Role of histamine receptors in the effects of histamine on the production of reactive oxygen species by whole blood phagocytes

Aims

The diverse physiological functions of histamine are mediated through distinct histamine receptors. In this study we investigated the role of H₂R and H₄R in the effects of histamine on the production of reactive oxygen species by phagocytes in whole blood.

Main methods

Changes in reactive oxygen species (ROS) production by whole blood phagocytes after treatment with histamine, H₄R agonists (4-methylhistamine, VUF8430), H₂R agonist (dimaprit) and their combinations with H₄R antagonist (JNJ10191584) and H₂R antagonist (ranitidine) were determined using the chemiluminescence (CL) assay. To exclude the direct scavenging effects of the studied compounds on the CL response, the antioxidant properties of all compounds were measured using several methods (TRAP, ORAC, and luminol–HRP–H₂O₂ based CL).

Key findings

Histamine, 4-methylhistamine, VUF8430 and dimaprit inhibited the spontaneous and OZP-activated whole blood CL in a dose-dependent manner. On the other hand, only VUF8430 was able to inhibit PMA-activated whole blood CL. Ranitidine, but not JNJ10191584, completely reduced the effects of histamine, 4-methylhistamine and dimaprit. The direct scavenging ability of tested compounds was negligible.

Significance

Our results demonstrate that the inhibitory effects of histamine on ROS production in whole blood phagocytes were caused by H₂R. Our results also suggest that H₄R agonists in concentrations higher than 10^− 6 M may also influence ROS production via binding to H₂R.     

Mouse anaphylactic shock is caused by reduced cardiac output,but not by systemic vasodilatation or pulmonary vasoconstriction,via PAF and histamine

Aims

Systemic anaphylaxis is life-threatening, and its pathophysiology is not fully clarified. Mice are frequently used for experimental study on anaphylaxis. However, the hemodynamic features and mechanisms of mouse anaphylactic hypotension remain unknown. Therefore, we determined mechanisms of systemic and pulmonary vascular response to anaphylactic hypotension in anesthetized BALB/c mice by using receptor antagonists of chemical mediators.

Main methods

Anaphylaxis was actively induced by an intravenous injection of the ovalbumin antigen into open-chest artificially ventilated sensitized mice. Mean arterial pressure (MAP), pulmonary arterial pressure (PAP), left atrial pressure, central venous pressure, and aortic blood flow (ABF) were continuously measured.

Key findings

In sensitized control mice, MAP and ABF showed initial, transient increases, followed by progressive decreases after the antigen injection. Total peripheral resistance (TPR) did not decrease, while PAP initially and transiently increased to 18.5 ± 0.5 mm Hg and pulmonary vascular resistance (PVR) also significantly increased. The antigen-induced decreases in MAP and ABF were attenuated by pretreatment with either a platelet-activating factor (PAF) receptor antagonist, CV6209, or a histamine H₁ receptor antagonist, diphenhydramine, and were abolished by their combination. Diphenhydramine augmented the initial increases in PAP and PVR, but did not affect the decrease of the corresponding MAP fall. The antagonists of either leukotriene C₄ or serotonin, alone or in combination with CV6209, exerted no significant effects.

Significance

Mouse anaphylactic hypotension is caused by a decrease in cardiac output but not vasodilatation, via actions of PAF and histamine. The slight increase in PAP is not involved in mouse anaphylactic hypotension.     

The (2′S,7′S)-O-(2-methylbutanoyl)-columbianetin as a novel allergic rhinitis-control agent

Aims

The (2′S,7′S)-O-(2-methylbutanoyl)-columbianetin (OMC) is a novel secondary metabolite extracted from Corydalis heterocarpa, which has long been used as a folk medicine for various inflammatory diseases in Korea. We examined the effect of OMC on allergic rhinitis (AR).

Main methods

We assessed the therapeutic effects and regulatory mechanisms of OMC on the phorbol 12-myristate 13-acetate plus A23187-stimulated mast cell line, HMC-1 cells and ovalbumin (OVA)-induced AR models.

