Publications by authors named "Yumi Kawahara"

39 Publications

Osteoblastic adherence regulates hematopoietic stem cell self-renewal and differentiation: a conceptional and study.

Stem Cell Investig 2021 11;8:21. Epub 2021 Oct 11.

Department of Pediatrics, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima, Japan.

Background: Intrinsic factors related to self-renewal regulatory factors in hematopoietic stem cells are well known; however, limited information is available on extrinsic factors, such as the cell environment. Therefore, in this study, we analyzed the regulatory mechanism of hematopoietic stem cell self-renewal, focusing on the osteoblastic niche, and examined how adherence to osteoblasts affects stem cell differentiation.

Methods: For this experimental study, we developed a co-culture system for hematopoietic stem cells and osteoblasts, such that cells adhered to osteoblasts can be separated from those that do not. Murine Sca1-positive cells were separated into groups according to whether they were attached to osteoblasts or detached from osteoblasts, and each group was then subjected to colony assays and bone marrow transplantation experiments.

Results: Adhered Sca1-positive cells developed more secondary colonies than non-adhered Sca1-positive cells. Furthermore, in bone marrow transplantation experiments, adhered Sca1-positive cells showed successful engraftment. We explored the role of Polycomb genes in the regulation of cell fate and found that self-renewing cells attached to osteoblasts had high expression and low expression, while this expression was reversed in differentiating cells.

Conclusions: Our results suggest that hematopoietic stem cells self-renew when they remain in osteoblastic niches after cell division. Further, when stem cells leave the niches, they undergo differentiation.
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http://dx.doi.org/10.21037/sci-2021-019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8578737PMC
October 2021

Nutritional intervention after an early assessment by a flexible endoscopic evaluation of swallowing is associated with a shorter hospital stay for patients with acute cerebral infarction: A retrospective study.

Asia Pac J Clin Nutr 2021 Jun;30(2):199-205

Nutrition Support Team, National Hospital Organization Takasaki General Medical Center, Takasaki, Gunma, Japan. Email:

Background And Objectives: It is important to evaluate the swallowing function of patients with acute cerebral infarction. The effects of nutritional intervention after an early assessment by a flexible endoscopic evaluation of swallowing (FEES) were evaluated.

Methods And Study Design: This retrospective study included 274 patients who were hospitalized for acute cerebral infarction and underwent a FEES between 2016 and 2018. The effects of early nutritional intervention after an assessment by a FEES within 48 h from admission were evaluated. The patients were divided into a shorter hospital stay group (<30 days) and a longer group (≥30 days). A multivariate analysis was performed to identify the predictive factors for a shorter hospital stay.

Results: The overall patient characteristics were as follows: 166 men; median age, 81 years old; and median body mass index (BMI), 21.1 kg/m2. No significant differences in the age, sex, or BMI were found between the shorter and longer hospital stay groups. A FEES within 48 h of admission (odds ratio [OR], 2.040; 95% confidence interval [CI], 1.120-3.700; p=0.019), FILS level ≥6 at admission (OR, 2.300; 95% CI, 1.190-4.440; p=0.013), and an administered energy dose of ≥18.5 kcal/kg on hospital day 3 (OR, 2.360; 95% CI, 1.180-4.690; p=0.015) were independently associated with a hospital stay <30 days.

Conclusions: Patients with acute cerebral infarction are more likely to have a shorter hospital stay (<30 days) if they undergo a FEES early after admission and receive optimal nutritional intervention.
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http://dx.doi.org/10.6133/apjcn.202106_30(2).0003DOI Listing
June 2021

Comparisons of Neurotrophic Effects of Mesenchymal Stem Cells Derived from Different Tissues on Chronic Spinal Cord Injury Rats.

Stem Cells Dev 2021 Sep 27;30(17):865-875. Epub 2021 Jul 27.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

Cell-based therapies with mesenchymal stem cells (MSCs) are considered as promising strategies for spinal cord injury (SCI). MSCs have unique characteristics due to differences in the derived tissues. However, relatively few studies have focused on differences in the therapeutic effects of MSCs derived from different tissues. In this study, the therapeutic effects of adipose tissue-derived MSCs, bone marrow-derived MSCs, and cranial bone-derived MSCs (cMSCs) on chronic SCI model rats were compared. MSCs were established from the collected adipose tissue, bone marrow, and cranial bone. Neurotrophic factor expression of each MSC type was analyzed by real-time PCR. SCI rats were established using the weight-drop method and transplanted intravenously with MSCs at 4 weeks after SCI. Hindlimb motor function was evaluated from before injury to 4 weeks after transplantation. Endogenous neurotrophic factor and neural repair factor expression in spinal cord (SC) tissue were examined by real-time PCR and western blot analyses. Although there were no differences in the expression levels of cell surface markers and multipotency, expression of , , and () was relatively higher in cMSCs. Transplantation of cMSCs improved motor function of chronic SCI model rats. Although there was no difference in the degree of engraftment of transplanted cells in the injured SC tissue, transplantation of cMSCs enhanced , , and messenger RNA expression and synaptophysin protein expression in injured SC tissue. As compared with MSCs derived other tissues, cMSCs highly express many neurotrophic factors, which improved motor function in chronic SCI model rats by promoting endogenous neurotrophic and neural plasticity factors. These results demonstrate the efficacy of cMSCs in cell-based therapy for chronic SCI.
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http://dx.doi.org/10.1089/scd.2021.0070DOI Listing
September 2021

Efficient preservation of sprouting vegetables under simulated microgravity conditions.

PLoS One 2020 15;15(10):e0240809. Epub 2020 Oct 15.

Space Bio-Laboratories Co., Ltd., Hiroshima, Japan.

