Effects Of Low-intensity Pulsed Ultrasound Research Articles

Low-intensity pulsed ultrasound regulates bone marrow mesenchymal stromal cells differentiation and inhibits bone loss by activating the IL-11-Wnt/β-catenin signaling pathway

BackgroundOsteoporosis (OP) is a common metabolic bone disease. Low-intensity pulsed ultrasound (LIPUS) can effectively promote bone formation and fracture healing. The Wnt/β-catenin signaling pathway is crucial for regulating bone homeostasis and bone diseases, and its downregulation is one of the main mechanisms of osteoporosis pathogenesis. Interleukin-11 (IL-11), which is regulated by mechanical stress, is a key factor in bone remodeling. Here, we investigated the optimal intervention parameters for LIPUS, the relationships among LIPUS, IL-11, and the Wnt/β-catenin signaling pathway, and the effects of LIPUS on bone loss and potential molecular mechanisms in ovariectomized (OVX) mice. MethodsBone marrow mesenchymal stromal cells (BMSCs) were subjected to LIPUS intervention for 0, 10, or 20 min to determine the optimal intervention time. The mediating role of IL-11 in LIPUS intervention was explored through IL-11 knockdown and overexpression. Finally, animal experiments were conducted to investigate the in vivo therapeutic effects of LIPUS. ResultsThe optimal intervention time for LIPUS was 20 min. LIPUS promoted IL-11 expression and upregulated the Wnt/β-catenin signaling pathway, thereby promoting osteogenic differentiation and inhibiting adipogenic differentiation of BMSCs. IL-11 mediates the regulation of the Wnt/β-catenin signaling pathway by LIPUS. Additionally, LIPUS effectively improved the bone microstructure in ovariectomized mice, inhibited bone loss, promoted IL-11 expression in bone tissue, and activated the Wnt/β-catenin signaling pathway in the femur. ConclusionLow-intensity pulsed ultrasound can regulate BMSCs differentiation and inhibit bone loss by promoting IL-11 expression and activating the Wnt/β-catenin signaling pathway.

Low-intensity pulsed ultrasound improves metabolic dysregulation in obese mice by suppressing inflammation and extracellular matrix remodeling

Chronic inflammation in white adipose tissue is crucial in obesity and related metabolic disorders. Low-intensity pulsed ultrasound (LIPUS) is renowned for its anti-inflammatory effects as a non-invasive treatment, yet its precise role in obesity has been uncertain. Our study investigates the therapeutic effect of LIPUS and its underlying mechanism on obesity in mice, thereby offering a novel approach for non-invasive treatment of obesity and associated metabolic disorders for human. Male C57BL/6J mice aged 10 weeks were fed a high-fat diet (HFD) for 8 weeks to establish obesity model, then underwent 8 weeks of LIPUS (frequency: 1.0 MHz, duty cycle: 20 %, Isata: 58–61 mW/cm2, 20 min per day) stimulation of the epididymal white adipose tissue. Fat and lean mass were measured using nuclear magnetic resonance (NMR), while energy homeostasis was evaluated using metabolic cages. Insulin resistance was assessed using glucose tolerance tests (GTT) and insulin tolerance tests (ITT). Regulatory mechanisms were explored using RNA sequencing. Results showed that LIPUS significantly reduced obesity markers in obese mice, including body and adipose tissue weight, and improved insulin resistance, without affecting food intake. RNA sequencing showed 250 up-regulated and 351 down-regulated genes between HFD-LIPUS group and HFD-Sham group, suggesting anti-inflammatory action. Quantitative PCR confirmed reduced pro-inflammatory gene expression and macrophage infiltration in eWAT. Gene set enrichment analysis showed decreased NF-κB signaling and extracellular matrix-receptor interactions in LIPUS-treated mice. Thus, LIPUS effectively mitigates metabolic dysregulation in HFD-induced obesity through inflammation suppression and extracellular matrix remodeling, which provides a potential physical therapy for metabolic syndrome in clinic.

3D-Cultured MC3T3-E1-Derived Exosomes Promote Endothelial Cell Biological Function under the Effect of LIPUS.

