Impact of low intensity pulsed ultrasound on volumetric root resorption of maxillary incisors in patients treated with clear aligner therapy: A retrospective study

ABSTRACT Objective: The aim of this study was to evaluate the volumetric root resorption in maxillary incisors following clear aligner therapy (CAT) with low-intensity pulsed ultrasound (LIPUS), and compare the results to CAT alone. Material and Methods: This retrospective study evaluated pretreatment (T0) and post-treatment (T1) cone-beam computed tomography imaging of 42 adult patients. Twenty-one patients (14 females, 7 males, mean age= 38.1±12.96 years) were treated using CAT with LIPUS device, whereas the other twenty-one matching controls patients (15 females, 6 males, mean age= 35.6±11.7 years) were treated using CAT alone. Images were analyzed and a segmentation protocol was applied on the maxillary incisors. Each segmented tooth volume was exported as a surface mesh in the Visualization Toolkit (VTK) file format. The VTK files for all maxillary incisors were coded and corresponding teeth volumes from T0 and T1 were superimposed. Clipping the crown of each tooth was done, then measurements of root volumes and differences between groups were performed. Changes in root volumes were assessed (p<0.05). Results: Root loss was evident in all teeth in both groups, but was significantly increased in all maxillary incisors of the control group (p<0.001) and in upper left central incisor of LIPUS group (p=0.009). When both groups were compared, there was statistically significant minimal volumetric root loss in LIPUS group (3.50-7.32 mm3), when compared to control group (11.48-12.95 mm3) (p<0.05). Conclusion: LIPUS group showed less volumetric root resorption compared to control group during the studied treatment time using clear aligners.

RESUMO Objetivo: O objetivo deste estudo foi avaliar volumetricamente a reabsorção radicular em incisivos superiores após tratamento com alinhadores transparentes (CAT) com e sem uso adjuvante de ultrassom de baixa intensidade (LIPUS). Material e Métodos: Esse estudo retrospectivo avaliou imagens de tomografia computadorizada de feixe cônico pré-tratamento (T0) e pós-tratamento (T1) de 42 pacientes adultos: 21 pacientes (14 mulheres, 7 homens, idade média= 38,1±12,96 anos) foram tratados com CAT e LIPUS, enquanto os outros 21 pacientes controles correspondentes (15 mulheres, 6 homens, idade média= 35,6±11,7 anos) foram tratados apenas com CAT. As imagens foram analisadas e foi aplicado um protocolo de segmentação dos incisivos superiores. Os volumes de cada dente segmentado foram exportados como malhas de superfície, em arquivos no formato Visualization Toolkit (VTK). Os arquivos VTK de todos os incisivos superiores foram codificados e foram sobrepostos os volumes dos dentes correspondentes a T0 e T1. Foi realizada a clipagem da coroa de cada dente e, em seguida, foram realizadas medições dos volumes radiculares e comparadas as diferenças entre os grupos, avaliando-se as alterações nos volumes de raízes (p<0,05). Resultados: A perda radicular foi evidente em todos os dentes em ambos os grupos, mas foi significativamente maior em todos os incisivos superiores do grupo controle (p<0,001) e no incisivo central superior esquerdo do grupo LIPUS (p=0,009). Quando ambos os grupos foram comparados, houve perda volumétrica mínima estatisticamente significativa no grupo LIPUS (3,50-7,32 mm3), em comparação ao grupo controle (11,48-12,95 mm3) (p<0,05). Conclusão: O grupo LIPUS apresentou menor volume de reabsorção radicular, em comparação ao grupo controle, durante o tempo de tratamento estudado usando alinhadores transparentes.

Efficacy of low intensity pulsed ultrasound in the treatment of erectile dysfunction / 现代泌尿外科杂志

【Objective】 To investigate the efficacy of low intensity pulsed ultrasound (LIPUS) in the treatment of erectile dysfunction (ED). 【Methods】 A total of 121 ED patients treated during June 2020 and June 2022 were selected as the research objects. According to the International Erectile Index Score (IIEF-EF), the patients were divided into three subgroups:mild (17-25 points), moderate (11-16 points), and severe (0-10 points). The total effective rate, erectile hardness scale (EHS), sex life log questions (SEP), general assessment questionnaire (GAQ), peak systolic velocity (PSV), end diastolic velocity (EDV), and adverse reactions of the three groups before treatment, 4 weeks and 12 weeks after treatment were analyzed. 【Results】 A total of 119 patients completed the follow-up. There were significant increases in IIEF-EF and EHS at week 4 and 12 (P<0.05), and the total effective rate was 69.75% and 76.47%, respectively. The total effective rate was significantly higher in the mild and moderate groups than in the severe group at week 4 and 12 (P<0.05). The patients who answered "yes" to SEP2 and SEP3 accounted for 91.60% and 71.43%, respectively at week 4, and 92.44% and 78.15% at week 12, both significantly higher than the rates before treatment (52.10% and 27.73%, P<0.05). The proportion of patients who answered "yes" to GAQl and GAQ2 at week 4 were 84.87% and 71.43%, respectively, and were 82.35% and 70.59% respectively at week 12, with no significant difference. The PSV level significantly increased at week 12 compared to that before treatment [(48.85±14.11) cm/s vs. (41.42±14.90) cm/s] (P<0.05), while the EDV level significantly decreased [(-0.57±7.01) cm/s vs. (2.25± 5.68)cm/s] (P<0.05). 【Conclusion】 LIPUS can improve erectile function in ED patients without obvious adverse reactions.

