text:
.class Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;
.super Lcom/google/common/collect/Maps$MapDifferenceImpl;
.source "Maps.java"

# interfaces
.implements Lcom/google/common/collect/SortedMapDifference;

# annotations
.annotation system Ldalvik/annotation/EnclosingClass;
    value = Lcom/google/common/collect/Maps;
.end annotation

.annotation system Ldalvik/annotation/InnerClass;
    accessFlags = 0x8
    name = "SortedMapDifferenceImpl"
.end annotation

.annotation system Ldalvik/annotation/Signature;
    value = {
        "<K:",
        "Ljava/lang/Object;",
        "V:",
        "Ljava/lang/Object;",
        ">",
        "Lcom/google/common/collect/Maps$MapDifferenceImpl",
        "<TK;TV;>;",
        "Lcom/google/common/collect/SortedMapDifference",
        "<TK;TV;>;"
    }
.end annotation

# direct methods
.method constructor <init>(ZLjava/util/SortedMap;Ljava/util/SortedMap;Ljava/util/SortedMap;Ljava/util/SortedMap;)V
    .locals 0
    .param p1, "areEqual"    # Z
    .annotation system Ldalvik/annotation/Signature;
        value = {
            "(Z",
            "Ljava/util/SortedMap",
            "<TK;TV;>;",
            "Ljava/util/SortedMap",
            "<TK;TV;>;",
            "Ljava/util/SortedMap",
            "<TK;TV;>;",
            "Ljava/util/SortedMap",
            "<TK;",
            "Lcom/google/common/collect/MapDifference$ValueDifference",
            "<TV;>;>;)V"
        }
    .end annotation

    .prologue
    .line 588
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    .local p2, "onlyOnLeft":Ljava/util/SortedMap;, "Ljava/util/SortedMap<TK;TV;>;"
    .local p3, "onlyOnRight":Ljava/util/SortedMap;, "Ljava/util/SortedMap<TK;TV;>;"
    .local p4, "onBoth":Ljava/util/SortedMap;, "Ljava/util/SortedMap<TK;TV;>;"
    .local p5, "differences":Ljava/util/SortedMap;, "Ljava/util/SortedMap<TK;Lcom/google/common/collect/MapDifference$ValueDifference<TV;>;>;"
    invoke-direct/range {p0 .. p5}, Lcom/google/common/collect/Maps$MapDifferenceImpl;-><init>(ZLjava/util/Map;Ljava/util/Map;Ljava/util/Map;Ljava/util/Map;)V

    .line 589
    return-void
.end method

# virtual methods
.method public bridge synthetic entriesDiffering()Ljava/util/Map;
    .locals 1

    .prologue
    .line 583
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-virtual {p0}, Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;->entriesDiffering()Ljava/util/SortedMap;

    move-result-object v0

    return-object v0
.end method

.method public entriesDiffering()Ljava/util/SortedMap;
    .locals 1
    .annotation system Ldalvik/annotation/Signature;
        value = {
            "()",
            "Ljava/util/SortedMap",
            "<TK;",
            "Lcom/google/common/collect/MapDifference$ValueDifference",
            "<TV;>;>;"
        }
    .end annotation

    .prologue
    .line 592
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-super {p0}, Lcom/google/common/collect/Maps$MapDifferenceImpl;->entriesDiffering()Ljava/util/Map;

    move-result-object v0

    check-cast v0, Ljava/util/SortedMap;

    return-object v0
.end method

.method public bridge synthetic entriesInCommon()Ljava/util/Map;
    .locals 1

    .prologue
    .line 583
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-virtual {p0}, Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;->entriesInCommon()Ljava/util/SortedMap;

    move-result-object v0

    return-object v0
.end method

.method public entriesInCommon()Ljava/util/SortedMap;
    .locals 1
    .annotation system Ldalvik/annotation/Signature;
        value = {
            "()",
            "Ljava/util/SortedMap",
            "<TK;TV;>;"
        }
    .end annotation

    .prologue
    .line 596
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-super {p0}, Lcom/google/common/collect/Maps$MapDifferenceImpl;->entriesInCommon()Ljava/util/Map;

    move-result-object v0

    check-cast v0, Ljava/util/SortedMap;

    return-object v0
.end method

.method public bridge synthetic entriesOnlyOnLeft()Ljava/util/Map;
    .locals 1

    .prologue
    .line 583
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-virtual {p0}, Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;->entriesOnlyOnLeft()Ljava/util/SortedMap;

    move-result-object v0

    return-object v0
.end method

.method public entriesOnlyOnLeft()Ljava/util/SortedMap;
    .locals 1
    .annotation system Ldalvik/annotation/Signature;
        value = {
            "()",
            "Ljava/util/SortedMap",
            "<TK;TV;>;"
        }
    .end annotation

    .prologue
    .line 600
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-super {p0}, Lcom/google/common/collect/Maps$MapDifferenceImpl;->entriesOnlyOnLeft()Ljava/util/Map;

    move-result-object v0

    check-cast v0, Ljava/util/SortedMap;

    return-object v0
.end method

.method public bridge synthetic entriesOnlyOnRight()Ljava/util/Map;
    .locals 1

    .prologue
    .line 583
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-virtual {p0}, Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;->entriesOnlyOnRight()Ljava/util/SortedMap;

    move-result-object v0

    return-object v0
.end method

.method public entriesOnlyOnRight()Ljava/util/SortedMap;
    .locals 1
    .annotation system Ldalvik/annotation/Signature;
        value = {
            "()",
            "Ljava/util/SortedMap",
            "<TK;TV;>;"
        }
    .end annotation

    .prologue
    .line 604
    .local p0, "this":Lcom/google/common/collect/Maps$SortedMapDifferenceImpl;, "Lcom/google/common/collect/Maps$SortedMapDifferenceImpl<TK;TV;>;"
    invoke-super {p0}, Lcom/google/common/collect/Maps$MapDifferenceImpl;->entriesOnlyOnRight()Ljava/util/Map;

    move-result-object v0

    check-cast v0, Ljava/util/SortedMap;