Porous Ti-6Al-4V scaffold materials can be used to heal massive bone defects because they can provide space for vascularisation and bone formation. During new bone tissue development, rapid vascular ingrowth into scaffold materials is very important. Osteoblast-derived exosomes are capable of facilitating angiogenesis-osteogenesis coupling. Low-intensity pulsed ultrasound (LIPUS) is a physical therapy modality widely utilised in the field of bone regeneration and has been proven to enhance the production and functionality of exosomes on two-dimensional surfaces. The impact of LIPUS on exosomes derived from osteoblasts cultured in three dimensions remains to be elucidated. In this study, exosomes produced by osteoblasts on porous Ti-6Al-4V scaffold materials under LIPUS and non-ultrasound stimulated conditions were co-cultured with endothelial cells. The findings indicated that the exosomes were consistently and stably taken up by the endothelial cells. Compared to the non-ultrasound group, the LIPUS group facilitated endothelial cell proliferation and angiogenesis. After 24 h of co-culture, the migration ability of endothelial cells in the LIPUS group was 17.30% higher relative to the non-ultrasound group. LIPUS may represent a potentially viable strategy to promote the efficacy of osteoblast-derived exosomes to enhance the angiogenesis of porous Ti-6Al-4V scaffold materials.

Therapeutic effects of LIPUS on the elasticity of injured tendons: animal experiment

Abstract This paper aims to explore the efficacy and duration of low-intensity pulsed ultrasound (LIPUS) treatment in improving the elastic properties of injured tendons through animal experiments. 48 healthy 3-month-old New Zealand white rabbits (weight 2.5-3.0 kg) were randomly divided into the Control group (n = 9) and model group (n = 39). The rabbits in the model group were injected with 0.2ml PGE2 mixed solution (concentration: 500 ng/0.2ml) into the Achilles tendon 2cm above the calcaneus, once a week, for 4 weeks, to build the Achilles tendon injury model. The Control group was injected with the same volume of PBS solution. Three rabbits were randomly selected from the Control and model groups for model validation, the model group was randomly divided into the Model (n = 6) and LIPUS (n = 30) groups. Then the LIPUS was randomly divided into the LIPUS1 (n = 6), LIPUS4 (n = 6), LIPUS7 (n = 6), LIPUS14 (n = 6), and LIPUS28 (n = 6) subgroups. Following interventions on days 1, 4, 7, 14, and 28, tendon elasticity was assessed by shear wave elastography (SWE). The elastic modulus of the Model, LIPUS1, and LIPUS4 groups exhibited a significant decrease compared to the Control group during the early stages of treatment. With prolonged treatment duration, the elastic modulus of LIPUS groups gradually increased. By day 7, no statistically significant difference in elastic modulus existed between the LIPUS7 and Control groups, indicating that LIPUS treatment had successfully restored the Achilles tendon’s elasticity to a healthy state. In summary, LIPUS therapy demonstrates the potential to effectively ameliorate the elastic properties of injured Achilles tendons.

Study on the Effects of Low-Intensity Pulsed Ultrasound and Iron Ions for Proliferation and Differentiation of Osteoblasts

ObjectiveThis study involved the proliferation and differentiation of osteoblasts treated with low-intensity pulsed ultrasound (LIPUS) and iron (Fe3+) ions, respectively. The biological effects of LIPUS and Fe3+ ions on the proliferation and differentiation of osteoblasts were also evaluated. MethodsMC3T3-E1 cells were seeded in six-well plates with the medium, which contained different concentrations of Fe3+ (0, 100, 200, 300, 400, 500, 600 and 700 μg L-1, respectively). LIPUS treatment was directed at the bottom of the plate for 20 min at an intensity of 80 mW cm-2 every day. ResultsViability results showed that a dose of 400 μg L-1 Fe3+ ions had the best effect at promoting osteogenic proliferation in cell culture. The results of alkaline phosphatase staining and mineralization indicated that the differentiation of osteoblasts was promoted by LIPUS and Fe3+ ions. Fluorescence staining results showed that the number of cell nuclei in the LIPUS, Fe3+ and LIPUS-Fe groups increased by 37.20%, 55.81% and 89.76%, respectively. Migration data indicated that migration and proliferation rates were increased by LIPUS and Fe3+, and the results of protein expression indicated that LIPUS and Fe3+ may increase the expression of Wnt, β-catenin, and Runx2, hence promoting normal bone regeneration and development. ConclusionThe combination of LIPUS (1.5 MHz, 80 mW cm-2) and Fe3+ accelerates the proliferation and differentiation of osteoblasts significantly compared with single-factor treatment (stimulated by LIPUS and Fe3+ ions, respectively). This study could establish a foundation for LIPUS-responsive biomaterials in the repair and regeneration of bone tissues.

