Granulocyte-macrophage colony-stimulating factor (GM-CSF) shows therapeutic effect on dimethylnitrosamine (DMN)-induced liver fibrosis in rats.

This study was undertaken to investigate the inhibitory effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) on dimethylnitrosamine (DMN)-induced liver fibrosis in rats. Liver fibrosis was induced in Sprague-Dawley rats by injecting DMN intraperitoneally (at 10 mg/kg of body weight) daily for three consecutive days per week for 4 weeks. To investigate the effect of GM-CSF on disease onset, GM-CSF (50 µg/kg of body weight) was co-treated with DMN for 2 consecutive days per week for 4 weeks (4-week groups). To observe the effect of GM-CSF on the progression of liver fibrosis, GM-CSF was post-treated alone at 5-8 weeks after the 4 weeks of DMN injection (8-week groups). We found that DMN administration for 4 weeks produced molecular and pathological manifestations of liver fibrosis, that is, it increased the expressions of collagen type I, alpha-smooth muscle actin (α-SMA), and transforming growth factor-ß1 (TGF-ß1), and decreased peroxisome proliferator-activated receptor gamma (PPAR-γ) expression. In addition, elevated serum levels of aspartate aminotransferase (AST), total bilirubin level (TBIL), and decreased albumin level (ALB) were observed. In both the 4-week and 8-week groups, GM-CSF clearly improved the pathological liver conditions in the gross and histological observations, and significantly recovered DMN-induced increases in AST and TBIL and decreases in ALB serum levels to normal. GM-CSF also significantly decreased DMN-induced increases in collagen type I, α-SMA, and TGF-ß1 and increased DMN-induced decreases in PPAR-γ expression. In the DMN groups, survival decreased continuously for 8 weeks after DMN treatment for the first 4 weeks. GM-CSF showed a survival benefit when co-treated for the first 4 weeks but a marginal effect when post-treated for 5-8 weeks. In conclusion, co-treatment of GM-CSF showed therapeutic effects on DMN-induced liver fibrosis and survival rates in rats, while post-treatment efficiently blocked liver fibrosis.

Low-intensity ultrasound (LIUS) differentially modulates mitochondrial reactive oxygen species (mtROS) generation by three different chemicals in PC12 cells.

We have previously shown that low-intensity ultrasound (LIUS) can modulate mitochondrial complex I activity and the generation of mitochondrial reactive oxygen species (mtROS) in PC12 cells. This study investigated the mechanism of LIUS by comparing its effect on mitochondrial dysfunction by three different pathways. LIUS was shown to reverse the effects of rotenone, a Q-site blocker, on the complex I inhibition, mtROS generation, and drop of mitochondrial membrane potential (Δψm). In contrast, common antioxidants, N-acetyl cysteine (NAC), and uric acid (UA) blocked rotenone-induced mtROS generation and Δψm drop without recovering the complex I activity, which suggested that Δψm drop is correlated with mtROS generation rather than complex I inhibition itself. Ionomycin, an ionophore for Ca2+, and L-buthionine-S,R-sulfoximine (BSO), an inhibitor of glutathione (GSH) biosynthesis, induced mtROS generation and Δψm drop without inhibiting complex I activity via different mechanisms. LIUS showed no effect on ionomycin-induced Δψm drop but showed partial inhibition on the other effects of ionomycin and BSO. These results suggest that LIUS might have redundant mechanisms but acted mainly on the complex I activity thereby modulating mtROS and Δψm levels. LIUS appeared to act on the Q-module of complex I because it showed no inhibitory effect on Zn2+, an inhibitor of the proton transporting P-module of complex I. Interestingly, pretreatment of LIUS for up to an hour in advance blocked the rotenone effect as efficiently as the co-treatment. Further studies are needed to reveal the exact mechanism of LIUS to inhibit complex I activity.

Low-intensity ultrasound attenuates paw edema formation and decreases vascular permeability induced by carrageenan injection in rats.

BACKGROUND: Therapeutic potential of low-intensity ultrasound (LIUS) has become evident in various musculoskeletal diseases. We have previously shown that LIUS has an inhibitory effect on local edema in various diseases including the arthritis and brain injury. In this study, we examined whether LIUS can attenuate paw edema formation vis-à-vis vascular permeability and inflammation in rats induced by carrageenan. LIUS with a frequency of 1 MHz and the intensities of 50, 100, or 200 mW/cm2 were exposed on rat paws for 10 min immediately after carrageenan injection. RESULTS: Carrageenan injection induced paw edema which was peaked at 6 h and gradually decreased nearly to the initial baseline value after 72 h. LIUS showed a significant reduction of paw edema formation at 2 and 6 h at all intensities tested. The highest reduction was observed at the intensity of 50 mW/cm2. Histological analyses confirmed that LIUS clearly decreased the carrageenan-induced swelling of interstitial space under the paw skin and infiltration of polymorphonuclear leukocytes. Moreover, Evans Blue extravasation analyses exhibited a significant decreases of vascular permeability by LIUS. Finally, immunohistochemical staining showed that expression of pro-inflammatory proteins, namely, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) induced by carrageenan injection was reduced back to the normal level after LIUS stimulation. CONCLUSIONS: These results provide a new supporting evidence for LIUS as a therapeutic alternative for the treatment of edema in inflammatory diseases such as cellulitis.

Low-Intensity Ultrasound Decreases Ischemia-Induced Edema by Inhibiting N-Methyl-d-Aspartic Acid Receptors.

