Low-level Laser Irradiation Treatment Research Articles

LLLI promotes BMSC proliferation through circRNA_0001052/miR-124-3p.

Osteoporosis (OP) is a multifactorial bone disease that occurs worldwide. The treatment of OP is still unsatisfactory. Bone mesenchymal stem cell (BMSC) differentiation is a key process in OP pathogenesis. Low-level laser irradiation (LLLI) has been reported to regulate BMSC proliferation, but the role of circRNAs in the LLLI-based promotion of BMSC proliferation remains unclear. CircRNAs are essential molecular regulators that participate in numerous biological processes and have therapeutic potential. miR-124-3p is an essential microRNA (miRNA), and its expression changes are related to BMSC proliferation ability. In the present study, gain-loss function of experiments demonstrated that circRNA_0001052 could regulate the proliferation of BMSCs by acting as a miR-124-3p sponge through the Wnt4/β-catenin pathway. The results of this study strongly suggest that circRNA_0001052 plays an essential role in BMSC proliferation in response to LLLI treatment, which is a potential therapeutic manipulation with clinical applications.

Nrf2 played an important role in radiation protection effect of low-level laser exposed on umbilical cord mesenchymal stem cell

To investigate the protective function of low-level laser irradiation (LLLI) against ionizing irradiation and explore the molecular mechanism of photomodulation of Nrf2 protein, the impact of LLLI (635 nm, 5.7 J/cm2) before 2 Gy gamma ray radiation of radio-sensitive tissue hematopoietic stem cells was evaluated. As a result, reduced levels of reactive oxygen species and increased expression of antioxidant enzymes were detected. Moreover, increased expression of Nrf2 was observed after LLLI, whereas brusatol pretreatment before LLLI abolished this effect. In vivo, transplantation of human umbilical cord mesenchymal stem cells (hUC-MSCs) was employed for therapy of hematopoietic function in an acute radiation sickness (H-ARS) mouse model, which was induced by 6-Gy ionizing irradiation; different hUC-MSC pretreatments including LLLI and Nrf2 RNAi were accounted for during experimental grouping. LLLI treatment of cells significantly increased the erythrocyte count and number of myelopoiesis clones (P < 0.05), but such improvements were reduced by Nrf2 RNAi pretreatment compared with cells transplanted without intervention. Therefore, LLLI may improve the radiation protection effect through molecular mechanisms related to the Nrf2 antioxidant pathway.

Low Level Laser Irradiation Increases Anabolism and Inhibits Extrinsic Apoptosis Pathway in Cultured Chondrocytes with Rectified Chondrogenic Phenotype

Background: Effective clinical treatments of cartilage lesions in defected joints essentially require great number of chondrogenic cells by ex vivo expansion of chondrocytes obtained from biopsy specimen. The conventional passaged chondrocytes provide required cells for treatments but with altered phenotype, which form fibrocartilaginous tissue in articular repairs. Methods: Passaged chondrocytes were treated with inflammatory cytokine TNF-α to induce apoptosis and stimulated by low level laser irradiation (LLLI) to investigate the effect on cellular metabolism using molecular, cellular and microscopic analyses. Findings: LLLI treated chondrocytes show enhanced synthetic features and gene expression pattern, favoring cell growth and reducing apoptosis, particularly through attenuating IL-1β-induced upregulation of mediators of extrinsic apoptosis pathway. The large number of LLLI-treated chondrocytes maintain, for the first time, ameliorated differentiated phenotype simultaneously. Interpretation: The LLLI treatment not only generates large number of viable chondrocytes in culture, potentiating their application in tissue engineering of cell-based therapies for functional repair of cartilage lesions, but also potentiates a modulatory mechanism enhancing chondrogenic properties applicable in therapeutic approaches. Funding Statement: This work was financially supported by the research grants of Guangdong Provincial Department of Science and Technology (2014A020211001 and 2016A050503003), and the National Natural Science Foundation of China (grant No. 61575065). Declaration of Interests: The authors declare that they have no conflicts of interest with the content of this article. Ethics Approval Statement: Three-week-old New Zealand white rabbits were sacrificed with the approval of the governing authority of Guangzhou Red Cross Hospital, Jinan University School of Medicine [SYXK (YUE) 2011-0072] for collection of fresh cartilage samples.

