Emerging Roles of Spatial Transcriptomics in Liver Research.

Spatial transcriptomics, leveraging sequencing- and imaging-based techniques, has emerged as a groundbreaking technology for mapping gene expression within the complex architectures of tissues. This approach provides an in-depth understanding of cellular and molecular dynamics across various states of healthy and diseased livers. Through the integration of sophisticated bioinformatics strategies, it enables detailed exploration of cellular heterogeneity, transitions in cell states, and intricate cell-cell interactions with remarkable precision. In liver research, spatial transcriptomics has been particularly revelatory, identifying distinct zonated functions of hepatocytes that are crucial for understanding the metabolic and detoxification processes of the liver. Moreover, this technology has unveiled new insights into the pathogenesis of liver diseases, such as the role of lipid-associated macrophages in steatosis and endothelial cell signals in liver regeneration and repair. In the domain of liver cancer, spatial transcriptomics has proven instrumental in delineating intratumor heterogeneity, identifying supportive microenvironmental niches and revealing the complex interplay between tumor cells and the immune system as well as susceptibility to immune checkpoint inhibitors. In conclusion, spatial transcriptomics represents a significant advance in hepatology, promising to enhance our understanding and treatment of liver diseases.

Airway inflammation in a novel mouse model of asthma-COPD overlap induced by co-exposure to papain and tobacco smoke.

Asthma and chronic obstructive pulmonary disease (COPD) are respiratory diseases associated with airway inflammation, which is the main pathogenesis. Although their causes and characteristics differ, in some cases, asthma and COPD may coexist in the same patient in a condition called asthma-COPD overlap (ACO). The prognosis of ACO is more unfavourable than those of asthma or COPD alone, without any treatment strategies demonstrating efficacy. Owing to its intricate spectrum of features, the detailed pathogenesis of how ACO exacerbates respiratory features remains unclear. In this study, we exposed papain-induced asthma model mice to tobacco smoke to establish an ACO mouse model, in which features of airway inflammation observed in both asthma and COPD were incorporated. This model exhibited distinctive mixed and corticosteroid-resistant airway inflammation and emphysematous changes that are characteristic of ACO. The novel mouse model established here is expected to significantly contribute to elucidating the mechanisms of the broad pathologies of ACO and identifying potential therapeutic targets.

Anti-fungal activity of a novel triazole, PC1244, against emerging azole-resistant Aspergillus fumigatus and other species of Aspergillus.

OBJECTIVES: The growing emergence of azole-resistant Aspergillus fumigatus strains worldwide is a major concern for current systemic antifungal treatment. Here we report antifungal activities of a novel inhaled triazole, PC1244, against a collection of multi-azole-resistant A. fumigatus strains. METHODS: MICs of PC1244 were determined for A. fumigatus carrying TR34/L98H (n = 81), TR46/Y121F/T289A (n = 24), M220 (n = 6), G54 (n = 11), TR53 (n = 1), TR463/Y121F/T289A (n = 2), G448S (n = 1), G432C (n = 1) and P216S (n = 1) resistance alleles originating from either India, the Netherlands or France. The effects of PC1244 were confirmed in an in vitro model of the human alveolus and in vivo in temporarily neutropenic, immunocompromised mice. RESULTS: PC1244 exhibited potent inhibition [geometric mean MIC (range), 1.0 mg/L (0.125 to >8 mg/L)] of growth of A. fumigatus strains carrying cyp51A gene mutations, showing much greater potency than voriconazole [15 mg/L (0.5 to >16 mg/L)], and an effect similar to those on other azole-susceptible Aspergillus spp. (Aspergillus flavus, Aspergillus terreus, Aspergillus tubingensis, Aspergillus nidulans, Aspergillus niger, Aspergillus nomius, Aspergillus tamarii) (0.18-1 mg/L). In TR34/L98H and TR46/Y121F/T289A A. fumigatus-infected in vitro human alveolus models, PC1244 achieved superior inhibition (IC50, 0.25 and 0.34 mg/L, respectively) compared with that of voriconazole (IC90, >3 mg/L and >10 mg/L, respectively). In vivo, once-daily intranasal administration of PC1244 (0.56-70 µg/mouse) to the A. fumigatus (AF91 with M220V)-infected mice reduced pulmonary fungal load and serum galactomannan more than intranasal posaconazole. CONCLUSIONS: PC1244 has the potential to become a novel topical treatment of azole-resistant pulmonary aspergillosis.