    return-object v0
.end method


text:
of Oncology
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About Spandidos
IL‑17A and GDF15 are able to induce epithelial‑mesenchymal transition of lung epithelial cells in response to cigarette smoke
Gang Jiang
Chen‑Tao Liu
Wei‑Dong Zhang
View Affiliations
Affiliations: Department of Respiration, Hunan Provincial People's Hospital, Changsha, Hunan 410005, P.R. China, Department of Paediatrics, Xiang Ya Hospital, Central South University, Changsha, Hunan 410008, P.R. China
Published online on: May 10, 2018 https://doi.org/10.3892/etm.2018.6145
Copyright: © Jiang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
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Smoking is one of the primary causes of chronic obstructive pulmonary disease (COPD). Sustained active epithelial‑mesenchymal transition (EMT) in COPD may explain the core pathophysiology of airway fibrosis and why lung cancer is so common among smokers. Interleukin (IL)‑17A and growth/differentiation factor (GDF)15 have been reported to be biomarkers of COPD; however, the role of IL‑17A and GDF15 in EMT remains unclear. The aim of the present study was to investigate the role of IL‑17A and GDF15 in the pathogenesis of COPD. It was demonstrated that IL‑17A and GDF15 are upregulated in patients with COPD, particularly those with a history of smoking. The results also revealed that IL‑17A and GDF15 expression was negatively correlated with the epithelial marker epithelial‑cadherin and positively correlated with the mesenchymal marker vimentin. Furthermore, treatment with cigarette smoke extract or IL‑17A induced GDF15 expression. Combined treatment with IL‑17A and GDF15 induced EMT in human small epithelial HSAEpiC cells in vitro. Collectively, the results of the present study suggest that IL‑17A and GDF15‑induced EMT serves an important role in the pathology of COPD.
Chronic obstructive pulmonary disease (COPD), which is characterised by poor reversible airflow limitation, is a major chronic lung disease and a global health problem (1,2). The alveolar response to noxious substances, including cigarette smoke, is considered to be a risk factor for the development of COPD (3); however, the underlying molecular mechanisms remain to be elucidated. Airway fibrosis has been reported to contribute to physiological airway dysfunction in COPD (4). Previous studies have suggested that epithelial-mesenchymal transition (EMT), a feature of lung fibrosis, may be important for the development of COPD (5–8). De-differentiation of the respiratory epithelium occurs via active EMT, however the underlying mechanism and the association between EMT and peribronchial fibrosis remain unclear (9). It was therefore hypothesised that de-differentiation of the COPD respiratory epithelium via EMT may serve a role in airway fibrosis and thereby in airway obstruction.
It has been reported that the airway epithelium provides frontline innate defence mechanisms as a physical barrier and via the secretion of protective factors. Interleukin (IL)-17A is an important cytokine that serves a vital role in a number of chronic inflammatory diseases (10,11). Previous studies have reported a greater number of IL-17A+ cells and increased IL-17A secretion in the submucosa of patients with COPD (12–14) and have also investigated the expression of IL-17A and IL-17F in the lung tissues of patients with stable COPD (15). Increased levels of growth differentiation factor 15 (GDF15) have been reported in the airway epithelium of smokers with COPD and in human airway epithelial cells exposed to cigarette smoke; as such, elevated GDF15 expression may contribute to the progression of COPD.
The aim of the present study was to elucidate the role of IL-17A and GDF15 in the pathogenesis of COPD. The results demonstrated that IL-17A and GDF15 are upregulated in patients with COPD, particularly in those with a history of smoking. IL-17A and GDF15 expression is negatively correlated with epithelial (E)-cadherin levels and positively correlated with the mesenchymal marker vimentin in clinical specimens. The results also revealed that treatment with cigarette smoke extract (CSE) and IL-17A was able to induce GDF15 expression, while increased IL-17A and GDF15 expression caused a corresponding increase in EMT in human small epithelial HSAEpiC cells in vitro. The results of the present study demonstrate that IL-17A and GDF15-induced EMT serves an important role in the pathology of COPD.
A total of 143 patients with COPD (57 non-smokers, 86 smokers) and 75 patients without COPD (45 non-smokers, 30 smokers) were recruited from Hunan Provincial People's Hospital during (Changsha, Hunan Province, China) from March 2014 to September 2016. Patients were excluded from the present study if they exhibited acute and chronic pulmonary diseases, including bronchial asthma, bronchiectasis, interstitial lung disease, heart disease and autoimmune disease. Patients were also excluded if they possessed other systemic disorders, including orthopedic, neurologic or unstable cardiac diseases, which may interfere with results. Patients were recruited into the present study if they had not received glucocorticoid and antibiotics within the last 3 months. A total of 99 male and 44 female patients with COPD and average age of 56.4 years (range, 39–74 years) were recruited. In addition, 43 male and 32 female patients without COPD were recruited (average age, 52.5 years; range, 33–72 years). Lung tissue samples were obtained following lobectomy or pneumonectomy for various medical reasons, including solitary pulmonary nodules, peripheral space-occupying lesions and peripheral lung cancer. COPD was diagnosed according to the guidelines of the Global Initiative for Chronic Obstructive Lung Disease (16) as follows: Non-COPD, forced expiratory volume in 1 sec (FEV1)/forced vital capacity (FVC) ≥70% and FEV1 ≥70%; COPD, FEV1/FVC <70% and FEV1 <70%. Patients with a history of any other significant respiratory diseases were excluded. Non-smokers were defined as those who had smoked on average <1 cigarette/day for <1 year or had never smoked. Smokers were defined as patients who had a smoking history of ≥1 year and had smoked on average ≥300 cigarettes/year. None of the patients received corticosteroids prior to surgery. All experiments were approved by the Ethics Committee of Hunan Provincial People's Hospital and informed consent was obtained from all patients prior to specimen collection.
Immunohistochemical (IHC) staining for IL-17A and GDF15 proteins was performed using the labeled streptavidin-biotin method with an LSAB kit (Dako; Agilent Technologies, Inc., Santa Clara, CA, USA) according to the manufacturer's protocol. Surgical specimens were fixed using 10% neutral-buffered formalin. Following fixation for 24–48 h at room temperature, an initial 3–5 mm thick section was routinely processed, paraffin embedded, deparaffinized and rehydrated. 5-µm sections were then cut, placed on charged slides and dried in an oven for 1 h at 56–60°C. Sections were then stored in the dark at 2–8°C and used for the IHC assay. Sections were then immersed in 0.01 Mcitric buffer (pH 6.0) and preheated to 97°C, for 30 min. The slides were incubated with 3% hydrogen peroxide for 10 min at room temperature, followed by incubation in normal goat serum (cat. no. ab7481; Abcam, Cambridge, UK) for 10 min at room temperature. The slides were incubated with antibodies against IL-17A (ab217359; 1:200) and GDF15 (ab206414; 1:100; both Abcam) at 37°C for 1 h, followed by incubation in Biotin-SP (long spacer) AffiniPure Goat Anti-Rabbit Immunoglobulin G (IgG; 1:2,000; cat. no. 111-065-008; Jackson ImmunoResearch Laboratories, Inc., West Grove, PA, USA) for 10 min at room temperature and subsequently with streptavidin peroxidase for 10 min at room temperature. Diaminobenzidene was employed as the chromogen for 5 min at room temperature. Samples were then counterstained with hematoxylin at room temperature for 30 sec and coverslipped for microscopic examination (50I; light microscope; Nikon Corporation, Tokyo, Japan).
Human small airway epithelial cells (HSAEpiCs) were obtained from ScienCellResearch Laboratories, Inc. (San Diego, CA, USA; 3230) and cultured at 37°C in Small Airway Epithelial Cell Medium (cat. no. 3131; ScienCell Research Laboratories, Inc.) in a humidified atmosphere containing 5% carbon dioxide. To induce GDF15 overexpression, the recombinant expression vector pReceiver-Lv154 which contains the coding sequence of GDF15 (NM_004864) was bought from GeneCopoeia, Inc. (Rockville, MD, USA). Lipofectamine™ 3000 (Thermo Fisher Scientific, Inc., Waltham, MA, USA; cat. no. L3000015) was used to transfect cells with DNA complexes according to the manufacturer's protocol. Cells were harvested for RNA and protein extraction 72 h following transfection.
CSE preparation
CSE was prepared according to a previously published protocol (17). CSE was made from one brand of commercial cigarette (Baisha; Hunan Changsha Tobacco Industrial Co., Ltd., Changsha, China) bubbled through 12.5 ml of bronchial epithelium basal medium (cat. no. CC-3171; Lonza Group Ltd., Basel, Switzerland), which was filtered through a 0.2-mm pore filter. To ensure standardization between experiments and batches of CSE, the absorbance was measured at 320 nm on a spectrophotometer. An optical density of 1 was defined as 100%. CSE was frozen in single use aliquots at −20°C. According to previous reports, 1% CSE is the equivalent of 5 cigarettes/day, 3% is 15 cigarettes/day, 6% is 30 cigarettes/day and 10% is equal to 50 cigarettes/day, which corresponds to human daily exposure to cigarette smoke (18,19). The cytotoxicity of CSE on primary basal cells was measured using a Cytotoxicity Detection kit 1644793001; Roche Diagnostics GmbH, Mannheim, Germany).
Cell stimulation
