Glabridin improves autoimmune disease in Trex1-deficient mice by reducing type I interferon production.

BACKGROUND: The cGAS-STING signaling pathway is an essential section of the natural immune system. In recent years, an increasing number of studies have shown a strong link between abnormal activation of the cGAS-STING signaling pathway, a natural immune pathway mediated by the nucleic acid receptor cGAS, and the development and progression of autoimmune diseases. Therefore, it is important to identify an effective compound to specifically downregulate this pathway for disease. METHODS: The effect of Glabridin (Glab) was investigated in BMDMs and Peripheral blood mononuclear cell (PBMC) by establishing an in vitro model of cGAS-STING signaling pathway activation. An activation model stimulated by DMXAA was also established in mice to study the effect of Glab. On the other hand, we investigated the possible mechanism of action of Glab and the effect of Glab on Trex1-deficient mice. RESULTS: In this research, we report that Glab, a major component of licorice, specifically inhibits the cGAS-STING signaling pathway by inhibiting the level of type I interferon and inflammatory cytokines (IL-6 and TNF-α). In addition, Glab has a therapeutic effect on innate immune diseases caused by abnormal cytoplasmic DNA in Trex1-deficient mice. Mechanistically, Glab can specifically inhibit the interaction of STING with IRF3. CONCLUSION: Glab is a specific inhibitor of the cGAS-STING signaling pathway and may be used in the clinical therapy of cGAS-STING pathway-mediated autoimmune diseases.

Glycyrrhiza uralensis polysaccharides ameliorates cecal ligation and puncture-induced sepsis by inhibiting the cGAS-STING signaling pathway.

Ethnopharmacological relevance: G. uralensis Fisch. (Glycyrrhiza uralensis) is an ancient and widely used traditional Chinese medicine with good efficacy in clearing heat and detoxifying action. Studies suggest that Glycyrrhiza Uralensis Polysaccharides (GUP), one of the major components of G. uralensis, has anti-inflammatory, anti-cancer and hepatoprotective effects., but its exact molecular mechanism has not been explored in depth. Aim of the study: Objectives of our research are about exploring the anti-inflammatory role of GUP and the mechanisms of its action. Materials and methods: ELISA kits, Western blotting, immunofluorescence, quantitative real-time PCR, immunoprecipitation and DMXAA-mediated STING activation mice models were performed to investigate the role of GUP on the cGAS-STING pathway. To determine the anti-inflammatory effects of GUP, cecal ligation and puncture (CLP) sepsis models were employed. Results: GUP could effectively inhibit the activation of the cGAS-STING signaling pathway accompany by a decrease the expression of type I interferon-related genes and inflammatory factors in BMDMs, THP-1, and human PBMCs. Mechanistically, GUP does not affect the oligomerization of STING, but affects the interaction of STING with TBK1 and TBK1 with IRF3. Significantly, GUP had great therapeutic effects on DMXAA-induced agonist experiments in vivo as well as CLP sepsis in mice. Conclusion: Our studies suggest that GUP is an effective inhibitor of the cGAS-STING pathway, which may be a potential medicine for the treatment of inflammatory diseases mediated by the cGAS-STING pathway.

[Moxibustion improves learning-memory ability by regulating apoptosis related proteins of hippocampus in diabetic rats with cognitive impairment].

OBJECTIVE: To investigate the effect of moxibustion on the proteins related with apoptosis and nuclear transcription factor kappa B (NF-κB) in hippocampus of diabetic rats with cognitive impairment (CI), so as to explore its mechanism underlying improvement of learning-memory ability. METHODS: Thirty SD rats were randomly divided into normal, model and moxibustion groups (n=10 rats/group). The diabetic model was established by i.p. injection of streptozotocin solution (25 mg·kg-1·d-1), followed by high-fat diet raising for 4 weeks, and the CI model was confirmed by Morris water maze test. The rats in the moxibustion group were given moxibustion at "Shenting" (GV24), "Baihui" (GV20) and "Dazhui" (GV14) for 20 min each time, the treatment was conducted 6 times a week for 4 weeks. The learning-memory ability was detected by Morris water maze test, the random blood glucose of rats was measured by glucometer and test strips. The protein and mRNA expression levels of Bcl-2, Bax, Caspase-3 and NF-κB p65 in hippocampus were detected by Western blot and quantitative real-time PCR, separately. RESULTS: After modeling, the random blood glucose, escape latency, and the expression levels of Bax, Caspase-3 and NF-κB p65 proteins and mRNAs in the model group were significantly increased, while the expression levels of Bcl-2 protein and mRNA were decreased (P<0.001，P<0.01, P<0.05) in comparison with the normal group. Following the treatment, the modeling induced increase of blood glucose, escape latency, and the expression levels of Bax, Caspase-3 and NF-κB p65 proteins and mRNAs, as well as decrease of Bcl-2 protein and mRNA expression levels were reversed (P<0.05, P<0.01, P<0.001). CONCLUSION: Moxibustion can improve learning-memory ability in diabetic rats with cognitive impairment, which may be related to its function in regulating the expression levels of hippocampal Bcl-2, Bax, Caspase-3 and NF-κB.

