Pulsed Low Intensity Research Articles

Global sonication of the human intracranial space via a jumbo planar transducer

Contrary to conditioning a Focused Ultrasound (FUS) beam to sonicate a localized region of the human brain, the goal of this investigation was to explore the prospect of distributing homogeneous ultrasound energy over the entire brain space with a large cranium-wide ultrasound beam. Recent ultrasound preclincal studies utilizing large or whole brain stimulation regions create a demand for expanding the treatment envelope of transcranial pulsed-low intensity ultrasound towards Global Brain Sonication (GBS) for potential human investigation. Here, we conduct ultrasound field characterizations when transmitting pulsed ultrasound through human skull specimens using a 1–3 piezocomposite planar transducer operating at 464 kHz with an active single-element surface of 30 × 30 cm. Through computational simulation and hydrophone scanning methodology, ultrasound wave behavior and dose homogeneity in the brain space were evaluated under various trajectories of sonication using the planar transducer. Clinically relevant pulse parameters used for transcranial therapeutic ultrasound applications were used in the experiments. Simulations and empirical testing revealed that dose homogeneity and acoustic intensity over the brain space are influenced by sonication trajectory, skull lens effects, and acoustic wave reflections. The transducer can emit a spatial peak pulse average intensity of 4.03 W/cm2 (0.24 MPa) measured in the free-field at 464 kHz with electrical power of 1 kW. The simulation showed that approximately 99 % of the cranial volume was exposed with <30 % of the maximum external acoustic intensity being transmitted into the skull. The transmission loss across all sonication trajectories is similar to previously reported FUS studies. A marker for GBS dose homogeneity is introduced to score the ultrasound pressure field uniformity in the intracranial space. Results of this study identify the initial challenges of exposing the entire human brain space with ultrasound using a large cranium-wide sonication beam intended for global brain therapeutic modulation.

Pulsed low-intensity laser treatment stimulates wound healing without enhancing biofilm development in vitro.

Treating infected or chronic wounds burdened with biofilms still is a major challenge in medical care. Healing-stimulating factors lose their efficacy due to bacterial degradation, and antimicrobial substances negatively affect dermal cells. Therefore, alternative treatment approaches like the pulsed low intensity laser therapy (LILT) require consideration. The effect of pulsed LILT (904nm, in three frequencies) on relevant human cells of the wound healing process (fibroblasts (BJ), keratinocytes (HaCaT), endothelial cells (HMEC), monocytes (THP-1)) were investigated in in-vitro and ex-vivo wound models with respect to viability, proliferation and migration. Antimicrobial efficacy of the most efficient frequency in cell biological analyses of LILT (3200Hz) was determined in a human biofilm model (lhBIOM). Quantification of bacterial load was evaluated by suspension method and qualitative visualization was performed by scanning electron microscopy (SEM). Pulsed LILT at 904nm at 3200Hz±50% showed the most positive effects on metabolic activity and proliferation of human wound cells in vitro (after 72h - BJ: BPT 0.97±0.05 vs. 0.75±0.04 (p=0.0283); HaCaT: BPT 0.79±0.04 vs. 0.59±0.02 (p=0.0106); HMEC: 0.74±0.02 vs. 0.52±0.04 (p=0.009); THP-1: 0.58±0.01 vs. 0.64±0.01 (p>0.05) and ex vivo. Interestingly, re-epithelialization was stimulated in a frequency-independent manner. The inhibition of metabolic activity after TNF-α application was abolished after laser treatment. No impact of LILT on monocytes was detected. Likewise, the tested LILT regimens showed no growth rate reducing effects on three bacterial strains (after 72h - PA: -1.03%; SA: -0.02%; EF: -1,89%) and one fungal (-2.06%) biofilm producing species compared to the respective untreated control. Accordingly, no significant morphological changes of the biofilms were observed after LILT treatment in the SEM. Frequent application of LILT (904nm, 3200Hz) seems to be beneficial for the metabolism of human dermal cells during wound healing. Considering this, the lack of disturbance of the behavior of the immune cells and no growth-inducing effect on bacteria and fungi in the biofilm can be assigned as rather positive. Based on this combined mode of action, LILT may be an option for hard to heal wounds infected with persistent biofilms.

