Notch signaling involved in ductus arteriosus remodeling
Inhibition of the Notch signaling pathway by DAPT prevents proliferation and migration of ductus arteriosus smooth muscle cells (DASMCs) following induction by angiotensin-II (Ang-II), a new study presented at the recently concluded 21st Asian Pacific Society of Cardiology Congress (APSC 2017) has shown.
Incubation of DASMCs in Ang-II with increasing concentrations of DAPT yielded dose-dependent reductions in cell proliferation. At DAPT concentrations of 0.5, 1 and 10 µM (p<0.01 for all), cell viability was significantly lower than in set-ups where cells were incubated with Ang-II alone. [APSC 2017, abstract MP-02]
Similarly, 48-hour incubations with 0.1 (p<0.01), 0.5 (p<0.01), 1 (p<0.001) and 10 (p<0.001) µM of DAPT resulted in significant and dose-dependent reductions in DNA synthesis in the DASMCs.
Compared with cells incubated with Ang-II alone, DASMCs that were incubated with 1 (p<0.05) and 10 (p<0.01) µM of DAPT showed significantly lower migration in the Boyden chamber assay.
In the 2016 paper, researchers “demonstrated that in DASMCs, DAPT could not only inhibit proliferation by promoting cell cycle arrest in the G0/G1-phase but also attenuate migration induced by Ang-II.” [Int J Biol Sci 2016;12:1063-1073]
“Even though the mechanisms involved in DASMC proliferation and migration are not fully elucidated, a growing body of evidence suggests that calcium influx has an important role,” they added.
Indeed, while treatment of 10 nM of Ang-II caused a spike in intracellular calcium levels, treatment with 1 (p<0.05) and 10 (p<0.01) µM of DAPT attenuated this in a dose-dependent manner. Similarly, 0.1 (p<0.05), 1 (p<0.05) and 10 (p<0.01) µM of DAPT decreased production of reactive oxygen species (ROS) compared with cells treated with Ang-II alone.
“Even though little is known regarding the role of ROS in DA remodeling, it is well established that ROS has an important role in proliferation and migration of aortic SMCs or [pulmonary arterial] SMCs,” researchers explained.
Confocal microscopy showed that Ang-II treatment promoted the translocation of the intracellular domain (ICD) of Notch 3 while subsequent treatment with DAPT disrupted this process.
Moreover, reverse transcription-polymerase chain reaction (RT-PCR) showed that DAPT treatment prevented the Ang-II induced upregulation of the Notch target genes HES1/2/5.
HES1 was significantly downregulated after treatment of 0.1 (p<0.05), 0.5 (p<0.01), 1 (p<0.01) and 10 (p<0.01) µM of DAPT, while 1 (p<0.05) and 10 (p<0.01) µM treatments significantly reduced HES2 expression. HES5 expression was significantly reduced by 1 (p<0.05), 0.5 (p<0.05), 1 (p<0.05) and 10 (p<0.01) µM DAPT treatments.
Notch inhibition leads to reduced DASMC migration and proliferation, indicating that the Notch signaling pathway is involved in the remodeling and closure of the DA in infants, researchers said.
“These effects are potentially mediated by attenuated calcium overload, reduced ROS production, and deactivations of ERK1/2, JNK and Akt signal transduction through the Notch3-HES1/2/5 pathway,” they added.