Flavonoids derived from licorice suppress LPS-induced acute lung injury in mice by inhibiting the cGAS-STING signaling pathway.

In recent years, we have found that the dysregulation of the cyclic-GMP-AMP synthase (cGAS)‒stimulator of interferon genes (STING) pathway leads to the development of immune and inflammatory diseases, therefore, finding compounds that can specifically regulate this pathway is essential for effective regulation of the immune pathway for addressing inflammatory diseases. Licorice flavonoids (LFs), are active ingredients extracted from the Chinese herb licorice, which has been reported to have strong anti-inflammatory activity in previous studies. Here, we report that LFs inhibit the activation of the cGAS-STING pathway evidenced by the inhibition of the expression of type I interferons and related downstream genes such as interferon-stimulated gene 15 (ISG15) and C-X-C motif chemokine ligand 10 (CXCL10), as well as inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). Notably, LFs markedly improve the LPS-induced acute lung injury by inhibiting the excessive activation of cGAS-STING signaling pathway. Mechanistically, LFs treatment leads to the blocking of 2'3'-cyclic GMP-AMP (cGAMP) synthesis resulting in an inhibition of the activation of the cGAS-STING pathway. Our results indicate that LFs is a specific inhibitor of the cGAS-STING pathway, which is suggested to be a potential candidate for the treatment of cGAS-STING pathway-mediated inflammatory diseases.

Icariside I reduces breast cancer proliferation, apoptosis, invasion, and metastasis probably through inhibiting IL-6/STAT3 signaling pathway.

OBJECTIVES: Breast cancer is a common malignancy in women. More than 90% of breast cancer deaths are caused by metastasis. Epimedii Folium (EF) is a commonly used herb with anti-tumor benefits, but its underlying mechanisms and active components for breast cancer prevention are little understood. This study assessed the therapeutic role of Icariside I (ICS I) in Epimedium flavonoids (EF) on lung metastasis of breast cancer, including the underlying mechanism. METHODS: Western blot, RT-qPCR, wound healing assay, colony formation assay, and flow cytometry were used to investigate the inhibition of breast cancer cells growth and migration by EF and ICS I through disrupting the IL-6/STAT3 pathway. Combined with 4T1 breast cancer model in mice, Western blot, RT-qPCR, Hematoxylin and Eosin staining, immunohistochemistry were used to evaluate the therapeutic role of ICS I in proliferation, apoptosis, invasion, and metastasis of breast cancer. KEY FINDINGS: EF can inhibit STAT3 phosphorylation and reduce the colony formation and migration of breast cancer cells. Detecting the active ingredients in EF, we found ICS I can reduce the activation of STAT3 in 4T1 breast cancer cells, impair colony formation and migration. Moreover, ICS I induced cells G1 phase arrest and modulated Cyclin D1, CDK4, bcl-2, and bax to inhibit proliferation and survival of breast cancer cells. Similarly, the in vivo studies demonstrated that ICS I significantly suppressed tumor development and lung metastasis in the 4T1 mouse model. Tumor cells in vehicle group were arranged in a spoke-like pattern with obvious heterogeneity, and multinucleated tumor giant cells were seen. But, the tumor cells in the ICS I group were disorganized and necrotic lysis was seen in some areas. In ICS I-treated group, tumors' STAT3 phosphorylation level, IL-6, Cyclin D1, CDK4, bcl-2, and vimentin expression were downregulated, bax and cleaved caspase 3 expression were upregulated. In the lung tissue, we could find less metastasis of breast cancer cells and less lung injury in the ICS I group. Besides, the expression of metastasis-related genes MMP9 and vimentin was decreased in the lung tissue of ICS I group. CONCLUSIONS: These findings suggest that ICS I can inhibit breast cancer proliferation, apoptosis, invasion and metastasis probably via targeting IL-6/STAT3 pathway. Therefore, ICS I has the potential to become an innovative therapeutic candidate to breast cancer prevention and treatment.

