C－肽是一个有生物活性的分子

Administration of C-peptide in type 1 diabetes patients results in increased blood flow in several tissues. Skeletal muscle perfusion (forearm) at rest and during exercise is increased in a concentration-dependent manner across the 0–1 nmol/l range, but higher concentrations of C-peptide elicit no further rise in blood flow [16]. In type 1 patients without signs of cardiac disease, C-peptide infusion results in augmented left ventricular blood flow. The patients also show improved rates of myocardial contraction, increased ejection fraction, plus a shortening of the QT interval [17,18]. There is also augmented skin capillary blood cell velocity and redistribution of blood flow to nutritive capillaries following C-peptide administration [19]. Direct measurements of sciatic endoneurial blood flow in diabetic animals demonstrated augmented nerve perfusion in response to C-peptide administration [13,20]. It is likely that the observed circulatory effects of C-peptide are mediated by stimulation and increased expression of endothelial nitric oxide synthase (eNOS) (see below), since the effect may be prevented by a NOS inhibitor [13]. An antithrombotic effect of C-peptide in diabetic mice has also been reported [21].

The nature of C-peptide cellular interactions has not been fully determined. Fluorescence correlation spectroscopy has shown the stereospecific binding of C-peptide to renal tubular cells, endothelial cells, skin fibroblasts, mesangial cells and neuroblastoma cells; the association constant was 3 × 10⁹ l/mol and the binding curve indicated saturation at 0.9 nmol/l [22]. C-peptide binding did not show cross-reactivity with insulin, IGF-1 or IGF-2. Binding of the peptide to detergent-solubilised cell fractions and cell lysates has also been demonstrated using fluorescence correlation spectroscopy and surface plasmon resonance [23], but there are no data based on conventional radioligand binding. It is unlikely that C-peptide interacts directly with lipid cell membrane components [24]. The identity of the cellular binding structure has so far proven elusive and a specific receptor has not been identified.

Figure 2provides an overview of intracellular responses to C-peptide. Most of the intracellular actions can be blocked by pre-incubation of cells with pertussis toxin, indicating the involvement of a G_i/G_o-linked protein in C-peptide signalling [25–28], as confirmed by direct studies assessing guanosine triphosphate γS binding to a G_iprotein after C-peptide exposure [29]. Exposure of renal tubular and endothelial cells to C-peptide at physiological concentrations results in a prompt elevation of intracellular Ca²⁺concentrations [27,30]. C-peptide also elicits phosphorylation of several protein kinase C isoforms and phosphatidylinositol 3-kinase [26,28,31]. Activation of one or several components of the mitogen-activated protein (MAP) kinase cascade in a concentration-dependent manner is consistently observed in all examined cell types following exposure to C-peptide [26,28,32,33]. C-peptide has also been found to mimic the effects of insulin in muscle cells [32].

Fig. 2 Preliminary model of C-peptide signalling pathways. The influence of C-peptide on eNOS and Na⁺/K⁺-ATPase include both activation and induction.Dashed linesindicate insulinomimetic signalling demonstrated in muscle cells.ATF1, activating transcription factor 1;CREB, cyclic AMP response element-binding protein;ERK1/2, extracellular signal-regulated kinase 1/2; GTPγS, guanosine triphosphate γS;JNK, c-Jun N-terminal kinase;PI3-K, phosphatidylinositol 3-kinase;PKB, protein kinase B;PKC, protein kinase C;PLC, phospholipase C;PPARγ, peroxisome proliferator-activated receptor γ;ZEB, zinc finger homeodomain enhancer-binding protein

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Interactions between C-peptide and insulin oligomers have been identified using surface plasmon resonance. Unexpectedly, it was discovered that C-peptide influences the disaggregation of insulin, probably by binding to insulin oligomers, with dissociation constants in the micromolar range [41]. Mass spectrometry revealed that insulin hexamers in solution became undetectable in the presence of C-peptide, leading to the conclusion that C-peptide binds to and causes disaggregation of hexameric insulin, increasing the availability of biologically active, monomeric insulin. Accordingly, subcutaneous injection of an insulin and C-peptide mixture in type 1 diabetes patients has been found to result in a more rapid appearance of insulin in plasma and more marked stimulation of glucose utilisation compared with injection of insulin alone [41].

The physiological effects of C-peptide can be demonstrated only in patients or animals deficient in C-peptide and not in healthy individuals. The nature of the C-peptide binding curve may help explain this unusual feature. The curve indicates saturation of cellular binding at approximately 0.9 nmol/l [22], which is within the physiological range. Thus, in healthy subjects and animals with normal beta cell function, receptor saturation is probably achieved at the ambient C-peptide concentration, so that no further effects can be expected in response to exogenous administration.