Effect of 650-nm low-level laser irradiation on c-Jun, c-Fos, ICAM-1, and CCL2 expression in experimental periodontitis

This study was designed to investigate the effect of 650-nm low-level laser irradiation (LLLI) as an adjunctive treatment of experimental periodontitis. To investigate possible LLLI-mediated anti-inflammatory effects, we utilized an experimental periodontitis (EP) rat model and analyzed c-Jun, c-Fos, ICAM-1, and CCL2 gene expressions on PB leukocytes and in the gingival tissue. Total RNA was isolated from the gingivae and peripheral blood (PB) leukocytes of normal, EP, scaling, and root planing (SRP)-treated EP and LLLI + SRP-treated EP rats, and gene expressions were analyzed by real-time PCR. The productions of c-Jun, c-Fos, ICAM-1, and CCL2 in gingivae were analyzed immunohistochemically. Tartrate-resistant acid phosphatase (TRAP) staining was used to determine osteoclast activity in alveolar bone. The c-Jun and ICAM-1 messenger RNA (mRNA) levels were significantly decreased in the EP rat gingival tissue treated by SRP + LLLI than by SRP, the c-Jun, ICAM-1, and c-Fos mRNA levels on PB leukocytes reduced after LLLI treatment but did not show any significant differences in both groups. There was no significant difference in CCL2 mRNA levels on PB leukocytes and in gingivae between the SRP + LLLI and the SRP groups. The c-Fos mRNA levels in gingivae did not show significant difference in both groups. Immunohistochemistry showed that the CCL2, ICAM-1, c-Jun, and c-Fos productions were significantly reduced in rats of the SRP + LLLI group compared with the only SRP group. LLLI significantly decreased the number of osteoclasts as demonstrated by TRAP staining. The 650-nm LLLI might be a useful treatment modality for periodontitis.

Effects of low-level laser irradiation on microRNA-214 expression in rats with myocardial infarction

Objective To observe the effect and significance of low-level laser irradiation (LLLI) on microRNA (miRNA, miR)-214 expression in myocardial infarct area and the edge of the organization. Methods The 110 SD rats were randomly divided into sham group, control group and treatment group. The control and treatment groups by ligation of the left anterior descending coronary artery produced myocardial infarction model, sham group wearing the same site without ligation, three weeks after the treatment group LLLI treatment infarct region, the control group and the sham group irradiating the same area but treatment device does not power on, each group 1, 24, 48, 72 h, take infarct tissue 7 d, using real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) was used to detect expression of miR-214, respectively, after irradiation. Results The expression treatment group, the infarct zone miR-214 from the rear L starts to rise for 1 h (36.15±1.21), 48 h peak (58.95±1.34), the expression of close control when 7 d (44.12±2.17), in 1, 24, 48, 72 h, 7 d expression were higher (36.15±1.21 vs. 14.55±1.51, P=0.000), (52.14±1.32 vs. 21.22±0.46, P=0.000), (58.95±1.34 vs. 36.08±2.24, P=0.000), (53.85±1.34 vs. 38.01±1.95, P=0.000), (44.12±2.17 vs. 39.41±3.13, P=0.000). Conclusion LLLI treatment can significantly increase the expression of myocardial infarction, infarct tissue miR-214, delaying myocardial fibrosis, reduce pathological remodeling, better myocardial protection. Key words: Low-level laser irradiation; Rat; Myocardial infarction; MicroRNA-214; Cardioprotection

Exosomes secreted by mice adipose-derived stem cells after low-level laser irradiation treatment reduce apoptosis of osteocyte induced by hypoxia.