Cells were seeded at a density of 1×106 onto 6-well plates and grown in bronchial epithelial growth medium (BEGM; Lonza Group Ltd.) supplemented with a BEGM™ BulletKit™ (cat. no. CC-3170; Lonza Group Ltd.) containing bovine pituitary extract, insulin, hydrocortisone, gentamycin/amphotericin, retinoic acid, transferrin, epinephrine and human epidermal growth factor (Lonza Group Ltd.). The medium was supplemented with heat-inactivated foetal bovine serum (cat. no. 10099141; Gibco; Thermo Fisher Scientific, Inc.; 10% in the growth medium or 1% in starvation medium). At confluence, cells were starved for 24 h (BEGM + 1% FBS), then treated daily with IL-17A 00 ng/ml) and CSE (10%) for 4 days, or treated with varying concentrations of IL7A (10, 25, 50, 100 ng/ml) or CSE (1, 2.5, 5, 10, 15%) for 3 days.
Reverse transcription-quantitative polymerase chain reaction (RT-qPCR)
Total RNA was extracted using TRIzol reagent (Invitrogen; Thermo Fisher Scientific, Inc., Waltham, MA, USA) according to the manufacturer's protocol. First strand cDNA was synthesised using Superscript II reverse transcriptase (Invitrogen; Thermo Fisher Scientific, Inc.). The reverse transcription temperature protocol was 42°C for 50 min, followed by 70°C for 15 min. Real-time PCR reactions were performed using the SYBR-Green Real time PCR Master Mix (Invitrogen; Thermo Fisher Scientific, Inc.) and PCR was performed using an ABI PRISM 7900 HT Sequence Detection System (Thermo Fisher Scientific, Inc.). The thermocycling conditions were as follows: 95°C for 5 min, followed by 95°C for 30 sec, 58°C for 30 sec and 40 cycles of 72°C for 10 sec. All reactions were performed in triplicate. Primers used for the amplification of the indicated genes are listed in Table I. Results were normalised to endogenous GAPDH and quantified using the 2−∆∆Cq method (20).
Table I.
Primers used in reverse transcription-quantitative polymerase chain reaction.
Primer sequences (3′-5′)
Interleukin 17A Forward ACAACCGATCCACCTCACCTT
Reverse CCCACGGACACCAGTATCTTCT
Growth/differentiation factor 15 Forward GTTGCGGAAACGCTACGAGGA
Reverse CGGAACAGAGCCCGGTGAAG
Epithelial-cadherin Forward CGCATTGCCACATACACTCTCT
Reverse GAGCACCTTCCATGACAGACC
Neural-cadherin Forward TCAGTGGCGGAGATCCTACTG
Reverse TTGACTGAGGCGGGTGCTGAA
Vimentin Forward AACTAATCTGGATTCACTCCCTCTGC
Reverse GAGAAGTTTCGTTGATAACCTGTC
β-actin Forward AGGGGCCGGACTCGTCATACT
Reverse GGCGGCACCACCATGTACCCT
Cells were washed with PBS and lysed with RIPA lysis buffer (cat. no. 89900; Invitrogen; Thermo Fisher Scientific) containing a 10% protease inhibitor cocktail (cat. no. 78429; Thermo Fisher Scientific, Inc.) on ice for 30 min. Protein concentrations were determined using a BCA Protein Assay Reagent kit (Thermo Fisher Scientific, Inc.). Aliquots of cell lysates containing 20 µg protein were separated using 10% SDS-polyacrylamide gel and transferred to polyvinylidene difluoride membranes (Invitrogen; Thermo Fisher Scientific, Inc.). Subsequent to blocking with bovine serum albumin (Sigma-Aldrich; Merck KGaA; Darmstadt, Germany) at room temperature for 1 h, membranes were incubated at 4°C overnight with primary antibodies against the following: E-cadherin (1:500; cat. no. ab15148), N-cadherin (1:1,000; cat. no. ab18203), Vimentin (1:1,500; cat. no. ab137321). The anti-E-cadherin, N-cadherin and Vimentin antibodies were obtained from Abcam. Subsequently, membranes were washed with Tris-buffered saline and Tween-20 (Sigma-Aldrich; Merck KGaA) and incubated with horseradish peroxidase-conjugated anti-Rabbit IgG secondary antibodies (1:1,000; cat. no. A21253; Thermo Fisher Scientific, Inc.). Immunoreactive proteins were detected using an ECL kit (Pierce; Thermo Fisher Scientific, Inc.). The relative protein expression was analyzed using Image-Pro plus software 6.0 (Media Cybernetics Inc., Rockville, MD, USA). The expression of β-actin (1:5,000; cat. no. ab227387; Abcam) was used as an internal control to normalize the expressions of other proteins.
Data were analysed using SPSS v.17.0 (SPSS, Inc., Chicago, IL, USA) and are presented as the mean ± standard deviation. The non-parametric Mann Whitney U test was used for comparisons-between two groups. One-way analysis of variance with a post-hoc Tukey's test was used for comparisons between more than two groups. The correlation between gene expression levels was analysed using Pearson's correlation. P<0.05 was considered to indicate a statistically significant difference.
IL-17A and GDF15 are upregulated in the lung tissues of patients with COPD and a history of smoking
IHC staining and RT-qPCR were performed in 143 samples from patients with COPD and 75 samples from patients without COPD. The results of IHC staining demonstrated that IL-17A and GDF15 were markedly upregulated in lung tissues from patients with COPD compared with non-COPD tissues (Fig. 1A). RT-qPCR revealed that IL-17A and GDF15 expression was significantly increased in patients with COPD compared with patients who did not have COPD (Fig. 1B and C). Furthermore, the expression of IL-17A and GDF15 was significantly increased in smokers compared with non-smokers in the COPD and non-COPD groups (Fig. 1B and C). A significant positive correlation was identified between IL-17A and GDF15 expression (Fig. 1D). These results suggest that upregulated IL-17A and GDF15 expression may contribute to COPD development, particularly in patients with a history of smoking.
Expression of IL-17A and GDF15 in patients with COPD. (A) Immunohistochemical staining of IL-17A and GDF15 in lung tissues. Reverse transcription-quantitative polymerase chain reaction was performed to assess the expression of (B) IL-17A and (C) GDF15 in lung tissue samples obtained from patients with COPD and patients without COPD. (D) IL-17A expression was positively correlated with GDF15 expression.*P<0.05 vs. non-COPD smokers and #P<0.05 vs. non-COPD non-smokers. IL, interleukin; GDF, growth/differentiation factor; COPD, chronic obstructive pulmonary disease.
Correlation between IL-17A, GDF15 and EMT markers
A correlation analysis was performed to assess the association between IL-17A, GDF15 and EMT markers [E-cadherin, neural (N)-cadherin and vimentin] in clinical specimens (Fig. 2). IL-17A expression was significantly negatively correlated with E-cadherin (Fig. 2A), whereas it was significantly positively correlated with N-cadherin (Fig. 2C) and vimentin (Fig. 2E). GDF15 expressionwas significantly negatively correlated with E-cadherin (Fig. 2B) and significantly positively correlated with vimentin (Fig. 2F). No significant correlation was observed between GDF15 and N-cadherin expression (Fig. 2D). These results suggest a possible link between increased IL-17A and GDF15 expression and EMT during the development of COPD.
Correlation between IL-17A, GDF15 and epithelial-mesenchymal transition biomarkers. (A) IL-17A and (B) GDF15 were negatively correlated with E-cadherin. (C) IL-17A was positively correlated with N-cadherin and (D) no significant correlation was observed between GDF15 and N-cadherin expression. (E) IL-17A and (F) GDF15 were positively correlated with vimentin expression. IL, interleukin; GDF, growth/differentiation factor; E-cadherin, epithelial cadherin; N-cadherin, neural cadherin.
The expression of E-cadherin was significantly downregulated in patients with COPD compared to those without COPD, whereas N-cadherin and vimentin were significantly upregulated (Fig. 3).
Expression of epithelial-mesenchymal transition biomarkers in patients with COPD. (A) E-cadherin, (B) N-cadherin and (C) vimentin expression in smokers and non-smokers with or without COPD. *P<0.05 vs. non-COPD smokers and #P<0.05 vs. non-COPD non-smokers. COPD, chronic obstructive pulmonary disease; E-cadherin, epithelial cadherin; N-cadherin, neural cadherin.
GDF15 is upregulated in CSE- and IL-17A-treated HSAEpiC cells
To address whether smoking affects GDF15 expression, HSAEpiCs were stimulated with CSE and assessed. The results demonstrated that the expression GDF15 mRNA significantly increased in a dose-dependent manner following CSE exposure for 3 days (Fig. 4A). Additionally, GDF15 expression increased significantly with exposure to 10% CSE in a time-dependent manner, reaching a peak following 3 days of exposure (Fig. 4B). It was also demonstrated that GDF15 expression was significantly upregulated by IL-17A in a dose- and time-dependent manner (Fig. 4C and D), suggesting that GDF15 expression is affected by both cigarette smoke exposure and IL-17A treatment in HSAEpiC cells.
IL-17A and CSE induce GDF15 expression. (A) Cells were treated with 1, 2.5, 5, 10 or 15% CSE for 3 days. (B) Cells were treated with 10% CSE for 1, 2, 3 or 4 days. (C) Cells were treated with 10, 25, 50 or 100 ng/ml IL-17A for 3 days. (D) Cells were treated with 100 ng/ml IL-17A for 1, 2, 3 or 4 days. *P<0.05 vs. control. IL, interleukin; CSE, cigarette smoke extract; GDF, growth/differentiation factor.
IL-17A and GDF15 induce EMT in HSAEpiC cells
To investigate the role of IL-17A and GDF15 in EMT, HSAEpiC cells were transfected with a GDF15-overexpressing lentivirus and treated with IL-17A (100 ng/ml) for 3 days. The results revealed that HSAEpiC cells lost their honeycomb-like epithelial cell morphology and became spindle shape, which is indicative of EMT (Fig. 5A). RT-qPCR and western blotting were performed to assess the expression of EMT markers and the results demonstrated that E-cadherin was downregulated while N-cadherin and vimentin were upregulated in cells GDF15-overexpressing cells with or without IL-17A treatment compared with control cells (Fig. 5B and C). Furthermore, the combination of IL-17A treatment and GDF15 overexpression had a significantly greater effect on E-cadherin, N-cadherin and vimentin expression compared with GDF15 overexpression alone (Fig. 5B and C). These data suggest that IL-17A, in conjunction with GDF15, is able to induce EMT in HSAEpiC cells.
IL-17A and GDF15 induce epithelial-mesenchymal transition in HSAEpiC cells. (A) Representative images of cell morphology following exposure to IL-17A and/or GDF15 for 3 days (magnification, ×200). Cells were exposed to IL-17A and/or GDF15 for 3 days and the expression of E-cadherin, N-cadherin and vimentin was measured using (B) reverse transcription-quantitative polymerase chain reaction and (C) western blotting. *P<0.05 vs. control; #P<0.05 vs. GDF15 alone. IL, interleukin; GDF, growth/differentiation factor; E-cadherin, epithelial cadherin; N-cadherin, neural cadherin.
