Role of Nrf2 in host defense against influenza virus in cigarette smoke-exposed mice

Influenza virus is a common respiratory tract viral infection. Although influenza can be fatal in patients with chronic pulmonary diseases such as chronic obstructive pulmonary disease, its pathogenesis is not fully understood. The Nrf2-mediated antioxidant system is essential to protect the lungs from oxidative injury and inflammation. In the present study, we investigated the role of Nrf2 in protection against influenza virus-induced pulmonary inflammation after cigarette smoke exposure with both in vitro and in vivo approaches. For in vitro analyses, peritoneal macrophages isolated from wild-type and Nrf2-deficient mice were treated with poly(I:C) and/or cigarette smoke extract. For in vivo analysis, these mice were infected with influenza A virus with or without exposure to cigarette smoke. In Nrf2-deficient macrophages, NF-κB activation and the induction of its target inflammatory genes were enhanced after costimulation with cigarette smoke extract and poly(I:C) compared with wild-type macrophages. The induction of antioxidant genes was observed for the lungs of wild-type mice but not those of Nrf2-deficient mice after cigarette smoke exposure. Cigarette smoke-exposed Nrf2-deficient mice showed higher rates of mortality than did wild-type mice after influenza virus infection, with enhanced peribronchial inflammation, lung permeability damage, and mucus hypersecretion. Lung oxidant levels and NF-κB-mediated inflammatory gene expression in the lungs were also enhanced in Nrf2-deficient mice. Our data indicate that the antioxidant pathway controlled by Nrf2 is pivotal for protection against the development of influenza virus-induced pulmonary inflammation and injury under oxidative conditions.     

uPA deficiency exacerbates muscular dystrophy in MDX mice

Duchenne muscular dystrophy (DMD) is a fatal and incurable muscle degenerative disorder. We identify a function of the protease urokinase plasminogen activator (uPA) in mdx mice, a mouse model of DMD. The expression of uPA is induced in mdx dystrophic muscle, and the genetic loss of uPA in mdx mice exacerbated muscle dystrophy and reduced muscular function. Bone marrow (BM) transplantation experiments revealed a critical function for BM-derived uPA in mdx muscle repair via three mechanisms: (1) by promoting the infiltration of BM-derived inflammatory cells; (2) by preventing the excessive deposition of fibrin; and (3) by promoting myoblast migration. Interestingly, genetic loss of the uPA receptor in mdx mice did not exacerbate muscular dystrophy in mdx mice, suggesting that uPA exerts its effects independently of its receptor. These findings underscore the importance of uPA in muscular dystrophy.     

Chemokine receptor CCR5 deficiency exacerbates cerulein-induced acute pancreatitis in mice

Acute pancreatitis (AP) is an inflammatory disease involving the production of different cytokines and chemokines and is characterized by leukocyte infiltration. Because the chemokine receptor CCR5 and its ligands [the CC chemokines CCL3/MIP-1alpha, CCL4/MIP-1beta, and CCL5/regulated upon activation, normal T cell expressed and secreted (RANTES)] regulate leukocyte chemotaxis and activation, we investigated the expression of CCR5 ligands and the role of CCR5 and its ligands in experimental AP in mice. AP was induced by hourly intraperitoneal injections of cerulein in CCR5-deficient (CCR5(-/-)) or wild-type (WT) mice. Induction of AP by cerulein resulted in an early increase of pancreatic CCL2, CCL3, and CCL4 mRNA expression, whereas CCL5 mRNA expression occurred later. CCR5(-/-) mice developed a more severe pancreatic injury than WT mice during cerulein-induced AP, as assessed by a more pronounced increase in serum amylase and lipase levels and by more severe pancreatic edema, inflammatory infiltrates (mainly neutrophils), and necrosis. CCR5(-/-) mice also exhibited increased production of CCL2/MCP-1, CCL3/MIP-1alpha, and CCL4/MIP-1beta during the course of cerulein-induced AP. In vivo simultaneous neutralization of CC chemokines with monoclonal antibodies in CCR5(-/-) mice reduced the severity of cerulein-induced AP, indicating a role of CC chemokines in exacerbating the course of AP in the absence of CCR5. Moreover, simultaneous neutralization of CCR5 ligands in WT mice also reduced the severity of cerulein-induced AP. In conclusion, lack of the chemokine receptor CCR5 exacerbates experimental cerulein-induced AP and leads to increased levels of CC chemokines and a more pronounced pancreatic inflammatory infiltrate, suggesting that CCR5 expression can modulate severity of AP.     

