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SITE OF INSULIN RESISTANCE IN TYPE 2 DIABETES

The maintenance of whole-body glucose homeostasis is dependent upon a normal insulin secretory response by the pancreatic beta cells and normal tissue sensitivity to the independent effects of hyperinsulinemia and hyperglycemia (i.e., the mass-action effect of glucose) to augment glucose uptake. In turn, the combined effects of insulin and hyperglycemia to promote glucose disposal are dependent on three tightly coupled mechanisms (Table 1): (i) suppression of endogenous (primarily hepatic) glucose production; (ii) stimulation of glucose uptake by the splanchnic (hepatic plus gastrointestinal) tissues; and (iii) stimulation of glucose uptake by peripheral tissues, primarily muscle (1,4,14). Muscle glucose uptake is regulated by flux through two major metabolic pathways: glycolysis (of which ~90% represents glucose oxidation) and glycogen synthesis.

Hepatic Glucose Production

In the overnight fasted state the liver of healthy subjects produces glucose at the rate of ~1.8-2.0 mgokg-1omin-1 (1,3,4,6,18,54). This glucose flux is essential to meet the needs of the brain and other neural tissues, which utilize glucose at a constant rate of ~1-1.2 mgokg-1omin-1 (2,169). Brain glucose uptake accounts for ~50-60% of glucose disposal during the postabsorptive state and this uptake is insulin independent. Therefore, brain glucose uptake occurs at the same rate during absorptive and postabsorptive periods and is not altered in type 2 diabetes (214). Following glucose ingestion, insulin is secreted into the portal vein and carried to the liver, where it suppresses hepatic glucose output. If the liver does not perceive this insulin signal and continues to produce glucose, there will be two inputs of glucose into the body, one from the liver and another from the gastrointestinal tract, and marked hyperglycemia will ensue.

In type 2 diabetic subjects with mild to moderate fasting hyperglycemia (140-200 mg/dl, 7.8-11.1 mmol/L), basal hepatic glucose production is increased by ~0.5 mg/kgomin . Consequently, during the overnight sleeping hours (2200 h to 0800 h), the liver of a 80-kg diabetic individual with modest fasting hyperglycemia adds an additional 35 g of glucose to the systemic circulation. The increase in basal HGP is closely correlated with the severity of fasting hyperglycemia (1,3,4,6,18,54,157-159,162) . Thus, in type 2 diabetics with overt fasting hyperglycemia (>140 mg/dl, 7.8 mmol/l), an excessive rate of hepatic glucose output is the major abnormality responsible for the elevated fasting plasma glucose concentration. The close relationship between fasting plasma glucose concentration and HGP has been demonstrated in numerous studies (164-166,170-174).

In the postabsorptive state, the fasting plasma insulin concentration in type 2 diabetics is 2-4 fold greater than in nondiabetic subjects. Because hyperinsulinemia is a potent inhibitor of HGP (1,3,4-6,16,18,164,165,175), hepatic resistance to the action of insulin must be present in the postabsorptive state to explain the excessive output of glucose by the liver. Hyperglycemia per se also exerts a powerful suppressive action on HGP (15,167,175-177). Therefore, the liver also must be glucose resistance with respect to the inhibitory effect of hyperglycemia to suppress hepatic glucose output, and this has been well documented (15,167,178,179).

Using the euglycemic insulin clamp technique in combination with tritiated glucose, the dose response relationship between hepatic glucose production and the plasma glucose concentration has been defined by Groop, DeFronzo, et al (18) . The following points should be emphasized: (i) first, the dose-response curve relating inhibition of HGP to the plasma insulin concentration is quite steep, with an effective dose for half-maximal insulin concentration (ED50) of ~30-40 µU/ml; (ii) second, in type 2 diabetic individuals the dose response curve is shifted to the right, indicating the presence of hepatic resistance to the inhibitory effect of insulin on hepatic glucose production. However, at plasma insulin concentrations within the high physiologic range (~100 uU/ml), the hepatic insulin resistance can be largely overcome and a near normal suppression of HGP can be achieved; (iii) third, the severity of the hepatic insulin resistance is related to the severity of the diabetic state. In type 2 diabetic individuals with mild fasting hyperglycemia, an increment in plasma insulin concentration of 100 µU/ml causes a complete suppression of HPG. However, in diabetic subjects with more severe fasting hyperglycemia, the ability of the same plasma insulin concentration to suppress HGP is impaired (18). These results suggest that there is an acquired component of hepatic insulin resistance and that this defect becomes progressively worse as the diabetic state decompensates over time.