The results of the present study demonstrate that IL-17A and GDF15 are upregulated in patients with COPD, particularly in those with a history of smoking, and that IL-17A and GDF15 expression levels are positively correlated with EMT progression. Furthermore, IL-17A treatment in conjunction with CSE-induced GDF15 overexpression triggered EMT in HSAEpiC cells in vitro.
It has been reported that cigarette smoking contributes to lung remodelling and peribronchiolar fibrosis in the small airways, leading to airway obstruction (5). The formation of peribronchiolar fibrosis is associated with EMT (8), while CSE exposure is able to induce EMT in human bronchial epithelial cells (6). COPD has a complex aetiology and is characterised by a combination of airway and lung parenchymal damage (21). The earliest changes in the small airways appear to be due to active EMT, which is associated with a cascade of changes in the expression of regulators, including Smads, Twist and β-catenin (7). Proinflammatory cytokines, including tumour necrosis factor-α, interferon-γ and IL-17, serve an important role in the pathobiology of COPD (22). There is also evidence that high levels of the aforementioned cytokines are responsible for the clinical manifestations of COPD (22,23). Elevated IL-17 expression has been reported in the lungs of patients with COPD; this elevation was attributed to cigarette smoke exposure-dependent effects on the nuclear factor-κB and the phosphoinositide3-kinase pathways (24). Additionally, a combination of IL-17A and CSE stimulation has been reported to affect the proliferation of human epithelial cells and cigarette smoke increases T helper-17 immunity in the lung tissue of patients with COPD (15). In IL-17A knockout mice exposed to cigarette smoke, lymphoid neogenesis was attenuated and chemokine C-X-C motif ligand (CXCL)12 expression was reduced, suggesting that IL-17A contributes to COPD disease progression and the development of lymphoid follicles by activating CXCL12 (25).
It has been reported that the number of CD4+IL-17+ cells is higher in patients with COPD compared with non-smokers, as well as in healthy smokers compared with non-smokers (26). The increase in CD4+IL-17+ cells was positively correlated with pathological changes and airflow limitations (27,28). However, Freeman et al (23) reported a decrease in CD4+ and CD8+ T cells in the peripheral blood during acute exacerbations of COPD, indicating T cell extravasation into inflammatory sites. They also reported that GDF15 is a sensitive marker of cardiopulmonary stress that greatly increases during acute exacerbations of COPD (29). Elevated serum GDF15 levels have been demonstrated to independently predict adverse outcomes in patients with exacerbated COPD (30); furthermore, circulating GDF15 is inversely correlated with rectus femora's cross-sectional area and exercise capacity in patients with COPD (31). This suggests that GDF15 overexpression may be associated with a higher frequency of exacerbations and increased mortality in patients with COPD (32). Importantly, a GDF15 deficiency attenuated cigarette smoke-induced pulmonary inflammation (33) and disruption of GDF15 expression significantly inhibited CSE-induced airway epithelial senescence via activation of the Smad1 pathway (34). The results of the present study revealed that GDF15 expression in HSAEpiC cells was significantly upregulated by IL-17A in a dose- and time-dependent manner. Furthermore, IL-17A in conjunction with GDF15 activated EMT in HSAEpiC cells, which may explain the correlation between IL-17A, GDF15 and EMT markers in clinical specimens. In future studies, knocking out GDF15 and IL-17A is necessary to further determine whether GDF15 and IL-17A are effectors of smoking on COPD. Furthermore, whether monoclonal antibodies for GDF15 and IL-17A diminish the effects of tobacco on airway and lung parenchymal damage in vivo should be assessed in further studies.
In summary, the results of the present study highlight the role of IL-17A and GDF15 in the development of COPD following exposure to cigarette smoke. IL-17A and GDF15-induced EMT was demonstrated to serve an important role in the etiopathology of COPD, which suggests that IL-17A and GDF15 suppression may be an effective therapeutic treatment for COPD. However, more in vivo studies should be conducted in the future.
The authors would like to thank Mrs Merissa E. Garvey for language modifications.
The present study was supported by the Hunan Provincial Natural Science Foundation of China (grant no. 2015JJ2091), the Scientific Research Foundation from Ministry Education of Hunan Province of China (grant no. 15C0832) and the Hunan Provincial Health and Family planning commission of China (grant no. B2017078).
All data generated or analyzed during this study are included in this published article.
GJ conceived and designed the experiments and wrote the manuscript; CTL performed the experiments and analyzed the data; WDZ contributed in collecting clinical tissue samples, performing the IHC assay and revising the manuscript.
Ethical approval for the study was granted from the Hunan Provincial People's Hospital Ethics Committee and all patients gave written informed consent.
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text:
RV-8 and the Crater Lake
Saturday 5/29/21 turned out to be almost exactly as forecast, except maybe a little better. How often does that happen?
Thursday night Carl sent me an email asking f I would be interested in taking a trip to Crater Lake on Saturday if the weather cooperated. I said sure, not really expecting the weather to cooperate. It is something we had been wanting to do for years, but it isn't that often that conditions are favorable over such a large distance. The total trip was about 650 miles. The plan was to fly from Arlington to Albany Oregon for lunch and gas, then fly around the lake, back to Albany for gas and a pit stop, then home.
The weather actually did cooperate. There were virtually no clouds anywhere along the route, the winds aloft were relatively light, and the temperatures were moderate (Albany got to about 78 degrees as we left).
The leg from Arlington to Albany was relatively uneventful and, even though we had a slight head wind, we arrived a few minutes ahead of schedule. In fact, the restaurant had only opened about 10 minutes before we got there.
As we were walking back to the planes, we saw a couple that had come in in a polished RV-7 and we stopped to chat with them for a few minutes. After that we taxied across the runway to the fuel island to top off. Fortunately it was on the way to the take off end of the runway anyway.
About 1:15 we took off headed for the lake. The terrain around the lake is around 9,000' so we started of at 9,500 intending to go higher as we got there. It was a bit bumpy at 9,500' so we went up to 11,500'. It was smoother there. I also noted that at 11,500' the Outside Air Temperature (OAT) was 38 degrees. Because of that, I was surprised to see as much snow as we did.
As we approached the rim we thought the lake would be just beyond the peaks. Turned out, it was surrounded by the peaks. The first thing we both noticed was that it was much larger than we expected.
It is truly a beautiful area and the photos and video I took don't come close to capturing the grandeur of the area. A friend had told me there was a lodge there, but I had no idea it was such a large lodge. There are roads going almost all the way around the crater. It would easily be possible to occupy ones self for a couple of days there at least.
The lodge was much bigger than I expected.
When we got there we were the only airplanes in the vicinity. When we were about halfway through our first orbit of the lake I noted 3 mores planes coming in. With the mild turbulence making picture taking difficult and the fact that there were now other airplanes in the area we decided to head out after just one orbit.
It was a bit bumpier heading out so we went to 12,500' for the first part of the trip back. It was very smooth at that altitude and it was a pleasant trip back.
We could have made it home with the fuel we had remaining, but that would have been another hour and forty five minutes at least. We were ready for a break and a pit stop, and it never hurts to have more fuel on board (unless you're on fire).
The length of this trip was a little more than my cameras could bear and by the time we got back to Albany both of them had crapped out. The wing camera battery died just after I landed. I had extra batteries to bring, but they had been sitting so long they were dead (did someone say lack of planning?). The other camera would not stay on. There was plenty of room on the SD card and plenty of battery power left, but every time I turned it on, it turned itself right off. That is why the video ends at Albany. On the other hand, not much happened on the way home and it looked just like the way down, but in the other direction.
We took off for home a little after 4:00 and I landed a little after 5:30. We had a head wind down to Albany, we had a head wind to Crater Lake from Albany at 9,500' and 11,500', and we had a headwind from Crater Lake back to Albany at 12,500'. I was hoping we would finally get a tail wind going home and we did, though is wasn't that strong a tail wind, though it was enough to give me ground speeds of about 199 MPH most of the way back.
It was a very enjoyable trip though it was a long day. It was 5.3 hrs on the Hobbs, 4.7 hours on the tach and about 8 hours on the clock. I would like to do it again sometime, maybe in the winter with more snow on everything. Now, if there was a runway near the lodge...
Arlington to Albany track here.
Albany to Crater Lake and back track here.
Albany back to Arlington track here.
Still photos are here.
Video is here.
RV-8 and the New Year's Richland Run
RV-8 and the Tale of the Toasty Coastal Trip
RV-8 and the Cloudy Weekend
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text:
Медаль Дарвіна () — медаль, що вручається Лондонським королівським товариством раз на два роки за видатні досягнення в біології в областях, в яких працював Чарлз Дарвін