Growth and regeneration of sympathetic neuron axons induced by pulsed low intensity ultrasound

Over a quarter million Americans are currently living with spinal cord injuries (SCIs), costing $3 billion a year to manage [1]. SCI results in myelin loss, axonal degradation, and death of sympathetic neurons, leading to lesion formation and disruption of the primary communication pathway in the spinal cord. Low intensity ultrasound has been shown to improve axonal regeneration in the peripheral nervous system, but its effect on cells in the central nervous systems is not yet studied. The objective of this work is to investigate the growth-stimulating effects of low to mid-intensity ultrasound on axons of sympathetic neurons. To find optimal ultrasound intensities, neuroendocrine-type PC-12 cells were cultured and treated with pulsed ultrasound at varying intensities. Cells were cultured in deprived states to model a post-SCI environment, while being treated with neurotrophic factor (NGF) to induce their neuron differentiation. Images of cells were analyzed for number of cells and neurite outgrowth. Experiments with PC-12 cells found that an ultrasound intensity of 1.02 W/cm2 led to optimal neurite outgrowth when analyzing the rate of neurite outgrowth and total neurite outgrowth at the final day of experimentation. Our ultimate goal is to develop an effective, noninvasive SCI treatment based on the ultrasound technology.

Blood-brain barrier opening with low intensity pulsed ultrasound for immune modulation and immune therapeutic delivery to CNS tumors.

Opening of the blood-brain barrier (BBB) by pulsed low intensity ultrasound has been developed during the last decade and is now recognized as a safe technique to transiently and repeatedly open the BBB. This non- or minimally invasive technique allows for a targeted and uniform dispersal of a wide range of therapeutic substances throughout the brain, including immune cells and antibodies. In this review article, we summarize pre-clinical studies that have used BBB-opening by pulsed low intensity ultrasound to enhance the delivery of immune therapeutics and effector cell populations, as well as several recent clinical studies that have been initiated. Based on this analysis, we propose immune therapeutic strategies that are most likely to benefit from this strategy. The literature review and trial data research were performed using Medline/Pubmed databases and clinical trial registry www.clinicaltrials.gov . The reference lists of all included articles were searched for additional studies. A wide range of immune therapeutic agents, including small molecular weight drugs, antibodies or NK cells, have been safely and efficiently delivered to the brain with pulsed low intensity ultrasound in preclinical models, and both tumor control and increased survival have been demonstrated in different types of brain tumor models in rodents. Ultrasound-induced BBB disruption may also stimulate innate and cellular immune responses. Ultrasound BBB opening has just recently entered clinical trials with encouraging results, and the association of this strategy with immune therapeutics creates a new field of brain tumor treatment.

Above-threshold ionization driven by few-cycle spatially bounded inhomogeneous laser fields

In this work, we study the main features of the photoelectrons generated when noble gas atoms are driven by spatially bounded inhomogeneous strong laser fields. These spatial inhomogeneous oscillating fields, employed to ionize and accelerate the electrons, result from the interaction between a pulsed low intensity laser and bow-tie shaped gold nanostructures. Under this excitation scheme, energy-resolved above-threshold ionization (ATI) photoelectron spectra have been simulated by solving the one-dimensional (1D) time-dependent Schrödinger equation (TDSE) within the single active electron (SAE) approximation. These quantum mechanical results are supported by their classical counterparts, obtained by the numerical integration of the Newton–Lorentz equation. By using near-infrared wavelengths (0.8–3 μm) sources, our results show that very high energetic electrons (with kinetic energies in the keV domain) can be generated, far exceeding the limits obtained by using conventional, spatially homogeneous fields. This new characteristic can be supported considering the non-recombining electrons trajectories, already reported by Neyra and coworkers (Neyra E, et al 2018 J. Opt. 20, 034002). In order to build a real representation of the spatial dependence of the plasmonic-enhanced field in an analytic function, we fit the generated ’actual’ field using two Gaussian functions. We have further analyzed and explored this plasmonic-modified ATI phenomenon in a model argon atom by using several driven wavelengths at intensities in the order of 1014 W cm−2. Throughout our contribution we carefully scrutinize the differences between the ATI obtained using spatially homogeneous and inhomogeneous laser fields. We present the various physical origins, or correspondingly distinct physical mechanisms, for the ATI generation driven by spatially bounded inhomogeneous fields.

