Related Experiment Videos
Correlation of carcinogen-induced unscheduled DNA synthesis and NAD reduction in fresh human lymphocytes
Abstract:
Incorporation of [3H]thymidine into the DNA of fresh human lymphocytes, treated with various chemical mutagens, was measured and correlated with cellular NAD levels before and after treatment. NAD levels in lymphocytes were significantly reduced following treatment with mutagenic chemicals. Reduction of cellular NAD pools was directly correlated with [3H]thymidine incorporation. As NAD levels decreased, [3H]thymidine incorporation increased. Theophylline, a known inhibitor of poly(ADP-ribose)polymerase, inhibited both the NAD reduction in cells treated with DNA damaging agents and the incorporation of [3H]thymidine into DNA. The inhibitory effect of theophylline on NAD depletion and on [3H]thymidine incorporation was dose and cell number dependent. Near normal responses to carcinogen exposure could be restored to theophylline-treated cells following the removal of theophylline. These data suggest that conversion of NAD to poly(ADP-ribose) may be necessary, or at least closely associated with, DNA repair in human lymphocytes.
Insights
Cellular nicotinamide adenine dinucleotide (NAD) levels decrease when human lymphocytes are exposed to mutagens. This reduction correlates with increased DNA repair, suggesting NAD
Area of Science:
- Molecular Biology
- Biochemistry
- Toxicology
Background:
- DNA damage triggers cellular responses aimed at repair.
- Nicotinamide adenine dinucleotide (NAD) is a crucial cellular metabolite.
- Poly(ADP-ribose)polymerase (PARP) utilizes NAD+ in DNA repair pathways.
Purpose of the Study:
- To investigate the relationship between cellular NAD levels and DNA repair in human lymphocytes.
- To determine the role of NAD conversion in DNA repair mechanisms following mutagen exposure.
Main Methods:
- Human lymphocytes were treated with chemical mutagens.
- Cellular NAD levels were measured before and after mutagen treatment.
- [3H]thymidine incorporation into DNA was quantified as a measure of DNA repair.
- The effect of theophylline, a PARP inhibitor, on NAD levels and DNA repair was assessed.
Main Results:
- Mutagenic chemical treatment significantly reduced cellular NAD levels in lymphocytes.
- A direct inverse correlation was observed between NAD levels and [3H]thymidine incorporation.
- Theophylline inhibited NAD reduction and [3H]thymidine incorporation in a dose-dependent manner.
- Reversal of theophylline's effects was observed upon its removal, restoring normal responses.
Conclusions:
- NAD depletion is closely associated with DNA repair processes in human lymphocytes.
- The conversion of NAD to poly(ADP-ribose) appears to be integral to DNA repair.
- PARP activity, dependent on NAD, plays a significant role in the cellular response to DNA damage.