Effect Of Low-intensity Pulsed Ultrasound Stimulation Research Articles

Low-intensity pulsed ultrasound regulates proliferation and differentiation of neural stem cells through notch signaling pathway

Low-intensity pulsed ultrasound (LIPUS) is widely used to regulate stem cell proliferation and differentiation. However, the effect of LIPUS stimulation on neural stem cells (NSCs) is not well documented. In this study, we have identified the optimal parameters, and investigated the cellular mechanisms of LIPUS to regulate the proliferation and differentiation of NSCs in vitro. NSCs were obtained and identified by nestin immunostaining. The proliferation of NSCs were measured by using Cell Counting Kit-8 (CCK-8). The expressions of nutritional factors (NTFs) were detected with immunoassay (ELISA). NSCs differentiation were detected by immunofluorescence and immunoblotting analysis. The expression level of proteins involved in the Notch signaling pathway was also measured by immunoblotting assay. Our results showed the intensity of 69.3 mW/cm2 (1 MHz, 8 V) was applicable for LIPUS stimulation. ELISA analysis demonstrated that LIPUS treatment promoted the expression of nutritional factors of NSCs in vitro. Immunofluorescence and immunoblotting analyses suggested that the LIPUS not only reduced the astrocyte differentiation, but also stimulated the differentiation to neurons. Additionally, LIPUS stimulation significantly upregulated expression level of Notch1 and Hes1. Results from our study suggest that LIPUS triggers NSCs proliferation and differentiation by modulating the Notch signaling pathway. This study implies LIPUS as a potential and promising therapeutic platform for the optimization of stem cells and enable noninvasive neuromodulation for central nervous system diseases.

Interaction of ultrasound waves with bone remodelling: a multiscale computational study.

Ultrasound stimulation is thought to influence bone remodelling process. But recently, the efficiency of ultrasound therapy for bone healing has been questioned. Despite an extensive literature describing the positive effect of ultrasound on bone regeneration-cell cultures, animal models, clinical studies-there are more and more reviews denouncing the inefficiency of clinical devices based on low-intensity pulsed ultrasound stimulation (LIPUS) of the bone healing. One of the reasons to cause controversy comes from the persistent misunderstanding of the underlying physical and biological mechanisms of ultrasound stimulation of bone repair. As ultrasonic waves are mechanical waves, the process to be studied is the one of the mechanotransduction. Previous studies on the bone mechanotransduction have demonstrated the key role of the osteocytes in bone mechano-sensing. Osteocytes are bone cells ubiquitous inside the bone matrix; they are immersed in the interstitial fluid (IF) inside the lacuno-canalicular network (LCN). They are considered as particularly sensitive to a particular type of mechanical stress: wall shear stress on osteocytes due to the IF flow in the LCN. Inspired from these findings and observations, the present work investigates the effect of LIPUS on the cortical bone from the tissue to the osteocytes, considering that the impact of the ultrasound stimulation applied at the tissue scale is related to the mechanical stress experimented by the bone cells. To do that simulations based on the finite element method are carried out in the commercial software Comsol Multiphysics to assess the wall shear stress levels induced by the LIPUS on the osteocytes. Two formulations of the wall shear stress were investigated based on two IF flow models inside the LCN and associated with two different values of the LCN permeability. The wall shear stress estimate is very different depending on the assumption considered. One of these two models provides wall shear stress values in accordance with previous works published on bone mechanotransduction. This study presents the preliminary results of a computational model that could provide keys to understanding the underlying mechanisms of the LIPUS.

Transcranial Low-Intensity Pulsed Ultrasound Modulates Structural and Functional Synaptic Plasticity in Rat Hippocampus.

Plasticity of synaptic structure and function play an essential role in neuronal development, cognitive functions, and degenerative diseases. Recently, low-intensity pulsed ultrasound (LIPUS) stimulation has been reported as a promising technology for neuromodulation. However, the effect of LIPUS stimulation on the structural and functional synaptic plasticity in rat hippocampus has not yet been addressed. The aim of this study was to investigate whether LIPUS stimulation could affect the dendritic structure, electrophysiological properties, and expression level of glutamate receptors GluN2A, GluN2B, and GluR1 subunits in rat hippocampus. Transcranial LIPUS was delivered to CA1 of the intact hippocampus of rats ( n = 40 ) for 10 days (10 min/day) with the following parameters: fundamental frequency of 0.5 MHz, pulse repetition frequency (PRF) of 500 Hz, peak negative pressure of 0.42 MPa, and Ispta of 360 mW/cm2. The effect of LIPUS on dendritic structure, electrophysiological properties, and the expression of neurotransmitter receptors was measured using Golgi staining, electrophysiological recording, and western blotting, respectively. Golgi staining and electrophysiological recordings showed that LIPUS stimulation significantly increased the density of dendritic spines (0.72 ± 0.17 versus 0.94 ± 0.19 spines/ [Formula: see text], ) and the frequency of spontaneous excitatory postsynaptic current (0.37 ± 0.14 versus 1.77 ± 0.37 Hz, ) of CA1 hippocampal neurons. Furthermore, the western blotting analysis demonstrated a significant increase in the expression level of GluN2A ( ). The results illustrated the effect of LIPUS on the dendritic structure, function, and neurotransmitter receptors, which may provide a powerful tool for treating neurodegenerative diseases.

