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The zebrafish (Danio rerio) is a valuable model organism for studying human biology due to its easy genetic manipulation and small size. It is optically transparent and shares genetic similarities with humans, making it ideal for studying developmental processes, diseases, and drug screening via imaging-based approaches. Solid malignant tumors often contain hypoxic areas that stimulate the release of extracellular vesicles (EVs), lipid-bound structures released by cells into the extracellular space, that facilitate short- and long-range intercellular communication and metastatization. Here we investigate the effects of EVs derived from neuroblastoma (NB), a pediatric solid tumor, on metastatic niche formation using the zebrafish as an in vivo model. Intravascular injection in zebrafish embryos allows a non-invasive visualization of EVs dispersion, uptake, and interactions with host cells. To improve repeatability of our results and ease the injection steps, we used an agarose device replica molded from a custom designed micromilled aluminum mold. We first demonstrated that EVs released under hypoxic conditions promote angiogenesis and are more easily internalized by endothelial cells than those purified from normoxic cells. We also showed that injection of with hypoxic EVs increased macrophages mobilization. We then focused on the caudal hematopoietic tissue (CHT) region of the embryo as a potential metastatic site. After hypoxic EVs injection, we highlighted changes in the expression of mmp-9 and cxcl8b genes. Furthermore, we investigated the ability of NB-derived EVs to prime a metastatic niche by a two-step injection of EVs first, followed by NB cells. Interestingly, we found that embryos injected with hypoxic EVs had more proliferating NB cells than those injected with normoxic EVs. Our findings suggest that EVs released by hypoxic NB cells alter the behavior of recipient cells in the zebrafish embryo and promote metastatic outgrowth. In addition, we demonstrated the ability of the zebrafish embryo to be a suitable model for studying the interactions between EVs and recipient cells in the metastatic process. [ABSTRACT FROM AUTHOR]

Abstract Background The endozepine triakontatetraneuropeptide (TTN) induces intracellular calcium ([Ca++]i) changes followed by activation in human polymorphonuclear leukocytes (PMNs). The present study was undertaken to investigate the role of protein kinase (PK) C in the modulation of the response to TTN by human PMNs, and to examine the pharmacology of TTN-induced Ca++ entry through the plasma membrane of these cells. Results The PKC activator 12-O-tetradecanoylphorbol-13-acetate (PMA) concentration-dependently inhibited TTN-induced [Ca++]i rise, and this effect was reverted by the PKC inhibitors rottlerin (partially) and Ro 32-0432 (completely). PMA also inhibited TTN-induced IL-8 mRNA expression. In the absence of PMA, however, rottlerin (but not Ro 32-0432) per se partially inhibited TTN-induced [Ca++]i rise. The response of [Ca++]i to TTN was also sensitive to mibefradil and flunarizine (T-type Ca++-channel blockers), but not to nifedipine, verapamil (L-type) or ω-conotoxin GVIA (N-type). In agreement with this observation, PCR analysis showed the expression in human PMNs of the mRNA for all the α1 subunits of T-type Ca++ channels (namely, α1G, α1H, and α1I). Conclusions In human PMNs TTN activates PKC-modulated pathways leading to Ca++ entry possibly through T-type Ca++ channels.

Background: The endozepine triakontatetraneuropeptide (TTN) induces intracellular calcium ([Ca++]i) changes followed by activation in human polymorphonuclear leukocytes (PMNs). The present study was undertaken to investigate the role of protein kinase (PK) C in the modulation of the response to TTN by human PMNs, and to examine the pharmacology of TTN-induced Ca++ entry through the plasma membrane of these cells. Results: The PKC activator 12-O-tetradecanoylphorbol-13-acetate (PMA) concentrationdependently inhibited TTN-induced [Ca++]i rise, and this effect was reverted by the PKC inhibitors rottlerin (partially) and Ro 32-0432 (completely). PMA also inhibited TTN-induced IL-8 mRNA expression. In the absence of PMA, however, rottlerin (but not Ro 32-0432) per se partially inhibited TTN-induced [Ca++]i rise. The response of [Ca++]i to TTN was also sensitive to mibefradil and flunarizine (T-type Ca++-channel blockers), but not to nifedipine, verapamil (L-type) or ω-conotoxin GVIA (N-type). In agreement with this observation, PCR analysis showed the expression in human PMNs of the mRNA for all the α1 subunits of T-type Ca++ channels (namely, α1G, α1H, and α1I). Conclusions: In human PMNs TTN activates PKC-modulated pathways leading to Ca++ entry possibly through T-type Ca++ channels. [ABSTRACT FROM AUTHOR]

1. Rifamazine (AF/RP) a dimeric rifamycin, is active against bacterial DNA-dependent RNA polymerase and against viral RNA-dependent DNA polymerase. 2. Rifamazine is active also against DNA-dependent RNA polymerase extracted from rifampicin-resistant mutants of Escherichia coli. It does not interfere with enzyme-template interaction or with RNA elongation. It blocks initiation. 3. A comparison is made between the mechanism of action of rifamazine and that of rifampicin, and of AF/013 (octyloxime of 3-formylrifamycin SV), a C-class rifamycin. Our results show that the mechanism of action of rifamazine is more similar to that of rifampicin than to that of the octyloxime derivative. 4. Activity of rifamazine against RNA polymerase from rifampicin-resistant mutants is thought to be due to binding of the dimer to both the rifamycin-specific binding site and to a second weak site. [ABSTRACT FROM AUTHOR]