Effect of low-intensity pulsed ultrasound on postoperative rehabilitation of rotator cuff tears: Protocol for a systematic review and meta-analysis.

Rotator cuff tears are a common shoulder injury that significantly impacts patients' daily lives and work abilities. Although surgical treatment methods for rotator cuff tears have been continuously improved with advances in medical technology, postoperative rehabilitation remains challenging. Therefore, finding effective rehabilitation treatments is crucial for improving patient prognosis and enhancing quality of life. This study will aim to systematically evaluate the impact of low-intensity pulsed ultrasound (LIPUS) on postoperative rehabilitation of rotator cuff tears, comprehensively assessing the efficacy and safety of LIPUS in postoperative recovery. This protocol will search multiple databases including PubMed/MEDLINE, Embase, Cochrane Library, CNKI, Scopus, and Web of Science to identify randomized controlled trials related to LIPUS for postoperative rehabilitation of rotator cuff tears. The search will encompass literature published from the inception of the databases up to April 2024. Methodological quality assessment and data extraction will be conducted using the Cochrane Handbook for Systematic Reviews of Interventions and PRISMA guidelines. Meta-analysis will be performed on appropriate studies using either random-effects or fixed-effects models, and subgroup analyses will be conducted to explore potential heterogeneity. Studies meeting the inclusion criteria will be included in the analysis. All analyses will be performed using Stata version 16.0. The incidence of rotator cuff tear rates will be assessed by imaging techniques such as MRI or ultrasound. Pain intensity will be scored using standardized pain assessment scales, such as the Visual Analog Scale (VAS). Improved range of motion (ROM) in shoulder flexion, abduction, and rotation. Functional outcomes will be evaluated using effective measures such as Constant-Murley scores (CMS) and shoulder joint scores by American Shoulder and Elbow Surgeons (ASES). Adverse events associated with LIPUS therapy, including skin irritation, increased pain, or any other complications. Subgroup analysis will also be carried out if possible. Following the meta-analysis, we will assess the overall effect of LIPUS on postoperative rehabilitation of rotator cuff tears, and further explore its impact on aspects such as pain relief, functional improvement, and postoperative complications. It is anticipated that this study will provide comprehensive evidence regarding the role of LIPUS in postoperative rehabilitation of rotator cuff tears, guiding clinical practice and future research. The resultant manuscript will be submitted for publication in a peer-reviewed journal. CRD42024530798.

Effects of Low-intensity Ultrasound on Root Resorptions Induced by a System of Associated Forces

Introduction The bio-stimulatory effects of low-intensity ultrasound on dental tissues which were clinically introduced in 2004 demonstrated a reduction in root resorption in maxillary premolars subjected to a force level of 50 g. Similar results by applying a torque of 50 g to the maxillary premolars were obtained in 2016; however, the effects of low-intensity pulsed ultrasound (LIPUS) on the presence of high-intensity forces are not yet known. Aim The purpose of this study was to examine the effects of LIPUS on a sample of 40 patients. Methods The analysis was conducted by applying two forces to teeth no. 14 and 24, each with an intensity greater than 50 g, and applied at two different points on the coronal surface. Subsequently, LIPUS was applied daily for 20 minutes to tooth no. 14, while tooth no. 24 served as a control. The study lasted for 28 days. Results Overall, the group treated with LIPUS demonstrated an improvement in the average total volume and percentage of root resorption compared to the control group. The same results emerged for the diameter, depth, and surface of the gaps of resorption. Conclusion This study showed that it was possible to reduce root resorption in the presence of forces with intensities greater than 50 g. Nevertheless, a longer period of study will make it possible to obtain more rational results as to the effectiveness of low-intensity ultrasound on root resorption induced by orthodontic treatment.

Low-intensity pulsed ultrasound improves myocardial ischaemia‒reperfusion injury via migrasome-mediated mitocytosis.

