The relative roles of obesity, insulin resistance, insulin secretory dysfunction, and excess hepatic glucose production in the development of non-insulin-dependent diabetes mellitus (NIDDM) are controversial. We conducted a prospective study to determine which of these factors predicted the development of the disease in a group of Pima Indians.
Non-insulin-dependent diabetes mellitus (NIDDM) is a genetic disorder characterized by two major pathogenic processes: reduced insulin action and a relative or absolute decrease in plasma insulin concentrations. We studied 116 nondiabetic siblings from 45 families to determine if in vivo insulin action showed any aggregation among siblings. Subjects were Pima Indians from the Gila River Indian Community in Arizona who, as a group, have the highest reported incidence and prevalence of NIDDM in the world. In vivo insulin action was determined by the euglycemic-clamp technique at two rates of insulin infusion in each subject with resulting mean plasma insulin concentrations of 119 and 1938 microU/ml. After adjustment for age, sex, and degree of obesity, there was significant aggregation among siblings of in vivo insulin action at the high insulin infusion rate (P less than or equal to .0001). Family membership independently accounted for approximately 34% of the variance in this measure of insulin action. Glucose uptake at the lower insulin infusion rate also showed familial aggregation (P less than .01), with family membership independently accounting for approximately 15% of the variance of this measurement. We conclude that in vivo insulin action is a familial characteristic. The familial component of insulin action occurs in addition to the effects of obesity, age, and sex on insulin action. Therefore it is not sufficient to simply know that an individual is lean or obese to predict his/her in vivo insulin resistance, because it must also be known whether he/she is from an insulin-resistant or insulin-sensitive family.(ABSTRACT TRUNCATED AT 250 WORDS)
Increased in vivo resistance to insulin-mediated glucose disposal has been observed in obese subjects with normal glucose tolerance and in nonobese subjects with glucose intolerance. To determine whether the insulin resistance of glucoseintolerant obese subjects can be accounted for by obesity alone, insulin-mediated glucose disposal was measured in 14 glucoseintolerant and 21 nondiabetic Southwestern American Indians with similar degrees of obesity. A mixture of insulin, glucose, and somatostatin was infused which delivered the same quantity of glucose and achieved similar plasma insulin concentrations in all subjects. Despite similar steady state plasma insulin levels, the mean steady state plasma glucose concentration was higher in the glucose-intolerant subjects than in weight-matched subjects with normal glucose tolerance (226 ± 10 vs. 136 ± 13 mg/dl; P < 0.0001). This increased resistance to insulin action was found in the presence of similar insulin binding to mononuclear cells (measured in 8 glucose-intolerant subjects and 9 subjects with normal glucose tolerance). In obese Southwestern American Indians with glucose intolerance, abnormalities beyond the site of insulin binding to its receptor may explain the observed increase in in vivo insulin resistance.
Twenty-three structural analogues of AMP were tested for their ability to activate and bind phosphorylase b in the presence and absence of substrate or protamine sulfate (or both). Michaelis constants, maximal velocities, and inhibition constants were measured by nucleotide activation of phosphorylase b and compared with the corresponding dissociation and inhibition constants measured by nucleotide quenching of a pyridoxal 5'-phosphate-associated enzyme fluorescence. The excitation maximum was 347 mµ and the emission maximum was 522 mµ. A sigmoidal saturation curve was obtained for AMP quenching of enzyme fluorescence. This, coupled with the ATP inhibition and the glucose 1-phosphate enhancement of AMP quenching of enzyme fluorescence, identified the site of AMP binding for enzyme fluorescence quenching with the AMP binding site for activation of phosphorylase b. Protamine enhanced the binding of some nucleotides to phosphorylase b but apparently affected only the Vmax of others as measured by nucleotide activation of the enzyme. An increase in enzyme fluorescence associated with addition of ATP and a quenching effect by added AMP or glucose-1-P suggested that the observed quenching was the result of the protein conformational state changes associated with the allosteric properties of the enzyme. Nucleotide analogues of AMP exhibited different heterotropic binding properties with phosphorylase b. Binding studies in the presence and absence of substrate revealed that for some nucleotides the affinity was unchanged while others showed less affinity for enzyme in the presence of substrate. Enzyme activation by bound nucleotide was sensitive to the integrity of both the adenine ring and the 5'-phosphate, but the latter appeared to be the most critical structural feature. Both the 5'-phosphate and purine ring were involved in nucleotide binding.
It has been variously suggested that chemical carcinogens interact with a protein regulator of cell growth, ultimately causing neoplastic cells to arise in which the protein target of the carcinogen is reduced or absent. As a test of the target reduction aspect of these hypotheses, the amount of the principal protein target of a hepatic carcinogen was determined immunologically in liver tumors caused by that carcinogen.