In Vitro and In Vivo Efficacy of a Novel and Long-Acting Fungicidal Azole, PC1244, on Aspergillus fumigatus Infection.

The antifungal effects of the novel triazole PC1244, designed for topical or inhaled administration, against Aspergillus fumigatus were tested in a range of in vitro and in vivo studies. PC1244 demonstrated potent antifungal activities against clinical A. fumigatus isolates (n = 96) with a MIC range of 0.016 to 0.25 µg/ml, whereas the MIC range for voriconazole was 0.25 to 0.5 µg/ml. PC1244 was a strong tight-binding inhibitor of recombinant A. fumigatus CYP51A and CYP51B (sterol 14α-demethylase) enzymes and strongly inhibited ergosterol synthesis in A. fumigatus with a 50% inhibitory concentration of 8 nM. PC1244 was effective against a broad spectrum of pathogenic fungi (MIC range, <0.0078 to 2 µg/ml), especially Aspergillus terreus, Trichophyton rubrum, Candida albicans, Candida glabrata, Candida krusei, Cryptococcus gattii, Cryptococcus neoformans, and Rhizopus oryzae PC1244 also proved to be quickly absorbed into both A. fumigatus hyphae and bronchial epithelial cells, producing persistent antifungal effects. In addition, PC1244 showed fungicidal activity (minimum fungicidal concentration, 2 µg/ml) which indicated that it was 8-fold more potent than voriconazole. In vivo, once-daily intranasal administration of PC1244 (3.2 to 80 µg/ml) to temporarily neutropenic, immunocompromised mice 24 h after inoculation with itraconazole-susceptible A. fumigatus substantially reduced the fungal load in the lung, the galactomannan concentration in serum, and circulating inflammatory cytokine levels. Furthermore, 7 days of extended prophylaxis with PC1244 showed in vivo effects superior to those of 1 day of prophylactic treatment, suggesting accumulation of the effects of PC1244. Thus, PC1244 has the potential to be a novel therapy for the treatment of A. fumigatus infection in the lungs of humans.

In Vivo Biomarker Analysis of the Effects of Intranasally Dosed PC945, a Novel Antifungal Triazole, on Aspergillus fumigatus Infection in Immunocompromised Mice.

PC945 is a novel triazole optimized for lung delivery, and the objective of this study is to determine the effects of intranasally dosed PC945 on Aspergillus fumigatus infection and associated biomarkers in immunocompromised mice. PC945, posaconazole, or voriconazole was administered intranasally once daily on days 0 to 3 (early intervention) or days 1 to 3 (late intervention) postinfection in temporarily neutropenic A/J mice infected intranasally with A. fumigatus, and bronchoalveolar lavage fluid (BALF) and serum were collected on day 3. The effects of extended prophylaxis treatment (daily from days -7 to +3 or days -7 to 0) were also compared with those of the shorter treatment regimens (days -1 to +3 or days -1 and 0). Early and late interventions with PC945 (2.8 to 350 µg/mouse; approximately 0.11 to ∼14 mg/kg of body weight) were found to inhibit lung fungal loads and to decrease the concentrations of galactomannan (GM) in both BALF and serum as well as several biomarkers in BALF (interferon gamma [IFN-γ], interleukin-17 [IL-17], and malondialdehyde) and serum (tumor necrosis factor alpha [TNF-α] and IL-6) in a dose-dependent manner and were >3- and >47-fold more potent than intranasally dosed posaconazole and voriconazole, respectively. Furthermore, extended prophylaxis with low-dose PC945 (0.56 µg/mouse; 0.022 mg/kg) was found to inhibit fungal loads and to decrease the concentrations biomarkers more potently than did the shorter treatment regimens. Thus, PC945 dosed intranasally once daily showed potent antifungal effects, and the effects of PC945 accumulated upon repeat dosing and were persistent. Therefore, PC945 has the potential to be a novel inhaled therapy for the treatment of A. fumigatus infection in humans.