Vitamin D deficiency in undifferentiated connective tissue disease

Osteoporosis and disorders of bone fragility are highly heritable, but despite much effort the identities of few of the genes involved has been established. Recent developments in genetics such as genome-wide association studies are revolutionizing research in this field, and it is likely that further contributions will be made through application of next-generation sequencing technologies, analysis of copy number variation polymorphisms, and high-throughput mouse mutagenesis programs. This article outlines what we know about osteoporosis genetics to date and the probable future directions of research in this field.     

Effect of vitamin E and selenium on antioxidant enzymes in brain,kidney and liver of cigarette smoke-exposed mice

The present study was conducted to evaluate the protective effects of vitamin E and selenium (Se) application on alteration of antioxidant enzyme activities against cigarette smoking induced oxidative damage in brains, kidneys and liver of mice. Male mice (balb/c) were exposed to cigarette smoke and treated with Se and/or vitamin E. Glutathione transferase (GST), glutathione peroxidase (GPX), glutathione reductase (GRX), superoxide dismutase (SOD) and catalase (CAT) enzyme activities in mice brain, kidney and liver were measured spectrophotometrically. GST, GPX, GRX, SOD and CAT enzyme activities in the brains of smoke-exposed mice were found lower than the enzymes activities of control mice and Se-and vitamin E-treated mice at the end of the three and five months. Opposite to brain, enzyme activities in kidneys and livers of smoke-exposed mice were found higher than the enzymes activities of control mice and Se-and vitamin E-treated mice at the end of the three and five months. Activities of GST, GPX, GRX SOD and CAT in the livers, kidneys and brains of smoke-exposed mice were found statistically different (p < 0.01) compared to control mice and Se-and vitamin E-treated mice. Combined application of vitamin E and Se had an additive protective effect against changing enzymes activities in smoke-exposed mice livers, kidneys and brains at the end of the both application periods. These results suggest that cigarette smoke exposure enhances the oxidative stress, thereby disturbing the tissue antioxidant defense system and combined application of vitamin E and Se protects the brain, kidney and liver from oxidative damage through their antioxidant potential.     

Vitamin D receptor deficiency and low vitamin D diet stimulate aortic calcification and osteogenic key factor expression in mice

Low levels of 25-hydroxy vitamin D (25(OH)D) are associated with cardiovascular diseases. Herein, we tested the hypothesis that vitamin D deficiency could be a causal factor in atherosclerotic vascular changes and vascular calcification. Aortic root sections of vitamin D receptor knockout (VDR(-/-)) mice that were stained for vascular calcification and immunostained for osteoblastic differentiation factors showed more calcified areas and a higher expression of the osteogenic key factors Msx2, Bmp2, and Runx2 than the wild-type mice (P<0.01). Data from LDL receptor knockout (LDLR(-/-)) mice that were fed western diet with either low (50 IU/kg), recommended (1,000 IU/kg), or high (10,000 IU/kg) amounts of vitamin D(3) over 16 weeks revealed increasing plasma concentrations of 25(OH)D (P<0.001) with increasing intake of vitamin D, whereas levels of calcium and phosphorus in plasma and femur were not influenced by the dietary treatment. Mice treated with the low vitamin D diet had more calcified lesions and a higher expression of Msx2, Bmp2, and Runx2 in aortic roots than mice fed recommended or high amounts of vitamin D (P<0.001). Taken together, these findings indicate vitamin D deficiency as a risk factor for aortic valve and aortic vessel calcification and a stimulator of osteogenic key factor expression in these vascular areas.     

