Increased FGF1-FGFRc expression in idiopathic pulmonary fibrosis

Background

Recent clinical studies show that tyrosine kinase inhibitors slow the rate of lung function decline and decrease the number of acute exacerbations in patients with Idiopathic Pulmonary Fibrosis (IPF). However, in the murine bleomycin model of fibrosis, not all tyrosine kinase signaling is detrimental. Exogenous ligands Fibroblast Growth Factor (FGF) 7 and 10 improve murine lung repair and increase survival after injury via tyrosine kinase FGF receptor 2b-signaling. Therefore, the level and location of FGF/FGFR expression as well as the exogenous effect of the most highly expressed FGFR2b ligand, FGF1, was analyzed on human lung fibroblasts.

Methods

FGF ligand and receptor expression was evaluated in donor and IPF whole lung homogenates using western blotting and qPCR. Immunohistochemistry for FGF1 and FGFR1/2/3/4 were performed on human lung tissue. Lastly, the effects of FGF1, a potent, multi-FGFR ligand, were studied on primary cultures of IPF and non-IPF donor fibroblasts. Western blots for pro-fibrotic markers, proliferation, FACS for apoptosis, transwell assays and MetaMorph analyses on cell cultures were performed.

Results

Whole lung homogenate analyses revealed decreased FGFR b-isoform expression, and an increase in FGFR c-isoform expression. Of the FGFR2b-ligands, FGF1 was the most significantly increased in IPF patients; downstream targets of FGF-signaling, p-ERK1/2 and p-AKT were also increased. Immunohistochemistry revealed FGF1 co-localization within basal cell sheets, myofibroblast foci, and Surfactant protein-C positive alveolar epithelial type-II cells as well as co-localization with FGFR1, FGFR2, FGFR3, FGFR4 and myofibroblasts expressing the migratory marker Fascin. Both alone and in the presence of heparin, FGF1 led to increased MAPK-signaling in primary lung fibroblasts. While smooth muscle actin was unchanged, heparin + FGF1 decreased collagen production in IPF fibroblasts. In addition, FGF1 + heparin increased apoptosis and cell migration. The FGFR inhibitor (PD173074) attenuated these effects.

Conclusions

Strong expression of FGF1/FGFRs in pathogenic regions of IPF suggest that aberrant FGF1-FGFR signaling is increased in IPF patients and may contribute to the pathogenesis of lung fibrosis by supporting fibroblast migration and increased MAPK-signaling.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0242-2) contains supplementary material, which is available to authorized users.     

Cytofluorometric detection of wine lactic acid bacteria: application of malolactic fermentation to the monitoring

In this study we report for the first time a rapid, efficient and cost-effective method for the enumeration of lactic acid bacteria (LAB) in wine. Indeed, up to now, detection of LAB in wine, especially red wine, was not possible. Wines contain debris that cannot be separated from bacteria using flow cytometry (FCM). Furthermore, the dyes tested in previous reports did not allow an efficient staining of bacteria. Using FCM and a combination of BOX/PI dyes, we were able to count bacteria in wines. The study was performed in wine inoculated with Oenococcus oeni (10⁶ CFU ml^?1) stained with either FDA or BOX/PI and analyzed by FCM during the malolactic fermentation (MLF). The analysis show a strong correlation between the numbers of BOX/PI-stained cells determined by FCM and the cell numbers determined by plate counts (red wine: R ² ≥ 0.97, white wine R ² ≥ 0.965). On the other hand, we found that the enumeration of O. oeni labeled with FDA was only possible in white wine (R ² ≥ 0.97). Viable yeast and LAB populations can be rapidly discriminated and quantified in simultaneous malolactic-alcoholic wine fermentations using BOX/PI and scatter parameters in a one single measurement. This rapid procedure is therefore a suitable method for monitoring O. oeni populations during winemaking, offers a detection limit of <10⁴ CFU ml^?1 and can be considered a useful method for investigating the dynamics of microbial growth in wine and applied for microbiological quality control in wineries.     

Synthesis of some pyrazolyl benzenesulfonamide derivatives as dual anti-inflammatory antimicrobial agents

Four series of pyrazolylbenzenesulfonamide derivatives were synthesized and evaluated for their anti-inflammatory activity using cotton pellet induced granuloma and carrageenan-induced rat paw edema bioassays. Moreover, COX-1 and COX-2 inhibitory activity, ulcerogenic effect and acute toxiCIT000y were also determined. Furthermore, the target compounds were screened for their in-vitro antimicrobial activity against Eischerichia coli, Staphylococcus aureus and Candida albicans. Compounds 4-(3-Phenyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9a and 4-(3-Tolyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9b were not only found to be the most active dual anti-inflammatory antimicrobial agents in the present study with good safety margin and minimal ulcerogenic effect but also exhibited good selective inhibitory activity towards COX-2. A docking pose for 9a and 9b separately in the active site of the human COX-2 enzyme was also obtained. Therefore, these compounds would represent a fruitful matrix for the development of dual anti-inflammatory antimicrobial candidates with remarkable COX-2 selectivity.     

Genotypic and phenotypic characteristics of tunisian isoniazid-resistant <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis> strains

Forty three isoniazid (INH)-resistant Mycobacterium tuberculosis isolates were characterized on the basis of the most common INH associated mutations, katG315 and mabA −15C→T, and phenotypic properties (i.e. MIC of INH, resistance associated pattern, and catalase activity). Typing for resistance mutations was performed by Multiplex Allele-Specific PCR and sequencing reaction. Mutations at either codon were detected in 67.5% of isolates: katG315 in 37.2, mabA −15C→T in 27.9 and both of them in 2.4%, respectively. katG sequencing showed a G insertion at codon 325 detected in 2 strains and leading to amino acid change T326D which has not been previously reported. Distribution of each mutation, among the investigated strains, showed that katG S315T was associated with multiple-drug profile, high-level INH resistance and loss or decreased catalase activity; whereas the mabA −15C→T was more prevalent in mono-INH resistant isolates, but it was not only associated with a low-level INH resistance. It seems that determination of catalase activity aids in the detection of isolates for which MICs are high and could, in conjunction with molecular methods, provide rapid detection of most clinical INH-resistant strains.     