In Vitro and In Vivo Antifungal Profile of a Novel and Long-Acting Inhaled Azole, PC945, on Aspergillus fumigatus Infection.

The profile of PC945, a novel triazole antifungal designed for administration via inhalation, was assessed in a range of in vitro and in vivo studies. PC945 was characterized as a potent, tightly binding inhibitor of Aspergillus fumigatus sterol 14α-demethylase (CYP51A and CYP51B) activity (50% inhibitory concentrations [IC50s], 0.23 µM and 0.22 µM, respectively) with characteristic type II azole binding spectra. Against 96 clinically isolated A. fumigatus strains, the MIC values of PC945 ranged from 0.032 to >8 µg/ml, while those of voriconazole ranged from 0.064 to 4 µg/ml. Spectrophotometric analysis of the effects of PC945 against itraconazole-susceptible and -resistant A. fumigatus growth yielded IC50 (determined based on optical density [OD]) values of 0.0012 to 0.034 µg/ml, whereas voriconazole (0.019 to >1 µg/ml) was less effective than PC945. PC945 was effective against a broad spectrum of pathogenic fungi (with MICs ranging from 0.0078 to 2 µg/ml), including Aspergillus terreus, Trichophyton rubrum, Candida albicans, Candida glabrata, Candida krusei, Cryptococcus gattii, Cryptococcus neoformans, and Rhizopus oryzae (1 or 2 isolates each). In addition, when A. fumigatus hyphae or human bronchial cells were treated with PC945 and then washed, PC945 was found to be absorbed quickly into both target and nontarget cells and to produce persistent antifungal effects. Among temporarily neutropenic immunocompromised mice infected with A. fumigatus intranasally, 50% of the animals survived until day 7 when treated intranasally with PC945 at 0.56 µg/mouse, while posaconazole showed similar effects (44%) at 14 µg/mouse. This profile affirms that topical treatment with PC945 should provide potent antifungal activity in the lung.

Effects of intranasally dosed posaconazole on fungal load and biomarkers in Aspergillus fumigatus infected immunocompromised mice.

Although anti-fungal triazoles are dosed orally or systemically for Aspergillus fumigatus infection, systemic adverse events and limited exposure of the lung cavity would make a topical treatment for the lung an attractive option. In this study, we examined the effects of intranasally dosed posaconazole on survival rates and biomarkers in A. fumigatus (itraconazole susceptible: ATCC13073 [Af]; or resistant: NCPF7100 [AfR]) infected, temporarily neutropenic A/J mice. Once daily treatment produced a dose-dependent improvement of survival of Af-infected mice (ED50 : 0.019 mg/mouse [approx. 0.755 mg/kg, in]), similar to its potency (ED50 : 0.775 mg/kg, po) after once daily oral dosing. For AfR infection, either intranasal or oral posaconazole was largely ineffective on survival, although the highest dose of intranasal treatment (0.35 mg/mouse) achieved 75% survival rate. Early intervention (treated on days 0, 1, 2 and 3 postinfection) and late intervention (treated on days 1, 2 and 3) with intranasal posaconazole (0.014-0.35 mg/mouse) demonstrated potent inhibition of lung fungal load and galactomannan levels in both bronchoalveolar lavage fluid (BALF) and serum as well as inflammatory cells, IFN-γ, IL-17 and malondialdehyde (MDA) in BALF. Thus, posaconazole when dosed intranasally once daily showed an improvement of survival equivalent to or better than oral treatment, and produced potent inhibition of fungal load and biomarkers.

[Anti-inflammatory Effects of a Src Inhibitor on the Murine Model of Asthma Exacerbation Induced by Ovalbumin and Lipopolysaccharide].