Key findings

OMC significantly decreased the releases of histamine and tryptase from stimulated HMC-1 cells. The degranulation process, characterized by morphological extension of the filopodia on the surface and membrane ruffling, was strongly induced in the stimulated-HMC-1 cell, however OMC suppressed the morphological changes in stimulated-HMC-1 cells. OMC reduced the production and mRNA expression of inflammatory cytokines. These inhibitory actions by OMC were dependent on the regulation of mitogen-activated protein kinases, nuclear factor-κB, and caspapase-1 signaling pathways. In the AR animal model, the increased rub scores and AR biomarkers (histamine and IgE) in ovalbumin (OVA)-sensitized mice were significantly reduced by the administration of OMC. Furthermore, eosinophils and mast cell infiltrations in nasal mucosa tissue were also blocked through the regulation of macrophage-inflammatory protein and intercellular adhesion molecule-1 levels.

Significance

OMC showed the possibility to regulate AR in activated mast cells and OVA-induced AR models. Hence, we suggest that OMC is a powerful and feasible new agent to suppress AR.     

Azoxystrobin,a mitochondrial complex III Qo site inhibitor,exerts beneficial metabolic effects in vivo and in vitro

Background

Several anti-diabetes drugs exert beneficial effects against metabolic syndrome by inhibiting mitochondrial function. Although much progress has been made toward understanding the role of mitochondrial function inhibitors in treating metabolic diseases, the potential effects of these inhibitors on mitochondrial respiratory chain complex III remain unclear.

Methods

We investigated the metabolic effects of azoxystrobin (AZOX), a Q_o inhibitor of complex III, in a high-fat diet-fed mouse model with insulin resistance in order to elucidate the mechanism by which AZOX improves glucose and lipid metabolism at the metabolic cellular level.

Results

Acute administration of AZOX in mice increased the respiratory exchange ratio. Chronic treatment with AZOX reduced body weight and significantly improved glucose tolerance and insulin sensitivity in high-fat diet-fed mice. AZOX treatment resulted in decreased triacylglycerol accumulation and down-regulated the expression of genes involved in liver lipogenesis. AZOX increased glucose uptake in L6 myotubes and 3T3-L1 adipocytes and inhibited de novo lipogenesis in HepG2 cells. The findings indicate that AZOX-mediated alterations to lipid and glucose metabolism may depend on AMP-activated protein kinase (AMPK) signaling.

Conclusions

AZOX, a Q_o inhibitor of mitochondrial respiratory complex III, exerts whole-body beneficial effects on the regulation of glucose and lipid homeostasis in high-fat diet-fed mice.

General significance

These findings provide evidence that a Q_o inhibitor of mitochondrial respiratory complex III could represent a novel approach for the treatment of obesity.     

Prostaglandin E2 produced by inducible COX-2 and mPGES-1 promoting cancer cell proliferation in vitro and in vivo

Aim

Many cancers originate and flourish in a prolonged inflammatory environment. Our aim is to understand the mechanisms of how the pathway of prostaglandin E₂ (PGE₂) biosynthesis and signaling can promote cancer growth in inflammatory environment at cellular and animal model levels.

Main methods

In this study, a chronic inflammation pathway was mimicked with a stable cell line that over-expressed a novel human enzyme consisting of cyclooxygenase isoform-2 (COX-2) linked to microsomal (PGE₂ synthase-1 (mPGES-1)) for the overproduction of pathogenic PGE₂. This PGE₂-producing cell line was co-cultured and co-implanted with three human cancer cell lines including prostate, lung, and colon cancers in vitro and in vivo, respectively.

Key findings

Increases in cell doubling rates for the three cancer cell types in the presence of the PGE₂-producing cell line were clearly observed. In addition, one of the four human PGE₂ subtype receptors, EP₁, was used as a model to identify PGE₂-signaling involved in promoting the cancer cell growth. This finding was further proven in vivo by co-implanting the PGE₂-producing cells line and the EP₁-positive cancer cells into the immune deficient mice, after that, it was observed that the PGE₂-producing cells promoted all three types of cancer formation in the mice.

Significance

This study clearly demonstrated that the human COX-2 linked to mPGES-1 is a pathway that, when mediated by the EP, is linked to promoting cancer growth in a chronic inflammatory environment. The identified pathway could be used as a novel target for developing and advancing anti-inflammation and anti-cancer interventions.     

Carvacryl acetate,a derivative of carvacrol,reduces nociceptive and inflammatory response in mice

Aims

The present study aimed to investigate the potential anti-inflammatory and anti-nociceptive effects of carvacryl acetate, a derivative of carvacrol, in mice.