Efficacy of low-intensity pulsed ultrasound therapy in elderly patients with coronary heart disease and angina pectoris / 中华老年医学杂志

Objective:To investigate the practical efficacy of low-intensity pulsed ultrasound(LIPUS)and its effects on hemodynamics in patients with coronary heart disease and angina pectoris.Methods:As a single-center randomized controlled study, 66 elderly patients with coronary heart disease and angina pectoris treated at Qilu Hospital of Shandong University between November 2021 and August 2022 were consecutively recruited.Participants were divided into an ultrasound group and a control group via the random number table method, with 33 in each group.Both groups were given conventional drug treatment, and the ultrasound group was supplemented with LIPUS treatment.After 20 sessions of treatment, the clinical efficacy, blood lipid levels, inflammatory factors and hemodynamics from noninvasive monitoring before and after treatment were compared.Results:There were no statistically significant differences in baseline data between the two groups( P>0.05). After treatment, marked effectiveness was seen in 12 cases(36.4%), effectiveness in 18 cases(54.5%), ineffectiveness in 2 cases(6.1%)and aggravation in 1 case(3%)in the ultrasound group, with a total effectiveness rate of 90.9 %.In the control group, marked effectiveness was seen 2 cases(6.1%), effectiveness in 19 cases(57.6%)and ineffectiveness in 9 cases(27.3%), with a total effectiveness rate of 63.6%.Statistically significant favorable results were achieved in the ultrasound treatment group, compared with the control group( P<0.05). Also, compared with the control group, patients in the ultrasound group showed lower total cholesterol[(2.31±1.03)mmol/L vs.(4.36±0.76)mmol/L, P<0.01], decreased low-density lipoprotein cholesterol[(1.24±0.70)mmol/L vs.(1.74±0.44)mmol/L, P<0.01], decreased triglycerides[(1.04±0.40)mmol/L vs.(1.28±0.49)mmol/L, P<0.05], decreased apolipoprotein B[(0.67±0.25)g/L vs.(0.90±0.14)g/L, P<0.01], decreased interleukin-6[(2.87±2.52)ng/L vs.(4.66±3.30)ng/L, P<0.05], and decreased high-sensitivity C-reactive protein[(1.04±1.41)mg/L vs.(3.80±5.78)mg/L, P<0.05]. Concerning hemodynamics, there was an increase in cardiac output[(4.92±1.05)L/min vs.(4.39±0.97)L/min, P<0.05], stroke volume[(75.85±17.50)ml/beat vs.(66.97±15.57)ml/beat, P<0.05], cardiac index[(37.50±7.14)ml·beat -1·m -2vs.(43.29±7.96)ml·beat -1·m -2, P<0.01], and stroke volume index[(3.05±0.45)L/min 2vs.(2.51±0.43)L/min 2, P<0.01], but a decrease in systemic vascular resistance[(1 358.29±325.23)dyne·s -1·cm -5vs.(1 617.94±526.27)dyne·s -1·cm -5, P<0.05], total vascular resistance[(4.07±24.30)% vs.(21.32±31.94)%, P<0.05], and average heart rate[(63.43±7.42)beats/min vs.(69.11±10.89)beats/min, P<0.05]. Conclusions:LIPUS treatment can improve symptoms, blood lipid profiles, inflammatory factors and hemodynamics in patients with coronary heart disease and angina pectoris.It is a safe and effective adjuvant therapy for elderly patients with coronary heart disease and angina pectoris.

Effect of low intensity ultra sound on bone regeneration and healing: a systematic review

ABSTRACT Objective: Low Intensity Pulsed Ultra Sound (LIPUS) is found to have stimulatory effect on bone healing and regeneration. This review aimed to assess whether LIPUS enhances bone regeneration and healing in terms of efficiency in improving clinical, radiographic, histologic parameters or serum and tissue biomarkers. Methods: A comprehensive search based on PRISMA guidelines with pre-determined eligibility criteria was conducted to identify randomized controlled clinical trials evaluating effectiveness of Low intensity pulsed ultrasound in bone regeneration and healing. The title and abstract of the entries in all languages yielded from the PubMed, Google scholar and Cochrane library were screened. Results: 14 eligible Randomized controlled trials testing the effectiveness of LIPUS was evaluated. More heterogeneity was seen in the screened studies with respect to sample characteristics, type of bone and outcome measures. The studies that screened histological parameters state that LIPUS is significantly beneficial than control. In terms of time for radiographic union, most of the studies stated that LIPUS was more effective than control but numberof studies are very few. Whereas studies which evaluated parameters such as healing time and radiographic union were showing highly inconsistent results regarding effectiveness of LIPUS. Conclusion: This review cannot give a definitive conclusion that LIPUS is effective in bone healing with respect to clinical parameters but a positive influence on radiographical and histological parameters in bone healing and regeneration is promising to pursue future research.

