Abstract:Abstract. To date, there is limited data on the biological effects of low-intensity pulsed ultrasound (LIPUS) on primary malignant bone tumors. The purpose of the present study was to investigate the antitumor effects of LIPUS on osteosarcoma cells. The effects of LIPUS on cell viability, induction of apoptosis, mitochondrial membrane potential and intracellular signaling molecules in the LM8 osteosarcoma cell line were investigated. LIPUS inhibited cell viability (P=0.0022) and mitochondrial membrane potentia… Show more
“…LIPUS also inhibits the proliferation of human hepatocellular carcinoma cells and osteosarcoma cells and promotes their apoptosis. 35,36 In this study, LIPUS promoted the proliferation of BMSCs when the cells were exposed to LIPUS for 5 or 10 minutes once a day. However, if BMSCs are irradiated by LIPUS for 20 minutes once a day, 50 mW/cm 2 or higher intensity could inhibit the growth of cells.…”
Low‐intensity pulsed ultrasound (LIPUS) is a promising therapy that is widely used in clinical applications and fundamental research. Previous research has shown that LIPUS exposure has a positive effect on stem cell proliferation. However, the impact of LIPUS exposure on human bone marrow mesenchymal stem cells (hBMSCs) remains unknown. In our study, the effect and mechanism of LIPUS exposure on the proliferation of hBMSCs were investigated, and the optimal parameters of LIPUS were determined. hBMSCs were obtained and identified by flow cytometry, and the proliferation of hBMSCs was measured using the Cell Counting Kit‐8 assay to determine cell cycle and cell count. Expression levels of the phosphoinositide 3‐kinase (PI3K)/protein kinase B (AKt) pathway proteins and cyclin D1 were determined by western blot analysis. Next, hBMSCs were successfully cultured and identified as multipotent mesenchymal stem cells. We found that LIPUS could promote the proliferation of hBMSCs when the exposure time was 5 or 10 minutes per day. Furthermore, 50 or 60 mW/cm2 LIPUS had a more significant effect on cell proliferation, but if cells were irradiated by LIPUS for 20 minutes once a day, an intensity of at least 50 mW/cm2 could markedly inhibit cell growth. Cell cycle analysis demonstrated that LIPUS treatment drives cells to enter S and G2/M phases from the G0/G1 phase. LIPUS exposure increased phosphorylation of PI3K/AKt and significantly upregulated expression of cyclin D1. However, these effects were inhibited when cells were treated with PI3K inhibitor (LY294002), which in turn reduced LIPUS‐mediated proliferation of hBMSCs. These results suggest that LIPUS exposure may be involved in the proliferation of hBMSCs via activation of the PI3K/AKt signaling pathway and high expression of cyclin D1, and the intensity of 50 or 60 mW/cm2 and exposure time of 5 minutes were determined to be the optimal parameters for LIPUS exposure.
“…LIPUS also inhibits the proliferation of human hepatocellular carcinoma cells and osteosarcoma cells and promotes their apoptosis. 35,36 In this study, LIPUS promoted the proliferation of BMSCs when the cells were exposed to LIPUS for 5 or 10 minutes once a day. However, if BMSCs are irradiated by LIPUS for 20 minutes once a day, 50 mW/cm 2 or higher intensity could inhibit the growth of cells.…”
Low‐intensity pulsed ultrasound (LIPUS) is a promising therapy that is widely used in clinical applications and fundamental research. Previous research has shown that LIPUS exposure has a positive effect on stem cell proliferation. However, the impact of LIPUS exposure on human bone marrow mesenchymal stem cells (hBMSCs) remains unknown. In our study, the effect and mechanism of LIPUS exposure on the proliferation of hBMSCs were investigated, and the optimal parameters of LIPUS were determined. hBMSCs were obtained and identified by flow cytometry, and the proliferation of hBMSCs was measured using the Cell Counting Kit‐8 assay to determine cell cycle and cell count. Expression levels of the phosphoinositide 3‐kinase (PI3K)/protein kinase B (AKt) pathway proteins and cyclin D1 were determined by western blot analysis. Next, hBMSCs were successfully cultured and identified as multipotent mesenchymal stem cells. We found that LIPUS could promote the proliferation of hBMSCs when the exposure time was 5 or 10 minutes per day. Furthermore, 50 or 60 mW/cm2 LIPUS had a more significant effect on cell proliferation, but if cells were irradiated by LIPUS for 20 minutes once a day, an intensity of at least 50 mW/cm2 could markedly inhibit cell growth. Cell cycle analysis demonstrated that LIPUS treatment drives cells to enter S and G2/M phases from the G0/G1 phase. LIPUS exposure increased phosphorylation of PI3K/AKt and significantly upregulated expression of cyclin D1. However, these effects were inhibited when cells were treated with PI3K inhibitor (LY294002), which in turn reduced LIPUS‐mediated proliferation of hBMSCs. These results suggest that LIPUS exposure may be involved in the proliferation of hBMSCs via activation of the PI3K/AKt signaling pathway and high expression of cyclin D1, and the intensity of 50 or 60 mW/cm2 and exposure time of 5 minutes were determined to be the optimal parameters for LIPUS exposure.