Перше нагородження відбулось в 1890 році. Медаль була створена в пам'ять Чарльза Дарвіна і надається разом з грошовим призом в £1000. Як і більшість медалей Королівського товариства, кандидат повинен походити з Співдружності націй, при цьому вони повинні бути громадянами співдружності або жити в одній з держав-членів принаймні три роки до номінації. З моменту свого створення медаль було присуджено 64 особам, у тому числі її одержав Френсіс Дарвін, син Чарльза Дарвіна, і дві подружні пари, Джек і Іоланда Хеслоп-Харрісон в 1982 році і Пітер та Розмарі Грант в 2002 році. Першою медаллю був нагороджений Альфред Рассел Воллес, відомий біолог і натураліст, який незалежно від Чарльза Дарвіна приблизно в один і той же час розробив теорію еволюції шляхом природного відбору.

Примітки

Нагороди Лондонського королівського товариства
Нагороди з біології
Нагороди, засновані 1890

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ROSTOCK, Germany Jan 17 2023
INDIAN OCEAN Jan 17 2023
SINGAPORE Jan 14 2023
230119-N-MW880-1110
ADRIATIC SEA (Jan. 19, 2023) Naval Air Crewman (Helicopter) 3rd Class Christian Pedro, assigned to Helicopter Sea Combat Squadron (HSC) 5, is raised out of the water during a combat search and rescue training evolution, Jan. 19, 2023. Carrier Air Wing (CVW) 7 is the offensive air and strike component of Carrier Strike Group (CSG) 10 and the George H.W. Bush CSG. The squadrons of CVW-7 are Strike Fighter Squadron (VFA) 143, VFA-103, VFA-86, VFA-136, Carrier Airborne Early Warning Squadron (VAW) 121, HSC-5, and Helicopter Maritime Strike Squadron (HSM) 46. The George H.W. Bush CSG is on a scheduled deployment in the U.S. Naval Forces Europe area of operations, employed by U.S. Sixth Fleet to defend U.S., allied and partner interests. (U.S. Navy photo by Mass Communication Specialist 3rd Class Samuel Wagner)
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Location: ADRIATIC SEA


Neue Normalitat Wie Unternehmen Sich Permanent Selbst Erneuern is good choice for you that looking for nice reading experience. We hope you glad to visit our website. Please read our description and our privacy and policy page.
Finally I get this ebook, thanks for all these Neue Normalitat Wie Unternehmen Sich Permanent Selbst Erneuern can get now!


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Murder probe launched after man is stabbed to death in Kingsbury
A 38-year-old man has been arrested on suspicion of murder and remains in custody
Doreen Avenue, Kingsbury, where a man was found stabbed to death (Image: Google)
A murder investigation is underway after a man was stabbed to death in Kingsbury .
Emergency services were called to Doreen Avenue on Sunday (August 7) at 2.51am following reports of an assault.
A 41-year-old man was found with stab wounds and was pronounced dead at the scene shortly afterwards.
The man's next of kin have been informed but police said officers were awaiting formal identification.
A 38-year-old man has been arrested on suspicion of murder and is being held at a north London police station.
This is the fourth fatal stabbing on west London's streets in the space of a week, following incidents in Southall , Kilburn and Russell Square .
Detectives from the Homicide and Major Crime Command are investigating, and police said a post-mortem examination would be held in due course.
Anyone with information is asked to call the incident room at Hendon on 020 8358 0300. To remain anonymous, call Crimestoppers on 0800 555 111.
Aldi's urgent warning to anyone who eats its chocolate
AldiThe shop had some pressing allergy information to share with its shoppers


text:
Harmanda medioimmicans is een hooiwagen uit de familie Sclerosomatidae.

Sclerosomatidae


Auckland, located on the North Island and built around two harbors, is the largest city in New Zealand. If you're looking for a vibrant city life that's within reach to hundreds of beaches and epic hikes, consider Auckland. There is adventure everywhere. Students usually spend weekends exploring local mountains and forests or meeting up with Program Directors Dan and Max for a Surf. To be honest, we surf a lot. Auckland is also home to the largest Polynesian population in the world and has amazing food! You will not be disappointed!
Dan is an expert in environmental sciences and earth systems, and has a unique ability to blend his scientific knowledge with his Maori heritage. Dan has designed and taught field geology and environmental science courses for Te Whare Wananga o Awanuiarangi in Whakatane and is team leader on several University of Auckland and Frontiers Abroad research initiatives in the Bay of Plenty region on New Zealand. He is currently leading research efforts to integrate indigenous knowledge into modern science practices for environmental restoration and geothermal energy. In addition to his research and Frontiers Abroad duties, Dan instructs courses for the School of the Environment, University of Auckland.


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Your Chosen Genres	[ Art House ]	[ Before Christ (BC) ]	[ Drama ]	[ Natural History (Wildlife) ]	[ Recommended ]	[ Romance ]	Can be Combined with Other Genres. Click here to Combine Genres!
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CLOUDS OF SILS MARIA (2014)
At the peak of her international career, Maria Enders is asked to perform in a revival of the play that made her famous twenty years ago. But back then she played the role of Sigrid, an alluring young girl who disarms and eventually drives her boss Helena to suicide. Now she is being asked to step into the other role, that of the older Helena. She departs with her assistant to rehearse in Sils Maria; a remote region of the Alps. A young Hollywood starlet with a penchant for scandal is to take	find out more...
LOVE IS STRANGE (2013)
It's the off-season at the lonely Beauregard Hotel in Bournemoth, and only the long-term tenants are still in residence. Life is stirred up, however, when the beautiful Ann Shankland arrives to see her alcoholic ex-husband, John Malcolm, who is secretly engaged to Pat Cooper, the woman who runs the hotel. Meanwhile, snobbish Mrs Railton-Bell discovers that the kindly if rather doddering Major Pollock, played by David Niven, who won an Oscar for his performance, a retired officer who likes to find out more...
THOU WAST MILD AND LOVELY (2014)
"Josephine Decker has created a new style of thriller that employs allegory, incorporates touches of David Lynch as well as Magritte -esque imagery. Decker's setting of a remote farm feels like a metaphor for what turns out to be hell. The raw and emotional (and yes, sometimes funny) dialog tells a story that can seem familiar at points but really is meant to keep you guessing and off balance. I really enjoyed how the undertones of this film came to life through her very deft contrast of the find out more...