[New beta-carboline alkaloids contained in Psammosilene tunicoides].

OBJECTIVE: To study the chemical constituents of Psammosilene tunicoides. METHOD: The two chemical constituents were separated by various chromatographic methods, and their structures were identified on the basis of spectroscopic analysis. RESULT: Two beta-carboline alkaloids were separated from normal butanol fraction of P. tunicoides, and identified as 1-acetyl-3-methoxycarbonyl-beta-carboline (1) and 1-acetyl-3-methoxycarbonyl-4-hydroxyl-beta-carboline (2). CONCLUSION: Compound 1 is separated from this plant as natural products for the first time, and compound 2 was a new compound.

The role of MIP-1 alpha in the development of systemic inflammatory response and organ injury following trauma hemorrhage.

Although MIP-1alpha is an important chemokine in the recruitment of inflammatory cells, it remains unknown whether MIP-1alpha plays any role in the development of systemic inflammatory response following trauma-hemorrhage (T-H). C57BL/6J wild type (WT) and MIP-1alpha-deficient (KO) mice were used either as control, subjected to sham operation (cannulation or laparotomy only or cannulation plus laparotomy) or T-H (midline laparotomy, mean blood pressure 35 +/- 5 mmHg for 90 min, followed by resuscitation) and sacrificed 2 h thereafter. A marked increase in serum alpha-glutathione transferase, TNF-alpha, IL-6, IL-10, MCP-1, and MIP-1alpha and Kupffer cell cytokine production was observed in WT T-H mice compared with shams or control. In addition lung and liver tissue edema and neutrophil infiltration (myeloperoxidase (MPO) content) was also increased following T-H in WT animals. These inflammatory markers were markedly attenuated in the MIP-1alpha KO mice following T-H. Furthermore, compared with 2 h, MPO activities at 24 and 48 h after T-H declined steadily in both WT and KO mice. However, normalization of MPO activities to sham levels within 24 h was seen in KO mice but not in WT mice. Thus, MIP-1alpha plays an important role in mediating the acute inflammatory response following T-H. In the absence of MIP-1alpha, acute inflammatory responses were attenuated; rapidly recovered and less remote organ injury was noted following T-H. Thus, interventions that reduce MIP-1alpha levels following T-H should be useful in decreasing the deleterious inflammatory consequence of trauma.

Two-body wear test of enamel against laboratory polished and clinically adjusted zirconia.

AIM: A two-body wear test experiment was performed on human enamel, in simulated chewing motion, against non-veneered zirconia ceramic. Aim-1 was to ascertain the effect of zirconia roughness on enamel wear. Aim-2 was to ascertain the relative enamel wear between enamel-zirconia wear pair and enamel-enamel control pair. MATERIALS: Six molar and premolar human enamel cusps per group were used for a dental wear test against laboratory polished (LP) zirconia and laboratory polished and clinically adjusted (LP + CA) zirconia. Enamel antagonists were tested against incisor teeth as a control group to demonstrate laboratory enamel wear. METHODOLOGY: Two-body wear tests were conducted in a dual-axis biomimetic dental wear simulator. 49N loading force was used for 120,000 cycles with 1 mm lateral movement of the test specimen at 1.6Hz frequency, under constant ambient temperature water flow. Surface roughness before testing was determined using 3D profilometry. Loss of enamel height and volume i.e. vertical wear and volumetric wear respectively, were measured by superimposition of before and after testing scans by 3D laser scanning. Scanning electron microscopy was used for surface morphology assessment. One-way ANOVA and Post Hoc Multiple Comparisons with Bonferroni corrections were used at the 5% significance level to determine whether surface finish affected volumetric and vertical enamel loss. The relationship between volumetric and vertical loss of enamel was assessed using Pearson's correlation test. RESULTS: No significant difference was found between LP and LP + CA zirconia in vertical and volumetric enamel wear results. Control enamel had significantly higher vertical and volumetric enamel wear than LP and LP + CA zirconia. Pearson correlation revealed a strong relationship between vertical wear and volumetric wear of enamel. CONCLUSION: Within the constraints of the test method in this experiment, zirconia irrespective of surface preparation, was found to cause less vertical and volumetric enamel wear compared to control enamel. No statistically significant difference was seen between LP zirconia and LP + CA zirconia.

