In vitro and in vivo insulinotropic properties of the multifunctional frog skin peptide hymenochirin-1B: a structure-activity study

Amino Acids. 2016 Feb;48(2):535-47. doi: 10.1007/s00726-015-2107-x. Epub 2015 Oct 6.


Hymenochirin-1b (Hym-1B; IKLSPETKDNLKKVLKGAIKGAIAVAKMV.NH2) is a cationic, α-helical amphibian host-defense peptide with antimicrobial, anticancer, and immunomodulatory properties. This study investigates the abilities of the peptide and nine analogues containing substitutions of Pro(5), Glu(6), and Asp(9) by either L-lysine or D-lysine to stimulate insulin release in vitro using BRIN-BD11 clonal β cells or isolated mouse islets and in vivo using mice fed a high-fat diet to produce obesity and insulin resistance. Hym-1B produced a significant and concentration-dependent increase in the rate of insulin release from BRIN-BD11 cells without cytotoxicity at concentrations up to 1 µM with a threshold concentration of 1 nM. The threshold concentrations for the analogues were: [P5K], [E6K], [D9K], [P5K, E6K] and [E6K, D9k] 0.003 nM, [E6K, D9K] and [D9k] 0.01 nM, [P5K, D9K] 0.1 nM and [E6k] 0.3 nM. All peptides displayed cytotoxicity at concentrations ≥1 µM except the [P5K] and [D9k] analogues which were non-toxic at 3 µM. The potency and maximum rate of insulin release from mouse islets produced by the [P5K] peptide were significantly greater than produced by Hym-1B. Neither Hym-1B nor the [P5K] analogue at 1 µM concentration had an effect on membrane depolarization or intracellular Ca(2+). The [P5K] analogue (1 µM) produced a significant increase in cAMP concentration in BRIN-BD11 cells and stimulated GLP-1 secretion from GLUTag cells. Down-regulation of the protein kinase A pathway by overnight incubation with forskolin completely abolished the insulin-releasing effects of [P5K]hym-1B. Intraperitoneal administration of the [P5K] and [D9k] analogues (75 nmol/kg body weight) to high-fat-fed mice with insulin resistance significantly enhanced glucose tolerance with a concomitant increase in insulin secretion. We conclude that [P5K]hym-1B and [D9k]hym-1B show potential for development into anti-diabetic agents.

Keywords: Amphibian skin peptide; Hymenochirin-1b; Insulin release; Type 2 diabetes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amphibian Proteins / pharmacology*
  • Animals
  • Antimicrobial Cationic Peptides / chemical synthesis
  • Antimicrobial Cationic Peptides / pharmacology*
  • Anura
  • Calcium / metabolism
  • Cell Line
  • Cyclic AMP / biosynthesis
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Diabetes Mellitus, Type 2 / drug therapy*
  • Diet, High-Fat
  • Glucagon-Like Peptide 1 / metabolism
  • Glucose / metabolism
  • Hypoglycemic Agents / pharmacology*
  • Insulin / metabolism*
  • Insulin Resistance
  • Insulin Secretion
  • Islets of Langerhans / metabolism
  • L-Lactate Dehydrogenase / metabolism
  • Membrane Potentials / drug effects
  • Mice
  • Mice, Inbred C57BL
  • Organ Culture Techniques
  • Protein Kinase C / metabolism
  • Rats
  • Structure-Activity Relationship


  • Amphibian Proteins
  • Antimicrobial Cationic Peptides
  • Hypoglycemic Agents
  • Insulin
  • hymenochirin-1B, Hymenochirus boettgeri
  • Glucagon-Like Peptide 1
  • Cyclic AMP
  • L-Lactate Dehydrogenase
  • Cyclic AMP-Dependent Protein Kinases
  • Protein Kinase C
  • Glucose
  • Calcium