During myocardial ischaemia‒reperfusion injury (MIRI), the accumulation of damaged mitochondria could pose serious threats to the heart. The migrasomes, newly discovered mitocytosis-mediating organelles, selectively remove damaged mitochondria to provide mitochondrial quality control. Here, we utilised low-intensity pulsed ultrasound (LIPUS) on MIRI mice model and demonstrated that LIPUS reduced the infarcted area and improved cardiac dysfunction. Additionally, we found that LIPUS alleviated MIRI-induced mitochondrial dysfunction. We provided new evidence that LIPUS mechanical stimulation facilitated damaged mitochondrial excretion via migrasome-dependent mitocytosis. Inhibition the formation of migrasomes abolished the protective effect of LIPUS on MIRI. Mechanistically, LIPUS induced the formation of migrasomes by evoking the RhoA/Myosin II/F-actin pathway. Meanwhile, F-actin activated YAP nuclear translocation to transcriptionally activate the mitochondrial motor protein KIF5B and Drp1, which are indispensable for LIPUS-induced mitocytosis. These results revealed that LIPUS activates mitocytosis, a migrasome-dependent mitochondrial quality control mechanism, to protect against MIRI, underlining LIPUS as a safe and potentially non-invasive treatment for MIRI.

Effects of insonification on repairing the renal injury of diabetic nephropathy rats

IntroductionProlonged hyperglycemia in diabetes mellitus can result in the development of diabetic nephropathy (DN) and increase the susceptibility to kidney failure. Low-intensity pulsed ultrasound (LIPUS) is a non-invasive modality that...

Low-intensity pulsed ultrasound protects from inflammatory dilated cardiomyopathy through inciting extracellular vesicles.

CD4+ T cells are activated during inflammatory dilated cardiomyopathy (iDCM) development to induce immunogenic responses that damage the myocardium. Low-intensity pulsed ultrasound (LIPUS), a novel physiotherapy for cardiovascular diseases, has recently been shown to modulate inflammatory responses. However, its efficacy in iDCM remains unknown. Here, we investigated whether LIPUS could improve the severity of iDCM by orchestrating immune responses and explored its therapeutic mechanisms. In iDCM mice, LIPUS treatment reduced cardiac remodelling and dysfunction. Additionally, CD4+ T-cell inflammatory responses were suppressed. LIPUS increased Treg cells while decreasing Th17 cells. LIPUS mechanically stimulates endothelial cells, resulting in increased secretion of extracellular vesicles (EVs), which are taken up by CD4+ T cells and alter their differentiation and metabolic patterns. Moreover, EVs selectively loaded with microRNA (miR)-99a are responsible for the therapeutic effects of LIPUS. The hnRNPA2B1 translocation from the nucleus to the cytoplasm and binding to caveolin-1 and miR-99a confirmed the upstream mechanism of miR-99a transport. This complex is loaded into EVs and taken up by CD4+ T cells, which further suppress mTOR and TRIB2 expression to modulate cellular differentiation. Our findings revealed that LIPUS uses an EVs-dependent molecular mechanism to protect against iDCM progression. Therefore, LIPUS is a promising new treatment option for iDCM.

Effect of Low-Intensity Pulsed Ultrasound on Macrophage Properties and Fibrosis in the Infrapatellar Fat Pad in a Carrageenan-Induced Knee Osteoarthritis Rat Model.

In the progression of knee osteoarthritis (KOA), fibrosis of the infrapatellar fat pad (IFP) is a key pathological change. Low-intensity pulsed ultrasound (LIPUS) inhibits IFP fibrosis by decreasing the gene expression and activity of hypoxia-inducible factor (HIF-1α), which is a protein involved in IFP fibrosis in KOA rat models. On the other hand, macrophages play an important role in the progression of fibrosis in various tissues, and LIPUS irradiation suppresses macrophage infiltration and inflammatory cytokine secretion. However, whether LIPUS suppresses macrophage polarity and IFP fibrosis in KOA remains unclear. Therefore, we investigated the effect of LIPUS on macrophage polarity and IFP fibrosis. A KOA model was created by injecting carrageenin into the bilateral knee joints of Wistar rats (eight weeks old). Tissues were harvested over time for histological and molecular biological analysis. The KOA model was also subjected to LIPUS irradiation for two weeks following the injection of carrageenin. RM-4-positive cells were widely distributed in IFP two weeks after carrageenin administration, but M2 macrophages were significantly increased, and the Sirius red area was decreased in the LIPUS-irradiated group compared with those in the non-irradiated group. The gene expression of M1 macrophage markers was significantly decreased and that of M2 macrophage markers was significantly increased in the LIPUS-irradiated group. The expression of transforming growth factor-β (TGF-β) and type 1 collagen was also significantly decreased. These results suggest that LIPUS may serve as a novel approach for the treatment of KOA through its effect on M1 macrophages and suppression of TGF-β expression.