Mechanism of salutary effects of estrogen on cardiac function following trauma-hemorrhage: Akt-dependent HO-1 up-regulation.

OBJECTIVES: Because administration of 17beta-estradiol following trauma-hemorrhage improves cardiovascular responses, we investigated whether the salutary effects of 17beta-estradiol on cardiac function are mediated via Akt-dependent heme oxygenase-1 up-regulation under those conditions. DESIGN: Experimental animal study. SETTING: University laboratory. SUBJECTS: Male Sprague-Dawley rats. INTERVENTIONS: Rats underwent trauma-hemorrhage (mean blood pressure approximately 40 mm Hg for 90 mins) followed by fluid resuscitation. Before resuscitation, rats received either vehicle, 17beta-estradiol (1 mg/kg), or 17beta-estradiol plus the phosphoinositide 3-kinase inhibitor wortmannin (1 mg/kg). At 2 hrs after trauma-hemorrhage or sham operation, the rats were killed. MEASUREMENTS AND MAIN RESULTS: Cardiac function, heart tissue myeloperoxidase activity, cardiac and circulatory cytokine levels, cardiac intercellular adhesion molecule-1, and chemokine levels were measured. Cardiac Akt and heme oxygenase-1 were also determined. We found that 17beta-estradiol prevented the trauma-hemorrhage-induced impairment in cardiac function and increase in cardiac myeloperoxidase activity. Cardiac and systemic interleukin-6 and tumor necrosis factor-alpha levels as well as cardiac intercellular adhesion molecule-1, cytokine-induced neutrophil chemoattractant-1, and macrophage inflammatory protein-2 contents were increased following trauma-hemorrhage, which were normalized by 17beta-estradiol. Administration of 17beta-estradiol following trauma-hemorrhage restored cardiac Akt phosphorylation and further increased heme oxygenase-1 expression. Coadministration of wortmannin following trauma-hemorrhage abolished the previous effects by 17beta-estradiol. CONCLUSIONS: These results suggest that the 17beta-estradiol-meditated improvement in cardiac function following trauma-hemorrhage occurs via Akt-dependent heme oxygenase-1 up-regulation.

Estrogen ameliorates trauma-hemorrhage-induced lung injury via endothelial nitric oxide synthase-dependent activation of protein kinase G.

OBJECTIVE: In this study, we tested the hypothesis that 17beta-estradiol (E2) administration after trauma-hemorrhage reduces lung injury through a mechanism involving estrogen receptor (ER)-dependent activation of the endothelial nitric oxide (NO) synthase (eNOS)/protein kinase G (PKG)/vasodilator-stimulated phosphoprotein (VASP) pathway. BACKGROUND: Estrogen provides protection after injury via activation of multiple signaling cascades, including the cyclic GMP-dependent PKG pathway. Phosphorylation of VASP at Ser239 (p-VASP) can be used to assess PKG signaling activity. METHODS: Male Sprague-Dawley rats (275-325 g) underwent soft tissue trauma (midline laparotomy) and hemorrhagic shock (mean blood pressure 35-40 mm Hg for 90 minutes) followed by fluid resuscitation. Animals were pretreated with a nonselective NOS inhibitor (N(omega)-nitro-L-arginine methyl ester; 30 mg/kg), a soluble guanylyl cyclase (sGC) inhibitor [1H-(1, 2, 4) oxadiazolo (3, 4-alpha) quinoxalin-1-one; 10 mg/kg] or an ER antagonist (ICI 182,780; 3 mg/kg) 30 minutes before E2 (100 microg/kg) or vehicle administration. Animals were killed at 2 hours after resuscitation. RESULTS: Lung injury induced by trauma-hemorrhage is evidenced by edema (wet/dry ratio), neutrophil infiltration (myeloperoxidase activity), and with an increased expression of cytokines, chemokines, and adhesion molecules. E2 treatment after trauma-hemorrhage resulted in an increase in eNOS expression/phosphorylation, PKG-I activation, and VASP/p-VASP expression, which paralleled a decrease in lung injury. Inhibition of NOS and sGC abolished the E2-induced increase in PKG-I activity, VASP/p-VASP expression. Blockade of eNOS, PKG-I, and ER exacerbated lung inflammation and injury. CONCLUSIONS: These results collectively suggest that activation of the eNOS-PKG/VASP pathway by E2 protects against trauma-hemorrhage-induced lung injury.

