In this study, although their having high glycemic levels, patients with T1DM under intensive insulin treatment showed a trend for lower LDL cholesterol, as well as faster removal of the LDE marker, 14C-cholesteryl ester as compared with the control subjects.
LDL cholesterol concentration in the plasma is determined by the balance between LDL production rates and the LDL removal from the plasma, which is largely dependent on the action of LDL receptors. In most clinical situations, slow LDL removal, rather than increased production rates, is the cause of hypercholesterolemia. Nonetheless, in a recent study [16
], we have shown that the plasmatic removal of LDL, as monitored by LDE cholesteryl ester FCR, was faster in athletes than in sedentary subjects, although both groups had equal levels of LDL cholesterol. Those results suggest that the increase in LDL removal was compensated by increased LDL production. LDL turnover in the plasma should be more often renewed in athletes, and consequently the LDL peroxidation should be diminished [16
]. In contrast in patients with familial hypercholesterolemia the plasma removal process of LDE is delayed [9
]. Delay in LDL clearance makes room for increased lipoprotein peroxidation, uptake of oxidized LDL by macrophage scavenger receptors and subsequent formation of foam cells [17
]. In the present study, in T1DM patients, the increased LDL removal, estimated by the LDE kinetics, resulted indeed in a trend for diminution of LDL, although not confirmed statistically. Non-HDL includes not only LDL but also other VLDL catabolic products such as intermediate density lipoprotein (IDL) and VLDL remnants that are also removed by LDL receptors or by LRP receptors [18
]. Apo B concentration, that also marks the non-HDL lipoproteins, also tended to be lower in our T1DM patients. Insulin can act on LDL receptors by inducing receptor overexpression and increase in receptor activity [19
]. Therefore, the fast removal of LDE, shown in the present study, was probably accounted for stimulation of the LDL receptors consequent to peripheral hyperinsulinemia in T1DM patients.
Insulin action on glycemia is exerted chiefly through glucose cellular uptake mediated by GLUT4 [20
] and inhibition of glycogenolysis and gluconeogenesis [21
]. The actions on lipoprotein metabolism are exerted mainly through increase in lipolysis of triglyceride-rich lipoproteins by stimulation of lipoprotein lipase [22
], and inhibition of lipolysis of fats stored in the tissues by inhibition of hormone-sensitive lipase [23
]. Apparently, the subcutaneous insulin dose scheme that was insufficient to optimize glycemia-related mechanisms of action had the ability to optimize the LDL pathway.
Depending on the route of insulin administration, the concentration of insulin in the peripheral circulation is different from that of the portal circulation, which conceivably has consequences on insulin availability and action and differences between subcutaneous and intraperitonial routes were documented [24
]. Administration of excess exogenous insulin by subcutaneous route increases the catabolic rates of apo B LDL [25
], similarly to our findings of increased LDE clearance in the T1DM patients. Accelerated LDL clearance diminishes the exposure of the lipoprotein to oxidation and other pro-atherogenic effects [25
H-free cholesterol removal from the plasma was similar in T1DM and controls, but the 14
C-cholesteryl ester label is more reliable as tracer of LDE decay in the plasma than 3
H-free cholesterol. In fact, 3
H-free cholesterol kinetics was designed to evaluate the cholesterol esterification process in cases compared to controls. In the plasma, LDE also gains apo A1, the LCAT co-factor so that free cholesterol contained in LDE may suffer the esterification reaction [11
]. Another possibility is that free cholesterol may partially shift from LDE to HDL, where it can also be esterified.
Cholesterol esterification may be related with CAD: in our previous study, we showed that in non-diabetic CAD patients cholesterol esterification was higher [26
]. Nonetheless, in this study, in all points of the LDE decaying curves, cholesterol esterification did not statistically differ between T1DM and controls. Cholesterol esterification and LCAT activity have been poorly explored in T1DM. Chang et al. observed that the LCAT activity was higher in T1DM with higher glycemic levels than in those with lower glycemia [27
], but comparisons with non-diabetic controls were not reported. Our data showing that LDE cholesterol esterification was equal in T1DM and controls may be helpful for future targeting the status of this reaction in T1DM with or without CAD.
T1DM patients treated with insulin also showed normal levels of fasting triglyceridemia. In diabetes, deficient function of insulin-dependent lipoprotein lipase may lead not only to hypertriglyceridemia but also to decreased HDL-cholesterol levels. This is the so-called “see-saw effect” triglycerides-HDL cholesterol consequent to increase in lipid exchanges between VLDL and HDL. In respect to HDL, not only the HDL-cholesterol and apo A1 levels were normal but also the metabolic step of lipid transfers to HDL was normal in the T1DM patients. Recently, it was shown that patients with documented coronary artery disease had alterations in lipid transfers to HDL such as decrease of free cholesterol transfers to the lipoprotein [28
In a previous study performed in women with polycystic ovary syndrome [29
], a correlation was found between insulin resistance, measured by the HOMA-IR index and that is associated with the syndrome, with the transfer of triglycerides to HDL. On the other hand, the action of both CETP and PLTP, that promote lipid transfer, was found to be altered in T1DM patients, and in another study CETP alterations were related with the presence of macrovascular disease [30
]. Thus, lack of differences in lipid transfer in T1DM patients and controls may suggest that intensive insulin treatment could have contributed to the normal values in T1DM. Due to the importance of lipid transfer for HDL remodeling and function, this result, together with the finding of normal HDL-cholesterol and apo A1 levels adds up for the protection of intensive insulin treatment against macrovascular disease by maintenance of normal plasma lipid metabolism pathways.
Native LDL is recognized by LDL receptors by means of apo B, the only apo present in this lipoprotein. Recognition of LDE by receptors through apo E, which has greater affinity for the receptors results in removal of LDE considerably faster than the native lipoprotein[8
], which facilitates the execution of the experimental protocols and the comfort of the study subjects by shortening the blood sampling period. In many studies, LDE kinetics was accelerated or slowed as previously reported for the native lipoprotein [31
]. Nonetheless, as a limitation of those studies, the differences in protein recognition by the receptors and the compositional differences between LDE and the native lipoprotein are to be taken into account in the interpretation of the LDE-generated data for extrapolation to LDL metabolism.
It is worthwhile to point out that despite the beneficial effect on lipoprotein metabolism, as documented in this study, patients under intensive insulin therapy have reportedly higher cardiovascular mortality risk [34
]. This may be ascribed to the short follow-up periods of the insulin-treated patients, since atherosclerosis is a long-run process. In fact, in studies with 17 and 20 year follow-up, intensive insulin therapy achieved reduction of events in both T1DM and type 2 diabetes mellitus; patients were younger and had no manifestations of cardiovascular disease when they begun the treatment [36
In conclusion, although intensive insulin treatment was not sufficient to normalize glycemia, no abnormalities were found in the aspects of lipid metabolism examined here; the LDE clearance was even increased. Presumably, this dichotomy could have resulted from different thresholds of insulin levels to optimize glucose, on one hand, and plasma lipid metabolism, on the other. Insulin bioavailability after subcutaneous administration was also conceivably key for this outcome. The fact that under insulin treatment optimal lipid parameters can be attained independently of the glycemic control could be fundamental because of the importance of dyslipidemias in the development of diabetic macrovascular disease.