Therapeutic effect of low intensity pulsed ultrasound on painful gait in mice with Achilles tendon injury and its possible mechanism / 中华创伤杂志

Objective:To investigate the therapeutic effect and possible mechanism of low intensity pulsed ultrasound（LIPUS）on painful gait in mice with Achilles tendon injury.Methods:Male C57BL/6 mice were selected to establish the Achilles tendon injury model by surgically full-thinkness tear of the right Achilles tendon. The mice were divided into Achilles tendon injury group and Achilles tendon injury+LIPUS group according to the random number table method，with 7 mice per group. The Catwalk gait analysis system was used to evaluate the gait function of the mice by measuring the following five parameters 14 days after operation，including print area，standing time，step cycle，max intensity and stride length. Morphological changes of the Achilles tendon were observed by HE staining. Immunofluorescence staining was used to detect the expression level of nitric oxide synthase（iNOS）in the Achilles tendon tissues. At the same time，Achilles tendon cells were isolated and cultured in vitro. The cells were induced by 100 ng/ml lipopolysaccharide（LPS）for 12 hours to establish in vitro model（LPS group），and treated with LIPUS（LPS+LIPUS group）. The control group was set as well（without any treatment）. The nuclear translocation of nuclear transcription factor-κB（NF-κB）P65 was detected by immunofluorescence staining，and the expression of iNOS and phosphorylation（p）-NF-κB p65 protein was detected by Western blotting. Results:Compared with Achilles tendon injury group［（0.14±0.10）cm 2］，the print area of the affected limb in Achilles tendon injury+LIPUS group［（0.28±0.13）cm 2］was increased（ P<0.05）. Compared with Achilles tendon injury group［（0.11±0.04）seconds］，the standing time of the affected limb in Achilles tendon injury+LIPUS group［（0.21±0.03）seconds］was increased（ P<0.05）. Compared with Achilles tendon injury group［（0.25±0.05）seconds］，the step cycle of the affected limb in Achilles tendon injury+LIPUS group［（0.40±0.05）seconds］was increased（ P<0.05）. There was no significant difference in the max intensity and stride length between Achilles tendon injury group and Achilles tendon injury+LIPUS group（ P>0.05）. HE staining showed obvious hyperplasia of Achilles tendon tissues in Achilles tendon injury group，with loosely and randomly arranged fibers，accompanied by neovascularization and inflammatory cell infiltration. Achilles tendon injury+LIPUS group showed more orderly arranged fibers in Achilles tendon tissues，and the degree of neovascularization and inflammatory cell infiltration were reduced. Compared with Achilles tendon injury group［（5.70±0.81）%］，the expression level of iNOS in Achilles tendon injury+LIPUS group［（2.84±0.94）%］was decreased 14 days after operation（ P<0.05）. Immunofluorescence results of Achilles tendon cells showed that NF-κB p65 protein entered the nucleus in LPS group compared with control group，and that LIPUS treatment inhibited nuclear translocation. In control group，Western blotting showed that iNOS was not expressed，and that the expression of phosphorylated NF-κB p65 was 0.63±0.16. Compared with control group，the expression levels of iNOS（0.99±0.22）and p-NF-κB P65（1.02±0.19）in LPS group were significantly increased（ P<0.05）. Compared with LPS group，the expression levels of iNOS（0.62±0.10）and p-NF-κB P65（0.65±0.21）in LPS+LIPUS group were decreased（ P<0.05）. Conclusion:LIPUS treatment can alleviate pain gait in mice with Achilles tendon injury and inhibit iNOS expression in vivo and in vitro，which may be related to inhibition of NF-κB signaling pathway.

Low-intensity pulsed ultrasound ameliorates angiotensin II-induced cardiac fibrosis by alleviating inflammation via a caveolin-1-dependent pathway / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

OBJECTIVES@#Cardiac hypertrophy and fibrosis are major pathological manifestations observed in left ventricular remodeling induced by angiotensin II (AngII). Low-intensity pulsed ultrasound (LIPUS) has been reported to ameliorate cardiac dysfunction and myocardial fibrosis in myocardial infarction (MI) through mechano-transduction and its downstream pathways. In this study, we aimed to investigate whether LIPUS could exert a protective effect by ameliorating AngII-induced cardiac hypertrophy and fibrosis and if so, to further elucidate the underlying molecular mechanisms.@*METHODS@#We used AngII to mimic animal and cell culture models of cardiac hypertrophy and fibrosis. LIPUS irradiation was applied in vivo for 20 min every 2 d from one week before mini-pump implantation to four weeks after mini-pump implantation, and in vitro for 20 min on each of two occasions 6 h apart. Cardiac hypertrophy and fibrosis levels were then evaluated by echocardiographic, histopathological, and molecular biological methods.@*RESULTS@#Our results showed that LIPUS could ameliorate left ventricular remodeling in vivo and cardiac fibrosis in vitro by reducing AngII-induced release of inflammatory cytokines, but the protective effects on cardiac hypertrophy were limited in vitro. Given that LIPUS increased the expression of caveolin-1 in response to mechanical stimulation, we inhibited caveolin-1 activity with pyrazolopyrimidine 2 (pp2) in vivo and in vitro. LIPUS-induced downregulation of inflammation was reversed and the anti-fibrotic effects of LIPUS were absent.@*CONCLUSIONS@#These results indicated that LIPUS could ameliorate AngII-induced cardiac fibrosis by alleviating inflammation via a caveolin-1-dependent pathway, providing new insights for the development of novel therapeutic apparatus in clinical practice.