Flutamide protects against trauma-hemorrhage-induced liver injury via attenuation of the inflammatory response, oxidative stress, and apopotosis.

Although studies have shown that administration of testosterone receptor antagonist, flutamide, following trauma-hemorrhage, improves hepatic, cardiovascular, and immune functions, the precise cellular/molecular mechanisms responsible for producing these salutary effects remain largely unknown. To study this, male C3H/HeN mice were subjected to a midline laparotomy and hemorrhagic shock (35+/-5 mmHg for approximately 90 min), followed by resuscitation with Ringer lactate. Flutamide (25 mg/kg) or vehicle was administered subcutaneously at the onset of resuscitation, and animals were killed 2 h thereafter. Hepatic injury was assessed by plasma alpha-glutathione S-transferase concentration, liver myeloperoxidase activity, and nitrotyrosine formation. Hepatic malondialdehyde and 4-hydroxyalkenals (lipid peroxidation indicators), cellular DNA fragmentation, and the expression of inducible nitric oxide synthase and hypoxia-inducible factor-1alpha were also evaluated. Cytokines (TNF-alpha, IL-6) and chemokines (keratinocyte-derived chemokine and monocyte chemoattractant protein-1) levels were determined by cytometric bead array. The results indicate that flutamide administration after trauma-hemorrhage reduced liver injury, which was associated with decreased levels of alpha-glutathione S-transferase, myeloperoxidase activity, nitrotyrosine formation, lipid peroxidation, and cytokines/chemokines (systemic, liver tissue, and intracellular cytokines/chemokines). Cellular apoptosis, hepatocyte hypoxia-inducible factor-1alpha, and inducible nitric oxide synthase expression were also decreased under such conditions. Thus administration of flutamide following trauma-hemorrhage protects against liver injury via reduced inflammation, cellular oxidative stress, and apoptosis.

Selective inhibition of iNOS attenuates trauma-hemorrhage/resuscitation-induced hepatic injury.

Although trauma-hemorrhage produces tissue hypoxia, systemic inflammatory response and organ dysfunction, the mechanisms responsible for these alterations are not clear. Using a potent selective inducible nitric oxide (NO) synthase inhibitor, N-[3-(aminomethyl) benzyl]acetamidine (1400W), and a nonselective NO synthase inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME), we investigated whether inducible NO synthase plays any role in producing hepatic injury, inflammation, and changes of protein expression following trauma-hemorrhage. To investigate this, male Sprague-Dawley rats were subjected to midline laparotomy and hemorrhagic shock (mean blood pressure 35-40 mmHg for approximately 90 min) followed by fluid resuscitation. Animals were treated with either vehicle (DMSO) or 1400W (10 mg/kg body wt ip), or L-NAME (30 mg/kg iv), 30 min before resuscitation and killed 2 h after resuscitation. Trauma-hemorrhage/resuscitation induced a marked hypotension and increase in markers of hepatic injury (i.e., plasma alpha-glutathione S-transferase, tissue myeloperoxidase activity, and nitrotyrosine formation). Hepatic expression of iNOS, hypoxia-inducible factor-1alpha, ICAM-1, IL-6, TNF-alpha, and neutrophil chemoattractant (cytokine-induced neutrophil chemoattractant-1 and macrophage inflammatory protein-2) protein levels were also markedly increased following trauma-hemorrhage/resuscitation. Administration of the iNOS inhibitor 1400W significantly attenuated hypotension and expression of these mediators of hepatic injury induced by trauma-hemorrhage/resuscitation. However, administration of L-NAME could not attenuate hepatic dysfunction and tissue injury mediated by trauma-hemorrhage, although it improved mean blood pressure as did 1400W. These results indicate that increased expression of iNOS following trauma-hemorrhage plays an important role in the induction of hepatic damage under such conditions.

Mechanism of estrogen-mediated attenuation of hepatic injury following trauma-hemorrhage: Akt-dependent HO-1 up-regulation.