“…Ultrasound can be selectively targeted on tumor cells using focused ultrasound beams and ultrasound may potentially be more toxic to tumor cells due to the higher metabolic rate in glioma cells 10,18. Previous studies showed that ultrasound inhibits cell proliferation in osteosarcoma cells through inhibiting cell viability, mitochondrial membrane potential, reducing phosphorylated mitogen-activated protein kinase 7 and so on 19. In this study, LFLIU inhibited the proliferation of C6 and U87 cells in an intensity-dependent manner, indicates it seems to be an effective and safe procedure in the treatment of glioma.…”
Purpose:
To explore the effect of curcumin and low-frequency and low-intensity ultrasound (LFLIU) on C6 and U87 cell, and whether LFLIU could inhibit multidrug resistance protein 1 (MRP1) by increasing the sensitivity of curcumin via vascular epithelial growth factor (VEGF)/PI3K/Akt signaling pathway targeting.
Methods:
C6 and U87 cells were treated with various doses of curcumin and/or different intensities of LFLIU for 60 s. After 24 hrs, the effects of curcumin and/or LFLIU on the proliferation of C6 and U87 cells were examined. Real-time PCR and western blot analysis were used to detect the expression of VEGF and MRP1 at both mRNA and protein levels. The expression of MRP1 in C6 and U87 cells was also determined following stimulation with recombinant human VEGF and/or LY294002.
Results:
Curcumin and LFLIU inhibited the proliferation of glioma cells in an intensity- or dose-dependent manner. Furthermore, survivin was significant after combined treatment compares with that of curcumin or LFLIU treatment alone. VEGF and MRP1 were highly expressed in C6 and U87 cells, curcumin and LFLIU alone or in combination could decrease the expression of both VEGF and MRP1. MRP1 expression was down-regulated following LY294002 treatment, which blocked after exposure to VEGF.
Conclusion:
The synergistic effects, such as a higher inhibition rate, and lower expressions of MRP1 and VEGF, of combined curcumin and LFLIU against glioma was much better than that of a single treatment. The down-regulation of MRP1 may be related with the VEGF/PI3K/Akt pathway in glioma.
“…After incubation for 24 h, acrofolione A or B (15 μ m ) was added and the cells were incubated for 24 and 48 h. The collected cells were rinsed with cold PBS and immediately solubilised in cell lysate buffer (Cell Signaling Technology) containing complete protease inhibitor cocktail (Roche Diagnostics Ltd.) and 1 m m phenylmethylsulfonyl fluoride (Sigma‐Aldrich) by blocking the lysates gently at 4 °C for 30 min. The supernatants obtained from microcentrifugation at 14000 g for 5 min were transferred to clean test tubes, the sample proteins (0.5 mg/mL) were diluted and then incubated with PathScan Stress and Apoptosis Signaling Antibody Array kit reagent (Cell Signaling Technology) as per the manufacturer's instructions . The expressed dots were visualised using LumiGlo reagent and detected using an ImageQuant LAS‐4000.…”
Section: Methodsmentioning
confidence: 99%
“…The supernatants obtained from microcentrifugation at 14000g for 5 min were transferred to clean test tubes, the sample proteins (0.5 mg/mL) were diluted and then incubated with PathScan Stress and Apoptosis Signaling Antibody Array kit reagent (Cell Signaling Technology) as per the manufacturer's instructions. [23] The expressed dots were visualised using LumiGlo reagent and detected using an ImageQuant LAS-4000. Dot densities were measured using Image J (National Institutes of Health, Bethesda, MD, USA) and corrected to the relative density of a-tubulin.…”
Section: Exploration Of Intracellular Signalling Moleculesmentioning
Objectives
We investigated the apoptotic activities of acrofolione A (1) and B (2) isolated from Acronychia pedunculata against a human pre‐B cell leukaemia cell line (NALM‐6) to explore the apoptosis‐related signalling molecules targeted by 1 and 2.
Methods
The apoptosis effects of 1 and 2 in NALM‐6 cells were investigated by TUNEL staining, annexin V, mitochondria membrane potential and caspase 3/7 activity. We carried out a protein array to explore the signalling molecules involved in apoptosis comprehensively.
Key findings
Acrofolione A (1) suppressed the growth of NALM‐6, K562 and HPB‐ALL cells (IC50 16.7 ± 1.9, 17.9 ± 0.3 and 10.1 ± 0.2 μm, respectively) more effectively than acrofolione B (2). Both compounds time‐dependently increased the number of NALM‐6 cells with abnormal nuclei, and increased the number of annexin V‐positive cells and decreased the mitochondrial membrane potential of NALM‐6 cells. Acrofolione A (1) markedly elevated caspase 3/7 activity and increased the number of TUNEL‐positive cells. Cells treated with either compound showed enhanced expression of cleaved PARP and cleaved caspase 3 and 7, and reduced survivin protein levels.
Conclusions
Acrofolione A (1) and B (2) may be useful in the treatment of various types of leukaemia.
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