*4.3. Diabetes Contributes to Diastolic Dysfunction via Abnormal Phosphorylation of Titin, Which Can Be Reversed by Treatment with Metformin, and Neuregulin-1 (NRG-1)*

Modification of titin's stiffness via phosphorylation and dephosphorylation represents a way to modify global cardiac function. There has been clinical association between development of diastolic dysfunction and diabetes. The exact mechanism for how diabetes contributes to diastolic dysfunction is not known, but there are data suggesting that it is related to abnormal protein phosphorylation. One common therapy for diabetes that has been associated with improvement in diastolic dysfunction is metformin. It was demonstrated in a mouse model of diastolic dysfunction that metformin increases activation of PKG leading to phosphorylation of the N2Bus element and reduced passive stiffness of the sarcomere resulting in improvement of diastolic dysfunction [86]. In another study, cardiomyocytes isolated from patients with diabetes who also had diastolic dysfunction demonstrated increased passive tension compared to controls. Concurrent with the mechanical changes, the cardiomyocytes from patients with diastolic dysfunction also showed changes to the phosphorylation of titin. This included increased activation of PKCα and phosphorylation in the PEVK element, as well as reduced activation of PKG and decreased phosphorylation of the N2Bus element. These changes in phosphorylation are both predicted to increase passive tension as was observed [87]. When metformin or insulin were applied to the cells in culture, there was increased activity of ERK2 leading to increased phosphorylation of the N2Bus element which reduces passive tension; however, this was partially counteracted by increased phosphorylation of the PEVK element by PKCα. Similar abnormalities in phosphorylation were seen in a diastolic dysfunction mouse model, and the abnormal phosphorylation was reversed by treating the animals with neuregulin-1 (NRG-1). NRG-1 increased phosphorylation of the N2Bus

element via ERK2 and decreased phosphorylation of the PEVK element via PKCα. The animals showed improvement in their diastolic dysfunction as measured by reduced end-diastolic pressure [87]. This study suggests that NRG-1 can reduce passive stiffness of cardiomyocytes by specifically altering phosphorylation of titin and can improve diastolic dysfunction.
