Variation in tumor pH affects pH-triggered delivery of peptide-modified magnetic nanoparticles.

Acidification of the extracellular matrix, an intrinsic characteristic of many solid tumors, is widely exploited for physiologically triggered delivery of contrast agents, drugs, and nanoparticles to tumor. However, pH of tumor microenvironment shows intra- and inter-tumor variation. Herein, we investigate the impact of this variation on pH-triggered delivery of magnetic nanoparticles (MNPs) modified with pH-(low)-insertion peptide (pHLIP). Fluorescent flow cytometry, laser confocal scanning microscopy and transmission electron microscopy data proved that pHLIP-conjugated MNPs interacted with 4T1 cells in two-dimensional culture and in spheroids more effectively at pH 6.4 than at pH 7.2, and entered the cell via clathrin-independent endocytosis. The accumulation efficiency of pHLIP-conjugated MNPs in 4T1 tumors after their intravenous injection, monitored in vivo by magnetic resonance imaging, showed variation. Analysis of the tumor pH profiles recorded with implementation of original nanoprobe pH sensor, revealed obvious correlation between pH measured in the tumor with the amount of accumulated MNPs.

Morphological and molecular characteristics of "difficult" asthma.

BACKGROUND: There are several clinical variants of severe difficult-to-treat asthma: asthma with persistent airflow limitation, brittle asthma, and fatal asthma; but the differences between the pathogenetic mechanisms underlying the disease heterogeneity are unknown. OBJECTIVES: The aim was to evaluate the morphological and molecular characteristics of brittle asthma type I and asthma with persistent airflow limitation compared to mild-to-moderate asthma, by the analysis of the cellular structure and gene expression in the bronchial mucosa. METHODS: Bronchoscopic evaluation was performed in 42 asthmatic patients: 10 with brittle asthma, 10 with severe asthma with persistent airflow limitation, and 22 with mild-to-moderate asthma. Morphometric and cytological analyses of the bronchial mucosa were performed. The mRNA levels for the ADRB2, HRH1, and CHRM3 genes in the bronchial mucosa were measured by quantitative real-time polymerase chain reaction (PCR). RESULTS: A predominance of eosinophils (29.48/mm(2), 95% confidence interval [CI] 25.24-33.72) and neutrophils (40.13/mm(2), 95% CI 32.77-47.49) was observed in patients with mild-to-moderate asthma; however, histiocytes-macrophages (65.80/mm(2), 95% CI 56.95-74.65) and lymphocytes (52.94/mm(2), 95% CI 42.83-63.06) were more common in patients with brittle asthma, and neutrophil counts (81.11/mm(2), 95% CI 58.33-103.89) were significantly increased in subjects with persistent airflow limitation. An increase in the expression of the M(3)-cholinoreceptor and the beta(2)-adrenoreceptor genes was demonstrated in severe asthmatics compared to mild-to-moderate asthma patients. Significantly higher levels of CHRM3 (57.17%, 95% CI 55.04-59.29) and HRH1 (38.82%, 95% CI 35.84-41.81) mRNAs were observed in patients with brittle asthma. The level of ADRB2 gene expression (71.41%, 95% CI 63.54-85.09) was maximal in patients with asthma with persistent airflow limitation. CONCLUSIONS: There is evidence of significantly different morphological characteristics and molecular mechanisms of inflammation and bronchoconstriction underlying the clinical heterogeneity of severe asthma.

Calcium Phosphate Coating Prepared by Microarc Oxidation Affects hTERT Expression, Molecular Presentation, and Cytokine Secretion in Tumor-Derived Jurkat T Cells.

Calcium phosphate (CaP) materials are among the best bone graft substitutes, but their use in the repair of damaged bone in tumor patients is still unclear. The human Jurkat T lymphoblast leukemia-derived cell line (Jurkat T cells) was exposed in vitro to a titanium (Ti) substrate (10 × 10 × 1 mm3) with a bilateral rough (average roughness index (Ra) = 2-5 µm) CaP coating applied via the microarc oxidation (MAO) technique, and the morphofunctional response of the cells was studied. Scanning electron microscopy (SEM), X-ray diffraction (XRD), and energy dispersive X-ray spectroscope (EDX) analyses showed voltage-dependent (150-300 V) growth of structural (Ra index, mass, and thickness) and morphological surface and volume elements, a low Ca/PaT ratio (0.3-0.6), and the appearance of crystalline phases of CaHPO4 (monetite) and ß-Ca2P2O7 (calcium pyrophosphate). Cell and molecular reactions in 2-day and 14-day cultures differed strongly and correlated with the Ra values. There was significant upregulation of hTERT expression (1.7-fold), IL-17 secretion, the presentation of the activation antigens CD25 (by 2.7%) and CD95 (by 5.15%) on CD4+ cells, and 1.5-2-fold increased cell apoptosis and necrosis after two days of culture. Hyperactivation-dependent death of CD4+ cells triggered by the surface roughness of the CaP coating was proposed. Conversely, a 3.2-fold downregulation in hTERT expression increased the percentages of CD4+ cells and their CD95+ subset (by 15.5% and 22.9%, respectively) and inhibited the secretion of 17 of 27 test cytokines/chemokines without a reduction in Jurkat T cell survival after 14 days of coculture. Thereafter, cell hypoergy and the selection of an hTERT-independent viable CD4+ subset of tumor cells were proposed. The possible role of negative zeta potentials and Ca2+ as effectors of CaP roughness was discussed. The continuous (2-14 days) 1.5-6-fold reductions in the secretion of vascular endothelial growth factor (VEGF) by tumor cells correlated with the Ra values of microarc CaP-coated Ti substrates seems to limit surgical stress-induced metastasis of lymphoid malignancies.

Carbonyl stress phenomena during chronic infection with Opisthorchis felineus.

Infection with the fish borne liver fluke Opisthorchis felineus is common in the Eastern Europe (Ukraine, European part of Russia), Northern Asia (Siberia) and Central Asia (Northern Kazakhstan). Better understanding of the molecular pathogenesis of the biliary tract and liver during chronic opisthorchiasis can be expected to improve protection against and management of complications of this disease. We hypothesize that infection with O. felineus associates with formation of methylglyoxal and carbonyl stress in the liver and hence here we investigated the glyoxalase system and the receptor for advanced glycated end products (RAGE) in the liver of hamsters infected with this liver fluke. Expression of mRNA encoding glyoxalase 1 decreased at 8weeks of the infection and catalytic activity as well decreased at 8 and 12weeks after infection, and the expression of the glyoxalase 2 decreased until 36week post-infection, which associated with the decreasing activity of the enzyme at 8 and 12weeks post-infection. Glutathione levels in infected livers had decreased at week 8, whereas up-regulation of RAGE at mRNA levels was seen for the extended duration of the experimental infection of the hamsters. This outcome supported the notion of hepatic dicarbonyl stress during chronic opisthorchiasis. The inhibition of the glyoxalase system and accumulation of methylglyoxal at the early stages of the infection may underpin development of insulin resistance during opisthorchiasis.