C-Peptide 2 (rat)

The repair capacity of progenitor skeletal muscle satellite cells (SC) in Type 1 diabetes mellitus (T1DM) is decreased. This is associated with the loss of skeletal muscle function. In T1DM, the deficiency of C-peptide along with insulin is associated with an impairment of skeletal muscle functions such as growth, and repair, and is thought to be an important contributor to increased morbidity and mortality. Recently, cholesterol-lowering drugs (statins) have also been reported to increase the risk of skeletal muscle dysfunction. We hypothesised that C-peptide activates key signaling pathways in myoblasts, thus promoting cell survival and protecting against simvastatin-induced myotoxicity. This was tested by investigating the effects of C-peptide on the L6 rat myoblast cell line under serum-starved conditions. Results: C-peptide at concentrations as low as 0.03 nM exerted stimulatory effects on intracellular signaling pathways-MAP kinase (ERK1/2) and Akt. When apoptosis was induced by simvastatin, 3 nM C-peptide potently suppressed the apoptotic effect through a pertussis toxin-sensitive pathway. Simvastatin strongly impaired Akt signaling and stimulated the reactive oxygen species (ROS) production; suggesting that Akt signaling and oxidative stress are important factors in statin-induced apoptosis in L6 myoblasts. The findings indicate that C-peptide exerts an important protective effect against death signaling in myoblasts. Therefore, in T1DM, the deficiency of C-peptide may contribute to myopathy by rendering myoblast-like progenitor cells (involved in muscle regeneration) more susceptible to the toxic effects of insults such as simvastatin.

Na+,K(+)-ATPase is an ubiquitous membrane enzyme that allows the extrusion of three sodium ions from the cell and two potassium ions from the extracellular fluid. Its activity is decreased in many tissues of streptozotocin-induced diabetic animals. This impairment could be at least partly responsible for the development of diabetic complications. Na+,K(+)-ATPase activity is decreased in the red blood cell membranes of type 1 diabetic individuals, irrespective of the degree of diabetic control. It is less impaired or even normal in those of type 2 diabetic patients. The authors have shown that in the red blood cells of type 2 diabetic patients, Na+,K(+)-ATPase activity was strongly related to blood C-peptide levels in non-insulin-treated patients (in whom C-peptide concentration reflects that of insulin) as well as in insulin-treated patients. Furthermore, a gene-environment relationship has been observed. The alpha-1 isoform of the enzyme predominant in red blood cells and nerve tissue is encoded by the ATP1A1 gene. A polymorphism in the intron 1 of this gene is associated with lower enzyme activity in patients with C-peptide deficiency either with type 1 or type 2 diabetes, but not in normal individuals. There are several lines of evidence for a low C-peptide level being responsible for low Na+,K(+)-ATPase activity in the red blood cells. Short-term C-peptide infusion to type 1 diabetic patients restores normal Na+,K(+)-ATPase activity. Islet transplantation, which restores endogenous C-peptide secretion, enhances Na+,K(+)-ATPase activity proportionally to the rise in C-peptide. This C-peptide effect is not indirect. In fact, incubation of diabetic red blood cells with C-peptide at physiological concentration leads to an increase of Na+,K(+)-ATPase activity. In isolated proximal tubules of rats or in the medullary thick ascending limb of the kidney, C-peptide stimulates in a dose-dependent manner Na+,K(+)-ATPase activity. This impairment in Na+,K(+)-ATPase activity, mainly secondary to the lack of C-peptide, plays probably a role in the development of diabetic complications. Arguments have been developed showing that the diabetes-induced decrease in Na+,K(+)-ATPase activity compromises microvascular blood flow by two mechanisms: by affecting microvascular regulation and by decreasing red blood cell deformability, which leads to an increase in blood viscosity. C-peptide infusion restores red blood cell deformability and microvascular blood flow concomitantly with Na+,K(+)-ATPase activity. The defect in ATPase is strongly related to diabetic neuropathy. Patients with neuropathy have lower ATPase activity than those without. The diabetes-induced impairment in Na+,K(+)-ATPase activity is identical in red blood cells and neural tissue. Red blood cell ATPase activity is related to nerve conduction velocity in the peroneal and the tibial nerve of diabetic patients. C-peptide infusion to diabetic rats increases endoneural ATPase activity in rat. Because the defect in Na+,K(+)-ATPase activity is also probably involved in the development of diabetic nephropathy and cardiomyopathy, physiological C-peptide infusion could be beneficial for the prevention of diabetic complications.

With the use of a rabbit polyclonal antiserum against a conserved region (54-118) of C-peptide of human preproinsulin-like peptide 7, referred to herein as C-INSL7, neurons expressing C-INSL7-immunoreactivity (irC-INSL7) were detected in the pontine nucleus incertus, the lateral or ventrolateral periaqueductal gray, dorsal raphe nuclei and dorsal substantia nigra. Immunoreactive fibers were present in numerous forebrain areas, with a high density in the septum, hypothalamus and thalamus. Pre-absorption of C-INSL7 antiserum with the peptide C-INSL7 (1 microg/ml), but not the insulin-like peptide 7 (INSL7; 1 microg/ml), also known as relaxin 3, abolished the immunoreactivity. Optical imaging with a voltage-sensitive dye bis-[1,3-dibutylbarbituric acid] trimethineoxonol (DiSBAC4(3)) showed that C-INSL7 (100 nM) depolarized or hyperpolarized a small population of cultured rat hypothalamic neurons studied. Ratiometric imaging studies with calcium-sensitive dye fura-2 showed that C-INSL7 (10-1000 nM) produced a dose-dependent increase in cytosolic calcium concentrations [Ca2+]i in cultured hypothalamic neurons with two distinct patterns: (1) a sustained elevation lasting for minutes; and (2) a fast, transitory rise followed by oscillations. In a Ca2+-free Hanks' solution, C-INSL7 again elicited two types of calcium transients: (1) a fast, transitory increase not followed by a plateau phase, and (2) a transitory rise followed by oscillations. INSL7 (100 nM) elicited a depolarization or hyperpolarization in a small population of hypothalamic neurons, and an increase of [Ca2+]i with two patterns that were dissimilar from that of C-INSL7. [125I]C-INSL7 bindings to rat brain membranes were inhibited by C-INSL7 in a dose-dependent manner; the Kd and Bmax. values were 17.7 +/- 8.2 nM and 45.4 +/- 20.5 fmol/mg protein. INSL7 did not inhibit [125I]C-INSL7 binding to rat brain membranes, indicating that C-INSL7 and INSL7 bind to distinct binding sites. Collectively, our result raises the possibility that C-INSL7 acts as a signaling molecule independent from INSL7 in the rat CNS.