Asthma is often exacerbated by airway infection, and some patients with severe asthma may be unresponsive to conventional corticosteroid treatment. Src family kinases (SFKs) were recently implicated in the inflammatory responses of mice induced by allergen and bacterial toxin lipopolysaccharide (LPS). Therefore, we examined the effects of dasatinib (DAS), a Src inhibitor, on airway inflammation in mice induced by ovalbumin (OVA) and LPS. Male A/J mice were sensitized to OVA Day -14 and -7, challenged with intranasal OVA on Day 0, 2, 4, 6 and 8, and on Day 10, mice were also challenged with OVA via inhalation. Mice were treated intranasally with DAS or fluticasone propionate (FP), a glucocorticoid, twice daily for 3 d starting 1 d after OVA inhalation. Moreover, some mice were also administrated LPS 2 h after DAS or FP treatment to model of asthma exacerbation. One day after the last intervention, lung tissue and bronchoalveolar lavage fluid (BALF) were collected. DAS attenuated the accumulation of inflammatory cells and cytokines/chemokines in BALF induced by both OVA and OVA+LPS, while FP did not reduce accumulations induced by OVA+LPS. Therefore, targeting SFKs may be a superior therapeutic approach for asthma exacerbation by infection.

Anterior condylar arteriovenous fistula mainly fed by peripheral branches of the bilateral internal maxillary arteries: illustrative case.

BACKGROUND: The main feeding artery of an anterior condylar arteriovenous fistula (AC-AVF) is the ascending pharyngeal artery and rarely the internal maxillary artery. OBSERVATIONS: A 58-year-old male with a history of sinusitis since adolescence presented with a 5-year history of bilateral pulsatile tinnitus and a 2-month history of right ocular symptoms. Angiography showed that the peripheral branches of the bilateral internal maxillary arteries were the main feeding arteries of the AC-AVF and that they gathered in the clivus with a relatively large shunted pouch in the left jugular tubercle. Shunt flow drained to the right external jugular vein via the right superior ophthalmic vein. A sheath was placed in the right external jugular vein, and a small distal access catheter was guided to the right superior ophthalmic vein to allow the microcatheter to reach the shunted pouch. Selective angiography of the contralateral sphenopalatine artery allowed us to confirm the gathering site of the feeding arteries and the shunted pouch and archive the complete occlusion. LESSONS: Selective angiography of the contralateral sphenopalatine artery may be useful to confirm the gathering site of the peripheral branches of the bilateral internal maxillary arteries in an AC-AVF.

Relationship between anti-fungal effects and lung exposure of PC945, a novel inhaled antifungal agent, in Aspergillus fumigatus infected mice: Pulmonary PK-PD analysis of anti-fungal PC945.

PC945 is a novel antifungal agent, optimised for inhaled treatment. In this study, the relationship between antifungal effects of PC945 and its exposure in the lungs was investigated in Aspergillus fumigatus intranasally infected, temporarily neutropenic mice. Mice were given prophylactic PC945 intranasally once daily (0.56 µg/mouse) on either Day -7 to 0 (8 doses) or Day -1 to 0 (2 doses). Lung tissue, plasma and bronchoalveolar lavage (BAL) fluid were collected 24 or 72 h post A. fumigatus inoculation for biomarker and pharmacokinetic analyses. BAL cell pellets and supernatants were prepared separately by centrifugation. 8 prophylactic doses of PC945 were found to demonstrate significantly stronger antifungal effects (lung fungal burden and galactomannan (GM) in BAL and plasma) than prophylaxis with 2 doses. PC945 concentrations were below the limit of detection in plasma but readily measured in lung extracts. The concentrations were much higher after extended prophylaxis (709 and 312 ng/g of lung) than short prophylaxis (301 and 195 ng/g of lung) at 24 and 72 h post last dose, respectively, suggesting PC945 accumulation in whole lung after repeat dosing although it was likely to be a mixture of dissolved and undissolved PC945, meaning that the data should be interpreted with caution. Interestingly, low concentrations of PC945 were detected in BAL supernatant (6.6 and 1.9 ng/ml) whereas high levels of PC945 were measured in BAL cell pellets (626 and 406 ng/ml) at 24 and 72 h post last dose, respectively, in extended prophylaxis. In addition, the PC945 concentrations in BAL cells showed a statistically significant correlation with measured anti-fungal activities. These observations will be pursued, and it is intended that BAL cell concentrations of PC945 be measured in future clinical studies rather than standard measurement in BAL itself. Thus, PC945's profile makes it an attractive potential prophylactic agent for the prevention of pulmonary fungal infections.