The effectiveness of a simulated microgravity environment as a novel method for preserving the freshness of vegetables was investigated. Three types of vegetables were selected: vegetable soybean, mung bean sprouts, and white radish sprouts. These selected vegetables were fixed on a three-dimensional rotary gravity controller, rotated slowly. The selected vegetables were stored at 25°C and 66% of relative humidity for 9, 6, or 5 d while undergoing this process. The simulated microgravity was controlled utilizing a gravity controller around 0 m s-2. The mung bean sprouts stored for 6 d under simulated microgravity conditions maintained higher thickness levels than the vegetable samples stored under normal gravity conditions (9.8 m s-2) for the same duration. The mass of all three items decreased with time without regard to the gravity environment, though the samples stored within the simulated microgravity environment displayed significant mass retention on and after 3 d for mung bean sprout samples and 1 d for white radish sprout samples. In contrast, the mass retention effect was not observed in the vegetable soybean samples. Hence, it was confirmed that the mass retention effect of microgravity was limited to sprout vegetables. As a result of analysis harnessing a mathematical model, assuming that the majority of the mass loss is due to moisture loss, a significant difference in mass reduction coefficient occurs among mung bean sprouts and white radish sprouts due to the microgravity environment, and the mass retention effect of simulated microgravity is quantitatively evaluated utilizing mathematical models. Simulated microgravity, which varies significantly from conventional refrigeration, ethylene control, and modified atmosphere, was demonstrated effective as a novel method for preserving and maintaining the freshness of sprout vegetables. This founding will support long-term space flight missions by prolonging shelf life of sprout vegetables.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0240809PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7561153PMC
December 2020

Early Transplantation of Human Cranial Bone-derived Mesenchymal Stem Cells Enhances Functional Recovery in Ischemic Stroke Model Rats.

Neurol Med Chir (Tokyo) 2020 Feb 18;60(2):83-93. Epub 2020 Jan 18.

Department of Neurosurgery, Graduate School of Biomedical and Health Sciences, Hiroshima University.

We analyzed the cell characteristics, neuroprotective, and transplantation effects of human cranial bone-derived mesenchymal stem cells (hcMSCs) in ischemic stroke model rats compared with human iliac bone-derived mesenchymal stem cells (hiMSCs). The expressions of brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF ) as neurotrophic factors were analyzed in both MSCs. hiMSCs or hcMSCs were intravenously administered into ischemic stroke model rats at 3 or 24 h after middle cerebral artery occlusion (MCAO) and neurological function was evaluated. The survival rate of neuroblastoma × glioma hybrid cells (NG108-15) after 3 or 24 h oxidative or inflammatory stress and the neuroprotective effects of hiMSCs or hcMSCs-conditioned medium (CM) on 3 or 24 h oxidative or inflammatory stress-exposed NG108-15 cells were analyzed. The expressions of BDNF and VEGF were higher in hcMSCs than in hiMSCs. hcMSCs transplantation at 3 h after MCAO resulted in significant functional recovery compared with that in the hiMSCs or control group. The survival rate of stress-exposed NG108-15 was lower after 24 h stress than after 3 h stress. The survival rates of NG108-15 cells cultured with hcMSCs-CM after 3 h oxidative or inflammatory stress were significantly higher than in the control group. Our results suggest that hcMSCs transplantation in the early stage of ischemic stroke suppresses the damage of residual nerve cells and leads to functional recovery through the strong expressions of neurotrophic factors. This is the first report demonstrating a functional recovery effect after ischemic stroke following hcMSCs transplantation.
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http://dx.doi.org/10.2176/nmc.oa.2019-0186DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7040434PMC
February 2020

"Microgravity" as a unique and useful stem cell culture environment for cell-based therapy.

Regen Ther 2019 Dec 22;12:2-5. Epub 2019 Apr 22.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

Cell-based therapy using mesenchymal stem cells or pluripotent stem cells such as induced pluripotent stem cells has seen dramatic progress in recent years. Part of cell-based therapy are already covered by public medical insurance. Recently, researchers have attempted to improve therapeutic effects toward various diseases by cell transplantation. Culture environment is considered to be one of the most important factors affecting therapeutic effects, in particular factors such as physical stimuli, because cells have the potential to adapt to their surrounding environment. In this review, we provide an overview of the research on the effects of gravity alteration on cell kinetics such as proliferation or differentiation and on potential therapeutic effects, and we also summarize the remarkable possibilities of the use of microgravity culture in cell-based therapy for various diseases.
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http://dx.doi.org/10.1016/j.reth.2019.03.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6933149PMC
December 2019

Kaposi's sarcoma-associated herpesvirus is cell-intrinsically controlled in latency in microgravity.

Virus Res 2020 01 19;276:197821. Epub 2019 Nov 19.

Division of Virology, Department of Microbiology and Immunology, Osaka University Graduate School of Medicine, Suita, Osaka 565-0871, Japan.

In the next several decades, humans will explore deep space, including Mars. During long-term space flight, astronauts will be exposed to various physical stressors. Among these stressors, microgravity may compromise the immune system. Consistently, the reactivation of several latent herpesviruses has been reported in astronauts. Although herpesvirus infection status is determined by both cell-intrinsic and -extrinsic factors, it remains unclear which factors play major roles in the virus reactivation in microgravity. Here, using Kaposi's sarcoma-associated herpesvirus (KSHV)-infected cells, we found that KSHV is cell-intrinsically controlled in latency in microgravity. Innate immunity appeared to be unaffected in microgravity, while the expression of some restriction factors against KSHV, such as CTCF and AMPK, was upregulated. Collectively, the infected cells in microgravity can control KSHV in latency, possibly by unimpaired innate immunity and upregulated KSHV restriction factors. This is the first pilot study of the conflicts between cell-intrinsic defense systems and viruses in microgravity and provides fundamental information regarding host-virus interactions in microgravity.
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http://dx.doi.org/10.1016/j.virusres.2019.197821DOI Listing
January 2020

Maintenance of Neurogenic Differentiation Potential in Passaged Bone Marrow-Derived Human Mesenchymal Stem Cells Under Simulated Microgravity Conditions.