Kienböck's disease is a commonly seen posttraumatic avascular necrosis characterized by avascular necrosis of the lunate bone of the wrist which involves the dominant hand. In our study, we aimed to present midterm outcomes of 12 cases treated with radial metaphyseal core decompression. In our clinic, 12 patients who applied to our outpatient clinic with intractable pain despite at least six weeks of conservative treatment were previously diagnosed and evaluated as Kienböck's disease between the years 2006 and 2014. Patients at early stage received radial metaphyseal core decompression. The patients were evaluated as postoperative grip strength, flexion-extension gap, ulnar-radial deviation gap, VAS, Quick DASH and MAYO wrist scoring and patient satisfaction. We determined that interventions performed for Kienböck's disease cannot halt radiological progression. We are of the opinion that radial metaphyseal core decompression, aiming at increasing blood perfusion, improve early diagnosis and treatment of Kienböck's disease, increasing the patient satisfaction.

Odontogenic differentiation and biomineralization potential of dental pulp stem cells inside Mg-based bioceramic scaffolds under low-level laser treatment

This study aimed to investigate the potential of low-level laser irradiation (LLLI) to promote odontogenic differentiation and biomineralization by dental pulp stem cells (DPSCs) seeded inside bioceramic scaffolds. Mg-based, Zn-doped bioceramic scaffolds, synthesized by the sol-gel technique, were spotted with DPSCs and exposed to LLLI at 660nm with maximum output power of 140mw at fluencies (a) 2 and 4J/cm2 to evaluate cell viability/proliferation by the MTT assay and (b) 4J/cm2 to evaluate cell differentiation, using real-time PCR (expression of odontogenic markers) and a p-nitrophenylphosphate (pNPP)-based assay for alkaline phosphatase (ALP) activity measurement. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) analysis were used for structural/chemical characterization of the regenerated tissues. Exposure of the DPSCs/scaffold complexes to the proposed LLLI scheme was associated with statistically significant increase of odontogenesis-related markers (bone morphogenetic protein 2 (BMP-2): 22.4-fold, dentin sialophosphoprotein (DSPP): 28.4-fold, Osterix: 18.5-fold, and Runt-related transcription factor 2 (Runx2): 3.4-fold). ALP activity was significantly increased at 3 and 7days inside the irradiated compared to that in the non-irradiated SC/DPSC complexes, but gradually decreased until 14days. Newly formed Ca-P tissue was formed on the SC/DPSC complexes after 28days of culture that attained the characteristics of bioapatite. Overall, LLLI treatment proved to be beneficial for odontogenic differentiation and biomineralization of DPSCs inside the bioceramic scaffolds, making this therapeutic modality promising for targeted dentin engineering.

Low-level laser therapy for beta amyloid toxicity in rat hippocampus

Beta amyloid (Aβ) is well accepted to play a central role in the pathogenesis of Alzheimer's disease (AD). The present work evaluated the therapeutic effects of low-level laser irradiation (LLI) on Aβ-induced neurotoxicity in rat hippocampus. Aβ 1–42 was injected bilaterally to the hippocampus CA1 region of adult male rats, and 2-minute daily LLI treatment was applied transcranially after Aβ injection for 5 consecutive days. LLI treatment suppressed Aβ-induced hippocampal neurodegeneration and long-term spatial and recognition memory impairments. Molecular studies revealed that LLI treatment: (1) restored mitochondrial dynamics, by altering fission and fusion protein levels thereby suppressing Aβ-induced extensive fragmentation; (2) suppressed Aβ-induced collapse of mitochondrial membrane potential; (3) reduced oxidized mitochondrial DNA and excessive mitophagy; (4) facilitated mitochondrial homeostasis via modulation of the Bcl-2-associated X protein/B-cell lymphoma 2 ratio and of mitochondrial antioxidant expression; (5) promoted cytochrome c oxidase activity and adenosine triphosphate synthesis; (6) suppressed Aβ-induced glucose-6-phosphate dehydrogenase and nicotinamide adenine dinucleotide phosphate oxidase activity; (7) enhanced the total antioxidant capacity of hippocampal CA1 neurons, whereas reduced the oxidative damage; and (8) suppressed Aβ-induced reactive gliosis, inflammation, and tau hyperphosphorylation. Although development of AD treatments has focused on reducing cerebral Aβ levels, by the time the clinical diagnosis of AD or mild cognitive impairment is made, the brain is likely to have already been exposed to years of elevated Aβ levels with dire consequences for multiple cellular pathways. By alleviating a broad spectrum of Aβ-induced pathology that includes mitochondrial dysfunction, oxidative stress, neuroinflammation, neuronal apoptosis, and tau pathology, LLI could represent a new promising therapeutic strategy for AD.