Protein kinase B (Akt) is known to be involved in proinflammatory and chemotactic events in response to injury. Akt activation also leads to the induction of heme oxygenase (HO)-1. Up-regulation of HO-1 mediates potent, anti-inflammatory effects and attenuates organ injury. Although studies have shown that 17beta-estradiol (E2) prevents organ damage following trauma-hemorrhage, it remains unknown whether Akt/HO-1 plays any role in E2-mediated attenuation of hepatic injury following trauma-hemorrhage. To study this, male rats underwent trauma-hemorrhage (mean blood pressure, approximately 40 mmHg for 90 min), followed by fluid resuscitation. At the onset of resuscitation, rats were treated with vehicle, E2 (1 mg/kg body weight), E2 plus the PI-3K inhibitor (Wortmannin), or the estrogen receptor (ER) antagonist (ICI 182,780). At 2 h after sham operation or trauma-hemorrhage, plasma alpha-GST and hepatic tissue myeloperoxidase (MPO) activity, IL-6, TNF-alpha, ICAM-1, cytokine-induced neutrophil chemoattractant-1, and MIP-2 levels were measured. Hepatic Akt and HO-1 protein levels were also determined. Trauma-hemorrhage increased hepatic injury markers (alpha-GST and MPO activity), cytokines, ICAM-1, and chemokine levels. These parameters were markedly improved in the E2-treated rats following trauma-hemorrhage. E2 treatment also increased hepatic Akt activation and HO-1 expression compared with vehicle-treated, trauma-hemorrhage rats, which were abolished by coadministration of Wortmannin or ICI 182,780. These results suggest that the salutary effects of E2 on hepatic injury following trauma-hemorrhage are in part mediated via an ER-related, Akt-dependent up-regulation of HO-1.

Evaluation of ADCratio on liver MRI diffusion to discriminate benign versus malignant solid liver lesions.

PURPOSE: The aim of this project is to investigate the usefulness of the absolute liver lesion ADC value and ratio of Apparent diffusion coefficient (ADC) values of a liver lesion and liver parenchyma to discriminate between a benign and malignant lesion. METHODS: Liver MRI scans performed between January 2009 and June 2015 were retrospectively analysed. Scans were performed on either a 1.5 T or 3 T MRI unit. The type of liver lesion (benign or malignant) was determined by its radiological appearance, histology result and clinical management. Lesions with undetermined diagnosis or MRI studies degraded by artifacts were excluded. Liver cysts were also excluded from the analysis. ADC value of a lesion and liver parenchyma was measured and ADCratio was calculated. The values were analysed using independent samples t-test Results:Data set contained 39 benign lesions and 36 malignant lesions. Mean ADC value for benign lesions was 1678, and the mean value for malignant lesions was 1097 with a statistically significant difference of p < 0.001. All lesions with ADC value below 955 were malignant, while all lesions with ADC value above 1880 were benign. ADC value of 1260 was identified as the best available cut-off value for differentiating benign and malignant lesions, achieving sensitivity of 92%, specificity of 80% and an overall accuracy of 89%. The mean lesion to liver ADCratio for benign lesions was 1.3467 and for malignant lesions was 0.9038 with a statistically significant difference of p < 0.001. All lesions with ADCratio measuring <0.9 were malignant while lesions with ADCratio>1.5 were benign. ADCratio of 1.1 was identified statistically as the best available cut-off value for differentiating benign from malignant lesions, with sensitivity of 82%, specificity of 86% and an overall accuracy of 92%. CONCLUSION: Our dataset indicates that lesion to background liver ADCratio is superior in discriminating between benign and malignant focal lesions compared to absolute ADC values of the hepatic lesions.

Fracture toughness testing using photogrammetry and digital image correlation.

Digital image correlation (DIC) is an optical technique commonly used for measuring displacement fields by tracking artificially applied random speckle patterns, which can sometimes be a problem for tracking small-scale displacements. DIC is particularly useful for tracking the crack mouth opening displacement (CMOD) of a notched metallic specimen subjected to three-point bending for fracture toughness determination because the edges of the notch provide the required textural features for DIC without the need for speckle patterns. This simplifies the set-up process as the specimen and stage geometries do not need to account for the placement of a strain gauge. To enhance the accuracy of DIC, this study then successfully downscaled a photogrammetry technique commonly used to track crack propagation in large scale concrete tests so that the pixel coordinates of the captured images can be automatically related to their real-world coordinates, allowing for small scale displacements to be accurately tracked.