Dragon Skull by bravach is licensed under the Creative Commons - Attribution - Share Alike license.
3D Scan of a decorative dragon skull originaly made in synthetic stone.
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The model is really small so it must be upscaled to your liking.
I printed it only one time at 30 mm height (original skull should be around 150 mm).
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Due to lack of support for our bus trips over the last few years, it was decided at our recent AGM to see if a reduction in cost would prompt more people to come along on our trips.
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Sarah Jeong
Gabrielle Bourne Media 3 January 2021 8 April 2021 Comments Off on Sarah Jeong
Sarah Jeong New York Times is a racist and here is why. If you want to discuss 'racism' to make a positive change, then more power to you. But Jeong never did that. Instead, she mocked people. She stated her hatred for cops and white people. Jeong is a public figure and thus should be a 'role model'. She is in a position to influence for good. Instead, she chose to promote racist thoughts. Her tweets were full of venom. She illustrated her hatred.
The role of the New York Times in the Race-baiting
All the whilst, The New York Times went along with it. We are lumping Sarah Jeong and New York times together because they are both race hustlers. The New York Times also condoned her actions. Here is what they stated:
Sarah Jeong didn't last long. She left the editorial board one year later in August 2019. But she still remained at the New York Times in some capacity. Deputy editorial page editor Kate Kingsbury told CNN, "Sarah decided to leave the editorial board in August. But we're glad to still have her journalism and insights around technology in our pages through her work as a contributor." Her remaining proved that the New York Times condoned her tweets.
Racists don't change spots. They get hired
Here is a piece we wrote in July 2020, titled, Sarah Jeong believes 'reverse racism' isn't real but she's still a racist. We also included Nick Cannon. He stated, "white people" are "closer to animals" and "savages". He was fired by ViacomCBS for his anti-Semitic rhetoric in July 2020 and rehired by them less than one year later in February 2021. Apparently Cannon showed enough remorse.
Priyamvada Gopal stated, "White Lives don't matter". She said that it was to counter 'White Lives matter'. She supports black lives matter. If she valued black lives, then she should also value white lives. A life, no matter the skin colour should have value. Only racists focus on skin colour.
This is the same woman that demanded that porters at King's College call her "Dr Gopal" instead of 'madam'. It's ironic, how she exercised her 'class privilege' whilst accusing the porters of discrimination. Kings Cross immediately shot back and defended the porters. They did not find any wrong doing.
Gopal is a Professor of Postcolonial Studies in the Faculty of English at the University of Cambridge
Sarah Jeong 'racist' tweets caught up to her
Sarah Jeong comes to the New York Times with the baggage of inflammatory and racist Twitter posts. Howard Kurtz has more for 'Special Report.'
~ Fox News 3-August-2018
"People of colour can be racist too"
gayle king – Race-baiter / Flying monkey
IN DENIAL – SoHo Karen Is A Person of Colour And CAN'T Be Racist? Miya Ponsetto, also known as "SoHo Karen", told Gayle King that she cannot be racist because she is a person of colour. 22-year-old Ponsetto got into an altercation with 14-year-old Keyon Harrold, Jr. over her cell phone at the swanky "Arlo" Hotel in Manhattan. The incident was, of course, recorded, and after the video went viral. ~ Anthony Brian Logan 12-JAN-21
Happiness is a Habit!
Suzanne Soto


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Béhagnies ist eine Gemeinde im französischen Département Pas-de-Calais in der Region Hauts-de-France. Sie gehört zum Kanton Bapaume im Arrondissement Arras. Sie grenzt im Norden an Ervillers, im Nordosten an Mory, im Südosten an Sapignies, im Süden an Biefvillers-lès-Bapaume, im Südwesten an Bihucourt und im Nordwesten an Gomiécourt.

Die ehemalige Route nationale 37 und heutige Départementsstraße 917 führt über Béhagnies.

Bevölkerungsentwicklung

Weblinks

Ort in Hauts-de-France
Träger des Croix de guerre 1914–1918 (Ort in Frankreich)


Yak — a reflection of all the good that represents the animal, power, ruggedness, endurance, generosity. The need for personal power. Reflection of a blind rage.
To see a yak in your dream, represents your uniqueness and dependability. The yak may also be pun on yakking too much. Maybe you should stop talking and listen more.


The Cave of Amarnath is a well known shrine in hindu religion, which is situated in Jammu & Kashmir and is dedicated to Lord Shiva. It is located at about a distance of 141 Kms from Srinagar, at 3888 ms from capital of J & K and is possible to be reached by passing through Pahalgam Town. Snowy mountains cover the region of the cave, which is itself covered by snow in most seasons, except for a small period in summer, during which devotees can travel to the shrine. There is a pilgrimage undertaken annually by millions of hindu devotees to Amarnath Cave, who have to pass through tough mountain terrain, to have a view of the ice stalagmite which has been formed in this place.
Inside of amaranth cave which is 40 m high, there is a formation of ice stalagmite, due to the water freezing while dropping from the cave's roof and there is also a vertical growth from floor of the cave. This is thought to be a Shiva Linga by hindu worshippers. From the period of May to August, the snow melts and it waxes so that water keeps on seeping through the rocks. There is also a belief that the lingam increases in size with moon reaching at its highest position in summer and then shrinks, even though scientific evidences are not found.
As per the observations of a legend in hindu culture, the importance of the cave is associated with the explanation of life's importance to his wife Parvati, and there are two more formations in ice which represent Ganesha and Parvati.
Many people visit Amarnath Caves because they want to have a glimpse of the lingam made of ice stalagmite.
Since time immemorial, Amarnath Caves have become a place of worship, which is evident from references to this cave in Aryaraja performing puja for a lingam also made from ice in Kashmir. The name of the book is Rajatarangini, where there is reference to Amarnath or Amareshwara. Another belief holds that Suryamathi queen used to gift banalingas, trishuls and various other emblems of sacred importance to this temple around 11th century. A reference known as Rajavalipataka written by Prjayabhatta also contains references about visiting Amarnath cave. Plenty of other resources also have references to this cave pilgrimage.
After the middle ages were over, people forgot about the cave. But in 15th century, it was discovered by a shepherd and there came up various other stories about this cave's discovery. According to one of these stories, Buta Malik who was a shepherd received a coal bag from a holy man. On reaching his home, he looked into the bag and found coins instead of coal. When he returned overjoyed to the place where holy man was found, he instead discovered a lingam and cave.
Nearest airport to Amarnath Cave is at Srinagar, which in itself is a beautiful valley with lots of world famous sightseeing places like Nagina Lake, Dal Lake, Mughal Gardens, Shankaracharya Temple and others. Among the tourist places, this is highly sought after, known by some as paradise on earth. It is also known as summer capital of the country. From delhi and jammu, daily flights reach Srinagar, with some weekday flights reaching the place from Amritsar and Chandigarh.
Nearest railway station is that in jammu, which is considered as winter capital of J & K. this is a beautiful place with lots of temples, giving it the name of City of Temples. Various old temples can be visited such as Mahadev Mandir, Raghunath Temple and few others. There are good connections to many other cities of India.
Good road communication is present between jammu and Srinagar, with many buses and taxis plying on this road, which can be used to travel on the journey towards Amarnath Cave.
It is being considered that hindu God Shiva gave an insight of life and eternity in front of his consort Goddess Parvati. There are two more formations in ice, representing Goddess parvati and their son, Ganesha.
Shiva Lingam in Amarnath is considered one among the holy trinity, where Lord Shiva is thought of as a god who is living. There are vedic myths, rituals, and hymns found in rig veda, which evoke and testify the presence of Lord Shiva since time of world's beginning. According to legends, it is thought that Lord Shiva told his wife Goddess Parvati about various secretes related to creation inside this cave. Annual yatra is undertaken every year to this cave, which is located at height of about 14000 ft and the yatra is managed by State Government of J & K, mostly during months of July and August. Devotees visiting the holy shrine are let to conduct darshan from the time of Ashard Purnimashi till the period of Shravan Purnimashi, which extends to about a month or so.
The Cave of Amarnath was among the places selected by Lord Shiva to explain theories of creation and immortality to Parvati, who was his consort. She was of the query as to how he started to wear beads made with heads. In answer to this question, Lord Shiva answered that whenever Parvati was born, he used to add another head to the beads. On such an answer, parvati queried that why was she born again and again, while Lord Shiva was living ever. In reply, he answered that such a phenomenon was because of Amar Katha.
On regular insistence and continually avoiding the answer to such a question by Maa Parvati, Lord Shiva gave in one day and decided to tell the truth behind the secret of immortality. He wanted to reveal all the secrets to Parvati, but didn't want anyone else to listen due to which he chose Amarnath Cave.
When going for this purpose, he left his ride Nandi in Pahalgam, which is also known as Bail gaon. On the way, at Chandanwari, the moon was released by him from his hairs while the snakes were released at the banks of Lake Seshanag. Ganesh was left in the Mahagunas parvat. And finally at Panjtarni, five elements of air, water, earth, sky and fire were left behind. It is also believed that at this juncture, in order to sacrifice various matters of earthly world, Shiva and parvati did a Tandav Dance. Then only Bholenath and Maa Parvati entered into the Holy Shrine of Amarnath Cave. After that, Lord Shiva Went into Samadhi by sitting over a skin of deer and concentrated in such a manner that his tale wouldn't be heard by any living creature in and around the holy cave of Amaranth. Then he created Kalagni named Rudra and asked to spread fire all around, so that every living creature would be destroyed. And then he narrated the story of his immortality secret. But there was a bird's egg, which was lying under the deer skin was not harmed by the fire. Then the pigeon pair which was born out of this egg were said to have become immortal as they had heard the immortality secret.
Climate in the region of Amarnath Cave and the path leading up to it is quite difficult and uncertain with snowfall and rain happening any time, sometimes without warning. Abrupt changes in temperature is common, with sun and rain being interchanged in a very short time. Sometimes in winter, temperature goes down below -5 deg C.
Amaranth Cave yatra is usually undertaken during the months from July to August, which is the month of Sharavan, when lakhs of devotees take the difficult path to have a view of the lingam made from ice stalagmite, which waxes and wanes through the different cycles of the moon.
Arrangement for accomdoations are done near the shrine by means of tents with beds, blanket, mattresses, pillow and bedsheets. Sharing meals are also provided at Sheshnag, Panchtarni, Chinaware and Baltal.
People can avail hotel accommodations at halts in pahalgam and in Srinagar on houseboats, as per their tour plans.
People can avail transportations with 6-8 passengers in non A/C tata sumo, which also takes them for sightseeing.
Devotees need to get registration before they go into the region near the holy shrine for which Amarnath Yatra registration is required to be done.
Improving physical well being – Since the yatra to Amarnath Cave requires walking for long distance, people should be prepared before hand with regular walking practices for 4-5 kms per day, by a month before the yatra is undertaken so that the body gets accustomed for such rough journey.
Body should also be massaged on a daily basis with mustard oil and bathing in luke warm water so that one will have flexible body which will not get tired easily. This is also beneficial for reducing any harm to the body due to the quickly changing weather.
Breathing efficiency has to be increased – this will allow pilgrims to have better supply of oxygen to the body and yoga exercises such as Pranayam should also be practiced.