The glucose released by the liver can be derived from either glycogenolysis or gluconeogenesis (6,16,176). Studies employing the hepatic vein catheter technique have shown that the uptake of gluconeogenic precursors, especially lactate, is increased in type 2 diabetic subjects (180). Consistent with this observation, radioisotope turnover studies, using lactate, alanine, and glycerol, have shown that ~90% of the increase in HGP above baseline can be accounted for by accelerated gluconeogenesis (181,182). More recent studies employing 13C-magnetic resonance imaging (183) and D2O (184,185) have confirmed the important contribution of accelerated gluconeogenesis to the increase in HGP. An increased rate of glutamine conversion to glucose also has been shown to contribute to the elevated rate of gluconeogenesis in type 2 diabetic subjects (186). The mechanisms responsible for the increase in hepatic gluconeogenesis include hyperglucagonemia (187), increased circulating levels of gluconeogenic precursors (lactate, alanine, glycerol) (181,188), increased FFA oxidation (18,162,189), enhanced sensitivity to glucagon (190) and decreased sensitivity to insulin (1,4.18,164,165). Although the majority of evidence indicates that increased gluconeogenesis is the major cause of the increase in HGP in type 2 diabetic subjects (181-186), it is likely that accelerated glycogenolysis also contributes (181,191).

Because of the inaccessibility of the liver in man, it has been difficult to assess the role of key enzymes involved in the regulation of gluconeogenesis (pyruvate carboxylase, phosphoenol- pyruvate carboxykinase), glycogenolysis (glycogen phosphorylase), and net hepatic glucose output (glucokinase, glucose-6-phosphatase). However, considerable evidence from animal models of type 2 diabetes and some evidence in humans have implicated increased activity of PEPCK and G-6-Pase in the accelerated rate of hepatic glucose production (192-194).

The kidney also has been shown to produce glucose and estimates of the renal contribution to total endogenous glucose production have varied from 5% to 20% (8,9,195). These varying estimates of the contribution of renal gluconeogenesis to total glucose production are largely related to the methodology employed to measure glucose production by the kidney (196). One unconfirmed study suggests that the rate of renal gluconeogenesis is increased in type 2 diabetics with fasting hyperglycemia (197). Arguing against this possibility are studies employing the hepatic vein catheter technique which have shown that all of the increase in total body endogenous glucose production (measured with 3-3H-glucose) in type 2 diabetics can be accounted for by increased hepatic glucose output (measured by the hepatic vein catheter technique) (3).

Peripheral (Muscle) Glucose Uptake

Muscle is the major site of glucose disposal in man (1,3-5,14). Under euglycemic hyperinsulinemic conditions, approximately 80% of total body glucose uptake occurs in skeletal muscle (1,3-5). Studies employing the euglycemic insulin clamp in combination with femoral artery/vein catheterization have examined the effect of insulin on leg glucose uptake in type 2 diabetic and control subjects (3) . Since bone is metabolically inert and adipose tissue takes up less than 5% of an infused glucose load (17,198,199), muscle represents the major tissue responsible for leg glucose uptake.

In response to a physiologic increase in plasma insulin concentration (~80-100 uU/ml), leg (muscle) glucose uptake increases linearly, reaching a plateau value of 10 mg/kg leg wt per minute (3) . In contrast, in lean type 2 diabetic subjects, the onset of insulin action is delayed for ~40 min and the ability of the hormone to stimulate leg glucose uptake is markedly blunted, even though the study is carried out for an additional 60 min in the type 2 diabetic group to allow insulin to more fully express its biological effects (3). During the last hour of the insulin clamp study, the rate of glucose uptake was reduced by 50% in the diabetic group (3). These results provide conclusive evidence that the primary site of insulin resistance during euglycemic insulin clamp studies performed in type 2 diabetic subjects resides in muscle tissue. Using the forearm and leg catheterization techniques (13,153,200,202), a number of investigators have demonstrated a decreased rate of insulin-mediated glucose uptake by peripheral tissues. The use of positron emission tomography (PET) scanning to quantitate leg glucose uptake in type 2 diabetic subjects has provided additional support for the presence of severe muscle resistance to insulin in diabetic subjects (203).