Main methods

The anti-inflammatory activity was evaluated using various phlogistic agents that induce paw edema, peritonitis model, myeloperoxidase (MPO) activity, pro and anti-inflammatory cytokine levels. Evaluation of antinociceptive activity was conducted through acetic acid-induced writhing, hot plate test, formalin test, capsaicin and glutamate tests, as well as evaluation of motor performance on rotarod test.

Key findings

Pretreatment of mice with carvacryl acetate (75 mg/kg) significantly reduced carrageenan-induced paw edema (P < 0.05) when compared to vehicle-treated group. Likewise, carvacryl acetate (75 mg/kg) strongly inhibited edema induced by histamine, serotonin, prostaglandin E₂ and compound 48/80. In the peritonitis model, carvacryl acetate significantly decreased total and differential leukocyte counts, and reduced levels of myeloperoxidase and interleukin-1 beta (IL-1β) in the peritoneal exudate. The levels of IL-10, an anti-inflammatory cytokine, were enhanced by carvacryl acetate. Pretreatment with carvacryl acetate also decreased the number of acetic acid-induced writhing, increased the latency time of the animals on the hot plate and decreased paw licking time in the formalin, capsaicin and glutamate tests. The pretreatment with naloxone did not reverse the carvacryl acetate-mediated nociceptive effect.

Significance

In conclusion, the current study demonstrated that carvacryl acetate exhibited anti-inflammatory activity in mice by reducing inflammatory mediators, neutrophil migration and cytokine concentration, and anti-nociceptive activity due to the involvement of capsaicin and glutamate pathways.     

Sex Differences in Itch Perception and Modulation by Distraction – an fMRI Pilot Study in Healthy Volunteers

Background

Even though itch is a common syndrome of many diseases there is only little knowledge about sex and gender differences in pruritus, especially in central itch perception and modulation. To our knowledge, this is the first fMRI study examining sex differences in perception and its modulation by distraction.

Methods

Experimental itch was induced by application of histamine (0.1 mM) via microdialysis fibers twice at the left forearm and twice at the left lower leg in 33 healthy volunteers (17 females, 16 males). The brain activation patterns were assessed by fMRI during itch without and with distraction (Stroop task). Between the various conditions, subjects were asked to rate itch intensity, desire to scratch and pain intensity. In a second experiment in 10 of the 33 volunteers histamine was replaced by saline solution to serve as control for the ‘Stroop’ condition.

Results

Women generally presented higher itch intensities compared to men during itch over the course of the experiment. A more specific analysis revealed higher itch intensities and desire to scratch in women during experimental induced itch that can be reduced by distraction at the lower legs when itch is followed by ‘Stroop’. In contrast, men depicted significant reduction of ‘itch’ by ‘Stroop’ at the forearms. Women depicted higher brain activation of structures responsible for integration of sensory, affective information and motor integration/planning during ‘itch’ and ‘Stroop’ condition when compared to men. No sex differences were seen in the saline control condition.

Conclusion

Women and men exhibited localisation dependent differences in their itch perception with women presenting higher itch intensities and desire to scratch. Our findings parallel clinical observations of women reporting higher itch intensities depending on itch localisation and suffering more from itch as compared to men.     

Cardioprotection of H2S by downregulating iNOS and upregulating HO-1 expression in mice with CVB3-induced myocarditis

Aims

To explore the effects and potential mechanisms of hydrogen sulfide (H₂S) in CVB3-induced mice with myocarditis.

Main methods

A total of 75 six-week-old inbred male Balb/c mice were divided randomly into four groups (N, C, P and S). Group N was the negative control. The others were inoculated intraperitoneally (i.p.) with CVB3. Subsequently, groups P and S were injected i.p. once a day with DL-Proparglygylcine (PAG) and NaHS respectively. Group C was the positive control. Inducible nitric oxide synthase (iNOS) and heme oxygenase-1(HO-1) expression on cardiac tissues were evaluated by histopathological examination, immunohistochemistry, RT-PCR and Western blot.

Key findings

The heart-weight to body-weight (HW/BW) ratio, the histologic scores and the iNOS mRNA and protein expression levels were higher, and the HO-1 mRNA and protein expression levels were lower in mice treated with PAG than those mice solely inoculated with CVB3. Mice in group S had a significant decreased in the HW/BW ratio, the histologic scores and the iNOS mRNA and protein expression levels, and had a significant increased in the HO-1 mRNA and protein expression levels compared to the mice in group C. H₂S can attenuate inflammatory cell infiltration, alleviate cardiac edema, and limit myocardial lesions.