Low-intensity pulsed ultrasound stimulates proliferation of stem/progenitor cells: what we need to know to translate basic science research into clinical applications / 亚洲男科学杂志(英文版)

Low-intensity pulsed ultrasound (LIPUS) is a promising therapy that has been increasingly explored in basic research and clinical applications. LIPUS is an appealing therapeutic option as it is a noninvasive treatment that has many advantages, including no risk of infection or tissue damage and no known adverse reactions. LIPUS has been shown to have many benefits including promotion of tissue healing, angiogenesis, and tissue regeneration; inhibition of inflammation and pain relief; and stimulation of cell proliferation and differentiation. The biophysical mechanisms of LIPUS remain unclear and the studies are ongoing. In recent years, more and more research has focused on the relationship between LIPUS and stem/progenitor cells. A comprehensive search of the PubMed and Embase databases to July 2020 was performed. LIPUS has many effects on stem cells. Studies show that LIPUS can stimulate stem cells in vitro; promote stem cell proliferation, differentiation, and migration; maintain stem cell activity; alleviate the problems of insufficient seed cell source, differentiation, and maturation; and circumvent the low efficiency of stem cell transplantation. The mechanisms involved in the effects of LIPUS are not fully understood, but the effects demonstrated in studies thus far have been favorable. Much additional research is needed before LIPUS can progress from basic science research to large-scale clinical dissemination and application.

Safety and efficacy of focused low-intensity pulsed ultrasound therapy for the treatment of knee osteoarthritis / 上海交通大学学报（医学版）

Objective • To investigate the safety and efficacy of focused low-intensity pulsed ultrasound (FLIPUS) therapy on the patients with knee osteoarthritis (KOA). Methods • A total of 100 subjects with KOA from August 2018 to August 2019 in the Department of Rehabilitation Medicine at the Second Affiliated Hospital of Chongqing Medical University were sequentially randomized into group A and group B. Among them, FLIPUS was adopted to treat the 50 patients in group A, and the diclofenac sodium was adopted to treat the other 50 patients in group B. Visual analogue scale (VAS), Western Ontario McMaster Universities Osteoarthritis Index (WOMAC) and Lequesne index (LI) were used to evaluate knee joint pain, the structure and function of the affected knee joint, and the severity of the affected knee joint at baseline, on day 3, day 6 and day 10 after treatment respectively. The knee range of motion (KROM) and maximum walking speed (MWS) were used to evaluate the knee joint movement and walking function at baseline and on day 10 after treatment respectively. Results • There were on significant differences between two groups with respect to KOA evaluation indexes (P>0.05). Compared with those in group B, patients in group A showed significant improvements in VAS and WOMAC scores on day 6 and day 10 after treatment (P=0.021, P=0.000; P=0.005, P=0.001). Meanwhile, patients in group A had higher LI scores than those in group B on day 10 after treatment (P=0.000). In addition, patients in group A showed significant improvements in MWS (P=0.006) and non-significant improvements in KROM (P=0.064) on day 10 after treatment. Conclusion • FLIPUS is a safe and effective treatment modality for relieving pain and improving the functions of patients with KOA.

Low-intensity pulsed ultrasound for regenerating peripheral nerves: Potential for penile nerve / 亚洲男科学杂志(英文版)

Peripheral nerve damage, such as that found after surgery or trauma, is a substantial clinical challenge. Much research continues in attempts to improve outcomes after peripheral nerve damage and to promote nerve repair after injury. In recent years, low-intensity pulsed ultrasound (LIPUS) has been studied as a potential method of stimulating peripheral nerve regeneration. In this review, the physiology of peripheral nerve regeneration is reviewed, and the experiments employing LIPUS to improve peripheral nerve regeneration are discussed. Application of LIPUS following nerve surgery may promote nerve regeneration and improve functional outcomes through a variety of proposed mechanisms. These include an increase of neurotrophic factors, Schwann cell (SC) activation, cellular signaling activations, and induction of mitosis. We searched PubMed for articles related to these topics in both in vitro and in vivo animal research models. We found numerous studies, suggesting that LIPUS following nerve surgery promotes nerve regeneration and improves functional outcomes. Based on these findings, LIPUS could be a novel and valuable treatment for nerve injury-induced erectile dysfunction.

Effects of low-intensity pulsed ultrasound on hematopoietic function in rats after combined chemotherapy with doxorubicin and cyclophosphamide / 南方医科大学学报