Splanchnic (Hepatic) Glucose Uptake

In humans, it is difficult to catheterize the portal vein, and glucose disposal by the liver has not been examined directly. Using the hepatic vein catheterization technique in combination with the euglycemic insulin clamp, the contribution of the splanchnic (liver plus gastrointestinal) tissues to overall glucose homeostasis has been examined in lean type 2 diabetic subjects with mild to moderate fasting hyperglycemia (3). In the postabsorptive state, there is a net release of glucose from the splanchnic area (i.e., negative balance) in both control and type 2 diabetic subjects , reflecting glucose production by the liver. In response to insulin, splanchnic glucose output is promptly suppressed (reflecting the inhibition of HGP) and, by 20 min, the net glucose balance across the splanchnic region declines to zero (i.e., there was no net uptake or release) (3). After 2 h of sustained hyperinsulinemia, there is a small net uptake of glucose (~0.5 mgokg-1omin-1) by the splanchnic area (i.e., positive balance). This uptake is virtually identical to the rate of splanchnic glucose uptake observed in the basal state, indicating that the splanchnic tissues, like the brain, are insensitive to insulin at least with respect to the stimulation of glucose uptake (3,5,6,175). There was no difference between diabetic and control subjects in the amount of glucose taken up by the splanchnic tissues at any time during the insulin clamp study (3).

The results of these studies illustrate another important point: namely, that under conditions of euglycemic hyperinsulinemia, very little of the infused glucose is taken up by the splanchnic (and therefore hepatic) tissues (3,5,6,175). During the insulin clamp, the rate of whole body glucose uptake averaged 7 mgokg-1omin-1, and of this, only 0.5 mgokg-1omin-1 or 7%, was disposed of by the splanchnic region. Because the difference in insulin-mediated total body glucose uptake between the type 2 diabetic and control groups during the euglycemic insulin clamp study was 2.5 mgokg-1omin-1, from a purely quantitative standpoint it is obvious that a defect in splanchnic (hepatic) glucose removal never could account for the magnitude of impairment in total body glucose uptake following intravenous glucose/insulin administration. However, after glucose ingestion, the oral route of administration and the resultant hyperglycemia conspire to enhance splanchnic (hepatic) glucose uptake (6,7,11,12,16,26,175) and, under these conditions, diminished hepatic glucose uptake has been shown to contribute to the impairment in glucose tolerance in type 2 diabetes (see discussion below) (6,204,205).

Summary: Whole Body Glucose Utilization

Insulin-mediated whole body glucose utilization during the euglycemic insulin clamp is summarized in Fig. 12 (coming soon). The total height of each bar represents the amount of glucose taken up by all tissues in the body during the insulin clamp in control and type 2 diabetic subjects. Net splanchnic glucose uptake, quantitated by the hepatic venous catheterization technique, is similar in both groups and averaged 0.5 mgokg-1omin-1. Adipose tissue glucose uptake accounts for less than 5% of total glucose disposal (17,198,199). Brain glucose uptake, estimated to be 1.0-1.2 mgokg-1omin-1 in the postabsorptive state (2,169,206), is unaffected by hyperinsulinemia (169). Muscle glucose uptake (extrapolated from leg catheterization data) in control subjects accounts for ~75-80% of the total glucose uptake (1,3,4). In type 2 diabetic subjects, the largest part of the impairment in insulin-mediated glucose uptake is accounted for by a defect in muscle glucose disposal. Even if adipose tissue of type 2 diabetic subjects took up absolutely no glucose, it could, at best, explain only a small fraction of the defect in whole body glucose metabolism.

Glucose Disposal During OGTT

In every day life, the gastrointestinal tract represents the normal route of glucose entry into the body. However, the assessment of tissue glucose disposal following glucose ingestion presents a challenge because of the difficulties in quantitating the rate of glucose absorption, suppression of hepatic glucose production, and organ (liver and muscle) glucose uptake. Moreover, because the plasma glucose and insulin concentrations are changing simultaneously, it is difficult to draw conclusions about insulin secretion or insulin sensitivity.