The structural variability of C-peptide in different species has been put forward as an argument that C-peptide is unlikely to possess biological activity. C-peptide is, however, not unique in this regard. Parathyroid hormone, gastrin-releasing peptide and relaxin all show similar inter-species variability. In addition, among mammalian species, nine residues (Glu1, Glu3, Gln6, Val7, Glu11, Leu12, Leu26, Glu27 and Gln31), localised primarily to the N- and C-terminal segments, show ≥90% conservation [42]. The exact functional correlates for these residues are not apparent, but Glu3, Glu11 and particularly Glu27 are all known to be important for the cellular effects of C-peptide [42].

The DCCT study demonstrated a decreased occurrence of complications in the group of type 1 diabetes patients that received intensive insulin therapy and achieved improved blood glucose control, in keeping with the view that hyperglycaemia is a major culprit in the pathogenesis of microvascular complications. Even though blood glucose levels were close to normal in the group that received intensive treatment, a large proportion of these patients still developed complications: 40% presented with clinical neuropathy or grossly abnormal nerve conduction after 5 years [43], and a similar fraction had developed signs of nephropathy or retinopathy. This suggests that factors in addition to hyperglycaemia may be involved in the development of microvascular complications in type 1 diabetes patients. In view of the data for C-peptide described above, we suggest that lack of C-peptide contributes to the pathogenesis of complications of type 1 diabetes. If this were the case, one would expect that the complications associated with type 1 diabetes would be different from those associated with type 2. Direct comparisons have been made in patients and in animal models of diabetes, particularly with respect to neuropathy. Thus, neuropathy caused by type 1 diabetes occurs more predictably and progresses more rapidly than in type 2 diabetes. Both patients and experimental animals with type 1 diabetes neuropathy show characteristic structural abnormalities of the nodal and paranodal myelin sheath and ion channel barrier, which affect the large myelinated fibres in particular; these derangements are not seen in type 2 diabetes [44,45]. In addition, type 1 diabetic animals show progressive axonal degeneration coupled with impaired regenerative capacity, resulting in the gradual loss of nerve fibres, and this is either not present or significantly milder in animal models of type 2 diabetes [45]. Finally, in type 1 diabetes, but not in type 2, key nodal and paranodal molecules, e.g. contactin, caspr, and ankyrin_Gare downregulated [46]. In view of the above data, we propose that, besides hyperglycaemia, C-peptide deficiency is an important factor in the pathogenesis of neuropathy and possibly other microvascular complications of type 1 but not type 2 diabetes. Accordingly, type 1 diabetes should be considered a dual hormone deficiency disease and C-peptide replacement therapy may be beneficial in its treatment.

Why is it that despite the extensive documentation of multifaceted effects (Table 1), C-peptide is not yet generally recognised as a bioactive peptide? The fact that the peptide has no effects when administered to healthy animals or individuals may be partly responsible for this. An explanation for this unusual feature, based on the binding characteristics of the peptide, is presented above. Another complicating factor in this regard may be the lack of immediate or major effects of C-peptide withdrawal, as, for example, at the onset of type 1 diabetes. This may indicate that C-peptide is one of several players in the multifactorial system that regulates tissue microcirculation and endothelial function. The C-peptide contribution may be seen as redundant, at least in the short term. The presence of multiple regulatory mechanisms often signifies the vital importance of the specific physiological process being regulated, and the presence of compensatory regulation may help explain why a lack of C-peptide gives rise only to late manifestations that primarily occur in specific tissues. A third objection has been that the biological and clinical importance of C-peptide has not been established. This objection is refuted on the basis of the wealth of information now presented. Hence, we consider C-peptide to be bioactive.

Clearly, there is much more to learn about C-peptide. Identification of the mechanism whereby C-peptide interacts with cell membranes and further delineation of its intracellular signalling pathways and transcriptional effects in different cell types would enhance our understanding of C-peptide bioactivity. On the clinical side, further studies of long duration will be required to document the robustness of its beneficial effects on the different types of long-term complications to define its possible therapeutic role in the therapy of type 1 diabetes, as also emphasised in the accompanying paper by Luzi et al. [47]. Nevertheless, despite the fact that our knowledge is still incomplete, there are several lines of evidence in support of the notion that C-peptide is a bioactive peptide and that its replacement in type 1 diabetes may be beneficial in the treatment of long-term complications. Specific cellular binding of the peptide, its intracellular signalling characteristics and end effects, including its action on eNOS, Na⁺/K⁺-ATPase and several transcription factors, are now established for many cell systems and by different investigators. Results from studies in type 1 diabetes patients and animal models demonstrate that, at replacement doses, C-peptide exerts beneficial effects on the early stage functional and structural abnormalities of both the kidneys and the peripheral nerves. Even a cautious evaluation of the available evidence thus presents the picture of a bioactive peptide with therapeutic potential.