Microassay for glucose-induced preproinsulin mRNA expression to assess islet functional potency for islet transplantation

Transplantation. 2010 Jan 27;89(2):146-54. doi: 10.1097/TP.0b013e3181c4218d.


Background: The capacity for insulin synthesis in islets is important for islet transplantation to succeed. We developed a microassay that evaluates the potency of human islets by measuring changes in glucose-induced human insulin gene (INS) expression using a single islet in octuplicate samples.

Methods: Poly (A) messenger RNA (mRNA) was purified from a set of single handpicked human islets. Glucose-induced mature (postspliced) and premature (prespliced) insulin mRNA were quantified by reverse-transcriptase polymerase chain reaction using several insulin mRNA primers designed at different locations including, intron, exon, and an exon-intron junction.

Results: The synthesis of premature INS mRNA was significantly increased in islets exposed to high glucose for 16 vs. 4 hr (P<0.01), whereas mature INS mRNA showed no difference. Glucose-induced premature INS mRNA synthesis was attenuated in heat-damaged islets. Stimulation index (SI) calculated by normalizing premature by mature INS mRNA (SI_INS mRNA) positively correlated with SI of insulin release (SI_16h insulin) from the same set of islets during 16-hr incubation in high or low glucose media, and SI of glucose-mediated insulin release obtained from the same islet lot in a perifusion system (n=12). Furthermore, linear multiple regression analysis using SI_INS mRNA and SI_16h insulin predicted islet transplantation outcome in nonobese diabetic (NOD) scid mice (n=8).

Conclusion: The measurement of glucose-induced premature INS mRNA normalized by mature INS mRNA can be used to assess the functional quality of human islets and may predict islet function after transplantation in type 1 diabetic patients.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis
  • DNA Primers
  • DNA, Complementary / genetics
  • Exons
  • Gene Amplification
  • Gene Expression Regulation / drug effects*
  • Glucose / pharmacology*
  • Humans
  • Insulin / genetics*
  • Insulin / metabolism
  • Insulin Secretion
  • Insulin-Secreting Cells / cytology
  • Insulin-Secreting Cells / physiology
  • Introns
  • Islets of Langerhans / cytology
  • Islets of Langerhans / drug effects
  • Islets of Langerhans / metabolism
  • Islets of Langerhans / physiology*
  • Islets of Langerhans Transplantation / methods*
  • Protein Precursors / genetics*
  • RNA, Messenger / genetics*


  • DNA Primers
  • DNA, Complementary
  • Insulin
  • Protein Precursors
  • RNA, Messenger
  • preproinsulin
  • Glucose