OBJECTIVE@#To investigate the effect of low-intensity pulsed ultrasound (LIPUS) on hematopoietic function in rats after combined chemotherapy with doxorubicin and cyclophosphamide.@*METHODS@#Eighty rats were randomized into control group and LIPUS group (=40) for treatment with intraperitoneal injection of doxorubicin (2 mg/kg)+cyclophosphamide (20 mg/kg) for 4 consecutive days and continuous irradiation with LIPUS for 7 days following the injections, respectively. The white blood cells, red blood cells and platelets counts in each group were measured at 0, 4, 7, 9, 11, 14 and 18 days after the start of drug administration. The pathological sections of the bone marrow were examined at 0, 4 and 11 days, and the flow cytometry was performed for detecting the cell apoptosis; qPCR was performed for detecting the expressions of SCF, ICAM-1, and VCAM-1 mRNAs, and ELISA was used to detect the expressions of IL-3 and GM-CSF.@*RESULTS@#The white blood cell count was significantly higher in LIPUS group than in the control group ( < 0.05). Histopathological examination of the bone marrow revealed significantly increased hematopoietic tissue in LIPUS group ( < 0.05). Flow cytometry demonstrated an obviously lower cell apoptosis rate in the bone marrow in LIPUS group than in the control group ( < 0.05). Compared with those in the control group, the mRNA expression levels of ICAM-1 and VCAM-1 as well as the protein levels of IL-3 and GM-CSF were significantly increased in LIPUS group ( < 0.05).@*CONCLUSIONS@#LIPUS can alleviate the hematopoietic damage after combined chemotherapy with doxorubicin with cyclophosphamide probably by increasing the expressions of ICAM- 1, VCAM-1, IL- 3, and GM-CSF.

Low-intensity pulsed ultrasound promotes proliferation and migration of HaCaT keratinocytes through the PI3K/AKT and JNK pathways

Although the effects of low-intensity pulsed ultrasound (LIPUS) on diverse cell types have been fully studied, the functional role of LIPUS in keratinocytes remains poorly understood. This study aimed to investigate the effects of LIPUS on proliferation and migration of HaCaT cells as well as the regulatory mechanisms associated with signaling pathways. Human HaCaT cells were exposed or not to LIPUS, and cell proliferation and migration were measured by BrdU incorporation assay and Transwell assay, respectively. Expression of proteins associated with proliferation and migration was evaluated by western blot analysis. Expression of key kinases in the PI3K/AKT and JNK pathways was also evaluated by western blot analysis. Effects of LIPUS on the PI3K/AKT and JNK pathways, and whether LIPUS affected HaCaT cells via these two pathways were finally explored. When the parameter of LIPUS (number of cycles) was set at 300, cell viability was the highest after LIPUS stimulation. We then found that the percentage of BrdU positive cells was enhanced by LIPUS, along with up-regulation of cyclinD1, CDK6, CDK4, and VEGF. LIPUS promoted migration, as well as up-regulation of MMP-2 and MMP-9. Phosphorylation levels of key kinases in the PI3K/AKT and JNK pathways were increased by LIPUS. Inhibition of either PI3K/AKT pathway or JNK pathway attenuated effects of LIPUS on HaCaT cells, and co-inhibition of these two pathways showed augmented effects. LIPUS promoted proliferation and migration of HaCaT cells through activating the PI3K/AKT and JNK pathways.

The effects of low intensity pulsed ultrasound on the expression of calcium transportation associated proteins during dentin-pulp complex injury and repair in rats / 实用口腔医学杂志

Objective: To investigate low intensity pulsed ultrasound (LIPUS) on the expression of L-type calcium ion channels(cav1. 2) and Na +-Ca2 + exchangers(NCX1) during dentin-pulp complex injury and repair in rats. Methods: Cavity preparation was made on the upper right first molar of 40 male adult SD rats,20 of them and the upper left first molar of the other 20 were randomly chosen for LIPUS irradiation(frequency: 1. 5 MHz,200 μs pulses,pulse repetition frequency: 1 KHz,ISATA 30 mW/cm2,20 min /d),so the animals were randomly allocated into 4 groups(n = 10): Control group,LIPUS group,cavity preparation group and cavity preparation + LIPUS group. At 1,3,7,14 d post-irradiation the rats were sacrificed respectively for HE stain and immunohistochemical analysis. Results: Reparative dentin formation was observed at 14 days after cavity preparation and LIPUS irradiation,the expression of Cav1. 2(L-type) and NCX1 in this group were increased significantly at day 1 and day 3. Compared with the control group, the expression of Cav1. 2 in LIPUS group increased at day 1 post-irradiation. Conclusion: LIPUS may enhance tertiary dentin formation and up-regulate the expression of Cav1. 2 and NCX1 at the early period of dentin injury.

Effects of low-intensity pulsed ultrasound on fracture healing and caveolin-1 gene expression in the radius defects of rabbits / 复旦学报（医学版）

Objective To explore the effect of low-intensity pulsed ultrasound (LIPUS) on long bone fracture healing and to examine caveolin-1 gene expression in the radius defects of rabbits.Methods A total of 24 New Zealand rabbits with 3-mm bone defects at lower 1/3 in both radii were randomly assigned to 4 groups (n=6).Daily LIPUS treatment was performed to the right fracture sites at a intensity of 30 mW/cm2 for 20 minutes,while the left sites received sham treatment with power off.To assess the effects of LIPUS on bone defects,X-ray imaging and hematoxylin-eosin staining were applied 7,14,21,28 days after the surgery.Additionally,the immunohistochemical staining was used to determine the subcllular localization of caveolin-1 and semi-quantify the caveolin-1 level,qPCR was performed to detect the mRNA level of caveolin-1,gene Col2a1 and Col10a1,and osteocalcin.Results On day 14,the radiological score of the right radii and mineralized callus area were significantly higher than that of the left ones,both of them were elevated with time flied.Histological examination suggested that the differentiation and apoptosis of chondrocytes along with the formation and bridging of the bone trabeculas appeared earlier in the right radius defects.The immunohistochemical staining showed that on day 7 and 14,the level of caveolin-1 increased with the proliferation and differentiation of condrocytes,and was significantly higher in callus tissues on the right sites.On day 21 and 28,the mesenchymal stem cells migrated to the surface of cartilage matrix started to differentiate into osteoblasts,the level of caveolin-1 decreased,and was significantly lower on the right sites.The result of qPCR indicated that compared with the left sites the caveolin-1 gene expression on the right sites was significantly higher on day 7,while significantly lower on day 21.The mRNA expression levels of Col2a1,Col10a1 and osteocalcin on the right sites were significantly higher on day 7 and 14,but they were significantly lower on day 21 and 28,except for Col10a1 on day 28.Conclusions Advancing endochondral ossification is considered to be a crucial mechanism during long bone fracture healing promoted by LIPUS.The caveolin-1 gene expression first increased in the chondrocytes then decreased in the mesenchymal stem cells during the process.