As you may have seen in a previous blog the lovely people at Decathlon sent me a few items to review and this is the second in a series of products I will be reviewing.
Decathlon cater for lots of different sports and when I opened a box of goodies from them, in addition to the cycling gear I was rather pleased to see a pair of running shoes - in a very favourable colour!
To give them their full title the 'Kaenji Eliorun Men's Wellness Running Shoes' in orange are vivid to say the least.
There is a lot of orange mixed in with yellow, white, grey and black and several parts that offer a reflective surface when jogging at dawn/dusk. The reflective elements are on the heels, sides and tongue and I have to say very effective.
These shoes are very light and when wearing them you really don't notice any weight at all.
There is cushioning in the heel called 'K-ring heel technology' and it really does offer a fair degree of shock absorption.
I really like these shoes - or course for the colour - but they are very comfortable and I have to report at least as good if not better than some of my jogging trainers costing a great deal more.
There are other colours available - blue and grey I think - but for me these are perfect. Again I suspect you are thinking about the price? Well they cost £39.99 and in addition to what feel is again another great price point from Decathlon, these shoes actually come with a 2 year guarantee!
As yet I am not sure whether Decathlon want these back after I have reviewed them? I can only say that with the summer hopefully with us, they are not getting these back!
B'Twin Orange Mountain Bike Shorts - Very Orange!


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Baki Season 4: Renewal Confirmed? Is It The Last Season? Everything To Know!
The second strongest fighter in the world has finally returned to the screens again. After the crazy finale of the recently released third season, fans are dying to watch Baki Season 4. As per the looks, the fourth installment can turn out to be much more exciting than ever before. However, has Netflix renewed it yet? Here's everything you need to know.
Baki the Grappler debuted three decades ago as a manga series. The 1991 manga ended up becoming one of the best sports manga of that era. In 2001, Japanese animation studio Group TAC produced an anime adaptation that is still praised. Years later, Netflix acquired the rights to create an ONA (Original Net Animation). The first season came out in June 2018 and was split into two parts (considered as Season 1 & 2 for the international audience).
The streamer confirmed a second installment (considered as Season 3), which is likely to be split into two parts as well. The third part recently released on June 4, 2020, and has been receiving immense praise from viewers around the globe. Fans have begun to demand a fourth season already.
Baki Season 4: Netflix Renewal
Netflix's famous martial arts series received much praise from viewers and critics alike. Amidst the Coronavirus pandemic, quarantined fans are now enjoying freshly released 13 episodes of the anime. Well, the majority of the audience as already binged the entire season by far. With every passing day, the demand for Baki Season 4 seems to be increasing. However, the streamer is yet to renew the fourth part.
Fortunately, chances for the renewal of another season are pretty high. The streamer has been releasing all Baki seasons in two 13-episode cours. Hence, the story of the third part will remain incomplete without Baki Season 4. Although even if the creators are planning to produce another installment, Netflix won't make it official anytime soon. The popular streaming giant takes at least a month to greenlight a series. Thus, fans might need to wait for a few weeks to get a formal announcement. We will update this section as soon as it comes out.
Is This The Last Season?
Does Netflix have enough source material for Baki Season 4?
While the 2001 anime adapted the plot of the Grappler Baki manga series, Netflix's 2018 series covers the New Grappler Baki (AKA Baki) manga. The first season of the anime (Part 1 & 2) ended up covering chapters 1 through 178 of the New Grappler Baki manga. The recently released third season of the show (2020) consumed more than 80 chapters after that. Thereby, Netflix now has only a few volumes left to cover in Baki Season 4.
Well, the remaining volumes of the New Grappler Baki won't be enough to another 13 episodes of the anime. Hence, it is speculated that Netflix might use the Baki Hanma manga series for the fourth part. The entire Baki manga series consists of five parts (including sequels). Hence, the streamer has enough source material for continuing the anime adaptation. However, the renewal of all future installments depends upon the performance of Baki Season 3 and 4.
The popular franchise centers around the story of its main protagonist Baki Hanma. The 17-year-old is known as the second strongest fighter in the world. After years of training, Baki gives a tough fight to his powerful opponents and intends to grab his father's first place as the strongest martial arts fighter worldwide.
The 2020 installment of the anime adaptation follows the Chinese Challenge story arc as Baki enters the Raitai Tournament. It begins where the second season left. After being poisoned, the lead character is rushed to China, seeking a cure. While his body is still weak, Baki participates in the biggest martial arts tournament in the country. Amongst several others, his father Yujiro Hanma (world's strongest fighter) and Mohammad Alai Jr are also going to participate in the battle.
The Baki vs. Yujiro fight is going to be the highlight of the upcoming season. It will be based on the Godlike Clash Of The Kids Saga. Hence, in the later episodes, we'll see Mohammad Alai Jr challenging the protagonist for a fight. However, upon being rejected, Alai Jr proposes to Baki's girlfriend so that he fights back.
Baki Season 4: Release Date
It hasn't been long since the brand new episodes of Baki Season 3 released. Yet, fans are already eager to watch more episodes. As of now, Netflix has not confirmed the renewal of Baki Season 4. However, speculations say that the fourth part might arrive sooner this year. Currently, an October 2020 premiere date seems most likely. Keep checking this corner for all future updates.
I'm Sorry Season 3: Will It Arrive In 2020? Plot Details, Release Date & More
Kengan Ashura Season 3: Release Delayed! Is It The Last Season? Details


BLANCHETON, Bertrand, and Samuel MAVEYRAUD. "Central Bank Independence and Inflation: The Government Debt and Inflation Targeting Channel". Finance Bulletin 1, no. 2 (July 18, 2018): 13-33. Accessed April 20, 2019. https://financebulletin.org/article/view/2128.