Significance

Our data support that H₂S can inhibit iNOS overexpression and induce HO-1 expression, both of which contribute to the cardioprotection of H₂S in CVB3-induced mice myocarditis.     

Oxygen-induced retinopathy in mice with retinal photoreceptor cell degeneration

Aims

It is reported that retinal neovascularization seems to rarely co-exist with retinitis pigmentosa in patients and in some mouse models; however, it is not widely acknowledged as a universal phenomenon in all strains of all animal species. We aimed to further explore this phenomenon with an oxygen-induced retinopathy model in mice with retinal photoreceptor cell degeneration.

Main methods

Oxygen-induced retinopathy of colored and albino mice with rapid retinal degeneration were compared to homologous wild-type mice. The retinas were analyzed using high-molecular-weight FITC-dextran stained flat-mount preparation, hematoxylin and eosin (H&E) stained cross-sections, an immunohistochemical test for vascular endothelial growth factor (VEGF) distribution and Western blotting for VEGF expression after exposure to hyperoxia between postnatal days 17 (P17) and 21.

Key findings

Leakage and areas of non-perfusion of the retinal blood vessels were alleviated in the retinal degeneration mice. The number of preretinal vascular endothelial cell nuclei in the retinal degeneration mice was smaller than that in the homologous wild-type mice after exposure to hyperoxia (P < 0.01). The degree of oxygen-induced retinopathy was positively correlated with the VEGF expression level. However, the VEGF expression level was lower in the retinal degeneration mice.

Significance

Proliferative retinopathy occurred in mice with rapid retinal degeneration, but retinal photoreceptor cell degeneration could partially restrain the retinal neovascularization in this rapid retinal degeneration mouse model.     

The acute antinociceptive effect of hyperbaric oxygen is not accompanied by an increase in markers of oxidative stress

Aims

Exposure to hyperbaric oxygen (HBO₂) causes an antinociceptive response in mice. However, breathing oxygen (O₂) at an elevated pressure can potentially cause oxygen toxicity. The aim of this study was to identify the determinants of HBO₂ antinociception and the toxicity profile of HBO₂.

Main methods

Male NIH Swiss mice were assessed for acute antinociceptive responsiveness under room air or 100% O₂ at 1.0 or 3.5 atmospheres absolute (ATA), using the acetic acid-induced abdominal constriction test. For the oxygen toxicity test, mice were exposed to 3.5 ATA oxygen for 11 min, 60 min, and 60 min daily for 2 days (120 min) or 60 min daily for 4 days (240 min), then assessed by analyzing the levels of two oxidative stress markers, MDA (malondialdehyde) and protein carbonyl in brain, spinal cord and lung.

Key findings

Only the combination of 100% O₂ and 3.5 ATA caused significant antinociception. The antinociceptive effect of 100% O₂ was pressure-dependent up to 3.5 ATA. In the oxygen toxicity test, mice exposed to HBO₂ for different time intervals had levels of brain, spinal cord and lung MDA and protein carbonyl that were comparable to that of control animals exposed to room air.

Significance

Treatment with 100% O₂ evokes a pressure-dependent antinociceptive effect. Since there was no significant increase in levels of the oxidative stress markers in the tested tissues, it is concluded that HBO₂ at 3.5 ATA produces antinociception in the absence of oxidative stress in mice.     

Perinatal inflammation results in decreased oligodendrocyte numbers in adulthood

Aims

Maternal inflammation is a risk factor for preterm birth, and premature infants are often exposed to supplemental oxygen as a life-sustaining therapy. While more immature neonates are surviving, rates of neurodevelopmental impairment are not improving. We developed a novel mouse model with clinically relevant exposures to test the hypothesis that systemic maternal inflammation with transient neonatal hyperoxia exposure will induce a phenotype similar to diffuse periventricular leukomalacia (PVL) like that observed in premature human infants.

Main methods

Timed-pregnant C3H/HeN mice received intraperitoneal injections of lipopolysaccharide (LPS) or saline on embryonic day 16. Newborn pups were placed in room air (RA) or 85% oxygen (O₂) for 14 days, followed by 14 days in RA recovery. Oligodendroglial and microglial populations were evaluated at 14 and 28 days.