Vitamin D receptor signaling inhibits atherosclerosis in mice

Although vitamin D has been implicated in cardiovascular protection, few studies have addressed the role of vitamin D receptor (VDR) in atherosclerosis. Here we investigate the effect of inactivation of the VDR signaling on atherogenesis and the antiatherosclerotic mechanism of vitamin D. Low density lipoprotein receptor (LDLR)(-/-)/VDR(-/-) mice exhibited site-specific accelerated atherogenesis, accompanied by increases in adhesion molecules and proinflammatory cytokines in the aorta and cholesterol influx in macrophages. Macrophages showed marked renin up-regulation in the absence of VDR, and inhibition of renin by aliskiren reduced atherosclerosis in LDLR(-/-)/VDR(-/-) mice, suggesting that the renin-angiotensin system (RAS) promotes atherosclerosis in the absence of VDR. LDLR(-/-) mice receiving LDLR(-/-)/VDR(-/-) BMT developed larger lesions than LDLR(-/-) BMT controls. Moreover, LDLR(-/-) mice receiving Rag-1(-/-)/VDR(-/-) BMT, which were unable to generate functional T and B lymphocytes, still had more severe atherosclerosis than Rag-1(-/-) BMT controls, suggesting a critical role of macrophage VDR signaling in atherosclerotic suppression. Aliskiren treatment eliminated the difference in lesions between Rag-1(-/-)/VDR(-/-) BMT and Rag-1(-/-) BMT recipients, indicating that local RAS activation in macrophages contributes to the enhanced atherogenesis seen in Rag-1(-/-)/VDR(-/-) BMT mice. Taken together, these observations provide evidence that macrophage VDR signaling, in part by suppressing the local RAS, inhibits atherosclerosis in mice.     

Caveolin-1 deficiency exacerbates cardiac dysfunction and reduces survival in mice with myocardial infarction

Caveolin (Cav)-1 has been involved in the pathogenesis of ischemic injuries. For instance, modulations of Cav-1 expression have been reported in animal models of myocardial infarction and cerebral ischemia-reperfusion. Furthermore, ablation of the Cav-1 gene in mice has been shown to increase the extent of ischemic injury in models of cerebral and hindlimb ischemia. Cav-1 has also been suggested to play a role in myocardial ischemic preconditioning. However, the role of Cav-1 in myocardial ischemia (MI)-induced cardiac dysfunction still remains to be determined. We determined the outcome of a permanent left anterior descending coronary artery (LAD) ligation in Cav-1 knockout (KO) mice. Wild-type (WT) and Cav-1 KO mice were subjected to permanent LAD ligation for 24 h. The progression of ischemic injury was monitored by echocardiography, hemodynamic measurements, 2,3,5-triphenyltetrazolium chloride staining, β-binding analysis, cAMP level measurements, and Western blot analyses. Cav-1 KO mice subjected to LAD ligation display reduced survival compared with WT mice. Despite similar infarct sizes, Cav-1 KO mice subjected to MI showed reduced left ventricular (LV) ejection fraction and fractional shortening as well as increased LV end-diastolic pressures compared with their WT counterparts. Mechanistically, Cav-1 KO mice subjected to MI exhibit reduced β-adrenergic receptor density at the plasma membrane as well as decreased cAMP levels and PKA phosphorylation. In conclusion, ablation of the Cav-1 gene exacerbates cardiac dysfunction and reduces survival in mice subjected to MI. Mechanistically, Cav-1 KO mice subjected to LAD ligation display abnormalities in β-adrenergic signaling.     

Formation of 8-nitroguanine in tobacco cigarette smokers and in tobacco smoke-exposed Wistar rats

Our previous studies have shown that 8-nitroguanine (8-NO(2)-G) could serve as a specific biomarker of DNA damage induced by gaseous nitrogen oxides (NO(x)) exposure. To evaluate the effect of tobacco cigarette smoking on the DNA damage in peripheral lymphocytes of cigarette smoke ones, we randomly collected and determined the level of 8-NO(2)-G in DNA extracted from peripheral lymphocyte of 15 each of light-smoking healthy volunteer (L-S, less than one pack per day), moderate-smoking healthy volunteers (M-S, one to two pack per day for 5-10 years), heavy-smoking healthy volunteers (H-S, over two packs per day for 10 years), lung cancer patients with heavy smoking (cancer H-S) and non-smoking healthy controls. Both of the mean level of the 8-NO(2)-G levels in peripheral lymphocyte (0.90+/-1.0, 1.23+/-1.14, 1.43+/-0.79, 3.62+/-1.38 ng per microg DNA) and serum nitrite (38.99+/-9.58, 46.70+/-9.38, 55.46+/-10.45, 70.1+/-18.54 microM) of L-S, M-S, H-S and cancer H-S groups were higher than that of non-smoking healthy controls (0.02+/-0.04 and 18.96+/-4.31 for 8-NO(2)-G level and serum nitrite, respectively). Furthermore, in animal experiment, a dose-dependent increase in 8-NO(2)-G was observed in rat lung and peripheral lymphocyte DNA of Wistar rats after tobacco cigarette smoke exposure twice a day, for 1 month. The level of 8-NO(2)-G is 0.17+/-0.41, 1.65+/-3.15, 23.50+/-20.75 and 37.58+/-17.55 ng per microg lung DNA for rat exposed with tobacco cigarette smoke from 0, 5, 10, 15 cigarettes per day, respectively. It was also found that count of peripheral lymphocytes and nitrite concentration in serum of rat increased after the tobacco smoke exposure. It is postulated that tobacco cigarette smoking could induce DNA damage (8-NO(2)-G formation) by exo- and endogenous NO(x).     