[Expression of intercellular adhesion molecule-1 in different organs of the mice with endotoxic shock induced by lipopolysaccharide].

To investigate and compare the expression of intercellular adhesion molecule-1 (ICAM-1) in different organs of the mice with endotoxic shock induced by lipopolysaccharide (LPS), protein and mRNA of ICAM-1 were measured by Western blotting and RT-PCR respectively in different organs of BALB/c mice administered intraperitoneally with 5 mg/kg LPS. The results showed that the constitutive expression of ICAM-1 protein and mRNA was the greatest in the lungs, followed by the spleen, kidney and intestine. After LPS stimulation, the upregulation of ICAM-1 was still greatest in the lungs, followed by the liver, spleen, heart, kidney and intestine. Compared with the normal mice, the expression of ICAM-1 protein in endotoxic shocked mice increased by 4.5-fold in the lungs, 3.0-fold in the kidney, 1.5-fold in the spleen; the expression in the liver and heart was negative under normal condition and changed into positive during endotoxic shock; but ICAM-1 expression in the intestine did not change significantly. The expression of ICAM-1 mRNA also increased consistently. These data highlight that LPS can up-regulate ICAM-1 protein and mRNA expression in different tissues of the mice with endotoxic shock. The difference in ICAM-1 expression among the organs may lead to different sensitivity of organ damage in endotoxic shock. This suggests that inhibition of ICAM-1 expression may be a useful principle for prevention and treatment of endotoxic shock.

Regulation of inducible nitricoxide synthase gene transcription by p38 mitogen-activated protein kinase in human embryonic kidney 293 cells.

OBJECTIVE: To study the transcriptional regulation of inducible nitric oxide synthase (iNOS) gene by p38 mitogen-activated protein kinase (MAPK). METHODS: With human embryonic kidney (HEK) 293 cells as the target and the assistance of lipofectamine-mediated co-transfection techniques and luciferase reporter gene systems, FLAG-tagged p38 isoforms (namely FLAG-p38 alpha, FLAG-p38 beta;, FLAG-p38 gamma and FLAG-p38 phi;) in pcDNA3, pcDNA3, piNOS-Luc and pCMV-beta; were transfected into HEK 293 cells, and the relative activity of luciferase was subsequently tested. RESULTS: Highest luciferase activity occurred only in p38 alpha group compared with the other three isoform groups under no stimulation. Under the stimulation by lipopolysaccharide (LPS), the luciferase activity of each group was obviously increased and the highest activity occurred in p38 beta group. CONCLUSION: LPS can induce transcription and activation of iNOS gene, and p38 MAPK is involved in the transcription regulation of iNOS gene in HEK 293 cells.

Role of p38 mitogen-activated protein kinase in lipopolysaccharide-induced expression of inducible nitric oxide synthase in human endothelial cells.

OBJECTIVE: To understand the role of p38 mitogen-activated protein kinase (p38MAPK) in the expression of inducible nitric oxide synthase (iNOS) and NO production in human endothelial cells under the stimulation by lipopolysaccharide (LPS). METHODS: NO level in the supernatant of the cell culture media was measured with Griess method, and iNOS protein and mRNA expressions by the cells were detected with immunofluorescence analysis and reverse transcriptase-polymerase chain reaction (RT-PCR) respectively. Immunoprecipitation assay was employed to examine p38 MAPK activity. RESULTS: It was shown that in comparison with the basal level of iNOS expression in cultured endothelial cells line ECV304, iNOS mRNA and protein expressions were significantly increased in the cells after LPS stimulation. In response to LPS treatment, obvious enhancement of p38 MAPK activity in ECV304 took place after the stimulation, with the peak level occurring at 15 min that maintained for approximately 45 min before gradual decline. When treated with SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl) imidazole], a highly specific inhibitor of p38 MAPK, significant inhibition of LPS-induced iNOS protein and mRNA expressions was observed. CONCLUSIONS: p38 MAPK plays an important role in iNOS expression and NO production in ECV304 cells, and, inhibition of the signal transduction pathway may consequently be an effective approach to reduce the production of iNOS and other cytokines for the treatment of septic shock.