To address these issues, Ferrannini, DeFronzo, and colleagues (7,11,12,205) administered oral glucose to healthy control subjects in combination with hepatic vein catheterization to examine splanchnic glucose metabolism. The oral glucose load and endogenous glucose pool were labeled with [1-14C]glucose and [3-3H]glucose, respectively, to quantitate total body glucose disposal (from tritiated glucose turnover) and endogenous HGP (difference between the total rate of glucose appearance, as measured with tritiated glucose, and the rate of oral glucose appearance, as measured with [1-14C]glucose).

During the 3.5 h after glucose (68 g) ingestion: (i) 19 g, or 28%, or the oral load was taken up by splanchnic tissues; (ii) 48 g, or 72%, was disposed of by peripheral (non-splanchnic) tissues; (iii) of the 48 g taken up by peripheral tissues, the brain (an insulin-independent tissue) accounted for ~15 g (~1 mgokg-1omin-1), or 22%, of the total glucose load (12); (iv) basal HGP declined by 53%. Similar percentages for splanchnic glucose uptake (24%-29%) and suppression of HGP (50%-60%) in normal subjects have been reported by other investigators (13,204,207-209). The contribution of skeletal muscle to the disposal of an oral glucose load has been reported to vary from a low of 26% (207) to a high of 56% (208), with a mean of 45% (11,13,207-209). These results emphasize several important differences between oral and intravenous glucose administration. After glucose ingestion: (i) HGP is less completely suppressed, most likely do to activation of local sympathetic nerves that innervate the liver (210); (ii) peripheral tissue (primarily muscle) glucose uptake is quantitatively less important; (3) splanchnic glucose uptake is quantitatively much more important.

In type 2 diabetic individuals (12,204,205,211,212) the disposition of an oral glucose load is significantly altered. The disturbance in glucose metabolism is accounted for by two factors: (i) decreased tissue glucose uptake and (ii) impaired suppression of HGP. Splanchnic glucose uptake is similar in diabetic and control groups. Inappropriate suppression of HGP accounted for approximately one-third of the defect in total-body glucose homeostasis, while reduced peripheral (muscle) glucose uptake accounted for the remaining two-thirds. Since hyperglycemia per se enhances splanchnic (hepatic) glucose uptake in proportion to the increase in plasma glucose concentration (24,175), the splanchnic glucose clearance (SGU ÷ plasma glucose concentration) is markedly reduced in all type 2 diabetic subjects following glucose ingestion. Using a combined insulin clamp/OGTT technique, an impairment in glucose uptake by the splanchnic tissues in type 2 diabetics has been demonstrated directly (213).

When viewed in absolute terms, most studies have demonstrated that the total amount of glucose taken up by all tissues of body over the 4 hour period following the ingestion of an oral glucose load is normal (13) or slightly decreased (204,205,211). However, this occurs at the expense of postprandial hyperglycemia. Thus, the efficiency of glucose disposal, i.e., the glucose clearance (tissue glucose uptake ÷ plasma glucose concentration), is severely reduced. It should be emphasized that it is not the absolute glucose disposal rate, but rather the increment in glucose disposal above baseline that determines the rise in plasma glucose concentration above the fasting value. Every published study (13,204,205,211) has demonstrated that the incremental response in whole-body glucose uptake is moderately to severely reduced in type 2 diabetic individuals. Similar results have been reported for forearm muscle glucose uptake (13,201,202,208,209) , pointing out the important contribution of diminished muscle glucose disposal to impaired oral glucose tolerance in type 2 diabetes.

In summary, results of the OGTT indicate that both impaired suppression of HGP and decreased tissue (muscle) glucose uptake contribute approximately equally to the glucose intolerance of type 2 diabetes. The efficiency of the splanchnic (hepatic) tissues to take up glucose (as reflected by the splanchnic glucose clearance) also is impaired in type 2 diabetic individuals.