Low-Intensity Pulsed Ultrasound: A Physical Stimulus with Immunomodulatory and Anti-inflammatory Potential.

Ultrasound has expanded into the therapeutic field as a medical imaging and diagnostic technique. Low-intensity pulsed ultrasound (LIPUS) is a kind of therapeutic ultrasound that plays a vital role in promoting fracture healing, wound repair, immunomodulation, and reducing inflammation. Its anti-inflammatory effects are manifested by decreased pro-inflammatory cytokines and chemokines, accelerated regression of immune cell invasion, and accelerated damage repair. Although the anti-inflammatory mechanism of LIPUS is not very clear, many in vitro and in vivo studies have shown that LIPUS may play its anti-inflammatory role by activating signaling pathways such as integrin/Focal adhesion kinase (FAK)/Phosphatidylinositol 3-kinase (PI3K)/Serine threonine kinase (Akt), Vascular endothelial growth factor (VEGF)/endothelial nitric oxide synthase (eNOS), or inhibiting signaling pathways such as Toll-like receptors (TLRs)/Nuclear factor kappa-B (NF-κB) and p38-Mitogen-activated protein kinase (MAPK). As a non-invasive physical therapy, the anti-inflammatory and immunomodulatory effects of LIPUS deserve further exploration.

Effects of Low-Intensity Pulsed Ultrasound on the Regulation of Free Fatty Acid Release in 3T3-L1 Cells.

To investigate the effects of low-intensity pulsed ultrasound (LIPUS) on the proliferation, differentiation, and tumor necrosis factor-α (TNF-α)-induced lipolysis of 3T3-L1 cells, and to explore the feasibility of regulating the release of free fatty acids (FFA) to prevent lipotoxicity. Different intensities (30, 60, 90, and 120 mW/cm2) of LIPUS were applied to 3T3-L1 preadipocytes for different durations (5, 10, 15, 20, 25, and 30 minutes). Appropriate parameters for subsequent experiments were selected by assessing cell viability. The effect of LIPUS on the proliferation and differentiation of 3T3-L1 cells was evaluated by microscope observation, flow cytometry, and lipid content determination. After treated with LIPUS and TNF-α (50 ng/mL), the degree of lipolysis was assessed by measuring the extracellular FFA content. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the mRNA expression of relevant genes. Different parameters of LIPUS significantly enhance the viability of 3T3-L1 cells (P < .05), with 20 minutes and 30 mW/cm2 as the most suitable settings. After LIPUS treatment, 3T3-L1 cell proliferation accelerated, apoptosis rate and G1 phase cell proportion decreased, the content of lipid droplets and TG was increased in differentiated cells, while FFA release decreased (P < .05). The expression of PCNA, PPARγ, C/EBPα, Perilipin A mRNA increased, and the expression of TNF-α, ATGL, HSL mRNA decreased (P < .05). LIPUS could promote the proliferation and differentiation of 3T3-L1 cells and inhibit TNF-α-induced lipolysis, indicating its potential as a therapy for mitigating lipotoxicity caused by decompensated adipocytes.

Low-intensity pulsed ultrasound promotes the osteogenesis of mechanical force-treated periodontal ligament cells via Piezo1.

Low-intensity pulsed ultrasound (LIPUS) can accelerate tooth movement and preserve tooth and bone integrity during orthodontic treatment. However, the mechanisms by which LIPUS affects tissue remodeling during orthodontic tooth movement (OTM) remain unclear. Periodontal ligament cells (PDLCs) are pivotal in maintaining periodontal tissue equilibrium when subjected to mechanical stimuli. One notable mechano-sensitive ion channel, Piezo1, can modulate cellular function in response to mechanical cues. This study aimed to elucidate the involvement of Piezo1 in the osteogenic response of force-treated PDLCs when stimulated by LIPUS. After establishing rat OTM models, LIPUS was used to stimulate rats locally. OTM distance and alveolar bone density were assessed using micro-computed tomography, and histological analyses included hematoxylin and eosin staining, tartrate-resistant acid phosphatase staining and immunohistochemical staining. GsMTx4 and Yoda1 were respectively utilized for Piezo1 functional inhibition and activation experiments in rats. We isolated human PDLCs (hPDLCs) in vitro and evaluated the effects of LIPUS on the osteogenic differentiation of force-treated hPDLCs using real-time quantitative PCR, Western blot, alkaline phosphatase and alizarin red staining. Small interfering RNA and Yoda1 were employed to validate the role of Piezo1 in this process. LIPUS promoted osteoclast differentiation and accelerated OTM in rats. Furthermore, LIPUS alleviated alveolar bone resorption under pressure and enhanced osteogenesis of force-treated PDLCs both in vivo and in vitro by downregulating Piezo1 expression. Subsequent administration of GsMTx4 in rats and siPIEZO1 transfection in hPDLCs attenuated the inhibitory effect on osteogenic differentiation under pressure, whereas LIPUS efficacy was partially mitigated. Yoda1 treatment inhibited osteogenic differentiation of hPDLCs, resulting in reduced expression of Collagen Ⅰα1 and osteocalcin in the periodontal ligament. However, LIPUS administration was able to counteract these effects. This research unveils that LIPUS promotes the osteogenesis of force-treated PDLCs via downregulating Piezo1.

