Elsevier

Journal of Diabetes and its Complications

Volume 27, Issue 6, November–December 2013, Pages 538-547
Journal of Diabetes and its Complications

C-peptide preserves the renal microvascular architecture in the streptozotocin-induced diabetic rat

https://doi.org/10.1016/j.jdiacomp.2013.07.002Get rights and content

Abstract

Aims

C-peptide is renoprotective in type 1 diabetes, however, the mechanisms of its actions are not completely understood. We hypothesized that C-peptide attenuates diabetes-associated renal microvascular injury.

Method

After 4 or 8 weeks of streptozotocin (STZ)-induced diabetes, rats received either vehicle or C-peptide in the presence of low or high doses of insulin. Urine albumin excretion (UAE) was measured prior to initiation of treatment (baseline) and 2 or 4 weeks after treatment (sacrifice). Glomerular hypertrophy, glomerular filtration rate (GFR) and renal microvascular density, quantified ex vivo by 3D micro-CT reconstruction, were measured at sacrifice.

Results

In rats receiving low doses of insulin, treatment with C-peptide reduced HbA1c levels by 24%. In these rats, the 107% increase in UAE rate from baseline to sacrifice in vehicle-treated rats was largely prevented with C-peptide. C-peptide also reduced diabetes-associated glomerular hyperfiltration by 30%, glomerular hypertrophy by 22% and increased the density of microvessels between 0 and 500 μm in diameter by an average of 31% compared with vehicle-treated groups. Similar renoprotective effects of C-peptide were observed in rats treated with higher doses of daily insulin, despite no differences in HbA1c levels.

Conclusions

The study suggests that C-peptide is renoprotective by preserving the integrity of the renal microvasculature irrespective of glucose regulation.

Introduction

C-peptide plays the important role of connecting the A and B chains of the pro-insulin molecule and once cleaved off, it is released from pancreatic beta-cells in equal molar amounts to insulin (Hills et al., 2010, Horwitz et al., 1975, Steiner, 2004). Due to the high degree of structural variability between species and a lack of a conserved sequence, C-peptide has historically been thought to have no functional significance independent of insulin production and has only been used clinically as a surrogate marker of insulin release and a measure of beta-cell activity in diabetes mellitus (Faber and Binder, 1977, Horwitz et al., 1975, Roth et al., 2012, Steiner, 2004). However, recent studies have shown that C-peptide may have therapeutic applications in the treatment of type 1 diabetes-associated end-organ complications, including diabetic nephropathy (Huang et al., 2002, Johansson et al., 2000, Nordquist et al., 2007, Samnegard et al., 2004, Sjoquist et al., 1998). Indeed, studies have shown that acute and chronic infusion of C-peptide reduces albuminuria, proteinuria, glomerular hyperfiltration and hypertrophy in streptozotocin (STZ)-induced diabetic rats and mice or alloxan-induced diabetic mice (Huang et al., 2002, Maezawa et al., 2006, Nordquist et al., 2009, Nordquist et al., 2008, Samnegard et al., 2001, Samnegard et al., 2005, Sjoquist et al., 1998). Furthermore, clinical studies show that long term administration of C-peptide to patients with type 1 diabetes decreases albuminuria and improves renal function (Johansson et al., 2000). However, the mechanisms by which C-peptide protects the diabetic kidney are unclear.

Most efforts to examine the potential mechanisms by which C-peptide exerts some of its actions in the diabetic kidney have focused on its role in regulating microvascular changes. Specifically, several studies have shown that C-peptide reduces glomerular hyperfiltration associated with early stages of diabetes by constricting afferent arterioles (Huang et al., 2002, Nordquist et al., 2008, Nordquist et al., 2009, Samnegard et al., 2001, Samnegard et al., 2005). In addition, diabetic nephropathy is also characterized by early and progressive renal microvascular rarefaction (Khavandi et al., 2009, Lindenmeyer et al., 2007, Maric-Bilkan et al., 2012). Thus, successful therapeutic treatments for diabetic renal disease should include preservation of the renal microvascular function and architecture. Therefore, the current study was designed to test the hypothesis that C-peptide protects the kidney in STZ-induced diabetic rat by preserving the structure and integrity of the renal microvasculature.

Section snippets

Effects of C-peptide in diabetic rats and non-diabetic rat in the presence of low doses of insulin

The study was performed in 12-week old male Sprague–Dawley rats (Harlan, Madison, WI) maintained on standard rat chow and tap water ad libitum. Nineteen rats were rendered diabetic with a single i.p. injection of streptozotocin (STZ, 55 mg/kg in 0.1 mM citrate buffer, pH 7.4) as previously described (Mankhey, Wells, Bhatti, & Maric, 2006) and given insulin (2–4 U every 3 days, Lantus, Aventis Pharmaceuticals Inc., Kansas City, MO) throughout the study to prevent excessive weight loss and mortality.

Effects of C-peptide on metabolic function in non-diabetic and diabetic rats treated with low doses of insulin

No differences in body weight, water or food intake were observed between the vehicle (diabetic) and C-peptide (diabetic)-treated groups (Table 1). Plasma levels of C-peptide (diabetic) were 135% higher in the C-peptide-treated compared with vehicle (diabetic)-treated animals (Table 1); however, these levels were approximately 2.5-fold lower than those observed in a non-diabetic rat (Sawyer et al., 2012).

For the first 8 weeks of untreated diabetes, no differences in blood glucose levels were

Discussion

The current study shows that, in animals receiving low doses of insulin, the 4-week treatment with C-peptide administered after an 8-week period of untreated diabetes, reduced UAE, glomerular hyperfiltration, glomerular hypertrophy and macrophage infiltration. Furthermore, treatment with C-peptide increased the density of renal microvessels up to 500 μm in diameter, which was accompanied by reduced renal protein expression of VEGFR2 and pro-apoptotic Bax and increased expression of

Conclusion

The current study shows that C-peptide in the STZ-induced diabetic rat is renoprotective independent of blood glucose regulation. The renoprotective effects of C-peptide were observed after a period of untreated diabetes, which supports C-peptide therapy as a potential intervention for slowing or possibly, reversing the progression of renal injury in patients with already established diabetic renal disease.

Acknowledgments

The authors would like to thank Ms. Stephanie Evans for technical assistance with paraffin sectioning and Ms. Haiyan Zhang for measurement of plasma insulin levels. This work was supported by National Institutes of Health Grants DK075832 (C. Maric-Bilkan), HL095638 (A. Chade) and HL51971 (J.E. Hall for the support of core facilities used for part of the study) as well as the Intramural Research grant (C. Maric-Bilkan).

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    Disclosures: None.

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