Stem Cells Dev 2019 12 11;28(23):1552-1561. Epub 2019 Nov 11.

Department of Biotechnology, Graduate School of Engineering, Osaka University, Osaka, Japan.

Human mesenchymal stem cells (hMSCs) are considered to be able to adapt to environmental changes induced by gravity during cell expansion. In this study, we investigated neurogenic differentiation potential of passaged hMSCs under conventional gravity and simulated microgravity conditions. Immunostaining, quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR), and western blot analysis of neurogenic differentiation markers, neurofilament heavy (NF-H), and microtubule-associated protein 2 (MAP2) revealed that differentiated cells from the cells cultured under simulated microgravity conditions expressed higher neurogenic levels than those from conventional gravity conditions. The levels of NF-H and MAP2 in the cells from simulated microgravity conditions were consistent during passage culture, whereas cells from conventional gravity conditions exhibited a reduction of the neurogenic levels against an increase of their passage number. In growth culture, cells under simulated microgravity conditions showed less apical stress fibers over their nucleus with fewer cells having a polarization of lamin A/C than those under conventional gravity conditions. The ratio of lamin A/C to lamin B expression in the cells under simulated microgravity conditions was constant; however, cells cultured under conventional gravity conditions showed an increase in the lamin ratio during passages. Furthermore, analysis of activating H3K4me3 and repressive H3K27me3 modifications at promoters of neuronal lineage genes indicated that cells passaged under simulated microgravity conditions sustained the methylation during serial cultivation. Nevertheless, the enrichment of H3K27me3 significantly increased in the passaged cells cultured under conventional gravity conditions. These results demonstrated that simulated microgravity-coordinated cytoskeleton-lamin reorganization leads to suppression of histone modification associated with neurogenic differentiation capacity of passaged hMSCs.
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http://dx.doi.org/10.1089/scd.2019.0146DOI Listing
December 2019

Alterations in Nuclear Lamina and the Cytoskeleton of Bone Marrow-Derived Human Mesenchymal Stem Cells Cultured Under Simulated Microgravity Conditions.

Stem Cells Dev 2019 09 17;28(17):1167-1176. Epub 2019 Jul 17.

Department of Biotechnology, Graduate School of Engineering, Osaka University, Osaka, Japan.

Cells sense and respond to environmental changes induced by gravity. Although reactions to conventional culture have been intensively studied, little is known about the cellular reaction to simulated microgravity conditions. Thus, in this study, we investigated the effects of simulated microgravity on human mesenchymal stem cells using a three-dimensional clinostat (Gravite), a recently developed device used to generate simulated microgravity condition in vitro. Our time-lapse analysis shows that cells cultured under conventional culture conditions have a stretched morphology and undergo unidirectional migration, whereas cells cultured under simulated microgravity conditions undergo multidirectional migration with directional changes of cell movement. Furthermore, cells cultured under conventional culture conditions maintained their spindle shape through fibronectin fibril formation in their bodies and focal adhesion stabilization with enriched stress fibers. However, cells cultured under simulated microgravity conditions were partially contracted and the fibril structures were degraded in the cell bodies. Additionally, paxillin phosphorylation in the cells cultured under simulated microgravity conditions was more intense at the cell periphery in regions near the leading and trailing edges, but was less expressed in the cell bodies compared with that observed in cells cultured under conventional culture conditions. Furthermore, lamin A/C, a major component of the nuclear lamina, was mainly located on the apical side in cells cultured under conventional culture conditions, indicating basal-to-apical polarization. However, cells cultured under simulated microgravity conditions showed lamin A/C localization on both the apical and basal sides. Taken together, these results demonstrate that simulated microgravity-driven fibronectin assembly affects nuclear lamina organization through the spatial reorganization of the cytoskeleton.
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http://dx.doi.org/10.1089/scd.2018.0229DOI Listing
September 2019

Erratum: Publisher Correction: Simulated microgravity attenuates myogenic differentiation via epigenetic regulations.

NPJ Microgravity 2018 18;4:15. Epub 2018 Jul 18.

1Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

[This corrects the article DOI: 10.1038/s41526-018-0045-0.].
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http://dx.doi.org/10.1038/s41526-018-0047-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6051999PMC
July 2018

Stem Cell Culture in Microgravity and Its Application in Cell-Based Therapy.

Stem Cells Dev 2018 09 7;27(18):1298-1302. Epub 2018 Aug 7.

1 Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University , Hiroshima, Japan.

Recent years have witnessed a rapid increase in space experiments. Initially, scientists focused on understanding the phenomenon of microgravity to discover countermeasures for preventing the adverse effects of microgravity on the astronauts' bodies. Lately, the application of microgravity environment has been gradually increasing with diverse objectives. Protein crystallization and three-dimensional cell culture are typical examples of microgravity application. Our recent studies suggested that microgravity is a useful tool for cell culture in cell-based therapy. In this review, we discussed microgravity-induced changes at cellular and molecular levels observed in experiments conducted during space flight or using simulated microgravity device. In addition, we summarized the utility of microgravity environment in cell-based therapy for central nervous system diseases.
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http://dx.doi.org/10.1089/scd.2017.0298DOI Listing
September 2018

Simulated microgravity attenuates myogenic differentiation via epigenetic regulations.

NPJ Microgravity 2018 23;4:11. Epub 2018 May 23.

1Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

The molecular mechanisms involved in myogenic differentiation are relatively well-known. Myogenic differentiation is regulated by the sequential activation of the basic helix-loop-helix myogenic regulatory transcription factors (MRFs), and biomechanical signals play an important role in the regulation of myogenesis. In this study, we sought to determine whether simulated microgravity culture using Gravite may affect myoblast differentiation and expression of MRF genes. Although rat myoblasts, L6 cells were differentiated to myotubes in an incubation period-dependent manner, myogenesis of L6 cells was significantly attenuated under simulated microgravity (10G) conditions. Real-time Reverse transcription polymerase chain reaction (RT-PCR) showed that expressions of , , , , and under 1 G conditions increase in an incubation period-dependent manner, and that expression was specifically observed to increase transiently in the early phase. However, expressions of and were significantly inhibited under simulated microgravity conditions. To clarify the molecular mechanisms, L6 cells were treated with 5-AzaC, and further incubated with differentiation medium under 1 G or 10 G conditions. The results showed differences in expression levels of , , and, as well as those of myotube thickness between 1 G and 10 G conditions, completely disappeared in this experimental condition. Modified HpaII tiny fragment enrichment by ligation-mediated PCR (HELP)-assay showed that kinetic changes of DNA methylation status were attenuated in simulated microgravity conditions. These results indicate that microgravity regulates myogenesis and expression by controlling DNA methylation.
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http://dx.doi.org/10.1038/s41526-018-0045-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5966377PMC
May 2018

Simulated Microgravity Culture Enhances the Neuroprotective Effects of Human Cranial Bone-Derived Mesenchymal Stem Cells in Traumatic Brain Injury.

Stem Cells Dev 2018 09 5;27(18):1287-1297. Epub 2018 Jul 5.

1 Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical & Health Sciences, Hiroshima University , Hiroshima, Japan .

Fundamental cures of central nervous system (CNS) diseases are rarely achieved due to the low regenerative ability of the CNS. Recently, cell-based therapy using mesenchymal stem cells (MSCs) has been explored as an effective treatment for CNS diseases. Among the various tissue-derived MSCs, we have isolated human cranial bone-derived MSCs (cMSCs) in our laboratory. In addition, we have focused on simulated microgravity (MG) as a valuable culture environment of MSCs. However, detailed mechanisms underlying functional recovery from transplantation of MSCs cultured under MG conditions remain unclear. In this study, we investigated the therapeutic mechanisms of transplantation of cMSCs cultured under MG conditions in traumatic brain injury (TBI) model mice. Human cMSCs were cultured under 1G and MG conditions, and cMSCs cultured under MG conditions expressed significantly higher messenger RNA (mRNA) levels of hepatocyte growth factor (HGF) and transforming growth factor beta (TGF-β). In TBI model mice, the transplantation of cMSCs cultured under MG conditions (group MG) showed greater motor functional improvement compared with only phosphate-buffered saline administration (group PBS). Moreover, the protein expression levels of tumor necrosis factor alpha (TNF-α) and the Bcl-2-associated X protein (Bax)/b cell leukemia/lymphoma 2 protein (Bcl-2) ratio were significantly lower at brain injury sites in mice of group MG than those of group PBS. In addition, an in vitro study showed that the conditioned medium of cMSCs cultured under MG conditions significantly suppressed the cell death of NG108-15 cells exposed to oxidative or inflammatory stress through anti-inflammatory and antiapoptosis effects. These findings demonstrate that culturing cMSCs under simulated MG increases the neuroprotective effects, suggesting that simulated MG cultures may be a useful method for cell-based therapy strategies for CNS diseases.
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http://dx.doi.org/10.1089/scd.2017.0299DOI Listing
September 2018

Rat Cranial Bone-Derived Mesenchymal Stem Cell Transplantation Promotes Functional Recovery in Ischemic Stroke Model Rats.

Stem Cells Dev 2018 08 29;27(15):1053-1061. Epub 2018 Jun 29.

1 Department of Neurosurgery, Graduate School of Biomedical and Health Sciences, Hiroshima University , Hiroshima, Japan .

The functional disorders caused by central nervous system (CNS) diseases, such as ischemic stroke, are clinically incurable and current treatments have limited effects. Previous studies suggested that cell-based therapy using mesenchymal stem cells (MSCs) exerts therapeutic effects for ischemic stroke. In addition, the characteristics of MSCs may depend on their sources. Among the derived tissues of MSCs, we have focused on cranial bones originating from the neural crest. We previously demonstrated that the neurogenic potential of human cranial bone-derived MSCs (cMSCs) was higher than that of human iliac bone-derived MSCs. Therefore, we presumed that cMSCs have a higher therapeutic potential for CNS diseases. However, the therapeutic effects of cMSCs have not yet been elucidated in detail. In the present study, we aimed to demonstrate the therapeutic effects of transplantation with rat cranial bone-derived MSCs (rcMSCs) in ischemic stroke model rats. The mRNA expression of brain-derived neurotrophic factor and nerve growth factor was significantly stronger in rcMSCs than in rat bone marrow-derived MSCs (rbMSCs). Ischemic stroke model rats in the rcMSC transplantation group showed better functional recovery than those in the no transplantation and rbMSC transplantation groups. Furthermore, in the in vitro study, the conditioned medium of rcMSCs significantly suppressed the death of neuroblastoma × glioma hybrid cells (NG108-15) exposed to oxidative and inflammatory stresses. These results suggest that cMSCs have potential as a candidate cell-based therapy for CNS diseases.
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http://dx.doi.org/10.1089/scd.2018.0022DOI Listing
August 2018

Mesenchymal Stem Cell-Based Therapy Improves Lower Limb Movement After Spinal Cord Ischemia in Rats.

Ann Thorac Surg 2018 05 12;105(5):1523-1530. Epub 2018 Jan 12.