[Inhibitory Effects of Dabigatran on Airway Inflammation Induced by Lipopolysaccharide in Mice].

Asthma and chronic obstructive pulmonary disease (COPD) are characterised by chronic inflammation in the lung that is associated with airway obstruction. Inhaled therapy with a combination of corticosteroid and a long-acting ß2-agonist is an effective anti-inflammatory medicine for asthma, but in patients with severe asthma and COPD fails to completely control these symptoms with current therapies. The inflammatory process in these diseases, which involves activation of the coagulation and fibrinolytic system in the lung, offers the opportunity for alternative anti-inflammatory therapies. In this study, we investigated the effects of anti-coagulants on lipopolysaccharide (LPS)-induced airway inflammation in mice. A/J mice were exposed to LPS, a bacterial endotoxin, intranasally and accumulation of inflammatory cells, TNF-α, C-X-C motif chemokine (CXCL) 1, and osteopontin in bronchoalveolar lavage fluid (BALF) was monitored by flow cytometry and an enzyme-linked immunosorbent assay. LPS exposure induced airway neutrophilia and accumulation of TNF-α, CXCL1, and osteopontin in BALF. This LPS-induced airway inflammation was not relieved using a corticosteroid, fluticasone propionate (FP), or a direct inhibitor of Factor Xa, rivaroxaban. In contrast, a direct thrombin inhibitor, dabigatran, inhibited LPS-induced airway neutrophilia and decreased inflammatory cytokine production in a dose dependent manner. Furthermore, combination of dabigatran and FP elicited stronger inhibition of LPS-induced airway inflammation. Therefore, these results suggest that dabigatran could be an effective new therapy for severe respiratory diseases.

[TNF-α Decreased Corticosteroid Responsiveness in Mice Models of Airway Inflammation Induced by Double Strand RNA and/or Tobacco Smoke Exposure].

Reduction of corticosteroid responsiveness is one of the important clinical problems in chronic obstructive pulmonary disease (COPD). In this study, we determined the effects of neutralization of tumor necrosis factor-α (TNF-α) on corticosteroid insensitivity in mice models of airway inflammation induced by poly(I:C) and tobacco smoke (TS) exposure. Mice (male A/J strain, 5 weeks old) were exposed to TS for 10 d, or TS for 11 d and poly(I:C) for 3 d. Anti-TNF-α antibody was intranasally treated once every other day 2 h before the TS exposure, and dexamethasone 21-phosphate (DEX) was treated 30 min before the TS or poly(I:C) exposure. On the next day of the last stimulation, mice were sacrificed. The combination treatment of DEX and TNF-α neutralization was significantly attenuated the increase of the numbers of inflammatory cells in BALF and the TNF-α mRNA expression levels induced by TS and poly(I:C) exposure, even though TNF-α neutralization alone had little effect. These data indicated that neutralization of TNF-α restores corticosteroid responsiveness. Therefore, our study suggests that targeting TNF-α signaling pathway provides a new therapeutic approach to corticosteroid refractory airway diseases.

Antifungal synergy of a topical triazole, PC945, with a systemic triazole against respiratory Aspergillus fumigatus infection.