Bone repair and ultrasound stimulation: An insight into the interaction of LIPUS with the bone callus through a multiscale computational study

In the 1950s, the effect of ultrasound stimulation on bone healing has been discovered. Nowadays, Low Intensity Pulsed Ultrasound Stimulation (LIPUS) is admitted to influence the mechanotransduction of bone. Nevertheless, despite a growing literature—cell cultures, animal models, and clinical studies—the underlying physical and biological mechanisms of LIPUS on bone healing are still misunderstood. Inspired from previous studies on the mechanotransduction induced by physiological loading, this work focuses on the effect of LIPUS on the osteocytes. These bone cells are thought to be the principal mechanosensors of bone. They are ubiquitous inside the bone matrix, immersed in the lacuno-canalicular network (LCN) filled with interstitial fluid (IF). The goal is to relate the ultrasound stimulation applied at the tissue scale, to the biological response at the cell scale. To tackle this question, two finite element models were implemented in the commercial software Comsol Multiphysics. The tissue-scale model considers an anisotropic poroelastic matrix to evaluate the IF pressure gradient induced by LIPUS into the LCN. Then, in the cell-scale model, the IF shear stress magnitude and the induced drag forces applied on osteocyte process are calculated and compared with levels of cell activation recorded in literature.

Effect of low intensity pulsed ultrasound stimulation on sinus augmentation in rabbits

ObjectivesThe objective of the present study was to evaluate the efficacy of low intensity pulsed ultrasound stimulation (LIPUS) and to determine the optimal frequency for enhancing bone regeneration in sinus augmentation using a rabbit model.Material and methodsThirty male rabbits underwent sinus augmentation. Two rectangular nasal bone windows were outlined bilaterally. LIPUS was applied at two different frequencies (1 MHz and 3 MHz) on experimental sites daily for 2, 4 and 8 weeks. Each histological area of the experimental and control sites was divided into upper and lower parts from the parietal region to a depth of 5 mm. Each area of new bone was measured.ResultsAt 2, 4 and 8 weeks, the experimental sites in the 1 MHz group exhibited significantly more new bone growth than the control sites in both the upper and lower parts. When the upper and lower parts of each area were measured in combination there was a statistical difference between the test and control sites in the 1 MHz group at 2, 4 and 8 weeks; however, there were no statistical differences between the test and control sites in the 3 MHz group.ConclusionsThe results suggest that clinical application of LIPUS for sinus augmentation may promote new bone formation, and that the effect of LIPUS for sinus augmentation at a frequency of 1 MHz was greater than at 3 MHz until 8 weeks after sinus augmentation.

Effects of low intensity pulsed ultrasound stimulation on bone regeneration in rat parietal bone defect model

Purpose: Low intensity pulsed ultrasound stimulation (LIPUS) has been clinically applied to promote bone fracture healing in the orthopedic field. Thus, it is likely that LIPUS also stimulates bone regeneration in bone defects in the cranial-maxillofacial area. However, this has not been clearly proved. Furthermore, optimal time point and period of the application after the surgery has not been reported. The purpose of the present study was to evaluate the effect of LIPUS on bone regeneration in the rat parietal bone defects especially focusing on time and period of the application. Materials and Methods: Eighteen Wistar rats (14 weeks old) were divided into 6 groups: 5 experimental groups and a control group. Bone defect of 5 mm diameter was prepared on each side of the parietal bone and customized gelatin membranes were placed over the bone defects. LIPUS (160 mW/cm2, 15 min/day) was applied to the defect area with an active transducer externally in the experimental groups according to the schedules of the applications: Group 1 (day 6 - 12), group 2 (day 13 - 19), group 3 (day 20 - 26), group 4 (day 6 - 19) and group 5 (day 6 - 26). All the animals were sacrificed at 28 days. The defects were analyzed with micro CT and then histologically. Results: In Group 1, new bone formation was significantly promoted and the newly-formed bone was thick and matured compared to the one of the control group. In other experimental groups there were tendencies of stimulation of new bone formation; however, they were not statistically significant. Discussion and Conclusion: The present study demonstrated that amount of new bone formation in the bone defect depended on the time and period of LIPUS application. It has been suggested that application of LIPUS at an early healing period, the second week after the surgery, effectively accelerated new bone formation.

Involvement of Calcitonin Gene-Related Peptide Innervation in the Promoting Effect of Low-Intensity Pulsed Ultrasound on Spinal Fusion Without Decortication

A prospective left-right comparison designed experiment using a rabbit posterolateral intertransverse process fusion model. To investigate the involvement of calcitonin gene-related peptide (CGRP) innervation in the promoting effect of low intensity pulsed ultrasound stimulation (LIPUS) on spinal fusion without decortication. Sensory neuropeptide CGRP is involved in bone repair and ectopic ossification. Comparison of CGRP innervations in ectopic bone between sham LIPUS and LIPUS sides can help us to understand the relationship between sensory nerve innervation and LIPUS. A total of 27 New Zealand white rabbits underwent bilateral posterolateral intertransverse process fusion with implantation of porous poly-D,L-lactic acid blocks loaded with 1.25 μg recombinant human bone morphogenetic protein-4 solution. One side was provided LIPUS daily whereas the other side served as control. Animals were killed and the operated lumbar vertebrae were harvested for histomorphologic evaluation at 3 days (n = 3), 1 week (n = 6), 3 weeks (n = 6), 7 weeks (n = 6), and 12 weeks (n = 6) following surgery, respectively. LIPUS accelerated the invasion of CGRP-positive nerve fibers during ectopic ossification spatially and temporally. Spatially, CGRP-positive nerve fibers were also observed in the new formed cartilage and bone tissues on LIPUS side, whereas they were only detected in the fibrous tissue and bone marrow on sham LIPUS side. Temporally, the density of CGRP-positive nerve fibers was significantly higher on the LIPUS side when compared with the sham LIPUS side. LIPUS promoted the invasion of CGRP sensory nerve in ectopic bone, which may in turn contribute to the promoting effect of LIPUS on ectopic ossification.