Severe Vitamin E deficiency exacerbates acute hyperoxic lung injury associated with increased oxidative stress and inflammation

Hyperoxia causes acute lung injury along with an increase of oxidative stress and inflammation. It was hypothesized that vitamin E deficiency might exacerbate acute hyperoxic lung injury. This study used alpha-tocopherol transfer protein knockout (alpha-TTP KO) mice fed a vitamin E-deficient diet (KO E(-) mice) as a model of severe vitamin E deficiency. Compared with wild-type (WT) mice, KO E(-) mice showed a significantly lower survival rate during hyperoxia. After 72 h of hyperoxia, KO E(-) mice had more severe histologic lung damage and higher values of the total cell count and the protein content of bronchoalveolar lavage fluid (BALF) than WT mice. IL-6 mRNA expression in lung tissue and the levels of 8-iso-prostaglandin F(2alpha) (8-iso-PGF(2alpha)) in both lungs and BALF were higher in KO E(-) mice than in WT mice. It was concluded that severe vitamin E deficiency exacerbates acute hyperoxic lung injury associated with increased oxidative stress or inflammation.     

Vitamin D deficiency in Europeans today and in Viking settlers of Greenland

The vast majority of the Earth’s population lives between the 20th and 40th parallel north and south. It seems that right here humans have found the best living conditions relating not only to temperature and food recourses, but also to UV radiation necessary for the production of vitamin D by human skin. An exception to this general rule is Europe. Nearly half a billion people live between the 40th and 60th parallel north of the equator despite the fact that the amounts of UV radiation there are much lower. Moreover, since the time of the Vikings, there has always been a part of the European population that lived even further north than the 60th parallel (the northern parts of Europe, including Greenland). In this work, we present the potential role that vitamin D deficiency might have played in the extinction of the Vikings of Greenland. We analyze factors that contribute to the discrepancy between the theoretical distribution of areas with vitamin D deficiency and today’s reality, like the impact of civilization, religious traditions, as well as vitamin D supplementation in food products and as a biologically active dietary additive. The global migration of people on a scale and speed never seen before is now even more important for this discrepancy.     

Associations with Vitamin D deficiency in "at risk" Australians

In a study of 185 elderly living in assisted care and 192 frail aged living in the community in the Sydney metropolitan area, nursing home residents were found to be at a 3-fold and hostel dwellers at a 2-fold risk of Vitamin D [25(OH)D] deficiency (<25 nmol/L) compared to self care residents. Middle Eastern people were found to be at 4-fold risk and Vietnamese a 3-fold risk of deficiency compared to their Australian counterparts. In recently arrived Chinese immigrants, Vitamin D deficiency, was found in 28%, and marginal levels (<37 nmol/L) in 60%, compared to the 34 and 76% found in our nursing home population, and 25 and 57% in hostel care residents. Of the Middle Eastern elderly, 58% were deficient and 83% marginal; although only 18% of Vietnamese were deficient, 68% had marginal Vitamin D status. Other factors associated with Vitamin D deficiency were mobility and sun exposure in assisted care, and low dietary Vitamin D and calcium intake, reduced exercise levels and high % body fat levels in the immigrant groups.     

Cyclophilin D deficiency prevents diet-induced obesity in mice

Mitochondrial coupling efficiency is pivotal in thermogenesis and energy homeostasis. Here we show that deletion of cyclophilin D (CypD), a key modulator of the mitochondrial permeability transition pore, demonstrated resistance to diet-induced obesity (DIO) in both male and female mice, due to increased basal metabolic rate, heat production, total energy expenditure and expenditure of fat energy, despite increased food consumption. Absorption of fatty acids is not altered between CypD(-/-) and wild-type mice. Adult CypD(-/-) developed hyperglycemia, insulin resistance and glucose intolerance albeit resistant to DIO. These data demonstrate that inhibition of CypD function could protect from HFD-IO by increasing energy expenditure in both male and female mice. Inhibition of CypD may offer a novel target to modulate metabolism.