Summary of Insulin Resistance in Type 2 Diabetes

Insulin resistance involving both muscle and liver are characteristic features of the glucose intolerance in type 2 diabetic individuals. In the basal state, the liver represents a major site of insulin resistance, and this is reflected by overproduction of glucose despite the presence of both fasting hyperinsulinemia and hyperglycemia. This accelerated rate of hepatic glucose output is the primary determinant of the elevated fasting plasma glucose concentration in type 2 diabetic individuals. Although tissue (muscle) glucose uptake in the postabsorptive state is increased when viewed in absolute terms, the efficiency with which glucose is taken up (i.e., the glucose clearance) is diminished. After glucose infusion or ingestion (i.e., in the insulin stimulated state), both decreased muscle glucose uptake and impaired suppression of HGP contribute to the insulin resistance. Following glucose ingestion, the defects in insulin-mediated glucose uptake by muscle and the suppression of HGP by insulin contribute approximately equally to the disturbance in whole-body glucose homeostasis in type 2 diabetes. However, under euglycemic hyperinsulinemic conditions, HPG is largely suppressed and impaired muscle glucose uptake is primarily responsible for the insulin resistance.

DYNAMIC INTERACTION BETWEEN INSULIN SENSITIVITY AND INSULIN SECRETION IN TYPE 2 DIABETES

Type 2 diabetic subjects manifest abnormalities both in tissue (muscle, fat, and liver) sensitivity to insulin and in pancreatic insulin secretion. To understand how these two metabolic disturbances interact to produce the full-blown diabetic condition, it is necessary to quantitate insulin action and insulin secretion in the same individual over a wide range of insulin sensitivity. This dynamic interaction is demonstrated graphically by results obtained in healthy, lean, young normal glucose tolerant women who received a euglycemic insulin clamp (1 mUokg-1omin-1) and were stratified into quartiles based upon the rate of insulin-mediated glucose disposal (49) . Insulin secretion was measured independently on a separate day with a +125 mg/dl hyperglycemic clamp . Insulin resistance and insulin secretion were strongly and positively correlated (r=0.79, p<0.001). Women who were the most insulin resistant (quartile 1) had the highest fasting plasma insulin concentrations and highest early and late phase plasma insulin responses . Similar results relating the plasma insulin response and the severity of insulin resistance have been reported in normal glucose tolerant subjects with the minimal model technique (46,47) and the insulin suppression test/oral glucose tolerance test (214).

A number of groups have examined the dynamic interaction between insulin secretion and insulin sensitivity in type 2 diabetic subjects (1,4,34,35,38,39,42,46-48,58-61,150,162). DeFronzo (4) studied lean (ideal body weight < 120%) and obese (ideal body weight > 125%) subjects with varying degrees of glucose tolerance as follows: Group I-obese subjects (n=24) with normal glucose tolerance; Group II-obese subjects (n=23) with impaired glucose tolerance; Group III-obese subjects (n=35) with overt diabetes, subdivided into those with a hyperinsulinemic response and those with a hypoinsulinemic response during a 100-gram OGTT; Group IV-normal weight type 2 diabetics (n=26); Group V-normal weight subjects (n=25) with normal glucose tolerance. All subjects ingested 100 g of glucose to provide a measure of glucose tolerance and insulin secretion. Whole-body insulin sensitivity was quantitated with the euglycemic insulin (~100 µU/ml) clamp technique, which was performed with indirect calorimetry to quantitate rates of glucose oxidation and nonoxidative glucose disposal. The later primarily reflects glycogen synthesis (215).

In normal weight type 2 diabetic subjects, insulin-mediated whole-body glucose uptake was reduced by 40-50% and the impairment in insulin action resulted from defects in both oxidative and nonoxidative glucose metabolism (4) . Obese nondiabetic individuals were as insulin resistant as the normal-weight diabetic subjects (4) . Defects in both glucose oxidation and glucose storage contributed to the insulin resistance in the obese nondiabetic group . From the metabolic standpoint, therefore, obesity and type 2 diabetes closely resemble each other. Similar results concerning reduced whole-body insulin sensitivity in obese and type 2 diabetic individuals have been reported by other investigators (160,161,166,216-218). Despite nearly identical degrees of insulin resistance, normal-weight diabetic subjects manifested fasting hyperglycemia and marked glucose intolerance, whereas the obese nondiabetic individuals had normal or only minimally impaired oral glucose tolerance (4). This apparent paradox is explained by the plasma insulin response during the OGTT . Compared with control subjects, the obese nondiabetic group secreted more than twice as much insulin, and this was sufficient to offset the insulin resistance. In contrast, in normal-weight diabetic subjects, the pancreas, when faced with the same challenge, was unable to augment its secretion of insulin sufficiently to compensate for the insulin resistance. This imbalance between insulin supply by the ß-cells and the insulin requirement by tissues resulted in a frankly diabetic state, with fasting hyperglycemia and marked glucose intolerance.