Impact on migration of SD rat bone marrow mesenchymal stem cells treated with different intensities of low intensity pulsed ultrasound / 中国介入影像与治疗学

Objective To investigate the impact of low intensity pulsed ultrasound (LIPUS) with different intensities on the migration of bone marrow mesenchymal stem cells (BMSCs) in vitro.Methods BMSCs were divided into control group,30 mW/cm2 group,60 mW/cm2 group and 90 mW/cm2 group.Control group was treated by sham LIPUS exposure,and the other three groups were treated by LIPUS with corresponding intensities.The impact of LIPUS on scratch healing was tested with scratch assay,and the interference of proliferation was eliminated with MTT assay.The migration of BMSCs were evaluated with transwell migration assay.The expression of F-actin was analyzed with fluorescein isothiocyanate (FITC) fluorescent coloration.Results 24 h and 48 h after LIPUS exposure,there were statistical differences of scratch area among groups (F=26.559,106.110,both P＜0.001),and the scratch area of control group was the largest ([0.93 ± 0.26)mm2 of 24 h after LIPUS exposure and [0.70 ± 0.11]mm2 of 48 h after LIPUS exposure),while that of 30 mW/cm2 group was the smallest ([0.47 ±0.21]mm2 of 24 h after LIPUS exposure and [0.19±0.10]mm2 of 48 h after LIPUS exposure).There was no statistical difference of scratch area among the four groups immediately after LIPUS exposure (F=2.921,P=0.063).MTT assay results showed there was no statistical difference of absorbance among the four groups immediately,nor 24 h,48 h after LIPUS exposure (F=1.616,0.720,1.408;P=0.196,0.544,0.378).Significant difference was found in the number of cells migrated through the transwell chamber among the four groups (F=43.145,P＜0.001),and the cell number of 30 mW/cm2 group was the largest (212.53±35.32),while that of the control group was the least (89.53±19.27).F-actin fluorescence staining results showed the morphology of F-actin was changed after LIPUS exposure.The cytoskeleton became narrow and elongated.Statistical difference of relative fluorescence intensity was found among the four groups (F 64.350,P＜0.001).The relative fluorescence intensity of 30 mW/cm2 group was the largest (125.43 ± 17.43),while that of control group was the least (51.94± 12.76).Conclusion LIPUS can promote the migration ability of BMSCs in vitro with the best intensity was 30 mW/cm2.

Effect of low intensity pulsed ultrasound on the morphology of microglia in hippocampus of mice / 局解手术学杂志

Objective To study the effect of low intensity pulsed ultrasound on the hippocampus and to study its possible mechanism in mice.Methods Totally 32 adult C57/BL6 mice were randomly divided into the sham group(n=16)and the LIPUS group(n=16).Mice in the sham group were treated with sham operative,while mice in the LIPUS group were treated by low intensity pulsed ultrasound(average power density of 150 mW/cm2).After 7 days of treatment,immunohistochemical(IHC)and western bolt(WB)were used to determine the expression of DCX,GFAP,and Iba1 in the hippocampus,and the behavioral changes of mice in open field test were observed.Results After 7 days of treatment,the results of IHC showed that the number of microglial cells in the LIPUS group were significantly increased,and the difference was statistically significant compared with that in the sham group(P<0.05).The WB results indicated that the expression of Iba 1 protein in hippocampus was significantly increased in LIPUS group compared with that in the sham group(P<0.05).In the meantime,the behavior score of LIPUS group in the open field test was increased significantly(P<0.05).Conclusion Low intensity pulsed ultrasound can increase the number of microglia in the hippocampus and reduce the anxiety of the mice.