Antioxidant status and oxidative stress at rest and in response to acute exercise in judokas and sedentary men

It is well recognized that acute strenuous exercise is accompanied by an increase in free-radical production and subsequent oxidative stress, in addition to changes in blood antioxidant status. Chronic exercise provides protection against exercise-induced oxidative stress by upregulating endogenous antioxidant defense systems. Little is known regarding the protective effect afforded by judo exercise. Therefore, we determined antioxidant and oxidative stress biomarkers at rest and in response to acute exercise in 10 competitive judokas and 10 sedentary subjects after mixed exercise (anaerobic followed by aerobic). The subjects performed a Wingate test, followed by 30 minutes of aerobic exercise performed at 60% of maximal aerobic power. Blood samples were taken, by an intravenous catheter, at rest (R), immediately after the physical exercise (P0), and at 5 (P5), 10 (P10), and 20 (P20) minutes postexercise. The measured parameters included the activity of the antioxidant enzymes superoxide dismutase, glutathione peroxidase, and glutathione reductase, in addition to α-tocopherol, and total antioxidant status. Malondialdehyde was measured as a representation of lipid peroxidation. At rest, the judokas had higher values for all antioxidant and oxidative stress markers as compared to the sedentary subjects (p < 0.05). Plasma concentrations of all parameters except for α-tocopherol increased significantly above resting values for both the judokas and sedentary subjects (p < 0.05) and remained elevated at 20 minutes postexercise. A significant postexercise decrease was observed for α-tocopherol (p < 0.05) at P20 for judokas and at P5 for sedentary subjects. These data indicate that competitive judo athletes have higher endogenous antioxidant protection compared to sedentary subjects. However, both groups of subjects experience an increase in exercise-induced oxidative stress that is not different.     

Primary cilium depletion typifies cutaneous melanoma in situ and malignant melanoma

Cutaneous melanoma is a lethal malignancy that arises spontaneously or via in situ precursor neoplasms. While melanoma in situ and locally invasive malignant melanoma can be cured surgically, these lesions can sometimes be difficult to distinguish from melanocytic nevi. Thus, the identification of histolopathologic or molecular features that distinguish these biologically distinct lesions would represent an important advance. To this end, we determined the abundance of melanocytic primary cilia in a series of 62 cases composed of typical cutaneous melanocytic nevi, melanoma in situ, invasive melanoma, and metastatic melanoma. Primary cilia are sensory organelles that modulate developmental and adaptive signaling and notably, are substantially depleted from the neoplastic epithelium of pancreatic carcinoma at a stage equivalent to melanoma in situ. In this series, we find that while nearly all melanocytes in 22 melanocytic nevi possessed a primary cilium, a near-complete loss of this organelle was observed in 16 cases of melanoma in situ, in 16 unequivocal primary invasive melanomas, and in 8 metastatic tumors, each associated with a cutaneous primary lesion. These findings suggest that the primary cilium may be used to segregate cutaneous invasive melanoma and melanoma in situ from melanocytic nevi. Moreover, they place the loss of an organelle known to regulate oncogenic signaling at an early stage of melanoma development.     

Growth establishment and phenology of four woody Saharan species

Henophyton deserti, Calligonum polygonoides, Ephedra alata and Anthyllis henoniana may be suitable Saharan species for the rehabilitation of degraded lands. In this context, the phenology, excavation of root systems and seedling establishment of these species were investigated in the southern Tunisia. Our data suggest that phenological differences between the studied species may depend on the rooting depth. The mean seedling emergence rate was 73% in H. deserti, 66% in A. henoniana, 60% in C. polygonoides and 50% in E. alata. Statistical analyses of seedling establishment showed a highly significant difference among the four species in both seasons (F = 1595.15, P < 0.0001), species (F = 781.57, P < 0.0001) and species/season interaction (F = 61.84, P < 0.0001). After 3 years, seedling mortality ranged from 73% to 89%. Seedling mortality reached a maximum in the summer of the first year of study (P < 0.001). This study provided further understanding of the ecological tolerance, adaptive strategy and the low density of these species in nature. Henophyton deserti appears to be better adapted to the harsh desert environment. These findings explain the very high density observed in the Sahara compared with other studied species. This species has therefore proven to be an excellent candidate for the ecological restoration of degraded desert rangelands and can be introduced successfully in to rehabilitation programmes.     

Investigation of Alteration in the Levels of Iron and Copper in Scalp Hair Samples of Patients Having Different Types of Viral Hepatitis

The aim of this study was to measure the alterations of copper and iron contents in scalp hair samples of hepatitis A–E patients of both genders, same age group, and socioeconomic status. For comparative study, the scalp hair samples of healthy individuals of the same age and socioeconomic status were collected. The concentrations of copper and iron were measured using an atomic absorption spectrophotometer, after microwave-assisted acid digestion. The validity and accuracy of methodology was checked using a certified reference material. The results of this study showed that the mean values of copper and iron were higher in scalp hair samples of hepatitis patients than those of age-matched control subjects, while the difference was significant in the cases of patients having viral hepatitis B, C, and D as compared to those who have viral hepatitis A and E (p?<?0.001). It was concluded that the overload of copper and iron in the human body may cause lipid peroxidation and eventually damage the hepatic system.