after HPLC separation was recorded at 280 nm. To detect substances possibly interfering with the 5-HMF peak we always recorded the ratio of the peak heights A280 nm/A254 nm which was a constant value of 4.4. For each sample the blank was obtained by reduction with NaBH4 before hydrolysis with oxalic acid 1 mol/l. The best NaBH4/protein ratio was found to be 4 mg/mg. With this method we measured the nonenzymatic glycosylation (glycation) as 5-HMF in samples with a protein concentration as low as 0.8 mg/ml. 5-HMF produced per milligram of protein was independent from protein concentration for a wide range (0.8-10 mg/ml). The mean coefficient of variation for within assay and between precision was 6.8 and 11.6%, respectively. The 5-HMF measured on plasma proteins from normal subjects (n = 7) was 0.16 +/- 0.04 nmol/mg. Protein from insulin-dependent diabetic patients was 0.31 +/- 0.07 nmol/mg. With this method we succeeded in detecting an excessive glycation of platelet membrane proteins in 13 type-I diabetic patients.
To assess the long-term clinical usefulness of measuring glycated hemoglobin (Hb A1), we carried out a two-year longitudinal study involving 234 diabetic patients (116 males, 118 females; 139 with type I diabetes mellitus, 95 with type II). Hb A1 values correlated significantly (p less than 0.001) with a score index based on plasma glucose in a specimen collected after overnight fasting, and urinary glucose, and ketones in a 24-h specimen. However, we found that one of every three well-controlled patients (both type I and II subjects) had high values for Hb A1. Among poorly controlled patients, only those with "brittle" diabetes had good values for Hb A1.
An increase in fetal and maternal complications has been documented in cases of gestational diabetes, but the glucose levels that predict an increased risk have not been clearly defined. We evaluated the frequency of several neonatal complications (macrosomia, congenital anomalies, perinatal mortality, and prematurity) and maternal complications (toxemia, cesarean section, or both) in relation to glucose tolerance in 249 women in the third trimester of pregnancy. None of the women had previous evidence of diabetes, and all had normal results on an oral glucose-tolerance test, according to accepted criteria. On the basis of their two-hour plasma glucose levels, women were divided into three groups: A (glucose less than 100 mg per deciliter), B (glucose 100 to 119 mg per deciliter), and C (glucose 120 to 164 mg per deciliter). The higher two-hour plasma glucose levels were associated with a significant increase in the incidence of macrosomia (9.9, 15.5, and 27.5 percent in Groups A, B, and C, respectively), congenital abnormalities (0.7, 3.5, and 5.0 percent), and toxemia, cesarean section, or both (19.9, 25.9, and 40.0 percent). A significant correlation between the infant's weight and the mother's two-hour plasma glucose level was also observed. These data indicate that even limited degrees of maternal hyperglycemia, which are currently considered to be within the normal range, may affect the outcome of pregnancy.
An extremely rare case of factitious hypoglycemia in a young insulin-dependent pregnant woman in the first trimester of pregnancy is reported. Pregnancy resulted in the birth of a normal child. The clinical and laboratory procedures used to ascertain this diagnosis together with the psychological dynamics of the case are discussed.