Humanin analogue, HNG, inhibits platelet activation and thrombus formation by stabilizing platelet microtubules

Abstract

HNG, a highly potent mutant of the anti‐Alzheimer peptide‐humanin, has been shown to protect against ischaemia‐reperfusion (I/R) injury. However, the underlying mechanism related to platelet activation remains unknown. We proposed that HNG has an effect on platelet function and thrombus formation. In this study, platelet aggregation, granule secretion, clot retraction, integrin activation and adhesion under flow conditions were evaluated. In mice receiving HNG or saline, cremaster arterial thrombus formation induced by laser injury, tail bleeding time and blood loss were recorded. Platelet microtubule depolymerization was evaluated using immunofluorescence staining. Results showed that HNG inhibited platelet aggregation, P‐selectin expression, ATP release, and αIIbβ3 activation and adhesion under flow conditions. Mice receiving HNG had attenuated cremaster arterial thrombus formation, although the bleeding time was not prolonged. Moreover, HNG significantly inhibited microtubule depolymerization, enhanced tubulin acetylation in platelets stimulated by fibrinogen or microtubule depolymerization reagent, nocodazole, and inhibited AKT and ERK phosphorylation downstream of HDAC6 by collagen stimulation. Therefore, our results identified a novel role of HNG in platelet function and thrombus formation potentially through stabilizing platelet microtubules via tubulin acetylation. These findings suggest a potential benefit of HNG in the management of cardiovascular diseases.

Molecular docking‐assisted screening reveals tannic acid as a natural protein disulphide isomerase inhibitor with antiplatelet and antithrombotic activities

“…Fasting venous blood was obtained from healthy donors and immediately anticoagulated with ACD buffer. Washed and gel‐filtered platelets were prepared as described previously 19,20 . Procedures utilizing human samples were approved by the University Ethical Committee of Soochow University and were in accordance with the Declaration of Helsinki.…”

“…Washed and gel-filtered platelets were prepared as described previously. 19,20 Procedures utilizing human samples were approved by the University Ethical Committee of Soochow University and were in accordance with the Declaration of Helsinki. Informed consent was obtained from all participants.…”

Mechanisms of protection of retinal pigment epithelial cells from oxidant injury by humanin and other mitochondrial-derived peptides: Implications for age-related macular degeneration

“… [ 91 ] Myocardial ischemia and reperfusion Myocardium cell line, cardiac tissue Mice HNG Cardio protection, decrease in infarct size and improvement in ventricular function [ 95 ] Cerebrovascular diseases Cardiac tissue. Neuronal cells Mice HNG Removal of abnormal proteins via chaperone-mediated autophagy [ 91 ] Cardiovascular disease Platelets Mice HNG Attenuated cremaster arterial thrombus formation [ 96 ] Aging C. elegans Worm HN Increased lifespan [ 97 ] Cancer Blood plasma, testes Mice HNG Cytoprotection of germ cells and leukocytes and non-cancerous cells in tumor-bearing mice. [ 98 ] Neuroblastoma or medulloblastoma Growth plate tissues/metatarsal bones/tumor xenografts Human, Mice HNG Prevention of bone growth impairment and chondrocyte apoptosis [ 99 ] Cancer Breast cancer cells Human, Mice HN Pro-tumoral effects [ 100 ] Cancer and fertility Blood, liver, spleen, testes Mice HNG Amelioration of chemotherapy-induced germ ...…”

Control of data variations in intravital microscopy thrombosis models

“…However, in a series of parallel studies with FeCl 3 ‐induced injury model, most of the data, if not all, are consistent between these two models 14‐16 . Given its powerful quantitative image analysis and capacity to visualize multiple thrombi across arteries of a single animal because of the minimal injuries induced by the laser, this state‐of‐the‐art model has been used and adopted in major medical institutes worldwide 17‐21 …”

“…[14][15][16] Given its powerful quantitative image analysis and capacity to visualize multiple thrombi across arteries of a single animal because of the minimal injuries induced by the laser, this state-of-the-art model has been used and adopted in major medical institutes worldwide. [17][18][19][20][21] One of the major challenges for this laser-induced injury model is the large degree of variability of the data. Platelet accumulation and fibrin formation varies not only between researchers in different institutes, but also the same researcher using the same equipment, even in the same animals and the same vessels.…”