Department of Cardiovascular Surgery, Hiroshima University Hospital, Hiroshima, Japan.

Background: Spinal cord ischemia is a devastating complication after thoracic and thoracoabdominal aortic operations. In this study, we aimed to investigate the effects of mesenchymal stem cells (MSCs), which have regenerative capability and exert paracrine actions on damaged tissues, injected into rat models of spinal cord ischemia-reperfusion injury.

Methods: Forty-five Sprague-Dawley rats were divided into sham, phosphate-buffered saline (PBS), and MSC groups. Spinal cord ischemia was induced in the latter two groups by balloon occlusion of the thoracic aorta. MSCs and PBS were then immediately injected into the left carotid artery of the MSC and PBS groups, respectively. Hindlimb motor function was evaluated at 6 and 24 hours. The spinal cord was removed at 24 hours after ischemia-reperfusion injury, and histologic and immunohistochemical analyses and real-time polymerase chain reaction assessments were performed.

Results: Rats in the MSC and PBS groups showed flaccid paraparesis/paraplegia postoperatively. Hindlimb function was significantly better at 6 and 24 hours after ischemia-reperfusion injury in the MSC group than in the PBS group (p < 0.05). The number of terminal deoxynucleotidyl transferase dUTP nick end labeling-positive neuron cells in the spinal cord and the ratio of Bax to Bcl2 were significantly larger (p < 0.05) in the PBS group than in the MSC group. The injected MSCs were observed in the spinal cord 24 hours after ischemia-reperfusion injury.

Conclusions: The MSC therapy by transarterial injection immediately after spinal cord ischemia-reperfusion injury may improve lower limb function by preventing apoptosis of neuron cells in the spinal cord.
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http://dx.doi.org/10.1016/j.athoracsur.2017.12.014DOI Listing
May 2018

Neuromagnetic evaluation of a communication support system for hearing-impaired patients.

Neuroreport 2017 Aug;28(12):712-719

aDivision of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health SciencesbSpace Regenerative Medical CentercDepartment of Neurosurgery, Graduate School of Biomedical and Health Sciences, Hiroshima UniversitydSpace Bio-Laboratories Co., Ltd., HiroshimaeUniversal Sound Design Inc., Tokyo, Japan.

Hearing-impaired patients often encounter obstacles in communication. Not all of them wear hearing aids, citing issues with usage difficulty and discomfort in wearing. To overcome these difficulties, a new endeavor was started to improve sound intelligibility from the speaker's side. The present study objectively evaluated an intelligible-hearing (IH) loudspeaker by means of magnetoencephalography. Magnetic counterparts of mismatch negativity (MMNm) to pronunciation ('mi' and 'ni') were recorded and compared when they were transmitted from the IH loudspeaker and from a normal-hearing loudspeaker. On using the IH loudspeaker, the peak latency was found to be significantly shortened. In the case of hearing-impaired participants, marked MMNm responses were observed only when the IH loudspeaker was used. These findings suggest that improving sound intelligibility may be a supportive and rehabilitative approach for hearing-impaired patients.
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http://dx.doi.org/10.1097/WNR.0000000000000817DOI Listing
August 2017

The characteristics of human cranial bone marrow mesenchymal stem cells.

Neurosci Lett 2015 Oct 3;606:161-6. Epub 2015 Sep 3.

Department of Neurosurgery, Graduate School of Biomedical and Health Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima City, Hiroshima 734-8551, Japan.

Recently, cell-based therapy has attracted attention for treatment of central nervous system (CNS) disorders. Bone marrow-derived mesenchymal stem cells (BMSCs) are considered to have good engraftment potential. Therefore, more efficient and less invasive methods to obtain donor cells are required. Here, we established human BMSCs from cranial bone waste (cBMSCs) obtained following routine neurosurgical procedures. cBMSCs and cells obtained from the iliac crest (iBMSCs, standard BMSCs) showed expression of cell surface markers associated with mesenchymal stem cells and multipotency traits such as differentiation into osteogenic and adipogenic lineages. cBMSCs showed higher expression of the neural crest-associated mRNAs p75, Slug, and Snail than iBMSCs. Neurogenic induced cells from cBMSCs expressed the neural markers nestin, Pax6, neurofilament (NF)-L, and NF-M as seen with RT-PCR, and NF-M protein as seen with western blotting at higher levels than cells from iBMSCs. Immunostaining showed a significantly greater proportion of NF-M-positive cells in the population of induced cBMSCs compared with the population of iBMSCs. Thus, cBMSCs showed a greater tendency to differentiate into neuron-like cells than iBMSCs.
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http://dx.doi.org/10.1016/j.neulet.2015.08.056DOI Listing
October 2015

The Krüppel-like zinc finger transcription factor, GLI-similar 1, is regulated by hypoxia-inducible factors via non-canonical mechanisms.

Biochem Biophys Res Commun 2013 Nov;441(2):499-506

GLI-similar 1 (GLIS1) is important for the reprogramming of fibroblasts into induced pluripotent stem cells (iPSCs). However, the molecular mechanisms of regulation of GLIS1 expression remain unclear. We have therefore examined GLIS1 expression in various cancer cell lines and demonstrated that GLIS1 expression was dramatically increased under hypoxic conditions. Importantly, GLIS1 expression was significantly attenuated in VHL-overexpressing renal cell carcinoma cells compared to the VHL-deficient parent control. Moreover, promoter analysis demonstrated that GLIS1 transcription was regulated by hypoxia through a hypoxia-inducible factors (HIFs)-dependent mechanism. Co-transfection experiments revealed that HIF-2α had greater potency on the GLIS1 promoter activation than HIF-1α. Subsequent studies using wild-type and mutant HIF-2α demonstrated that DNA binding activity was not necessary but TADs were critical for GLIS1 induction. Finally, co-transfection experiments indicated that HIF-2α cooperated with AP-1 family members in upregulating GLIS1 transcription. These results suggest that the hypoxic signaling pathway may play a pivotal role in regulating the reprogramming factor GLIS1, via non-canonical mechanisms involving partner transcription factor rather than by direct HIF transactivation.
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November 2013

Interactive effects of cell therapy and rehabilitation realize the full potential of neurogenesis in brain injury model.

