Long-lasting effect of perinatal exposure to L-tryptophan on circadian clock of primary cell lines established from male offspring born from mothers fed on dietary protein restriction

PLoS One. 2013;8(2):e56231. doi: 10.1371/journal.pone.0056231. Epub 2013 Feb 27.


Background aims: Maternal undernutrition programs metabolic adaptations which are ultimately detrimental to adult. L-tryptophan supplementation was given to manipulate the long-term sequelae of early-life programming by undernutrition and explore whether cultured cells retain circadian clock dysregulation.

Methods: Male rat pups from mothers fed on low protein (8%, LP) or control (18%, CP) diet were given, one hour before light off, an oral bolus of L-tryptophan (125 mg/kg) between Day-12 and Day-21 of age. Body weight, food intake, blood glucose along with the capacity of colonization of primary cells from biopsies were measured during the young (45-55 days) and adult (110-130 days) phases. Circadian clock oscillations were re-induced by a serum shock over 30 hours on near-confluent cell monolayers to follow PERIOD1 and CLOCK proteins by Fluorescent Linked ImmunoSorbent Assay (FLISA) and period1 and bmal1 mRNA by RT-PCR. Cell survival in amino acid-free conditions were used to measure circadian expression of MAP-LC3B, MAP-LC3B-FP and Survivin.

Results: Tryptophan supplementation did not alter body weight gain nor feeding pattern. By three-way ANOVA of blood glucose, sampling time was found significant during all phases. A significant interaction between daily bolus (Tryptophan, saline) and diets (LP, CP) were found during young (p = 0.0291) and adult (p = 0.0285) phases. In adult phase, the capacity of colonization at seeding of primary cells was twice lower for LP rats. By three-way ANOVA of PERIOD1 perinuclear/nuclear immunoreactivity during young phase, we found a significant effect of diets (p = 0.049), daily bolus (p<0.0001) and synchronizer hours (p = 0.0002). All factors were significantly interacting (p = 0.0148). MAP-LC3B, MAP-LC3B-FP and Survivin were altered according to diets in young phase.

Conclusions: Sequelae of early-life undernutrition and the effects of L-tryptophan supplementation can be monitored non-invasively by circadian sampling of blood D-glucose and on the expression of PERIOD1 protein in established primary cell lines.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aging / blood
  • Aging / metabolism
  • Animals
  • Animals, Newborn
  • Autophagy / drug effects
  • Biomarkers / metabolism
  • Blood Glucose / drug effects
  • Blood Glucose / metabolism
  • CLOCK Proteins / genetics
  • CLOCK Proteins / metabolism
  • Cell Adhesion / drug effects
  • Cell Line
  • Circadian Clocks / drug effects*
  • Colony-Forming Units Assay
  • Diet, Protein-Restricted*
  • Energy Metabolism / drug effects
  • Feeding Behavior / drug effects*
  • Female
  • Intra-Abdominal Fat / anatomy & histology
  • Intra-Abdominal Fat / drug effects
  • Lactation / drug effects
  • Male
  • Maternal Exposure*
  • Phenotype
  • Pregnancy
  • Rats
  • Serum / metabolism
  • Tryptophan / pharmacology*
  • Tryptophan Hydroxylase / metabolism
  • Weight Gain / drug effects


  • Biomarkers
  • Blood Glucose
  • Tryptophan
  • Tryptophan Hydroxylase
  • CLOCK Proteins

Grants and funding

The authors thank for financial support the Capes-Cofecub initiative (Me 657/09). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.