The Effect of Low-Intensity Pulsed Ultrasound on Temporomandibular Joint Arthritis in Juvenile Rats.

Juvenile idiopathic arthritis is an inflammatory disease that can affect the temporomandibular joint (TMJ) and lower jaw growth. Better treatment options are needed, so this study investigated the effect of low-intensity pulsed ultrasound (LIPUS) on TMJ arthritis. Seventy-two 3-week-old male Wistar rats were invivo microcomputed tomography (micro-CT) scanned and divided into eight groups (n = 9). These groups were Group 1-TMJ arthritis and immediate LIPUS treatment (20 min/day, 4 weeks); Group 2-immediate LIPUS treatment and no TMJ arthritis; Group 3-TMJ arthritis and no LIPUS; Group 4-no TMJ arthritis and no LIPUS; Group 5-TMJ arthritis and LIPUS treatment with a delayed start by 4 weeks; Group 6-Delayed LIPUS and no TMJ arthritis; Group 7-TMJ arthritis and no (delayed) LIPUS; and Group 8-no TMJ arthritis and no (delayed) LIPUS. Ex vivo micro-CT scanning was completed, and samples were prepared for tissue analysis. Synovitis was observed in the TMJ arthritis (collagen-induced arthritis [CIA]) groups, but the severity appeared greater in the groups without LIPUS treatment. Fibrocartilage and hypertrophic cell layer thicknesses in the CIA group without LIPUS treatment were significantly greater (p < 0.05). Proteoglycan staining appeared greater in the LIPUS groups. Immediate LIPUS treatment increased the expression of type II collagen, type X collagen, and transforming growth factor-beta 1 (TGF-β1) immunostaining, and CIA (no LIPUS) increased MMP-13, vascular endothelial growth factor, and interleukin-1 beta (IL-1β) immunostaining. LIPUS treatment prevented growth disturbances observed in the CIA groups (no LIPUS) (p < 0.005). Our results have contributed to the understanding of the uses and limitations of the CIA juvenile rat model and have demonstrated the effects of LIPUS on the TMJ and mandibular growth. This information will help in designing future studies for investigating LIPUS and TMJ arthritis, leading to the development of new treatment options for children with juvenile arthritis in their TMJs.

Low-intensity pulsed ultrasound reduces alveolar bone resorption during orthodontic treatment via Lamin A/C-Yes-associated protein axis in stem cells.

The bone remodeling during orthodontic treatment for malocclusion often requires a long duration of around two to three years, which also may lead to some complications such as alveolar bone resorption or tooth root resorption. Low-intensity pulsed ultrasound (LIPUS), a noninvasive physical therapy, has been shown to promote bone fracture healing. It is also reported that LIPUS could reduce the duration of orthodontic treatment; however, how LIPUS regulates the bone metabolism during the orthodontic treatment process is still unclear. To investigate the effects of LIPUS on bone remodeling in an orthodontic tooth movement (OTM) model and explore the underlying mechanisms. A rat model of OTM was established, and alveolar bone remodeling and tooth movement rate were evaluated via micro-computed tomography and staining of tissue sections. In vitro, human bone marrow mesenchymal stem cells (hBMSCs) were isolated to detect their osteogenic differentiation potential under compression and LIPUS stimulation by quantitative reverse transcription-polymerase chain reaction, Western blot, alkaline phosphatase (ALP) staining, and Alizarin red staining. The expression of Yes-associated protein (YAP1), the actin cytoskeleton, and the Lamin A/C nucleoskeleton were detected with or without YAP1 small interfering RNA (siRNA) application via immunofluorescence. The force treatment inhibited the osteogenic differentiation potential of hBMSCs; moreover, the expression of osteogenesis markers, such as type 1 collagen (COL1), runt-related transcription factor 2, ALP, and osteocalcin (OCN), decreased. LIPUS could rescue the osteogenic differentiation of hBMSCs with increased expression of osteogenic marker inhibited by force. Mechanically, the expression of LaminA/C, F-actin, and YAP1 was downregulated after force treatment, which could be rescued by LIPUS. Moreover, the osteogenic differentiation of hBMSCs increased by LIPUS could be attenuated by YAP siRNA treatment. Consistently, LIPUS increased alveolar bone density and decreased vertical bone absorption in vivo. The decreased expression of COL1, OCN, and YAP1 on the compression side of the alveolar bone was partially rescued by LIPUS. LIPUS can accelerate tooth movement and reduce alveolar bone resorption by modulating the cytoskeleton-Lamin A/C-YAP axis, which may be a promising strategy to reduce the orthodontic treatment process.