Biological effects of low-level laser irradiation on umbilical cord mesenchymal stem cells

Low-level laser irradiation (LLLI) can enhance stem cell (SC) activity by increasing migration and proliferation. This study investigated the effects of LLLI on proliferation, enzymatic activity, and growth factor production in human umbilical cord mesenchymal SCs (hUC-MSCs) as well as the underlying mechanisms. hUC-MSCs were assigned to a control group (non-irradiation group) and three LLLI treatment groups (635 nm group, 808 nm group, and 635/808 nm group). Laser power density and energy density of 20 mW/cm2 and 12 J/cm2, respectively, were used for each experiment. The proliferation rate was higher in the 635 nm as compared to the other groups. LLLI at 808 nm did not induce cell proliferation. ROS levels in cells exposed to 635, 808, and 635/808 nm radiation were increased by 52.81%, 26.89%, and 21.15%, respectively, relative to the control group. CAT, tGPx, and SOD activity was increased. LLLI at 808 nm increased the levels of IL-1, IL-6, and NFκB but not VEGF. LLLI improved hUC-MSCs function and increased antioxidant activity. Dual-wavelength LLLI had more potent effects on hUC-MSCs than single-wavelength treatment. LLLI has potential applications in the preconditioning of hUC-MSCs in vitro prior to transplantation, which could improve the regenerative capacity of cells.

Effect of LLLI on Expression of Micro RNA-21 and Ventricular Remodeling in Rats after AMI

Objective: To assess the effects of low-level laser irradiation (LLLI) on the expression of micro RNA-21 and ventricular remodelling in the model of rat myocardial infarction. Methods: 110 adult-female SD rats were randomly divided into sham operation (30), control (40), and treatment (40) groups. MI model was prepared by ligation of left anterior descending artery in the control and treatment groups while simply threading at the same site of MI model in sham group. LLLI treated using (635 nm, 6mW, 125s, 0.96 J/cm2) after three weeks of MI at infarct region in the treatment group. At 1 h, 24 h, 48 h, 72 h and 7 d after LLLI treatment, the expression of miR-21 at the infracted myocardial tissue were detected using qRT-PCR. At the end of 4 weeks after MI, hearts were harvested for histological analysis. Results: MiR-21 expression in the treatment group was lower than both sham group as well as the control group (P<0.05). LVEF (%) and LVFS (%) in the treatment group before and after LLLI; (39.37 ± 1.35 vs. 47.62 ± 2.75, P<0.05; (19.23 ± 3.12) vs (24.15± 2.53), P<0.05). Collagen fiber content in treatment group were significantly lower than control group (28.79 ± 2.06%) vs (69.22 ± 3.64%), (P<0.05). Conclusion: LLLI treatment of myocardial infarction can significantly down regulate the expression of tissue miR-21 in the infarct region, increase left ventricular function and decrease collagen fiber content suggesting the beneficial effect of LLLI on delaying myocardial fibrosis, reduce the pathological ventricular remodelling (VR) after MI.