Low-intensity pulsed ultrasound pretreatment inhibits HMGB1 expression and attenuates lung ischemia-reperfusion injury in rats the cholinergic anti-inflammatory pathway / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the effects of low-intensity pulsed ultrasound (LIPUS) pretreatment on pulmonary expression of high mobility group box-1 (HMGB1) in a rat model of lung ischemia-reperfusion (IR).</p><p><b>METHODS</b>Thirty-two male SpragueDawley rats weighing 250-300 g were randomly divided (=8) into sham-operated group, lung IR group, LIPUS pretreatment group and pretreatment with α7-nicotinic cholinergic receptor (α7nAChR) antagonist group. In the sham-operated group, the left pulmonary hilum was dissociated without occlusion; in the other 3 groups, the left pulmonary hilum was occluded for 45 min followed by reperfusion for 180 min; LIPUS pretreatment for 30 min and intraperitoneal injection of methyllycaconitine (2 mg/kg), an α7nAChR antagonist, were administered before the operation. The wet/dry weight ratio (W/D) and pulmonary permeability index (LPI) of the lung tissue were measured, and the lung histopathology was observed and scored. The contents of interleukin-1 (IL-1) and IL-6 in the lung tissues were measured using ELISA, and the pulmonary expression of HMGB1 protein was detected using immunofluorescence assay and Western blotting.</p><p><b>RESULTS</b>Compared with those in the sham-operated group, the W/D of the lung tissue, LPI, pathological scores, IL-1 and IL-6 contents in the lung tissue, and pulmonary HMGB1 expression all significantly increased in the other 3 groups ( < 0.05). LIPUS preconditioning significantly lowered the W/D values, LPI, pathological score, IL-1 and IL-6 contents and HMGB1 expression in the lung tissues following lung IR, and these effects were significantly inhibited by administration of methyllycaconitine.</p><p><b>CONCLUSIONS</b>LIPUS preconditioning can reduce lung IR injury possibly by activating α7nAChR-dependent cholinergic anti-inflammatory pathway to reduce lung tissue HMGB1 expression.</p>

Effects of underwater lipus on the morphohistology of the CF-1 mice growth plate in late lactation stage / Efectos del ultrasonido subacuático sobre la morfohistología de la placa de crecimiento de ratones CF-1 en etapa de lactancia tardía

SUMMARY: Ultrasound is one of the most used tools in physiotherapy, its LIPUS modality allows to treat musculoskeletal injuries, but like the conventional ultrasound it is contraindicated its application in areas close to growth cartilage by a possible closure of this one. In the literature there is no conclusive evidence of this contraindication, so the objective of the present study is to evaluate the effects of doses equivalent to underwater LIPUS on the histomorphometry of the proximal tibial epiphysis growth plate in an animal model. LIPUS was applied in underwater mode in the right pelvic limb to 8 mice in p 15 for 4 days, 2 times a day. On completion of 8 sessions the histology of the stimulated limb growth plate was sacrificed and the histology. When evaluating the histomorphometry of the plaque, a significant increase in the thickness and area of the proliferative zone stimulated with LIPUS (p = 0.048) was observed. In the hypertrophic area were observed no significant differences in thickness or areas, but in the hypertrophic area there were significant changes in the histological organization, observing a significant increase of the entire columns in the stimulated plaque (p = 0.04). LIPUS in underwater modality modifies the histomorphometry of the proximal epiphyseal PC of the tibia of rodents in the late postnatal stage, promoting proliferation and columnar organization in the proliferative and hypertrophic zone, respectively.

RESUMEN: El ultrasonido es una de las herramientas más utilizadas en fisioterapia, su modalidad LIPUS permite tratar lesiones músculo-esqueléticas, pero al igual que el ultrasonido convencional está contraindicada su aplicación en zonas próximas a cartílago de crecimiento por un posible cierre de éste. En la literatura no existe evidencia contundente de esta contraindicación, por lo cual el objetivo del presente estudio fue evaluar los efectos de dosis equivalentes a LIPUS subacuático sobre la histomorfometría de la placa de crecimiento de la epífisis proximal de tibia en un modelo animal. Metodología. Se aplicó LIPUS en modalidad subacuática en el miembro pélvico derecho a 8 ratones en p 15 por 4 días, 2 veces al día. Al cumplir 8 sesiones se sacrificaron y se evaluó la histología de la placa de crecimiento del miembro estimulado. Al evaluar la histomorfometría de la placa se observó un aumento significativo en el espesor y el área de la zona proliferiva estimulada con LIPUS (p=0.048). En la zona hipertrófica no se observaron diferencias significativas en el espesor ni en las áreas, pero si existieron cambios significativos en la organización histológica de la zona, observando un aumento significativo de las columnas completas en la placa estimulada (p=0,04). El LIPUS en modalidad subacuática modifica la histomorfometría de la PC epifisiaria proximal de la tibia de roedores en el etapa postnatal tardía, promoviendo la proliferación y la organización columnar en la zona proliferativa e hipertrófica, respectivamente.

Effect of Low-intensity Pulsed Ultrasound on the Proliferation and Differentiation of Osteoblasts Cultured on Two Piezoelectric Ceramic Porous Scaffolds / 中国医科大学学报