Neurosci Lett 2013 Oct 13;555:73-8. Epub 2013 Sep 13.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

The therapeutic effect of rehabilitation after cell therapy for brain injury remains unclear. Here, we report the neural stem/progenitor cells transplantation into a brain injury mouse model followed by treadmill exercise training. Among all experimental groups, mice that underwent transplantation and treadmill exercise demonstrated significant functional motor and electrophysiological improvement. Transplanted cells at the brain injury site were observed and differentiated into neurons and astrocytes. Transplanted cells significantly differentiated into neurons in the mice that underwent transplantation and treadmill exercise compared with those treated with only transplantation. Furthermore, the expression of brain-derived neurotrophic factor and growth-associated protein 43 mRNAs were significantly up-regulated in the mice that underwent transplantation and treadmill exercise than in those in other experimental groups during the early recovery stage. These results suggest that rehabilitation after neural stem/progenitor cell transplantation enhances neurogenesis and promotes the recovery of motor function in brain injury model mice.
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http://dx.doi.org/10.1016/j.neulet.2013.09.009DOI Listing
October 2013

Electrical stimulation accelerates neuromuscular junction formation through ADAM19/neuregulin/ErbB signaling in vitro.

Neurosci Lett 2013 Jun 19;545:29-34. Epub 2013 Apr 19.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

The mechanism by which electrical stimulation affects formation of neuromuscular junctions (NMJs) remains unknown. NG108-15, a neural cell line, is commonly used in in vitro co-culture models of myotubes to observe synapse formation; therefore, we employed this model to observe the effects of electrical stimulation on NMJ formation. Initially, L6 cells were differentiated and NG108-15 cells were then added to the same culture dish. After 2 and 3 days of co-culture, the cells were electrically stimulated at 50 V and 0.5 Hz for 0, 5, 30, and 60 min (C, ES5, ES30, and ES60 groups, respectively) and were analyzed after co-culture for 4 days. Immunofluorescence experiments showed significantly increased aggregation of acetylcholine receptors and inhibition of neural outgrowth in the ES30 and ES60 groups. Furthermore, ADAM19 and phospho-ErbB3 were found to be specifically localized in co-cultured NG108-15 cells. Immunoblotting demonstrated that synapsin 1, ADAM19 precursor and its activated form, phospho-ErbB3, and ERK1 protein levels had increased in an electrical stimulation period-dependent manner. Thus, we found that electrical stimulation accelerated NMJ formation, possibly through activation of ADAM19/neuregulin/ErbB signaling in NG108-15 cells.
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http://dx.doi.org/10.1016/j.neulet.2013.04.006DOI Listing
June 2013

Simulated microgravity facilitates cell migration and neuroprotection after bone marrow stromal cell transplantation in spinal cord injury.

Stem Cell Res Ther 2013 Apr 1;4(2):35. Epub 2013 Apr 1.

Introduction: Recently, cell-based therapy has gained significant attention for the treatment of central nervous system diseases. Although bone marrow stromal cells (BMSCs) are considered to have good engraftment potential, challenges due to in vitro culturing, such as a decline in their functional potency, have been reported. Here, we investigated the efficacy of rat BMSCs (rBMSCs) cultured under simulated microgravity conditions, for transplantation into a rat model of spinal cord injury (SCI).

Methods: rBMSCs were cultured under two different conditions: standard gravity (1G) and simulated microgravity attained by using the 3D-clinostat. After 7 days of culture, the rBMSCs were analyzed morphologically, with RT-PCR and immunostaining, and were used for grafting. Adult rats were used for constructing SCI models by using a weight-dropping method and were grouped into three experimental groups for comparison. rBMSCs cultured under 1 g and simulated microgravity were transplanted intravenously immediately after SCI. We evaluated the hindlimb functional improvement for 3 weeks. Tissue repair after SCI was examined by calculating the cavity area ratio and immunohistochemistry.

Results: rBMSCs cultured under simulated microgravity expressed Oct-4 and CXCR4, in contrast to those cultured under 1 g conditions. Therefore, rBMSCs cultured under simulated microgravity were considered to be in an undifferentiated state and thus to possess high migration ability. After transplantation, grafted rBMSCs cultured under microgravity exhibited greater survival at the periphery of the lesion, and the motor functions of the rats that received these grafts improved significantly compared with the rats that received rBMSCs cultured in 1 g. In addition, rBMSCs cultured under microgravity were thought to have greater trophic effects on reestablishment and survival of host spinal neural tissues because cavity formations were reduced, and apoptosis-inhibiting factor expression was high at the periphery of the SCI lesion.

Conclusions: Here we show that transplantation of rBMSCs cultured under simulated microgravity facilitates functional recovery from SCI rather than those cultured under 1 g conditions.
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http://dx.doi.org/10.1186/scrt184DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3706926PMC
April 2013

Electrical stimulation enhances neurogenin2 expression through β-catenin signaling pathway of mouse bone marrow stromal cells and intensifies the effect of cell transplantation on brain injury.