Pulsed high intensity laser versus low intensity laser on healing of full thickness wound in diabetic rats (histological and immunohistochemical study)

Pulsed high intensity laser versus low intensity laser on healing of full thickness wound in diabetic rats (histological and immunohistochemical study)

THE EFFECT OF LOW INTENSITY ULTRASOUND THERAPY ON OSSEOINTEGRATION OF TITANIUM DENTAL IMPLANT PLACED IN THE ESTHETIC ZONE

Background: Endosseous dental implants may be considered as the most popular treatment option for partially and fully edentulous patients. It is generally accepted that implant success is primarily dependent upon or achieved by osseointegration, a direct contact between the implant surfaces and living bone and gum. For decrease the time of osseointegration there is a different modalities used as altering the surface and/or shape of the implant has been frequently researched and it has been shown that rough implant surfaces allow a higher percentage of bone-to-implant contact compared to implants with smooth surfaces. Also dual acid etching of titanium, engineering of dental pulp cells on various implant surfaces and biomimetic implant coatings containing bone morphogenetic protein-2. Other methods studied to enhance endogenous bone healing around biomaterials are different forms of biophysical stimulations such as pulsed electromagnetic fields and low intensity pulsed ultrasounds (LIPU). Ultrasound is a form of energy that is transmitted through biological tissues as high-frequency acoustic waves, which is widely used in medicine as a diagnostic, therapeutic and operative tool. The aim of the present study was to evaluate The effect of low-intensity pulsed ultrasound on the osseointegration of titanium dental implants .Patient and methods: Ten implants were placed in ten patients each one receives one implant. The patients were 6 males and 4 females ranged in ages from 25 to 40 years. All the implants were placed in maxilla. Observations were made postoperatively at time of implant placement then after one month, 3 months, 6 months and one year follow up periods for crestal bone loss and stability, Data analysis was performed in several steps. Initially, descriptive statistics for each case results. One way ANOVA followed by pair-wise Tukey post-hoc tests were performed to detect significance between preparation cases. Statistical analysis was performed using Graph-Pad Prism-4 statistics software for Windows. P values ≤0.05 are considered to be statistically significant in all tests.Conclusion: From the a aforementioned review it became clear that Low-intensity pulsed ultrasound could be accelerate the osteointegration of dental implant .

Sonosensitive theranostic liposomes for preclinical in vivo MRI-guided visualization of doxorubicin release stimulated by pulsed low intensity non-focused ultrasound

The main goal of this study was to assess the theranostic performance of a nanomedicine able to generate MRI contrast as a response to the release from liposomes of the antitumor drug Doxorubicin triggered by the local exposure to pulsed low intensity non focused ultrasounds (pLINFU). In vitro experiments showed that Gadoteridol was an excellent imaging agent for probing the release of Doxorubicin following pLINFU stimulation. On this basis, the theranostic system was investigated in vivo on a syngeneic murine model of TS/A breast cancer. MRI offered an excellent guidance for monitoring the pLINFU-stimulated release of the drug. Moreover, it provided: i) an in vivo proof of the effective release of the liposomal content, and ii) a confirmation of the therapeutic benefits of the overall protocol. Ex vivo fluorescence microscopy indicated that the good therapeutic outcome was originated from a better diffusion of the drug in the tumor following the pLINFU stimulus. Very interestingly, the broad diffusion of the drug in the tumor stroma appeared to be mediated by the presence of the liposomes themselves. The results of this study highlighted either the great potential of US-based stimuli to safely trigger the release of a drug from its nanocarrier or the associated significant therapeutic improvement. Finally, MRI demonstrated to be a valuable technique to support chemotherapy and monitoring the outcome. Furthermore, in this specific case, the theranostic agent developed has a high clinical translatability because the MRI agent utilized is already approved for human use.