Key findings

Brain weight to body weight ratios were lower in mice exposed to LPS. Oligodendrocyte numbers were decreased significantly in the cerebral cortex and hippocampus in groups exposed to LPS or LPS/O₂ at 14 days, and persisted in the cerebral cortex at 28 days for LPS/O₂ mice. At day 14, cleaved caspase 3 was increased and numbers of microglia were elevated in the cerebral cortex and hippocampus of LPS/O₂ animals.

Significance

These data indicate that combining systemic maternal LPS and neonatal hyperoxic exposure impairs myelination, and suggests that this novel mouse model may represent a subtle, diffuse form of periventricular white matter injury that could provide a clinically relevant platform for further study of perinatal brain injury.     

Intracellular Ca2+ transients in delta-sarcoglycan knockout mouse skeletal muscle

Background

δ-Sarcoglycan (δ-SG) knockout (KO) mice develop skeletal muscle histopathological alterations similar to those in humans with limb muscular dystrophy. Membrane fragility and increased Ca²⁺ permeability have been linked to muscle degeneration. However, little is known about the mechanisms by which genetic defects lead to disease.

Methods

Isolated skeletal muscle fibers of wild-type and δ-SG KO mice were used to investigate whether the absence of δ-SG alters the increase in intracellular Ca²⁺ during single twitches and tetani or during repeated stimulation. Immunolabeling, electrical field stimulation and Ca²⁺ transient recording techniques with fluorescent indicators were used.

Results

Ca²⁺ transients during single twitches and tetani generated by muscle fibers of δ-SG KO mice are similar to those of wild-type mice, but their amplitude is greatly decreased during protracted stimulation in KO compared to wild-type fibers. This impairment is independent of extracellular Ca²⁺ and is mimicked in wild-type fibers by blocking store-operated calcium channels with 2-aminoethoxydiphenyl borate (2-APB). Also, immunolabeling indicates the localization of a δ-SG isoform in the sarcoplasmic reticulum of the isolated skeletal muscle fibers of wild-type animals, which may be related to the functional differences between wild-type and KO muscles.

Conclusions

δ-SG has a role in calcium homeostasis in skeletal muscle fibers.

General significance

These results support a possible role of δ-SG on calcium homeostasis. The alterations caused by the absence of δ-SG may be related to the pathogenesis of muscular dystrophy.     

LOX-1 deletion and macrophage trafficking in atherosclerosis

Background

Atherosclerosis is associated with macrophage accumulation. LOX-1 has been shown to induce macrophage attachment, and its deletion (LOX-1 knockout, KO) reduces atherosclerosis in LDLr KO mice fed a high cholesterol diet. We examined differences in macrophage trafficking in age-matched wild type, LOX-1 KO, LDLr KO, and LDLr/LOX-1 double KO mice.

Methods

Sections of aortas of mice fed high cholesterol diet were collected at weeks 0, 4, 8, 12 and 19 and analyzed by immunohistochemistry and flow cytometry.

Results

In the LDLr KO mice aorta, CD68 positivity (macrophage accumulation) increased over time up to 12 weeks, and then the accumulation fell modestly but significantly. The periaortal fat and adventitia showed more CD68 positivity than the media and intima. This pattern was also evident in the non-atherosclerotic areas. Importantly, LOX-1 KO and LDLr–LOX-1 double KO mice showed diminished CD68 positivity in comparison to wild type and LDLR KO mice, respectively. Further, macrophages from LOX-1 KO mice revealed a marked reduction in migration (vs. macrophages from wild type mice) in in vitro migration assay.

Conclusions

LOX-1 deletion translates into reduction in macrophage trafficking in the aorta of LDLr KO mice. Most of the macrophage trafficking appears in the subadventitial regions.     

Magainin-AM2 improves glucose homeostasis and beta cell function in high-fat fed mice

Background

Magainin-AM2, a previously described amphibian host-defense peptide, stimulates insulin- and glucagon-like peptide-1-release in vitro. This study investigated anti-diabetic effects of the peptide in mice with diet-induced obesity and glucose intolerance.

Methods

Male National Institute of Health Swiss mice were maintained on a high-fat diet for 12-weeks prior to the daily treatment with magainin-AM2. Various indices of glucose tolerance were monitored together with insulin secretory responsiveness of islets at conclusion of study.

