Long-lasting and fast methylglyoxal-scavenging peptide CycK(Myr)R4E alleviates chronic pain in type 2 diabetic mice

Abstract

Introduction: Pathological levels of methylglyoxal (MG), a reactive dicarbonyl product of glucose, contribute to major neurological complications associated with type II diabetes, including chronic neuropathic pain. Strategies to target elevated MG have included small molecule MG scavengers, but scavenger deficiencies in proteolytic stability and onset of scavenging activity have precluded clinical translation. To address this gap, we developed a long-lasting and highly reactive cyclic peptide CycK(Myr)R₄E, and here evaluated its antihyperalgesic efficacy in the db/db mouse model of type II diabetes painful diabetic neuropathy.

Objectives: To test the hypothesis that CycK(Myr)R₄E can reduce behavioral and molecular signs of painful diabetic neuropathy.

Methods: We assessed heat hypersensitivity as an index of hyperalgesia, and touch-evoked expression of phosphorylated extracellular signal-regulated kinase as a measure of neuronal activity in spinal cord dorsal horn.

Results: We report that a single systemic injection of CycK(Myr)R₄E (3 mg/kg) reversed heat hypersensitivity. Repeated systemic injection of CycK(Myr)R₄E (0.125 mg/kg, 3 times per week, 6-12 weeks of age) prevented heat hypersensitivity and reduced stimulus-evoked phosphorylated extracellular signal-regulated kinase.

Conclusion: These studies promote CycK(Myr)R₄E as the most promising MG scavenger for the prevention and treatment of hyperalgesia in type 2 diabetic neuropathic pain.

Keywords: CycK(Myr)R4E; Diabetes; Methylglyoxal; Methylglyoxal scavenger; Painful diabetic neuropathy; db/db.

Figure 1.

Repeated systemic administration of methylglyoxal (MG) (50 mg/kg body, i.p.—5 days a week from D1-D25) increases heat sensitivity in male (A) and female (B) mice. n = 6 per sex. Repeated MG administration did not change weight (n = 8 per group) (C), HbA1c level (n = 3–4 per group) (D), or blood glucose levels (n = 8 per group) (E) in either sex. Data represented as mean ± SEM. (P > 0.05). *P < 0.05 saline vs methylglyoxal (MG).

Figure 2.

Single intraperitoneal injection of CycK(Myr)R₄E (CycK) (0.3 and 3 mg/kg) dose-dependently reversed heat hypersensitivity in the (A) db/db mouse model of T2D, but not in db/+ (heterozygous controls). Effects began within hours and lasted several days. (B) Area under the curve (AUC) analyses revealed CycK(Myr)R₄E reduced heat hypersensitivity in db/db mice compared to the db/+ (heterozygous controls). n = 9 to 13 per group. Data represented as mean ± SEM. (P < 0.05). (A)–Two-way ANOVA; (B)–One-way ANOVA. *P < 0.05 db/db CycK(Myr)R₄E 3 mg/kg vs db/db saline. #P < 0.05 db/db CycK(Myr)R₄E 0.3 mg/kg vs db/db saline.

Figure 3.

Repeated intraperitoneal injections of CycK(Myr)R₄E (CycK) attenuated heat hypersensitivity in db/db mice. Line graph describing heat sensitivity at the hind paw after the systemic administration of CycK(Myr)R₄E (0.125 mg/kg, i.p.) in db/db and db/+ (heterozygous controls) mice. n = 7 to 8 per group, males and females aggregated. Data represented as mean ± SEM. *P < 0.05 db/db CycK(Myr)R₄E vs db/db saline.

Figure 4.

Repeated administration of CycK(Myr)R₄E (CycK) reduced touch-stimulated pERK expression in the spinal cord dorsal horn on the side ipsilateral to simulation. (A–L) Representative images from mice treated with either saline (A–C in db/+ and G–I in db/db mice) or CycK(Myr)R₄E (0.125 mg/kg, i.p.) (D–F in db/+ and J–L in db/db mice). Sections were colabeled with pERK (A, D, G, J) plus NeuN (B, E, H, K). Images were taken with a ×20 objective. Insets are magnified images of the boxed region to better show immunoreactivity of NeuN and pERK. (M) Histograms illustrating the number of pERK + profiles in lamina I–II. Images were taken with a ×10 objective. Scale bars: 100 μm. 3 to 4 mice per group. Data represented as mean ± SEM. *P < 0.05 db/db CycK(Myr)R₄E vs db/db saline.