Objective To evaluate the effects of low-intensity pulsed ultrasound (LIPUS) treatment on the proliferation and differentiation of osteoblasts cultured on barium titanate/titanium alloy (BaTiO3) and zinc oxide/titanium alloy (ZnO) piezoelectric ceramic composite scaffolds.Methods The BaTiO3 and ZnO porous composite scaffolds were randomly divided into ultrasound (UBaTiO3 and UZnO) and control (CBaTiO3 and CZnO)groups,and MC3T3-E1 cells were cultured on the surface of the materials in each group.Cells in the ultrasound groups were subjected to a 20-min treatment LIPUS session daily,and the control group was sham-treated without the power source.The proliferation and differentiation rates of osteoblasts were evaluated using methyl thiazolyl tetrazolium (MTT),alkaline phosphatase (ALP),and osteocalcin (OCN) assays.Results Significant differences in the MTF values were observed between the UBaTiO3 and CBaTiO3 groups at day 4 (P ＜ 0.05) and between the UBaTiO3 and UZnO groups at days 1,4,and 7 (P ＜ 0.05).Compared with the CBaTiO3 group,the UBaTiO3 group showed significantly higher ALP activity and OCN levels at days 4 and 7 (P ＜ 0.05).Further,a significant difference in the ALP and OCN values between the UBaTiO3 and UZnO groups (P ＜ 0.05) was observed.Conclusion LIPUS promotes the proliferation and differentiation of osteoblasts on BaTiO3 and ZnO scaffolds.BaTiO3 scaffolds enable better promotion of osteogenesis under LIPUS treatment than ZnO scaffolds do.

Effect of low intensity pulsed ultrasound on expression of TGF-β1 and Smads during dentin injury and repair / 中南大学学报(医学版)

Objective:To explore the effect of low intensity pulsed ultrasound (LIPUS) on TGF-β1/Smad 2,3 signal pathway during the dentin injury and repair.Methods:Among 25 Sprague-Dawley rats,5 rats served as a blank control group without treatment.The remaining 20 rats received modified caries preparation inbilateral maxillary first molars to establish a model of dentin-pulp injury and repair.The right maxillary first molars served as a LIPUS group,which received LIPUS irradiation (frequency:1.5 MHz,pulse width:200 μs,pulse repetition frequency:1 kHz,spatial averaged temporal averaged intensity:30 mW/cm2,20 min/d),and the left maxillary first molars served as a cavity-prepared group,which received fake LIPUS irradiation.The rats were sacrificed at 1,3,5,7 and 14 days after LIPUS irradiation.Immunohistochemical staining and Image-pro plus 6.0 were applied to detect the expression and distribution of transforming growth factor-beta1 (TGF-β1) and small mothers against decapentaplegic 2/3(Smad 2 and Smad 3).Results:Immunohistochemical staining showed that the expression of TGF-β1 and Smad 2,3were low innormal pulp,but they were increased in different degree after dentin injury.The result of image analysis showed that the expression of TGF-β1 in the cavity-prepared group gradually increased at the first day and peaked at day 5,and then it returned to normal level at day 14.However,the expression of TGF-β 1 in the LIPUS group were significantly higher than that in the cavity-prepared group at day 3 and 5 (bothP＜0.05).The expressions of Smad 2,3 in both the LIPUS group and the cavity-prepared group were consistently increased all the way, but the expressions in the LIPUS group were higher compared with that in the cavity-prepared group (P＜0.05).Conclusion:The TGF-β1/Smad 2,3 signal pathway can be activated during the dentin injury and repair.LIPUS can up-regulate the expression of TGF-β1 and Smad 2,3 in the early period,which may take part in the dentin-pulp complex injury and repair process.

Effects of Low Intensity Pulsed Ultrasound on Osteocyte Injuries Induced by Tricalcium Phosphate Wear Particles in the Calvaria of Mice / 中国运动医学杂志

Objectives To observe the effect of low intensity pulsed ultrasound (LIPUS) on osteocyte injuries induced by the tricalcium phosphate(TCP) wear particles in the calvaria of mice.Methods Thirty ICR male mice of 6 to 8 weeks were randomly divided into a normal control group(n=10),a model group (n=10) and a LIPUS-treated group(n=10).A murine calvarial model of osteolysis was established in the model and LIPUS-treated groups through injecting TCP particles onto the surface of bilateral parietal bones at week 1,3,5,7 and 11.Mice in the normal group received negative ultrasound probe pressing,while those in the LIPUS-treated received LIPUS radiation.Three months later,the calvarias were obtained.The micro-CT,HE staining,flow cytometry and Western blotting were performed to estimate the calvarial osteolysis,osteocyte death,apoptosis and proteins expression of the dentin matrix protein 1 (DMP-1),sclerosis protein (SOST),glucose-regulated protein78 (GRP78),inositol-requiring enzyme(IRE 1 α),spliced X-box binding protein 1 (XBP1 s),c-Jun N-terminal kinase (JNK) and phosphorylated c-Jun N-terminal kinase(p-JNK) respectively.Results Compared with the normal control group,in the model group the viability of prosthetic osteocytes decreased significantly,and cell apoptosis was more obvious(P＜0.05);the osteocytic marker protein DMP-1 down-regulated significantly,but another marker protein SOST up-regulated significantly,which caused the decline in DMP-1/SOST(P＜0.05).Moreover,the expression levels of GRP78,IRE1,XBPls and p-JNK of the model group increased significantly(P＜0.05) in the calvaria osteocytes compared to the control group.However,in the LIPUS treatment group,osteocyte injuries and endoplasmic reticulum(ER) stress both decreased significantly,shown by a significant increase in the number and activity of osteocytes,DMP-1/SOST,and significant inhibition of the IRE1α-XBP1-JNK activation(P＜0.05).Conclusion LIPUS prevents osteocyte injuries induced by TCP wear particles in the calvaria of mice,which may be due to the inhibition of IRE1α-XBP1-JNK pathway activation through ER stress reaction.