Results

Following twice daily treatment with magainin-AM2 for 15 days, no significant difference in body weight and food intake was observed compared with saline-treated high fat control animals. However, non-fasting blood glucose was significantly (P < 0.05) decreased while plasma insulin concentrations were significantly (P < 0.05) increased. Oral and intraperitoneal glucose tolerance and insulin secretion following glucose administration via both routes were significantly (P < 0.05) enhanced. The peptide significantly (P < 0.001) improved insulin sensitivity as well as the beta cell responses of islets isolated from treated mice to a range of insulin secretagogues. Oxygen consumption, CO₂ production, respiratory exchange ratio and energy expenditure were not significantly altered by sub-chronic administration of magainin-AM2 but a significant (P < 0.05) reduction in fat deposition was observed.

Conclusion

These results indicate that magainin-AM2 improves glucose tolerance, insulin sensitivity and islet beta cells secretory responsiveness in mice with obesity-diabetes.

General significance

The activity of magainin-AM2 suggests the possibility of exploiting this peptide for treatment of type 2 diabetes.     

Carbamoyl-PROXYL-enhanced MRI detects very small disruptions in brain vascular permeability induced by dietary cholesterol

Background

Gd-DTPA-enhanced magnetic resonance imaging (MRI) is a conventional method for non-invasive investigation of blood-brain-barrier (BBB) permeability in animal models. It allows the visualization of serious injury to the BBB. We developed a novel approach for detecting very small disruptions in BBB permeability induced by dietary cholesterol by using carbamoyl-PROXYL (CMP) as an MRI contrast probe.

Methods

Mice were separated into two groups: normal diet (ND-mice) and high cholesterol diet (CD-mice). MRI-signal dynamics, plasma cholesterol, matrix metalloproteinase (MMP-9, MMP-2), and the white blood cell profile were analyzed. For the MRI analysis, two regions-of-interest (ROI) were selected: brain (ROI-1) and surrounding area (ROI-2).

Results

In the ROI-2 of ND-mice, CMP- or Gd-enhanced MRI-signal followed typical kinetics with a half-life of signal decay (τ_1/2) ~ 8 or ~ 15 min, respectively. In CD-mice, the MRI-signal increased continuously without decay.In the ROI-1 of ND- and CD-mice, MRI-signal enhancement was not detected by Gd-DTPA. In the ROI-1 of ND-mice, CMP-induced MRI-signal enhancement was negligible, while in CD-mice, it was significant (τ_1/2 > 15 min).Hypercholesterolemia increased the plasma levels of MMP-9 and neutrophils.

Conclusions

Hypercholesterolemia increases vascular permeability, which is mediated by MMP-9 and neutrophils.

General significance

Even very small disruptions in brain vascular permeability could be detected by CMP-enhanced MRI but not by Gd-DTPA-enhanced MRI.     

Propranolol impairs the closure of pressure ulcers in mice

Aims

β-Adrenoceptors modulate acute wound healing; however, few studies have shown the effects of β-adrenoceptor blockade on chronic wounds. Therefore, this study investigated the effect of β₁-/β₂-adrenoceptor blockade in wound healing of pressure ulcers.

Main methods

Male mice were daily treated with propranolol (β₁-/β₂-adrenoceptor antagonist) until euthanasia. One day after the beginning of treatment, two cycles of ischemia–reperfusion by external application of two magnetic plates were performed in skin to induce pressure ulcer formation.

Key findings

Propranolol administration reduced keratinocyte migration, transforming growth factor-β protein expression, re-epithelialization, and necrotic tissue loss. Neutrophil number and neutrophil elastase protein expression were increased in propranolol-treated group when compared with control group. Propranolol administration delayed macrophage mobilization and metalloproteinase-12 protein expression and reduced monocyte chemoattractant protein-1 protein expression. Myofibroblastic differentiation, angiogenesis, and wound closure were delayed in the propranolol-treated animals. Propranolol administration increased neo-epidermis thickness, reduced collagen deposition, and enhanced tenascin-C expression resulting in the formation of an immature and disorganized collagenous scar.

Significance

β₁-/β₂-Adrenoceptor blockade delays wound healing of ischemia–reperfusion skin injury through the impairment of the re-epithelialization and necrotic tissue loss which compromise wound inflammation, dermal reconstruction, and scar formation.     