Invasive pulmonary Aspergillosis is a leading cause of morbidity and mortality in immunosuppressed patients and treatment outcomes using oral antifungal triazoles remain suboptimal. Here we show that combining topical treatment using PC945, a novel inhaled triazole, with systemic treatment using known triazoles demonstrated synergistic antifungal effects against Aspergillus fumigatus (AF) in an in vitro human alveolus bilayer model and in the lungs of neutropenic immunocompromised mice. Combination treatment with apical PC945 and either basolateral posaconazole or voriconazole resulted in a synergistic interaction with potency improved over either compound as a monotherapy against both azole-susceptible and resistant AF invasion in vitro. Surprisingly there was little, or no synergistic interaction observed when apical and basolateral posaconazole or voriconazole were combined. In addition, repeated prophylactic treatment with PC945, but not posaconazole or voriconazole, showed superior effects to single prophylactic dose, suggesting tissue retention and/or accumulation of PC945. Furthermore, in mice infected with AF intranasally, 83% of animals treated with a combination of intranasal PC945 and oral posaconazole survived until day 7, while little protective effects were observed by either compound alone. Thus, the combination of a highly optimised topical triazole with oral triazoles potentially induces synergistic effects against AF infection.

[The Involvement of Src in Airway Inflammation Induced by Repeated Exposure to Lipopolysaccharide in Mice].

Corticosteroid insensitive airway inflammation is one of major barrier to effective managements of chronic airway diseases, such as chronic obstructive pulmonary disease (COPD) and severe asthma. The role of nonreceptor tyrosine kinase Src is important in airway inflammation in mice models of atopic asthma and COPD. Thus, in this study, we determined the effects of Src inhibitor, dasatinib, on airway inflammation induced by repeated intranasal exposure to lipopolysaccharide (LPS). Male mice (A/J strain, 5 weeks old) were intranasally exposed to LPS twice daily for 3 d, and dasatinib was intranasally treated 2 h prior to each LPS exposure. A day after the last stimulation, lungs and bronchoalveolar lavage fluid (BALF) were collected. Dasatinib attenuated the accumulation of inflammatory cells in lungs, and the increase in the numbers of inflammatory cells and the accumulation of cytokines/chemokines in BALF in a dose dependent manner. Therefore, this study suggested that targeting the Src can provide a new therapeutic approach for corticosteroid insensitive pulmonary diseases.

Repeated lipopolysaccharide exposure causes corticosteroid insensitive airway inflammation via activation of phosphoinositide-3-kinase δ pathway.

Corticosteroid resistance is one of major barriers to effective management of chronic inflammatory respiratory diseases, such as chronic obstructive pulmonary disease (COPD) and severe asthma. These patients often experience exacerbations with viral and/or bacterial infection, which may cause continuous corticosteroid insensitive inflammation. In this study, we observed that repeated exposure of lipopolysaccharide (LPS) intranasally attenuated the anti-inflammatory effects of the corticosteroid fluticasone propionate (FP) on neutrophils and CXCL1 levels in bronchoalveolar lavage (BAL) fluid in an in vivo murine model. Histone deacetylase-2 (HDAC2) and NF-E2 related factor 2 (Nrf2) levels in lungs after LPS administration for 3 consecutive days were significantly decreased to 38.9±6.3% (mean±SEM) and 77.5±2.7% of the levels seen after only one day of LPS exposure, respectively. In addition, 3 days LPS exposure resulted in an increase of Akt phosphorylation, indicating activation of the phosphoinositide-3-kinase (PI3K) pathway by 4-fold in lungs compared with 1 day of exposure. Furthermore, combination treatment with theophylline and FP significantly decreased the neutrophil accumulation and CXCL1 concentrations in BAL fluid from 22.5±1.8×104 cells/mL and 214.6±20.6 pg/mL to 7.9±0.5×104 cells/mL and 61.9±13.3 pg/mL, respectively. Combination treatment with IC87114, a selective PI3Kδ inhibitor, and FP also significantly decreased neutrophils and CXCL1 levels from 16.8±0.7×104 cells/mL and 182.4±4.6 pg/mL to 5.9±0.3×104 cells/mL and 71.4±2.7 pg/mL, respectively. Taken together, repeated exposure of LPS causes corticosteroid-insensitive airway inflammation in vivo, and the corticosteroid-resistance induced by LPS is at least partly mediated through the activation of PI3Kδ, resulting in decreased levels of HDAC2 and Nrf2. These findings provide a potentially new therapeutic approach to COPD and severe asthma.

Activation of transcription factor Nrf2 signalling by the sphingosine kinase inhibitor SKI-II is mediated by the formation of Keap1 dimers.