The interaction between several antibiotics and either normal human serum or EGTA-chelated Mg-treated serum has been tested. Synergism has been observed with rifampicin, tetracycline and doxycycline, but not with minocycline, aminoglycosides (kanamycin, tobramycin, gentamicin, sisomicin and amikacin), chloramphenicol, fosfomycin and erythromycin. It has been demonstrated that K12 strains bearing plasmids, conferring resistance to tetracycline, were killed in the presence of serum. Evidence has been presented that this synergistic action depends on complement and can be abolished by serum treatment with ethyleneglycol-tetraacetic acid. [ABSTRACT FROM PUBLISHER]

The 'QuantitatEVs: multiscale analyses, from bulk to single vesicle' workshop aimed to discuss quantitative strategies and harmonized wet and computational approaches toward the comprehensive analysis of extracellular vesicles (EVs) from bulk to single vesicle analyses with a special focus on emerging technologies. The workshop covered the key issues in the quantitative analysis of different EV-associated molecular components and EV biophysical features, which are considered the core of EV-associated biomarker discovery and validation for their clinical translation. The in-person-only workshop was held in Trento, Italy, from January 31 st to February 2 nd , 2023, and continued in Milan on February 3 rd with "Next Generation EVs", a satellite event dedicated to early career researchers (ECR). This report summarizes the main topics and outcomes of the workshop., Competing Interests: Conflicts of interest: The authors declare no conflict of interest.

Background and Aim: Fibrosing alveolitis develops in up to 80% of systemic sclerosis patients (SSc) but progression to end stage fibrosis occurs in about 15% of cases. Mechanisms leading to the process remain mostly unknown. We compared cytokine profiles of broncho-alveolar lavage fluids (BAL-f) from patients with SSc associated interstitial lung disease (SSc-ILD) (n. 34), idiopathic pulmonary fibrosis (IPF) (n. 13), stage II sarcoidosis (n. 14) and 9 controls., Methods: Interleukin (IL) 8, monocyte chemoattractant protein 1 (MCP-1), gamma-interferon (IFN-gamma), IL12, IL18 and IL10 and transforming growth factor-beta (TGF-beta) were assessed by ELISA in concentrated BAL-f., Results: Levels of IL8 and MCP-1 were significantly elevated in SSc-ILD and in IPF as compared with controls (Mann Whitney test p < 0.05), while MCP-1 values were significantly lower in SSc-ILD than in IPF. A significant correlation between neutrophils and IL8 levels (p = 0.047), as well as between eosinophils and MCP-1 levels (p = 0.004) was also observed. IFN-gamma levels were slightly higher than normal only in sarcoidosis (p = 0.06), whereas IL12 levels increased both in sarcoidosis and SSc-ILD (p < 0.05). No differences were found in IL18 and TGF-beta levels. Finally, IL10 levels were higher in SSc-ILD and sarcoidosis than in controls and IPF (p < 0.05)., Conclusion: BAL-f cytokine profile differentiates ILD associated with SSc from IPF. The lower expression of MCP-1 and the higher expression of the anti-fibrotic IL12 and the anti-inflammatory IL10, observed both in sarcoidosis and in SSc-ILD, could account for the better prognosis of these ILDs. Further longitudinal studies are required to confirm whether a different cytokine phenotype may be considered predictive of clinical outcome in SSc-ILD.

Background: The aim of our study was to compare clinical and BAL features of patients with bronchiolitis obliterans organizing pneumonia (BOOP) with those of patients with usual interstitial pneumonia (UIP) and control subjects., Patients and Methods: This study reports on 14 patients with idiopathic BOOP. Diagnosis was made upon histology. Lung function tests were mostly normal. Chest X-ray and CT showed always a patchy consolidation, often associated with ground glass pattern. BAL was performed for cytology and for ELISA assessment of several cytokines (IL8, ILI0, IL12, gamma-interferon, IL 18, monocyte chemoattractant protein- 1)., Results: Cytology of BAL in BOOP showed a pattern of lymphocytic alveolitis (Lymphocytes: 0.36 x 10(6)/ml) associated with an increase in neutrophil and eosinophil counts (0.13 and 0.04 x 10(6)/ml respectively). Mean BALf levels in pg/ml of MCP-1, IL12 and IL18 were significantly increased in BOOP with respect to controls and UIP patients, while in UIP patients only a significant increase of IL8, MCP-1 and IL18 with respect to controls was detected. In addition, BALf levels of IL10, an anti-inflammatory cytokine, were significantly higher in BOOP patients with respect to controls and UIP patients., Conclusion: These findings are consistent with a marked degree of macrophage and lymphocyte activation in BOOP with an expansion of T helper-1 response. The concomitant increase of IL10 could be related to a limitation of the inflammatory process and the fibrotic evolution typical of this clinical picture.