Radiodensitometric Assessment of the Effect of Pulsed Electromagnetic Field Stimulation Versus Low Intensity Laser Irradiation on Mandibular Fracture Repair: A Preliminary Clinical Trial.

Closed reduction of mandibular fractures usually entails a relatively long period of immobilization, with the subsequent delay of rehabilitation. Therefore, shorter immobilization period with various approaches to protect or enhance bone healing have been investigated. The aim of this study was to analyze the effects of pulsed electromagnetic field (PEMF) and low intensity laser irradiation (LILI) on the fracture healing process, through radiodensitometric assessment of the bone callus. Eighteen patients with mandibular fractures at the tooth bearing area participated in this prospective study. They were treated by closed reduction using maxillo-mandibular fixation (MMF) and were consecutively assigned into 1 of 3 groups. In group A, the fracture sites were exposed to PEMF for 2h daily for 12days. In group B, the fracture sites were exposed to LILI on the tenth and twelfth postoperative days (2 sessions of 6min per day 2h apart). The fracture sites in group C acted as controls. MMF was maintained for 2weeks in group A and 4weeks in groups B and C. The bone fracture healing was evaluated clinically by investigating the union of the fractured segments and radiographically using computerized densitometry. The union of the fractured segments was tested by manual manipulation and the occlusion was assessed upon removal of MMF. Standardized digital panoramic radiographs were performed for each patient, immediately postoperatively as well as at 2 and 4weeks. The digital images were manipulated using the IDRISI software. A rectangular area of 10×15mm was drawn along the center of the fracture line. The obtained densitometry values were expressed in gray levels from 0 to 256. The collected data were then tabulated and statistically analyzed. After releasing the MMF, the bimanual mobility test of the fractured segments in all patients showed stability of the segments. The preinjury occlusion was maintained in all patients. The postoperative radiographs of all patients revealed good bony alignment of the bony segments. In all groups, comparison between the study intervals with respect to both means and changes percentages of the bone density values showed insignificant differences. At 2nd postoperative week, the mean bone density at the fracture sites decreased by 4.74, 6.6 and 27.89% in groups A, B and C respectively. The period from the 2nd to the 4th postoperative weeks showed increase in the bone density by 1.49, 1.95 and 14.12% in groups A, B and C respectively. Insignificant difference was found between the means of bone densities of group A and B throughout the study intervals. On the other hand, both groups showed insignificant difference with group C immediately postoperative and significant increase in bone density at the 2nd and 4th postoperative weeks. Short period immobilization of mandibular fractures for 2weeks supplemented with PEMF is recommended. Further studies are needed to evaluate the efficacy of LILI as a supplement to reduce the mandibular fracture immobilization period.

Pulsed-low intensity ultrasound enhances extracellular matrix production by fibroblasts encapsulated in alginate

In this study, the effect of pulsed-low intensity ultrasound on cell proliferation, collagen production and glycosaminoglycan deposition by 3T3 fibroblasts encapsulated in alginate was evaluated. Hoechst 33258 assay for cell number, hydroxyproline assay for collagen content and dimethylamine blue assay for glycosaminoglycan content were performed on samples from cell cultures treated with pulsed-low intensity ultrasound and a control group. Pulsed-low intensity ultrasound shows no effect on cell proliferation, while collagen and glycosaminoglycan contents were consistently higher in the samples treated with pulsed-low intensity ultrasound, showing a statistically significant difference (p < 0.05) on day 10. Alcian blue staining showed that glycosaminoglycans were deposited around the cells in both groups. These results suggest that pulsed-low intensity ultrasound shows no effect on cell proliferation but has potential for inducing collagen and glycosaminoglycan production in cells cultured in alginate gels.