BACKGROUND: Anti-oxidant capacity is crucial defence against environmental or endogenous oxidative stress. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that plays a key defensive role against oxidative and cytotoxic stress and cellular senescence. However, Nrf2 signalling is impaired in several aging-related diseases, such as chronic pulmonary obstructive disease (COPD), cancer, and neurodegenerative diseases. Thus, novel therapeutics that enhance Nrf2 signalling are an attractive approach to treat these diseases. METHODOLOGY/PRINCIPAL FINDINGS: Nrf2 was stabilized by SKI-II (2-(p-hydroxyanilino)-4-(p-chlorophenyl) thiazole), which is a known sphingosine kinase inhibitor, in human bronchial epithelial cell line, BEAS2B, and in primary human bronchial epithelial cells, leading to enhancement of anti-oxidant proteins, such as HO-1, NQO1 and GCLM. The activation of Nrf2 was achieved by the generation of inactive dimerized form of Keap1, a negative regulator of Nrf2 expression, which was independent of sphingosine kinase inhibition. Using mice that were exposed to cigarette smoke, SKI-II induced Nrf2 expression together with HO-1 in their lungs. In addition, SKI-II reduced cigarette smoke mediated oxidative stress, macrophages and neutrophil infiltration and markers of inflammation in mice. CONCLUSIONS/SIGNIFICANCE: SKI-II appears to be a novel activator of Nrf2 signalling via the inactivation of Keap1.

Toll-like receptor 3 stimulation causes corticosteroid-refractory airway neutrophilia and hyperresponsiveness in mice.

BACKGROUND: RNA virus infections, such as rhinovirus and respiratory syncytial virus, induce exacerbations in patients with COPD and asthma, and the inflammation is corticosteroid refractory. The main aim of this study is to establish a murine model induced by a Toll-like receptor 3 (TLR3) agonist, an RNA virus mimic, and investigate the response to corticosteroid. METHODS: A/J mice were given polyinosinic-polycytidylic acid (poly[I:C]), a TLR3 agonist, intranasally, in the presence or absence of cigarette smoke exposure. Inflammatory cell accumulation and C-X-C motif chemokine (CXCL) 1, interferon (IFN), and CXCL10 production in BAL fluid (BALF) were determined by flow cytometry and enzyme-linked immunosorbent assay, respectively, and airway hyperresponsiveness (AHR) to histamine/methacholine was determined by a two-chambered, double-flow plethysmography system. BALB/c and C57BL/6J mice were also used for comparisons. RESULTS: Intranasal treatment of poly(I:C) significantly induced airway neutrophilia; production of CXCL1, IFN-ß, and CXCL10; and necrotic cell accumulation in BALF. It also increased airway responsiveness to histamine or methacholine inhalation. This poly(I:C)-dependent airway inflammation and AHR was not inhibited by the corticosteroid fluticasone propionate (FP) (up to 0.5 mg/mL intranasal), although FP strongly inhibited lipopolysaccharide (TLR4 agonist)-induced airway neutrophilia. Furthermore, cigarette smoke exposure significantly increased TLR3 expression in murine lung tissue and exacerbated poly(I:C)-induced neutrophilia and AHR. CONCLUSIONS: These results suggest that TLR3 stimulation is involved in corticosteroid-refractory airway inflammation in lung, which is enhanced by cigarette smoking, and this may provide a model for understanding virus-induced exacerbations in COPD and their therapy.

Calcineurin-dependent mitochondrial disturbances in calcium-induced apoptosis of human immunodeficiency virus gp160-expressing CD4+ cells.

In CD4+ UE160 cells with inducible expression of gp160, mechanisms of apoptosis induced by human immunodeficiency virus (HIV) Env protein were analyzed. Induction of gp160 caused intracellular calcium increase followed by the release of cytochrome c from mitochondria, which was inhibited by calcineurin inhibitors. Association of BAD with Bcl-xL was observed, and a portion of BAD was dephosphorylated after induction of gp160. These data suggested that calcineurin plays a role in the HIV Env-induced apoptosis in a mitochondrion-dependent way.