Overexpression of human Argonaute2 inhibits cell and tumor growth

Background

Argonaute (Ago) proteins are essential for the biogenesis and function of ~ 20–30 nucleotide long RNAs such as microRNAs (miRNAs). Ago expression increases or decreases under various physiological conditions, although the functional consequences are unknown. In addition, while reduced global miRNA production was shown to enhance cellular transformation and tumorigenesis, how Ago proteins contribute to human diseases has not been reported.

Method

Ago2, an essential Ago isoform in mammals, was stably expressed in 293 T, the human embryonic kidney cell line, and H1299, the human lung adenocarcinoma cell line. miRNA and mRNA expression was investigated by quantitative PCR and microarray profiling. Cell proliferation and migration was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and scratch assay in the cell cultures, respectively. How Ago2 affected cell growth in vivo was determined by H1299 xenograft tumor growth in mice. Changes in Ago2 expression in human lung cancer samples were investigated by quantitative PCR and immunohistochemistry.

Results

Stable Ago2 overexpression elicited specific changes in miRNA and mRNA expression in both 293 T and H1299 cells. It also inhibited cell proliferation and migration in cell cultures as well as xenograft tumor growth in nude mice. Ago2 expression was lower in human lung adenocarcinomas than in the paired, non-cancerous tissues.

General significance

We concluded that changes in Ago2 expression might have significant physiological and pathological consequences in vivo.     

Selenium-binding protein 1: Its physiological function,dependence on aryl hydrocarbon receptors,and role in wasting syndrome by 2,3,7,8-tetrachlorodibenzo-p-dioxin

Background

Selenium-binding protein 1 (Selenbp1) is suggested to play a role in tumor suppression, and may be involved in the toxicity produced by dioxin, an activator of aryl hydrocarbon receptors (AhR). However, the mechanism or likelihood is largely unknown because of the limited information available about the physiological role of Selenbp1.

Methods

To address this issue, we generated Selenbp1-null [Selenbp1 (−/−)] mice, and examined the toxic effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in this mouse model.

Results

Selenbp1 (−/−) mice exhibited only a few differences from wild-type mice in their apparent phenotypes. However, a DNA microarray experiment showed that many genes including Notch1 and Cdk1, which are known to be enhanced in ovarian carcinoma, are also increased in the ovaries of Selenbp1 (−/−) mice. Based on the different responses to TCDD between C57BL/6J and DBA/2J strains of mice, the expression of Selenbp1 is suggested to be under the control of AhR. However, wasting syndrome by TCDD occurred equally in Selenbp1 (−/−) and (+/+) mice.

Conclusions

The above pieces of evidence suggest that 1) Selenbp1 suppresses the expression of tumor-promoting genes although a reduction in Selenbp1 alone is not very serious as far as the animals are concerned; and 2) Selenbp1 induction by TCDD is neither a pre-requisite for toxicity nor a protective response for combating TCDD toxicity.

General significance

Selenbp1 (−/−) mice exhibit little difference in their apparent phenotype and responsiveness to dioxin compared with the wild-type. This may be due to the compensation of Selenbp1 function by a closely-related protein, Selenbp2.     

Selenoprotein W serves as an antioxidant in chicken myoblasts

Background

Selenoprotein W (SelW) was thought to play an antioxidant role in mammals. Because chicken SelW has no cysteine (Cys) at the residue 37 (Cys37) that is required for the presumed antioxidant function in mammals, this study was conducted to determine whether chicken SelW possessed the same function.

Methods

Small interfering RNAs (siRNAs) technology was applied to suppress the SelW expression in chicken embryonic myoblasts. Thereafter, these myoblasts were treated with different concentrations of H₂O₂ and assayed for cell viability, apoptosis rate, reactive oxygen species (ROS) status, and expression levels of apoptosis-related genes and proteins (Bax, Bcl-2, and caspase-3).

Results

Silencing of the myoblast SelW gene decreased their cell viability, and increased their apoptosis rate and susceptibility to H₂O₂. While the knockout down of SelW up-regulated Bax and caspase-3 and down-regulated Bcl-2, the induced oxidative injuries were alleviated by treatment with a ROS scavenger, N-acetyl-l-cysteine (NAC).

Conclusion

Chicken SelW protected embryonic myoblasts against cell apoptosis mediated by endogenous and exogenous H₂O₂.

General significance

Chicken SelW possesses antioxidant function similar to the mammalian homologues despite the lack of Cys37 in the peptide.