L-Type Calcium Channel Modulates Low-Intensity Pulsed Ultrasound-Induced Excitation in Cultured Hippocampal Neurons.

As a noninvasive technique, ultrasound stimulation is known to modulate neuronal activity both in vitro and in vivo. The latest explanation of this phenomenon is that the acoustic wave can activate the ion channels and further impact the electrophysiological properties of targeted neurons. However, the underlying mechanism of low-intensity pulsed ultrasound (LIPUS)-induced neuro-modulation effects is still unclear. Here, we characterize the excitatory effects of LIPUS on spontaneous activity and the intracellular Ca2+ homeostasis in cultured hippocampal neurons. By whole-cell patch clamp recording, we found that 15min of 1-MHz LIPUS boosts the frequency of both spontaneous action potentials and spontaneous excitatory synaptic currents (sEPSCs) and also increases the amplitude of sEPSCs in hippocampal neurons. This phenomenon lasts for > 10min after LIPUS exposure. Together with Ca2+ imaging, we clarified that LIPUS increases the [Ca2+]cyto level by facilitating L-type Ca2+ channels (LTCCs). In addition, due to the [Ca2+]cyto elevation by LIPUS exposure, the Ca2+-dependent CaMKII-CREB pathway can be activated within 30 min to further regulate the gene transcription and protein expression. Our work suggests that LIPUS regulates neuronal activity in a Ca2+-dependent manner via LTCCs. This may also explain the multi-activation effects of LIPUS beyond neurons. LIPUS stimulation potentiates spontaneous neuronal activity by increasing Ca2+ influx.

Low-intensity pulsed ultrasound delays the progression of osteoarthritis by regulating the YAP-RIPK1-NF-κB axis and influencing autophagy.

Osteoarthritis (OA) is a degenerative disease characterized by chronic inflammation of the joint. As the disease progresses, patients will gradually develop symptoms such as pain, physical limitations and even disability. The risk factors for OA include genetics, gender, trauma, obesity, and age. Unfortunately, due to limited understanding of its pathological mechanism, there are currently no effective drugs or treatments to suspend the progression of osteoarthritis. In recent years, some studies found that low-intensity pulsed ultrasound (LIPUS) may have a positive effect on osteoarthritis. Nonetheless, the exact mechanism by which LIPUS affects osteoarthritis remains unknown. It is valuable to explore the specific mechanism of LIPUS in the treatment of OA. In this study, we validated the potential therapeutic effect of LIPUS on osteoarthritis by regulating the YAP-RIPK1-NF-κB axis at both cellular and animal levels. To verify the effect of YAP on OA, the expression of YAP was knocked down or overexpressed by siRNA and plasmid in chondrocytes and adeno-associated virus was injected into the knee joint of rats. The effect of LIPUS was investigated in inflammation chondrocytes induced by IL-1β and in the post-traumatic OA model. In this study, we observed that YAP plays an important role in the development of osteoarthritis and knocking down of YAP significantly inhibited the inflammation and alleviated cartilage degeneration. We also demonstrated that the expression of YAP was increased in osteoarthritis chondrocytes and YAP could interact with RIPK1, thereby regulating the NF-κB signal pathway and influencing inflammation. Moreover, we also discovered that LIPUS decreased the expression of YAP by restoring the impaired autophagy capacity and inhibiting the binding between YAP and RIPK1, thereby delaying the progression of osteoarthritis. Animal experiment showed that LIPUS could inhibit cartilage degeneration and alleviate the progression of OA. These results showed that LIPUS is effective in inhibiting inflammation and cartilage degeneration and alleviate the progression of OA. As a result, our results provide new insight of mechanism by which LIPUS delays the development of osteoarthritis, offering a novel therapeutic regimen for osteoarthritis.