Specific antiserum against the principal liver protein target of a hepatic azocarcinogen was obtained by immunization with the highly purified liver azoprotein isolated from rats fed 3′-methyl-4-dimethylaminoazobenzene for 15 to 18 days. The amount of the target protein in serially diluted liver and liver tumor cytosols was measured by gel double immunodiffusion with a precision of 10% (S.D.) or less.
Little difference in the relative content of the principal target protein was found in the liver cytosols of normal rats and of those fed a diet containing or lacking the azocarcinogen for 15 to 18 days. In contrast, in 15 cytosols of primary liver tumors of 15 rats fed the azocarcinogen until sacrifice, the content of protein target of azocarcinogen was 6 to 31% (average, 18 ± 8% S.D.) of that present in normal rat liver cytosol. In 7 cytosols of livers around tumors, the quantity of target protein was 66 to 116% (average, 83 ± 19% S.D.).
An additional series of assays examined 4 types of transplanted hepatomas that were originally caused by fluorenyl amide and biphenyl amide carcinogens. These carcinogens probably have principal target proteins that are different from those of azocarcinogens. Three kinds of well-differentiated hepatomas had mean levels (85 to 98%) of the protein target of azocarcinogens like that of normal liver. One type of poorly differentiated hepatoma had < 3%.
The finding of a reduction in amount of principal protein target of a carcinogen in primary tumors caused by that carcinogen, and not in transplanted differentiated tumors caused by other carcinogens, suggests two alternatives, namely, either the reduction need be specifically that of the target protein of the carcinogen which caused the neoplasm or it is due to other causes, e.g. , tumor progression, and therefore is not essential for neoplasia per se . These alternatives are separate from the more basic question of the role (if any) of the carcinogen-protein interaction in chemical carcinogenesis, possibly as part of a mechanism not involving target protein depletion in tumors.
If a single gene produced insulin resistance, with environmental effects creating some additional variance, insulin action might be distributed as a mixture of two normal distributions if the gene is dominant or recessive or as a mixture of three normal distributions if the gene is codominant. To estimate maximal insulin-stimulated glucose uptake rates (MaxMs), hyperinsulinemic-euglycemic clamps were performed on 245 nondiabetic Pima Indians (126 men, 119 women). Five models (for 1, 2, 3, 4, or 5 components each, normally distributed with a common variance) were fitted to the frequency distribution of MaxM by iterative maximum-likelihood estimation. The three-component model fit the data significantly better than a single normal distribution (chi 2 = 14.3 with 4 df P less than .01) or a mixture of two normal distributions (chi 2 = 9.9 with 2 df, P less than .01). Mixtures of four or five normal distributions did not fit the data significantly better than a mixture of three normal distributions. The first component of the distribution comprised 23%, the second 48%, and the third 29% of the total distribution. Similarly, the frequency distributions of fasting plasma insulin concentrations and a principal component score derived from MaxM and fasting insulin were best fitted by a mixture of three normal distributions. These results are consistent with the hypothesis that among Pimas, insulin resistance is determined by a single gene with a codominant mode of inheritance. Segregation analyses of studies performed in pedigrees are indicated to prove or disprove this genetic hypothesis.
Insulin receptor tyrosine kinase is an important step in insulin action. We examined the relationship between diet-induced changes in glucose metabolism and changes in skeletal muscle insulin-sensitive tyrosine kinase activity in 12 nondiabetic subjects. Subjects were fed a traditional, high carbohydrate Pima Indian diet and a modern, high fat western diet for 2 weeks in a randomized cross-over design. At the end of each dietary period, glucose tolerance was assessed, insulin sensitivity (S1) was estimated by Bergman's minimal model method, and insulin receptor concentration and tyrosine kinase activity were determined on lectin-purified extracts from quadriceps femoris muscle. Compared to the traditional diet, the modern diet was associated with a deterioration of glucose tolerance and an increase in glucose-induced plasma insulin levels. As expected, S1 changes were associated with opposite changes in plasma insulin levels. However, the changes in maximal tyrosine kinase activity were negatively correlated with changes in S1 (r = —0.69; P < 0.01) and positively correlated with changes in plasma glucose (r = 0.70; P < 0.01) and insulin response to glucose (r = 0.57; P < 0.025). These results suggest that the site of diet-induced changes in insulin action is beyond the insulinsensitive tyrosine kinase. The results further suggest that the kinase activity is modulated by prevailing plasma insulin levels.
