Magnetic Fe3O4 nanoparticles (MNPs) are often used to design agents enhancing contrast in magnetic resonance imaging (MRI) that can be considered as one of the efficient methods for cancer diagnostics. At present, increasing the specificity of the MRI contrast agent accumulation in tumor tissues remains an open question and attracts the attention of a wide range of researchers. One of the modern methods for enhancing the efficiency of contrast agents is the use of molecules for tumor acidic microenvironment targeting, for example, pH-low insertion peptide (pHLIP). We designed novel organosilicon MNPs covered with poly­(ethylene glycol) (PEG) and covalently modified by pHLIP. To study the specific features of the binding of pHLIP-modified MNPs to cells, we also obtained nanoconjugates with Cy5 fluorescent dye embedded in the SiO2 shell. The nanoconjugates obtained were characterized by transmission electron microscopy (TEM), attenuated total reflection (ATR), diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS), dynamic light scattering (DLS), UV and fluorescence spectrometry, thermogravimetric analysis (TGA), CHN elemental analyses, and vibrating sample magnetometry. Low cytotoxicity and high specificity of cellular uptake of pHLIP-modified MNPs at pH 6.4 versus 7.4 (up to 23-fold) were demonstrated in vitro. The dynamics of the nanoconjugate accumulation in the 4T1 breast cancer orthotopically grown in BALB/c mice and MDA-MB231 xenografts was evaluated in MRI experiments. Biodistribution and biocompatibility studies of the obtained nanoconjugate showed no pathological change in organs and in the blood biochemical parameters of mice after MNP administration. A high accumulation rate of pHLIP-modified MNPs in tumor compared with PEGylated MNPs after their intravenous administration was demonstrated. Thus, we propose a promising approach to design an MRI agent with the tumor acidic microenvironment targeting ability.

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Hsp67Bc in Drosophila melanogaster is a member of the small heat shock protein family, the main function of which is to prevent the aggregation of misfolded or damaged proteins. Hsp67Bc interacts with Starvin and Hsp23, which are known to be a part of the cold-stress response in the fly during the recovery phase. In this study, we investigated the role of the Hsp67Bc gene in the cold-stress response. We showed that in adult Drosophila, Hsp67Bc expression increases after cold stress and decreases after 1.5 h of recovery, indicating the involvement of Hsp67Bc in short-term stress recovery. We also implemented a deletion in the D. melanogaster Hsp67Bc gene using imprecise excision of a P-element and analyzed the cold tolerance of Hsp67Bc-null mutants at different developmental stages. We found that Hsp67Bc-null homozygous flies are viable and fertile but display varying cold-stress tolerance throughout the stages of ontogenesis: the survival after cold stress is slightly impaired in late 3rd instar larvae, unaffected in pupae, and notably affected in adult females. Moreover, the recovery from chill coma is delayed in Hsp67Bc-null adults of both sexes. In addition, the deletion in the Hsp67Bc gene caused more prominent up-regulation of Hsp70 following cold stress, suggesting the involvement of Hsp70 in compensation of the lack of the Hsp67Bc protein. Taken together, our results suggest that Hsp67Bc is involved in the recovery of flies from a comatose state and contributes to the protection of the fruit fly from cold stress.

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Hsp67Bc is a small heat shock protein found in Drosophila melanogaster. Apart from performing a function (common for all small heat shock proteins) of preventing aggregation of misfolded proteins, it is involved in macroautophagy regulation alongside the Starvin protein. Overexpression of the D. melanogaster Hsp67Bc gene has been shown to stimulate macroautophagy in S2 cell culture. Nonetheless, it has been unknown how the absence of the Hsp67Bc gene may affect it. Here, we studied the effect of Hsp67Bc gene deletion on the macroautophagy induced by the pathogenic Wolbachia wMelPop strain in D. melanogaster. We detected Wolbachia inside autophagic vacuoles in fly neurons, thereby proving that these endosymbionts were being eliminated via macroautophagy. Nevertheless, we did not register any difference in brain bacterial load between Hsp67Bc-null and control flies at all tested stages of ontogenesis. Moreover, the abundance of autophagic vacuoles was similar between neurons of the mutant and control flies, yet the cross-sectional area of autolysosomes on ultrathin sections was more than 1.5-fold larger in Hsp67Bc-null fly brains than in the control line. Our findings suggest that the product of the Hsp67Bc gene does not participate in the initiation of endosymbiont-induced macroautophagy but may mediate autophagosome maturation: the deletion of the Hsp67Bc gene leads to the increase in autolysosome size.

Drosophila melanogaster Hsp67Bc is a heat- and cold-inducible small heat shock protein that participates in the prevention of aggregation of misfolded proteins and in macroautophagy regulation. Overexpression of the Hsp67Bc gene has been shown to enhance macroautophagy in Drosophila S2 cells, and the deletion of this gene leads to the formation of a slightly increased number of autophagic vacuoles in the fruit f ly brain neurons. Recently, we found that Hsp67Bc-null D. melanogaster f lies have poor tolerance to cold stress (0 °C) of various durations. In the present work, we investigated how the Hsp67Bc gene deletion affects the f itness of fruit f lies under normal conditions and their tolerance to elevated temperatures at different developmental stages. Larvae and pupae were not adversely affected by the Hsp67Bc gene deletion, and adult Hsp67Bc-null f lies showed an extended lifespan in comparison with the control at normal (24–25 °C) and elevated temperature (29 °C), and after acute heat stress (37 °C, 2 h). At the same time, the fecundity of the mutant females was lower by 6–13 % in all tested environments, except for permanent maintenance at 29 °C, where the mean numbers of eggs laid by the mutant and control f lies were equal. We explain this phenomenon by a reduced number of ovarioles in Hsp67Bc-null females and enhanced macroautophagy in their germaria, which promotes the death of forming egg chambers. In addition, short heat stress (37 °C, 2 h), which increased the control line’s longevity (an effect common for a wide range of organisms), had a negative impact on the lifespan of Hsp67Bc-null f lies. Therefore, Hsp67Bc-null D. melanogaster have an extended lifespan under normal and elevated temperature conditions, and reduced fecundity and thermal stress tolerance.