The potential of pulsed low intensity ultrasound to stimulate chondrocytes matrix synthesis in agarose and monolayer cultures.

Pulsed low intensity ultrasound (PLIUS) has been used successfully for bone fracture repair and has therefore been suggested for cartilage regeneration. However, previous in vitro studies with chondrocytes show conflicting results as to the effect of PLIUS on the elaboration of extracellular matrix. This study tests the hypothesis that PLIUS, applied for 20 min/day, stimulates the synthesis of sulphated glycosaminoglycan (sGAG) by adult bovine articular chondrocytes cultured in either monolayer or agarose constructs. For both culture models, PLIUS at either 30 or 100 mW/cm2 intensity had no net effect on the total sGAG content. Although PLIUS at 100 mW/cm2 did induce a 20% increase in sGAG content at day 2 of culture in agarose, this response was lost by day 5. Intensities of 200 and 300 mW/cm2 resulted in cell death probably due to heating from the ultrasound transducers. The lack of a sustained up-regulation of sGAG synthesis may reflect the suggestion that PLIUS only induces a stimulatory effect in the presence of a tissue injury response. These results suggest that PLIUS has a limited potential to provide an effective method of stimulating matrix production as part of a tissue engineering strategy for cartilage repair.

Arthroscopic Ankle Arthrodesis: Effect of Pulsed Low Intensity Ultrasound (SS-68)

During the last decade, arthroscopic ankle arthrodesis has improved union rate and decreased union time as compared to open arthrodesis. However, there are still a small but significant number of patients who develop nonunion and require additional procedures. This prospective randomized study was designed to investigate whether postoperative use of non-invasive Pulsed Low Intensity Ultrasound results in further enhancement of arthroscopic ankle arthrodesis rate and/or decrease in the arthrodesis time.

Exposure to Pulsed Low Intensity Ultrasound Stimulates Extracellular Matrix Metabolism of Bovine Intervertebral Disc Cells Cultured in Alginate Beads

In vitro study on the effects of pulsed low intensity ultrasound on the cellular metabolism of bovine intervertebral disc cells. To determine whether pulsed low intensity ultrasound has effects on cell proliferation and extracellular matrix metabolism by bovine intervertebral disc cells. The application of pulsed low intensity ultrasound is known to be effective in stimulating fracture and cartilage repair. However, the effects of pulsed low intensity ultrasound on intervertebral disc cells are not known. Cells of the nucleus pulposus and inner and outer anulus fibrosus were enzymatically isolated from bovine coccygeal tissue and precultured in alginate beads for 14 days. In the ultrasound group, pulsed low intensity ultrasound was administered to the culture for 20 minutes daily for an additional 20 days. The control group was cultured in the same way but without administration of ultrasound. Cell viability, DNA content, proteoglycan and collagen synthesis, and proteoglycan content at days 10 and 20 after the initiation of treatment were evaluated. Characterization of newly synthesized collagen and proteoglycan was performed. No significant differences in cell viability and DNA content were observed between the two groups. On day 20, proteoglycan synthesis was increased by the application of pulsed low intensity ultrasound in nucleus pulposus and inner and outer anulus fibrosus cells (24%-26% increase, P < 0.001). The application of pulsed low intensity ultrasound increased proteoglycan content in alginate beads containing inner and outer anulus fibrosus cells (P < 0.05). Collagen synthesis by cells isolated from all three zones of the intervertebral disc was increased by the application of pulsed low intensity ultrasound (16%-19% increase, P < 0.05-0.0001). The application of pulsed low intensity ultrasound stimulated extracellular matrix metabolism in intervertebral disc cells. Pulsed low intensity ultrasound may prove useful for the physical stimulation of cell metabolism for tissue engineering of intervertebral disc tissue.

