RhoA rescues cardiac senescence by regulating Parkin-mediated mitophagy.

Heart failure is one of the leading causes of death worldwide. RhoA, a small GTPase, governs actin dynamics in various tissue and cell types, including cardiomyocytes; however, its involvement in cardiac function has not been fully elucidated. Here, we generated cardiomyocyte-specific RhoA conditional knockout (cKO) mice, which demonstrated a significantly shorter lifespan with left ventricular dilation and severely impaired ejection fraction. We found that the cardiac tissues of the cKO mice exhibited structural disorganization with fibrosis and also exhibited enhanced senescence compared with control mice. In addition, we show that cardiomyocyte mitochondria were structurally abnormal in the aged cKO hearts. Clearance of damaged mitochondria by mitophagy was remarkably inhibited in both cKO cardiomyocytes and RhoA-knockdown HL-1 cultured cardiomyocytes. In RhoA-depleted cardiomyocytes, we reveal that the expression of Parkin, an E3 ubiquitin ligase that plays a crucial role in mitophagy, was reduced, and expression of N-Myc, a negative regulator of Parkin, was increased. We further reveal that the RhoA-Rho kinase axis induced N-Myc phosphorylation, which led to N-Myc degradation and Parkin upregulation. Re-expression of Parkin in RhoA-depleted cardiomyocytes restored mitophagy, reduced mitochondrial damage, attenuated cardiomyocyte senescence, and rescued cardiac function both in vitro and in vivo. Finally, we found that patients with idiopathic dilated cardiomyopathy without causal mutations for dilated cardiomyopathy showed reduced cardiac expression of RhoA and Parkin. These results suggest that RhoA promotes Parkin-mediated mitophagy as an indispensable mechanism contributing to cardioprotection in the aging heart.

Vascular smooth muscle RhoA counteracts abdominal aortic aneurysm formation by modulating MAP4K4 activity.

Whether a small GTPase RhoA plays a role in the pathology of abdominal aortic aneurysm (AAA) has not been determined. We show here that RhoA expression is reduced in human AAA lesions, compared with normal areas. Furthermore, incidence of AAA formation is increased in vascular smooth muscle cell (VSMC)-specific RhoA conditional knockout (cKO) mice. The contractility of the aortic rings and VSMCs from RhoA cKO mice is reduced, and expression of genes related to the VSMC contractility is attenuated by loss of RhoA. RhoA depletion activates the mitogen-activated protein (MAP) kinase signaling, including MAP4K4, in the aorta and VSMCs. Inhibition of MAP4K4 activity by DMX-5804 decreases AAA formation. Set, a binding protein to active RhoA, functions as an activator of MAP4K4 by sequestering PP2A, an inhibitor of MAP4K4, in the absence of RhoA. In conclusion, RhoA counteracts AAA formation through inhibition of MAP4K4 in cooperation with Set.

Cardio- and reno-protective effects of dipeptidyl peptidase III in diabetic mice.

Diabetes mellitus (DM) causes injury to tissues and organs, including to the heart and kidney, resulting in increased morbidity and mortality. Thus, novel potential therapeutics are continuously required to minimize DM-related organ damage. We have previously shown that dipeptidyl peptidase III (DPPIII) has beneficial roles in a hypertensive mouse model, but it is unknown whether DPPIII has any effects on DM. In this study, we found that intravenous administration of recombinant DPPIII in diabetic db/db mice for 8 weeks suppressed the DM-induced cardiac diastolic dysfunctions and renal injury without alteration of the blood glucose level. This treatment inhibited inflammatory cell infiltration and fibrosis in the heart and blocked the increase in albuminuria by attenuating the disruption of the glomerular microvasculature and inhibiting the effacement of podocyte foot processes in the kidney. The beneficial role of DPPIII was, at least in part, mediated by the cleavage of a cytotoxic peptide, named Peptide 2, which was increased in db/db mice compared with normal mice. This peptide consisted of nine amino acids, was a digested fragment of complement component 3 (C3), and had an anaphylatoxin-like effect determined by the Miles assay and chemoattractant analysis. The effect was dependent on its interaction with the C3a receptor and protein kinase C-mediated RhoA activation downstream of the receptor in endothelial cells. In conclusion, DPPIII plays a protective role in the heart and kidney in a DM animal model through cleavage of a peptide that is a part of C3.

Stomatin-Mediated Inhibition of the Akt Signaling Axis Suppresses Tumor Growth.

The growth and progression of cancers are crucially regulated by the tumor microenvironment where tumor cells and stromal cells are mutually associated. In this study, we found that stomatin expression was markedly upregulated by the interaction between prostate cancer cells and stromal cells. Stomatin suppressed cancer cell proliferation and enhanced apoptosis in vitro and inhibited xenograft tumor growth in vivo. Stomatin inhibited Akt activation, which is mediated by phosphoinositide-dependent protein kinase 1 (PDPK1). PDPK1 protein stability was maintained by its binding to HSP90. Stomatin interacted with PDPK1 and interfered with the PDPK1-HSP90 complex formation, resulting in decreased PDPK1 expression. Knockdown of stomatin in cancer cells elevated Akt activation and promoted cell increase by promoting the interaction between PDPK1 and HSP90. Clinically, stomatin expression levels were significantly decreased in human prostate cancer samples with high Gleason scores, and lower expression of stomatin was associated with higher recurrence of prostate cancer after the operation. Collectively, these findings demonstrate the tumor-suppressive effect of stromal-induced stomatin on cancer cells. SIGNIFICANCE: These findings reveal that interactions with stromal cells induce expression of stomatin in prostate cancer cells, which suppresses tumor growth via attenuation of the Akt signaling axis.

The Age-dependent Elevation of miR-335-3p Leads to Reduced Cholesterol and Impaired Memory in Brain.

MiR-335-3p, a neuron-enriched microRNA, has been reported to be involved in aging and age-related neurological diseases. However, the role of miR-335-3p in cholesterol metabolism of astrocytes, and whether it affects neuronal functions, particularly during aging process, largely remains unknown. In this study, we uncover that miR-335-3p is significantly increased in aged cultured astrocytes and aged hippocampal brains, accompanied by decreased cellular cholesterol and diminished expression of HMGCR (3-hydroxy-3-methylglutaryl-CoA reductase) and 3-hydroxy-3-methylglutaryl-CoA synthase-1 (HMGCS1), both step-limiting enzymes in cholesterol synthesis pathway. We also demonstrate that miR-335-3p suppresses HMGCS1 post-transcriptionally by directly binding to its 3'UTR, and HMGCR through binding mediated by SFRS2. More importantly, aged mice with miR-335-3p deficiency in hippocampal brains exhibit improved learning and memory, accompanied by enhanced levels of postsynaptic density protein 95 (PSD95). We further reveal that the level change of PSD95 is resulted from altered cholesterol metabolism. Our findings provide a novel insight into the regulatory role of miR-335-3p in cholesterol metabolism in astrocytes, and consequently cognitive functions during aging.