BACKGROUND: We have previously shown that low-intensity ultrasound (LIUS), a noninvasive mechanical stimulus, inhibits brain edema formation induced by oxygen and glucose deprivation (OGD) or treatment with glutamate, a mediator of OGD-induced edema, in acute rat hippocampal slice model in vitro. METHODS: In this study, we treated the rat hippocampal slices with N-methyl-d-aspartic acid (NMDA) or (S)-3,5-dihydroxyphenylglycine (DHPG) to determine whether these different glutamate receptor agonists induce edema. The hippocampal slices were then either sonicated with LIUS or treated with N-methyl-d-aspartic acid receptor (NMDAR) antagonists, namely, MK-801 and ketamine, and observed their effects on edema formation. RESULTS: We observed that treatment with NMDA, an agonist of ionotropic glutamate receptors, induced brain edema at similar degrees compared with that induced by OGD. However, treatment with DHPG, an agonist of metabotropic glutamate receptors, did not significantly induce brain edema. Treatment with the NMDAR antagonists MK-801 or ketamine efficiently prevented brain edema formation by both OGD and NMDA in a concentration-dependent manner. N-Methyl-d-aspartic acid-induced brain edema was alleviated by LIUS in an intensity-dependent manner when ultrasound was administered at 30, 50, or 100 mW/cm2 for 20 minutes before the induction of the edema. Furthermore, LIUS reduced OGD- and NMDA-induced phosphorylation of NMDARs at Y1325. CONCLUSION: These results suggest that LIUS can inhibit OGD- or NMDA-induced NMDAR activation by preventing NMDAR phosphorylation, thereby reducing a subsequent brain edema formation. The mechanisms by which LIUS inhibits NMDAR phosphorylation need further investigation.

Low-Intensity Ultrasound Reduces High Glucose-Induced Nitric Oxide Generation in Retinal Pigment Epithelial Cells.

Diabetic retinopathy (DR) is a severe micro-vascular complication of diabetes. High glucose (HG)-evoked nitric oxide (NO) production mediated by increased oxidative stress is a key factor in DR pathogenesis. In this study, we examined whether low-intensity ultrasound (LIUS) stimulation can reduce HG-induced NO generation. We determined that LIUS stimulation decreased the HG-induced NO generation possibly via inhibition of reactive oxygen species (ROS) and subsequently diminished the associated pro-inflammatory pathway involving the induced expression of inducible nitric oxide synthase, cyclooxygenase-2 and vascular endothelial growth factor. In addition, we determined that LIUS stimulation reduced the quantity of NO produced by N-acetylcysteine, which was not mediated by ROS. These results indicate that LIUS can inhibit both ROS-dependent and -independent NO generation processes in ARPE-19 cells. We envision LIUS as a potential therapeutic alternative to treat DR. Further studies are required to understand the underlying mechanism of the LIUS-induced reduction of NO generation for DR therapy.

Low-Intensity Ultrasound Decreases α-Synuclein Aggregation via Attenuation of Mitochondrial Reactive Oxygen Species in MPP(+)-Treated PC12 Cells.

Many studies have shown that mitochondrial dysfunction and the subsequent oxidative stress caused by excessive reactive oxygen species (ROS) generation play a central role in the pathogenesis of Parkinson's disease (PD). We have previously shown that low-intensity ultrasound (LIUS) could reduce ROS generation by L-buthionine-(S,R)-sulfoximine (BSO) in retinal pigment epithelial cells. In this study, we studied the effects of LIUS stimulation on the ROS-dependent α-synuclein aggregation in 1-methyl-4-phenylpyridinium ion (MPP+)-treated PC12 cells. We found that LIUS stimulation suppressed the MPP+-induced ROS generation and inhibition of mitochondrial complex I activity in PC12 cells in an intensity-dependent manner at 30, 50, and 100 mW/cm2. Furthermore, LIUS stimulation at 100 mW/cm2 suppressed inhibition of mitochondrial complex activity by MPP+ and actually resulted in a decrease of α-synuclein phosphorylation and aggregation induced by MMP+ treatment in PC12 cells. LIUS stimulation also inhibited expression of casein kinase 2 (CK2) that appears to mediate ROS-dependent α-synuclein aggregation. Finally, LIUS stimulation alleviated the death of PC12 cells by MPP+ treatment in an intensity-dependent manner. We, hence, suggest that LIUS stimulation inhibits ROS generation by MPP+ treatment, thereby suppressing α-synuclein aggregation in PC12 cells.

Low intensity ultrasound inhibits brain oedema formation in rats: potential action on AQP4 membrane localization.

AIMS: Brain oedema is a major contributing factor to the morbidity and mortality of a variety of brain disorders. Although there has been considerable progress in our understanding of pathophysiological and molecular mechanisms associated with brain oedema so far, more effective treatment is required and is still awaited. Here we intended to study the effects of low intensity ultrasound (LIUS) on brain oedema. METHODS: We prepared the rat hippocampal slice in vitro and acute water intoxication in vivo models of brain oedema. We applied LIUS stimulation in these models and studied the molecular mechanisms of LIUS action on brain oedema. RESULTS: We found that LIUS stimulation markedly inhibited the oedema formation in both of these models. LIUS stimulation significantly reduced brain water content and intracranial pressure resulting in increased survival of the rats. Here, we showed that the AQP4 localization was increased in the astrocytic foot processes in the oedematous hippocampal slices, while it was significantly reduced in the LIUS-stimulated hippocampal slices. In the in vivo model too, AQP4 expression was markedly increased in the microvessels of the cerebral cortex and hippocampus after water intoxication but was reduced in the LIUS-stimulated rats. CONCLUSIONS: These data show that LIUS has an inhibitory effect on cytotoxic brain oedema and suggest its therapeutic potential to treat brain oedema. We propose that LIUS reduces the AQP4 localization around the astrocytic foot processes thereby decreasing water permeability into the brain tissue.