Preliminary Results of Tibial Bone Transports With Pulsed Low Intensity Ultrasound (Exogen???)

This study assessed whether the use of noninvasive, pulsed low intensity ultrasound (Exogen) reduced external fixation time for bone transport patients with large, segmental tibial defects. This was a prospective study compared to a previously treated control group. All surgeries and postoperative care were done at the senior author's hospital facilities. Eight patients with large tibial segmental defects, acute or chronic, were studied. All patients were male, mean age 34 years (range 18-50). All patients were treated with a single-level bone transport. Bone transports started 5 to 10 days after a metaphyseal corticotomy and progressed with one-quarter millimeter advancement 2 to 4 times per day. Exogen was applied to the regenerate site in the second postoperative week and to the docking site after docking was complete. The treatment was used for 20 minutes daily at each site. The patients were followed weekly for the first 4 weeks. They were then followed bimonthly for 2 months and then monthly until consolidation had occurred. The 2 main outcome measures used were the external fixation time measured in months and the external fixation index (time in the frame per cm of bone transported) measured in months per cm. The mean follow-up from frame removal to the time of the last clinic visit was 12.4 months (range 4-32). The mean external fixation time was 13.91. The mean external fixation index was 1.34 months per cm transported for the Exogen group and 2.02 months per cm for the control group. Although not statistically significant because of small numbers, the external fixation index was reduced by 17.21%. These data further confirm already published data on the acceleration of fracture healing by low intensity pulsed ultrasound and, in our case, regenerate consolidation. Shorter treatment periods reduce complications and expenses and return patients back to regular activities more quickly.

Current trends in the enhancement of biomaterial osteointegration: biophysical stimulation.

To enhance bone implant osteointegration, many strategies for improving biomaterial properties have been developed which include optimization of implant material, implant design, surface morphology and osteogenetic coatings. Other methods that have been attempted to enhance endogenous bone healing around biomaterials are different forms of biophysical stimulations such as pulsed electromagnetic fields (PEMFs) and low intensity pulsed ultrasounds (LIPUS), which were initially developed to accelerate fracture healing. To aid in the use of adjuvant biophysical therapies in the management of bone-implant osteointegration, the present authors reviewed experimental and clinical studies published in the literature over the last 20 years on the combined use of biomaterials and PEMFs or LIPUS, and summarized the methodology, and the possible mechanism of action and effectiveness of the different biophysical stimulations for the enhancement of bone healing processes around bone implanted biomaterials.

Biological Effects of Low Intensity Ultrasound: The Mechanism Involved, and its Implications on Therapy and on Biosafety of Ultrasound

The biological effects of low intensity ultrasound (US) in vitro; the mechanisms involved; and the factors that can enhance or inhibit these effects are reviewed. The lowest possible US intensities required to induce cell killing or to produce free radicals were determined. Following sonication in the region of these intensities, the effects of US in combination with either hyperthermia, hypotonia, echo-contrast agents (ECA), CO2, incubation time, high cell density or various agents were examined. The results showed that hyperthermia, hypotonia and microbubbles are good enhancers of the bioeffects, while CO2, incubation time and high cell density are good inhibitors. Cellular membrane damage is pivotal in the events leading to cell death, with the cellular damage-and-repair mechanism as an important determinant of the fate of the damaged cells. The optimal level of apoptosis (with minimal lysis) and optimal gene transfection efficiency were attained using a pulsed low intensity US. In summary, the findings suggest that low intensity US is potentially useful in therapy, while on the other hand, they also call for further investigation of such clinical scenarios as high-grade fever, edema or use of ECA which may lead to the lowering of the threshold for bioeffects with diagnostic US.