Low‑intensity pulsed ultrasound accelerates diabetic wound healing by ADSC‑derived exosomes via promoting theuptake of exosomes and enhancing angiogenesis.

Diabetic wounds remain a great challenge for clinicians globally as a lack of effective radical treatment often results in poor prognosis. Exosomes derived from adipose‑derived stem cells (ADSC‑Exos) have been explored as an appealing nanodrug delivery system in the treatment of diabetic wounds. However, the short half‑life and low utilization efficiency of exosomes limit their therapeutic effects. Low‑intensity pulsed ultrasound (LIPUS) provides a non‑invasive mechanical stimulus to cells and exerts a number of biological effects such as cavitation and thermal effects. In the present study, whether LIPUS could enhance ADSC‑Exo‑mediated diabetic wound repair was investigated and its possible mechanism of action was explored. After isolation and characterization, ADSC‑Exos were injected into mice with diabetic wounds, then the mice were exposed to LIPUS irradiation. The control mice were subcutaneously injected with PBS. Wound healing assays, laser Doppler perfusion, Masson's staining and angiogenesis assays were used to assess treatment efficiency. Then, ADSC‑Exos were cocultured with human umbilical vein endothelial cells (HUVECs), and the proliferation, migration and tube formation of HUVECs were assessed. Moreover, the cellular uptake of ADSC‑Exos invitro and invivo was assessed to explore the synergistic mechanisms underlying the effects of LIPUS. The invivo results demonstrated that LIPUS increased the uptake of exosomes and prolonged the residence of exosomes in the wound area, thus enhancing angiogenesis and accelerating wound repair in diabetic mice. The invitro results further confirmed that LIPUS enhanced the uptake efficiency of ADSC‑Exos by 10.93‑fold and significantly increased the proliferation, migration and tubular formation of HUVECs. Therefore, the present study indicates that LIPUS is a promising strategy to improve the therapeutic effects of ADSC‑Exos in diabetic wounds by promoting the cellular uptake of exosomes and enhancing angiogenesis.

Protective effect of low‐intensity pulsed ultrasound on immune checkpoint inhibitor-related myocarditis via fine-tuning CD4+ T-cell differentiation

PurposeImmune checkpoint inhibitors (ICIs) have transformed traditional cancer treatments. Specifically, ICI-related myocarditis is an immune-related adverse event (irAE) with high mortality. ICIs activate CD4+ T-lymphocyte reprogramming, causing an imbalance between Th17 and Treg cell differentiation, ultimately leading to myocardial inflammatory damage. Low-intensity pulsed ultrasound (LIPUS) can limit inflammatory responses, with positive therapeutic effects across various cardiovascular inflammatory diseases; however, its role in the pathogenesis of ICI-related myocarditis and CD4+ T-cell dysfunction remains unclear. Accordingly, this study investigated whether LIPUS can alleviate ICI-related myocarditis inflammatory damage and, if so, aimed to elucidate the beneficial effects of LIPUS and its underlying molecular mechanisms.MethodsAn in vivo model of ICI-related myocarditis was obtained by intraperitonially injecting male A/J mice with an InVivoPlus anti-mouse PD-1 inhibitor. LIPUS treatment was performed via an ultrasound-guided application to the heart via the chest wall. The echocardiographic parameters were observed and cardiac function was assessed using an in vivo imaging system. The expression of core components of the HIPPO pathway was analyzed via western blotting.ResultsLIPUS treatment reduced cardiac immune responses and inflammatory cardiac injury. Further, LIPUS treatment alleviated the inflammatory response in mice with ICI-related myocarditis. Mechanistically, in the HIPPO pathway, the activation of Mst1–TAZ axis improved autoimmune inflammation by altering the interaction between the transcription factors FOXP3 and RORγt and regulating the differentiation of Treg and Th17 cells.ConclusionLIPUS therapy was shown to reduce ICI-related myocarditis inflammatory damage and improve cardiac function, representing an exciting finding for irAEs treatment.