Effects of low-level laser irradiation on the tissue of rats after myocardial infarction

Objective To investigate the influence of low-level laser irradiation (LLLI) on myocardial infarction.Methods In LLLI preconditioning study,vascular endothelial growth factor (VEGF),superoxide dismutase (SOD) and malondialdehyde (MDA) in the infarcted myocardium were evaluated at 1 st h,1 st day and 1 st week after laser irradiation.Results LLLI significantly increased VEGF mRNA and protein expression (4.42 ±2.88 and 2.62 ± 1.83) in the irradiated region after myocardial infarction organization as compared with cntrol group (2.12 ± 0.58 and 1.02 ± 0.41) and sham group (2.42 ± 0.26 and 0.92 ± 0.37,P ＜ 0.05),and there was no significant difference between sham group and control group.After pretreatment for 1 h and 1 day,as compared with control group,the enzymatic activity of SOD in the LLLI infarct myocardial tissue [(0.84 ± 0.43) and (0.92 ± 0.34) U/mg] was significantly increased,and MDA production [(0.56 ±0.31) and (0.50 ± 0.29) nmol/mg] was significantly reduced (P ＜ 0.05).These indicators one week after pretreatment between LLLI group and control group showed no statistically significant difference (P ＞ 0.05).Conclusion LLLI treatment significantly raised VEGF expression and SOD activity in the infarction myocardial tissue,and reduced MDA content,which can improve microenvironment after myocardial infarction,promote cell regeneration and repair after myocardial infarction. Key words: Low-level laser irradiation; Myocardial infarction; Vascular endothelial growth factor; Superoxide dismutase; Malondialdehyde

MicroRNA-193 Pro-Proliferation Effects for Bone Mesenchymal Stem Cells After Low-Level Laser Irradiation Treatment Through Inhibitor of Growth Family, Member 5

The enhanced proliferation of mesenchymal stem cells (MSCs) can be helpful for the clinical translation of cell therapy. Low-level laser irradiation (LLLI) has been demonstrated as regulating MSC proliferation. MicroRNAs (miRNAs) are involved in various pathophysiologic processes in stem cells, but the role of miRNAs in the LLLI-based promotion of MSC proliferation remains unclear. We found that the proliferation level and cell cycle-associated genes in MSCs were increased after LLLI treatment in a time-dependent manner. Microarray assays revealed subsets of miRNAs to be differentially regulated, and these dynamic changes were confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) after LLLI. miR-193 was the most highly up-regulated miRNA, and the change in it was related with the proliferation level. Gain-loss function experiments demonstrated that miR-193 could regulate the proliferation of MSCs, including human's and rat's, but could not affect the apoptosis and differentiation level. Blockade of miR-193 repressed the MSC proliferation induced by LLLI. By qRT-PCR, we found that miR-193, in particular, regulated cyclin-dependent kinase 2 (CDK2) expression. Bioinformatic analyses and luciferase reporter assays revealed that inhibitor of growth family, member 5 (ING5) could be the best target of miR-193 to functionally regulate proliferation and CDK2 activity, and the mRNA and protein level of ING5 was regulated by miR-193. Furthermore, the ING5 inhibited by small interfering RNA (siRNA) could up-regulate the proliferation of MSCs and the expression of CDK2. Taken together, these results strongly suggest that miR-193 plays a critical part in MSC proliferation in response to LLLI stimulation, which is potentially amenable to therapeutic manipulation for clinical application.

Low-level laser irradiation treatment reduces CCL2 expression in rat rheumatoid synovia via a chemokine signaling pathway

Rheumatoid arthritis (RA) is an inflammatory joint disorder whose progression leads to the destruction of cartilage and bone. Although low-level laser irradiation (LLLI) is currently being evaluated for the treatment of RA, the molecular mechanisms underlying its effectiveness remain unclear. To investigate possible LLLI-mediated antiinflammatory effects, we utilized a collagen-induced arthritis (CIA) rat model and analyzed gene expression profiles in the synovial membranes of the knee joint. Total RNA was isolated from the synovial membrane tissue of the joints of untreated CIA rats or CIA rats treated with LLLI (830 nm Ga-Al-As diode), and gene expression profiles were analyzed by DNA microarray (41,000 rat genes), coupled with Ingenuity pathways analysis (IPA). DNA microarray analysis showed that CCL2 gene expression was increased in CIA tissue, and that LLLI treatment significantly decreased CIA-induced CCL2 mRNA levels. IPA revealed that chemokine signal pathways were involved in the activation of CCL2 production. These microarray data were further validated using real-time PCR and reverse transcription PCR. Immunohistochemistry confirmed that CCL2 production was decreased in CIA rats treated with LLLI. These findings suggest that decreased CCL2 expression may be one of the mechanisms involved in LLLI-mediated RA inflammation reduction.