From the red-rose 'city' of Petra to the luxurious spas lining the Dead Sea, Jordan offers something for every taste and budget. When are you going?
Jordan is best known for its true world wonder, the rose red city of Petra, carved from stone and long hidden in its desert canyon.
But Jordan is far from a one-trick pony (or should that be camel?). Elsewhere, Roman remains at Jerash and Azraq, Hellenic ruins at Umm Qais and crusader castles at Karak and Shobak draw history buffs.
And Jordan's natural charms are just as alluring: float in the briny Dead Sea, dive the dazzling depths to explore Red Sea reefs at Aqaba, spot wildlife in the valleys and heights of Dana Nature Reserve, and wander the desert expanses of Wadi Rum – by foot, 4WD or camel power.
Sprinkle in Jordan's tempting cuisine, great walking and canyoning, easily traversable distances and smiling people, and you've got the recipe for the perfect Middle East mezze.
The key to understanding Jordan is its people and its landscapes. Paul Morrison spent time with both, travelling with Bedouin across the country's vast deserts, unlocking the secrets of both Jordan's past and present. If you don't feel compelled to don a head scarf and head into the sand dunes after reading it, then perhaps Jordan isn't for you.
Amar Grover tackled the Malaqi Trail, a stunning trek which snakes its way through the dramatic canyons, waterfalls and rock pools of the Wadi Mujib Nature Reserve. Once a Bedouin hunting ground, it now hosts an Ibex breeding program and the odd hyena. Find out why Amar reckons it's a wadi wonderland.
Fancy a Middle East self-drive holiday? No? Well, perhaps Nick Redmayne can convince you to change your mind. Being mater of his own destiny opened up all kinds of opportunities he'd have otherwise missed – habibi hip-hop fusion, anyone – and helped him discover why Jordanian drivers are always honking their horns.
If being slathered in mud and wrapped in clingfilm is more your thing, swing by Gail Simmon's Guide to Dead Sea spas. It will do wonders to your complexion.
The 'rose-red city' of Petra is Jordan's biggest attraction and a genuinely breathtaking world wonder – the lost desert city of childhood imagination.
Spectacular location behind impenetrable mountains – check; lost for centuries – check; tales of hidden treasure – check; fascinating remnants of a long-forgotten culture – check. Petra delivers, with bells on.
But with thousands of visitors each day, it's vital you plan your visit accordingly.
Matthew Teller's Complete Guide to Petra is a great place to start, an exhaustive overview of everything the rose-red city has to offer. His 6 Petra Highlights piece is worth a look too, especially if you are pushed for time.
Ten Wanderlust readers joined Lyn Hughes and photographer Paul Harris in Jordan to research and produce the Ultimate Top 5 Guide to Petra and Wadi Rum.
And don't forget to check out Stanford's suggestions for the 7 books to take to Petra. From guidebooks to narratives, they'll ensure you'll return from Petra with a deeper and fuller understanding of history and lasting allure.
There's more to Jordan than Petra, of course, and our guide to off-beat Jordan will make sure you find it. From Crusader Castles to wetlands teeming with birdlife, no hidden corner is left unexplored.
David Symes was Director of Jordan Tourism and visited the country over 40 times. He reveals his favourite restaurant in the country and a nature reserve you must visit here.
Finally, Hossam Moussa lifts the lid on the Jordanian capital, Amman. A finalist in the Wanderlust World Guide Awards, he contends that Amman is seriously underrated as a tourist destination and explains why.
From sophisticated Lebanese styles to basic Bedouin fare, Jordanian cooking is a cosmopolitan mix of influences – and one of the great pleasures of travel in the Middle East.
Matthew Teller gives you the lowdown on Jordan's finest food and drink, from street stalls to its better restaurants, including a handy guide to the country's increasing well-regarded wines.
Sally Broom, on the other hand, visited Beit Sitti – 'My Grandmother's House' – a unique cooking school in Amman where she learned to prepare Mensaf like, well, Grandma, used to make. Find out how you can too.
The traditional chefs of Petra kitchen have kindly shared the recipes of their favourite soups, salads and mezzes so you can bring the tastes and aromas of Jordan to your dining room at home.
With its surreal landscapes and photogenic tribes, Jordan is photographic nirvana. But with that comes a particular set of problems. Petra is often crawling with tourists, and besides, even if you do manage to keep them out of your view finder, how do you get a shot that hasn't been taken a million times before?
Thankfully, our resident photographic guru, Paul Harris, has the answers. You'll find them in his piece entitled Photographing an icon: Petra.
You might also want to see some of the fantastic images taken by our readers.
Ready to plan your trip? Our Jordan travel guide is the perfect place to start. Our Petra Travel Guide is worth checking out too. If you have any questions, pop over to the myWanderlust forum where our knowledgeable community is ready to spring into action and share all that they know. Or check out the questions people have already asked about Jordan. The answer to yours might already be there.
Here's a selection of fantastic tours offered by our partners. From multi-activity adventures and to more leisurely explorations of Jordan's wonders, there's sure to be something that tickles your fancy. You'll find the full list of tours available here.


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An interview with two Chiang Mai activist artists
Activist artists from across Thailand are beginning to express themselves
By Will Langston | Mon 3 Aug 2020
With so many protests and forms of activism popping up around Chiang Mai recently, two young performance artists speak up.
Benz Pisuttisakand and Yossunthon Ruttapradid or 'Ten' as he's called, are both Fine Arts students at Chiang Mai University. They are in their fourth year of study and both are majoring in Media Art. Media Art is an interdisciplinary field that deals with current developments in digital art, new media art and electronic art.
Benz is 22 and Ten is 21 and they are both blossoming Thai performance artists. A slight rain began as we stood under a tree just outside Tha Pae Gate in between performances.
"I come with a friend, we study art together," said Benz about the performances, "I just want to practice my way to express freedom of speech and action in a public space. As an artist. As a citizen of this country."
Benz is short with patchy whiskers, shaggy hair and black round-frame glasses.
He continued, "There are a number of performers from The Lanyim Group, and I'm with my friend for today. We just come to observe and to practice our artwork." The performance began at 7am and ran until midnight with performances at every hour of the day at Tha Pae Gate. Multiple local art organisations unified for this event.
The name of the performance was entitled "EAT (I'M) ARE". The title is wordplay based on the famous steak eatery in Thailand called 'EAT AM ARE'. According to one CMU student the title alludes to, "Someone or something that has more power. And, that someone or something is trying to eat, limit or disqualify their freedom and rights." The concept echoes other recent public performances with the constant motif being 'freedom'.
Benz performed emotional drawings on a lime green easel. He said, "Today I chose the action of drawing as my self expression while using a public space. I don't expect people to feel anything specific because I can't really predict the direction it would be. I just expect them to see me there. That's enough for me. I want to make it recognisable that we have the right to free speech and the right of using public space. There are a lot of absurdities that happen these days."
Ten, who is taller and sported a white tee-shirt with red writing scribbled across it said about his motivations, "I wanna try my first time performing art in a public space and I have a message that I want to share with the public. I think this place is more impactful than any art gallery and I think the engagement of the audience is wider than any art gallery."
In Ten's performance, he laid with his head on a pillow looking up at a cardboard guillotine with the words "Lucid Dream" painted on it. "I want to tell people about the violence of the government. My body position and the pillow express irony with a sense of comfort and danger. When I perform, I really try to sleep, but I cannot because of my anxiety about the danger I see."
Both Benz and Ten are calm and cool as they speak. Surprising so, considering the police that were present during their performances. When asked about their concerns of law enforcement Ten commented, "I'm not scared," he slowly pulled a cigarette from a box, "But, I use the advantage of art to speak as the message. And, if I were to ever get in trouble, I just think that I choose this path by myself. I prepared my heart quite well."
The rain picked up just as the two artist friends finished packing up their performance gear. And, just as the three of us finished saying our goodbyes, another performance artist began laying a black blanket on the ground in preparation.
There would be another 14 hours of performances to go.
For more information about future events, please contact:
https://www.facebook.com/LanyimGroup
www.willlangston.com