Neurosci Lett 2013 Jan 6;533:71-6. Epub 2012 Nov 6.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

Bone marrow stromal cells (BMSCs) have received significant attention for its use in neural regeneration. However, neural replacement by transplanted BMSCs was not very effective. Recently, the gene transfection method has improved the capability of cell transplantation; however, this method results in canceration and immune rejection. We induced the differentiation of mouse BMSCs into neural cells using electrical stimulation and transplanted the cells into traumatic brain injury (TBI) model mice. We found that the electrically stimulated cells have good potential to differentiate into neural cells and contribute to recovery from TBI without differentiating into astrocytes. In addition, we found that electrical stimulation enhanced neurogenin2 (Ngn2) expression. Ngn2 is involved in neural differentiation and inhibits astrocytic differentiation during cell growth. Furthermore, we found that this enhancement of Ngn2 expression occurred through β-catenin signaling pathway. This study may contribute to the use of BMSCs for neural replacement in central nervous system diseases.
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http://dx.doi.org/10.1016/j.neulet.2012.10.023DOI Listing
January 2013

Influence of dual-task performance on muscle and brain activity.

Int J Rehabil Res 2013 Jun;36(2):127-33

Division of Bio-Environmental Adaptation Sciences, Graduate School of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan.

The simultaneous performance of multiple tasks is often required in daily life. The dual-task paradigm has been used extensively to evaluate the ability to perform simultaneous behavioral tasks. However, relatively few studies have been carried out to determine the muscle and brain activity underlying dual-task performance. This study determined the influence of single-task and dual-task conditions on muscle and brain activity by measuring ankle dorsiflexion, surface electromyography, and magnetoencephalography. Increased muscle activation and variability in the dorsiflexion interval was observed during dual-task performance compared with single-task performance, whereas brain activity in the contralateral motor and frontal areas was reduced. Incidental movement of the non-task-related ankle (contralateral ankle) showed a different pattern of brain activity during the dual-task performance in another experiment. These findings suggest that the activity decreased in several information-processing areas under dual-task conditions as a result of decentralization of attention. Activity in the right and left motor fields may play a role in the dual-task performance, as indicated by changes in brain activity during dual-task performance in two groups classified according to the activity level of the contralateral ankle.
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http://dx.doi.org/10.1097/MRR.0b013e32835acfb8DOI Listing
June 2013

Regulation of hematopoietic stem cells using protein transduction domain-fused Polycomb.

Exp Hematol 2012 Sep 24;40(9):751-760.e1. Epub 2012 May 24.

Department of Pediatrics, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima, Japan.

The Polycomb-group complex is a chromatin regulatory factor that is classified into two different complexes: Polycomb repressive complex 1 and 2. Components of Polycomb repressive complex 1 are involved in the self-renewal of hematopoietic stem cells. Bmi1, one of these components, maintains the immaturity of neural and cancer stem cells as well as that of hematopoietic stem cells. We constructed recombinant protein transduction domain (PTD)-Polycomb proteins and transduced them into murine bone marrow (BM) cells. We designed and fused the PTD-protein transduction domain to three proteins (i.e., green fluorescent protein, Bmi1, and Mel18). Murine BM cells were incubated for 48 h and each PTD-Polycomb protein was added. Then, we analyzed the function of hematopoiesis using the colony assay and transplantation. BM cells exposed to PTD-Bmi1 showed an increased number of colonies. In contrast, BM cells exposed to PTD-Mel18 or to both proteins showed a decreased number of colonies. Hematopoietic cells derived from PTD-Bmi1-transduced BM cells were significantly increased in the peripheral blood at 6 weeks after transplantation. Moreover, 80% of mice transplanted with PTD-Bmi1-transduced BM cells died at 8 to 24 weeks after transplantation. However, only a few early deaths were observed in the mice transplanted with BM cells exposed to both PTD-Bmi1 and PTD-Mel18. We expect that hematopoietic stem cells could proliferate after transduction with PTD-Bmi1, but this may generate undesirable effects, e.g., tumorigenesis. Thus, Bmi1 and Mel18 have opposing functions and are present in distinct complexes.
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http://dx.doi.org/10.1016/j.exphem.2012.05.005DOI Listing
September 2012

Neuromagnetic beta oscillation changes during motor imagery and motor execution of skilled movements.

Neuroreport 2011 Mar;22(5):217-22

Division of Bio-Environmental Adaptation Sciences, Graduate School of Health Sciences, Hiroshima University, Hiroshima, Japan.

We showed the differences in brain activities during motor imagery and motor execution when performing single skilled movements using magnetoencephalography. The tasks included finger tapping and chopstick usage with the dominant or nondominant hand. Chopstick usage with the nondominant hand was an unfamiliar task and required higher skill. Neuromagnetic data were processed by fast Fourier transformation, and β band event-related synchronization was evaluated. Beta oscillation changes were observed in the right and left sensorimotor cortices during both tasks; however, the ipsilateral changes were smaller during motor imagery than during motor execution. These results suggest that motor imagery of skilled movement tasks causes a smaller neuronal burden in the sensorimotor cortex.
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http://dx.doi.org/10.1097/WNR.0b013e328344b480DOI Listing
March 2011

Simulated microgravity maintains the undifferentiated state and enhances the neural repair potential of bone marrow stromal cells.

Stem Cells Dev 2011 May 7;20(5):893-900. Epub 2010 Nov 7.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Health Sciences, Hiroshima University, Hiroshima, Japan.