REDUCTION OF CXCR4 EXPRESSION IN RHEUMATOID ARTHRITIS RAT JOINTS BY LOW LEVEL DIODE LASER IRRADIATION

Rheumatoid arthritis (RA) is an inflammatory joint disorder, whose progression leads to the destruction of cartilage and bone. Chemokines and their receptors are potential therapeutic targets in RA. Among these, it has been suggested that CXC chemokine 4 (CXCR4) and its ligand CXC ligand 12 (CXCL12) are involved in RA pathogenesis. Low-level laser irradiation (LLLI) is currently being evaluated for the treatment of RA; however, the molecular mechanisms underlying its effectiveness remain unclear. To understand the anti-inflammatory effect of LLLI, we used the collagen-induced arthritis (CIA) rat as RA model and analyzed the gene expression profile in synovial membrane in the hindpaw joints of control, CIA and CIA + LLLI. Expression of CXCR4 and CXCL12 genes were also studied. Total RNA was isolated from the synovial membrane tissue of CIA rat joints or CIA joints treated with LLLI (830 nm Ga-Al-As diode), and gene expression profiles were analyzed by DNA microarray (41,000 rat genes). The mRNA levels were confirmed by reverse transcription polymerase chain reaction (RT-PCR) and real-time PCR. Immunohistochemical examination to examine CXCR4 protein expression was also carried out. DNA microarray analysis showed that CXCR4 gene expression was increased in CIA tissue and LLLI treatment significantly decreased CIA-induced CXCR4 mRNA levels. In contrast, CXCL12 did not show any significant changes. The microarray data of CXCR4 mRNA levels were further validated using RT-PCR and real-time PCR. Increased CXCR4 mRNA levels by CIA and its reduction following LLLI was successfully confirmed. CXCR4 production was increased in CIA joints and its production was decreased by LLLI. Decreased CXCR4 expression may be one of the mechanisms in LLLI-mediated reduction of RA inflammation.

The Effects of Low Level Laser Irradiation on Gingival Inflammation

The goal of this study was to analyze the effects of low level laser irradiation treatment and conservative treatment on gingival inflammation. It is widely accepted today that the primary etiological factor for the onset of periodontitis is dental plaque, although the exact mechanism of damage remains unknown. Inflammation is a basic response of periodontal tissue to damage and serves as a fast first line of defense against damage and infections. The treatment of gingivitis and periodontitis has gone through various stages: from the simplest, classical treatment methods, through improved radical interventions, to a new era marked by laser technology. Low level laser irradiation has an anti-inflammatory effect, both general and local. The research was done on patients who had chronic periodontal disease (mild periodontitis) with expressed clinical symptoms of gingival inflammation. All patients in the study underwent conservative treatment. After conservative therapy, the patients from the experimental group were subjected to 10 low level laser treatment sessions. Both groups underwent regular follow-up visits 1, 3, and 6 months after treatment, which involved only clinical examination using plaque index (PI), gingival index (GI), and bleeding on probing index (BOP index). A considerable decrease in all three indexes after the application of both therapies was noticed. The follow-up visits revealed the difference in index values. With laser therapy, the values of indexes decreased steadily, whereas with conservative therapy they increased up to a certain point, but did not reach the pre-therapy values. A general conclusion can be drawn that low level laser irradiation (semiconductor, 670 nm) can be used as a successful physical adjuvant method of treatment, which, together with traditional periodontal therapy, leads to better and longer-lasting therapeutic results.