When obesity and diabetes coexist in the same individual, the severity of insulin resistance is only slightly greater than that in either the normal-weight diabetic or nondiabetic obese groups (4) , and the magnitude of the defects in glucose oxidation and nonoxidative glucose disposal are similar in all obese and diabetic groups . Although hyperinsulinemic and hypoinuslinemic obese diabetic subjects were equally insulin resistant, the severity of glucose intolerance is worse in the hypoinsulinemic group, and this was related entirely to the presence of severe insulin deficiency .

An integrated summary of insulin action and insulin secretion in obese, diabetic, and lean groups is presented in figure 4 (coming soon). The plasma insulin response during the OGTT is shown in the top panel, along with the rate of insulin-mediated glucose disposal. In the obese nondiabetic subjects, tissue sensitivity to insulin is markedly reduced, but glucose tolerance remains perfectly normal because the ß-cells are able to augment their insulin secretory capacity appropriately to offset the defect in insulin action. As the obese individual develops impaired intolerance, there is a further reduction in insulin-mediated glucose disposal, which is due primarily to a decrease in glycogen synthesis. However, there is only a small additional impairment in glucose tolerance, because the ß-cells are able to augment further their secretion of insulin to counteract the deterioration in insulin sensitivity. The progression of the obese, glucose intolerant person to overt diabetes is heralded by a decline in insulin secretion without any worsening of insulin resistance . The obese diabetic has tipped over the top of Starling's curve of the pancreas and is now on the descending portion . Even though the plasma insulin response is increased compared to nondiabetic control subjects, it is not elevated appropriately for the degree of insulin resistance. In the normal-weight diabetic group, there is a further decline in glucose tolerance, which results from a greater impairment in insulin secretion without any additional deterioration in insulin sensitivity. Lastly, the obese diabetic group with a low insulin response manifests the greatest glucose intolerance, due to the presence of marked insulin deficiency without any further worsening of insulin sensitivity .

The natural history of type 2 diabetes described above is consistent with results in humans and monkeys published by other investigators (33-39,42,43,59-61,98,150). In lean subjects with a wide range of glucose tolerance, Reaven et al (42) demonstrated that the progression from normal to impaired glucose tolerance was marked by the development of severe insulin resistance, which was counterbalanced by a compensatory increase in insulin secretion. The onset of type 2 diabetes was associated with no (or only slight) further deterioration in tissue sensitivity to insulin . Rather, insulin secretion declined and the impairment in beta cell function was paralleled by a decrease in glucose tolerance . A similar sequence of events has been documented prospectively in Pima Indians (34-39,58,60). The sequence of events described in Caucasians (1,4,41,42,44,47,59,162,219), Pima Indians (34-39,58,60,219), and Pacific Islanders (33,62,220) is consistent with the development of type 2 diabetes in the rhesus monkeys (48). As monkeys grows older, they become obese and develop a diabetic condition closely resembling human type 2 diabetes. The earliest detectable abnormality in this primate model is a decrease in tissue sensitivity to insulin. Because of a compensatory increase in insulin secretion, the fasting plasma glucose concentration and glucose tolerance remain normal.

The studies detailed above indicate that insulin resistance is an early and characteristic feature of the natural history of type 2 diabetes in high risk populations. Overt diabetes develops only in those individuals whose beta cells are unable to appropriately augment their secretion of insulin to compensate for the defect in insulin action. It should be recognized, however, that there are well-described type 2 diabetic populations in whom insulin sensitivity is normal at the onset of diabetes, whereas insulin secretion is severely impaired (81-83). How frequently this occurs in typical type 2 diabetic patients remains to be determined. This insulinopenic variety of type 2 diabetes appears to be more common in African-Americans, elderly subjects, and lean Caucasians. In this later group, it is important to exclude type 1 diabetes, since ~10% of Caucasian individuals with older onset diabetes are islet cell antibody and/or GAD positive (220).