Recently, regenerative medicine with bone marrow stromal cells (BMSCs) has gained significant attention for the treatment of central nervous system diseases. Here, we investigated the activity of BMSCs under simulated microgravity conditions. Mouse BMSCs (mBMSCs) were isolated from C57BL/6 mice and harvested in 1G condition. Subjects were divided into 4 groups: cultured under simulated microgravity and 1G condition in growth medium and neural differentiation medium. After 7 days of culture, the mBMSCs were used for morphological analysis, reverse transcription (RT)-polymerase chain reaction, immunostaining analysis, and grafting. Neural-induced mBMSCs cultured under 1G conditions exhibited neural differentiation, whereas those cultured under simulated microgravity did not. Moreover, under simulated microgravity conditions, mBMSCs could be cultured in an undifferentiated state. Next, we intravenously injected cells into a mouse model of cerebral contusion. Graft mBMSCs cultured under simulated microgravity exhibited greater survival in the damaged region, and the motor function of the grafted mice improved significantly. mBMSCs cultured under simulated microgravity expressed CXCR4 on their cell membrane. Our study indicates that culturing cells under simulated microgravity enhances their survival rate by maintaining an undifferentiated state of cells, making this a potentially attractive method for culturing donor cells to be used in grafting.
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http://dx.doi.org/10.1089/scd.2010.0294DOI Listing
May 2011

Detrimental effects of microgravity on mouse preimplantation development in vitro.

PLoS One 2009 Aug 25;4(8):e6753. Epub 2009 Aug 25.

Laboratory for Genomic Reprogramming, RIKEN, Center for Developmental Biology, Kobe, Japan.

Sustaining life beyond Earth either on space stations or on other planets will require a clear understanding of how the space environment affects key phases of mammalian reproduction. However, because of the difficulty of doing such experiments in mammals, most studies of reproduction in space have been carried out with other taxa, such as sea urchins, fish, amphibians or birds. Here, we studied the possibility of mammalian fertilization and preimplantation development under microgravity (microG) conditions using a three-dimensional (3D) clinostat, which faithfully simulates 10(-3) G using 3D rotation. Fertilization occurred normally in vitro under microG. However, although we obtained 75 healthy offspring from microG-fertilized and -cultured embryos after transfer to recipient females, the birth rate was lower than among the 1G controls. Immunostaining demonstrated that in vitro culture under microG caused slower development and fewer trophectoderm cells than in 1G controls but did not affect polarization of the blastocyst. These results suggest for the first time that fertilization can occur normally under microG environment in a mammal, but normal preimplantation embryo development might require 1G.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0006753PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2727478PMC
August 2009

Effects of simulated microgravity on proliferation and chemosensitivity in malignant glioma cells.

Neurosci Lett 2009 Sep 21;463(1):54-9. Epub 2009 Jul 21.

Department of Neurosurgery, Graduate School of Biomedical Sciences, Japan.

A three-dimensional (3D) clinostat is a device for generating multidirectional G force, resulting in an environment with an average of 10(-3)G. We cultured human malignant glioma cell lines in a 3D-clinostat (CL group) and examined the growth properties and chemosensitivity of the cells compared to cells cultured under normal 1G conditions (C group). The growth rate was significantly inhibited in the CL group, but without cell cycle change. Mitochondrial activity was also inhibited in the CL group. Thus, inhibition of malignant glioma proliferation occurred that could be attributed to deceleration of mitosis. Chemosensitivity to cisplatin (cis-diamminedichloroplatinum(II), CDDP) in the CL group was significantly enhanced compared to the C group. This method has significant potential as a treatment of malignant gliomas and a tool for understanding developmental biology.
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http://dx.doi.org/10.1016/j.neulet.2009.07.045DOI Listing
September 2009

LIF-free embryonic stem cell culture in simulated microgravity.

PLoS One 2009 Jul 23;4(7):e6343. Epub 2009 Jul 23.

Department of Clinical Neuroscience and Therapeutics, Graduate School of Biomedical Sciences, Hiroshima University, Minami-ku, Hiroshima, Japan.

Background: Leukemia inhibitory factor (LIF) is an indispensable factor for maintaining mouse embryonic stem (ES) cell pluripotency. A feeder layer and serum are also needed to maintain an undifferentiated state, however, such animal derived materials need to be eliminated for clinical applications. Therefore, a more reliable ES cell culture technique is required.

Methodology/principal Findings: We cultured mouse ES cells in simulated microgravity using a 3D-clinostat. We used feeder-free and serum-free media without LIF.

Conclusions/significance: Here we show that simulated microgravity allows novel LIF-free and animal derived material-free culture methods for mouse ES cells.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0006343PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2710515PMC
July 2009

Low-intensity pulsed ultrasound accelerates osteoblast differentiation and promotes bone formation in an osteoporosis rat model.

Pathobiology 2009 May 19;76(3):99-107. Epub 2009 May 19.

Division of Bio-Environmental Adaptation Sciences, Graduate School of Health Sciences, Hiroshima University, Hiroshima, Japan.

Objective: We examined the effects of low-intensity pulsed ultrasound (LIPUS) on cell differentiation, bone mineralized nodule formation and core-binding factor A1 (Cbfa1) expression in a normal human osteoblast (NHOst) cell line and bone formation in an osteoporosis animal model.

Methods: NHOst cells were cultured in vitro in medium with or without LIPUS stimulation. The ultrasound stimulation frequency was 1.0 MHz at an intensity of 30 mW/cm(2) for 20 min. Rats were divided into a sham-operated group (Sham) and an ovariectomized group (OVX). The right femur was treated with LIPUS (Sham-LIPUS and OVX-LIPUS) and the left femur was left untreated (Sham-CON and OVX-CON).

Results: LIPUS stimulation accelerated bone nodule formation and enhanced alkaline phosphatase activity. The expression levels of Cbfa1 decreased and calcification occurred earlier and more frequently in the LIPUS than in the CON groups. The wet weight of the femur increased in OVX rats with LIPUS stimulation. Morphological images showed an increase in trabecular spongiosa in the OVX-LIPUS group.

Conclusion: LIPUS accelerated osteogenesis. Moreover, since LIPUS prevents bone loss, it may be a promising treatment for osteoporosis.
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http://dx.doi.org/10.1159/000209387DOI Listing
May 2009
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