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The differentiation of endothelial cells from human iPSC has incontestable advantages in diseases research and therapeutic applications. However, the safe use of iPSC derivatives in regenerative medicine requires an enhanced understanding and control of factors that optimize in vitro reprogramming and differentiation protocols. Shifts in cellular metabolism associated with intracellular pH changes affect the enzymes that control epigenetic configuration, which impact chromatin reorganization and gene expression changes during reprogramming and differentiation. FLIM-based metabolic imaging of NADH and FAD is a powerful tool for measuring mitochondrial metabolic state and widely used diagnostic method for identification of neoplastic diseases, skin diseases, ocular pathologies and stem cells differentiation. Therefore, in this study, we used the potential of FLIM-based metabolic imaging and fluorescence microscopy of NADH and FAD to study the metabolic changes during iPSC differentiation in endothelial cells. The evaluation of the intracellular pH was carried out with the fluorescent pH-sensor SypHer-2 and fluorescence microscopy to obtain complete information about metabolic status of iPSC and their endothelial derivatives. Based on the FAD/NAD(P)H optical redox ratios increase and the contributions rise of the NAD(P)H fluorescence lifetime in iPSC during endothelial differentiation, we demonstrated an contribution increase of OXPHOS to cellular metabolism. Based on the shift toward more acidic intracellular pH in endothelial cell derived from iPSCs we verified their oxidative state.

Biomeshes based on decellularized bovine pericardium (DBP) are widely used in reconstructive surgery due to their wide availability and the attractive biomechanical properties. However, their efficacy in clinical applications is often affected by the uncontrolled immunogenicity and proteolytic degradation. To address this issue, we present here in vivo multiparametric imaging analysis of epoxy crosslinked DBPs to reveal their fate after implantation. We first analyzed the structure of the crosslinked DBP using scanning electron microscopy and evaluated proteolytic stability and cytotoxicity. Next, using combination of fluorescence and hypoxia imaging, X-ray computed microtomography and histology techniques we studied the fate of DBPs after subcutaneous implantation in animals. Our approach revealed high resistance to biodegradation, gradual remodeling of a surrounding tissue forming the connective tissue capsule and calcification of crosslinked DBPs. These changes were concomitant to the development of hypoxia in the samples within 3 weeks after implantation and subsequent induction of angiogenesis and vascularization. Collectively, presented approach provides new insights on the transplantation of the epoxy crosslinked biomeshes, the risks associated with its applications in soft-tissue reconstruction and can be transferred to studies of other types of implants.

The paper briefly describes the concept of scanning transmission X-ray microscopy (STXM) and the Nanoscope station, which is planned to be installed on the 4th generation SKIF synchrotron. The microscope is designed to study the internal structure of cells and dynamic processes in them with nanometer spatial resolution. The microscope uses a unique absorption contrast between proteins and water, in the region of 15, in the spectral range of the "water window", λ=2.3 - 4.3 nm, which eliminates the need for contrasting and fluorophores, and minimizes the doses absorbed in the samples necessary for obtaining high-quality 3D images. The article presents an X-ray optical diagram of a microscope, its main technical characteristics and an estimate of absorbed doses depending on the resolution.

Stressed InGaAs/GaAs quantum-well laser structures are grown by gas-phase epitaxy from organometallic compounds on GaAs substrates and artificial Ge/Si substrates based on Si(001) with an epitaxial metamorphic Ge layer. To suppress the relaxation of elastic stresses during the growth of InGaAs quantum wells with a high fraction of In, strain-compensating GaAsP layers are applied. The structural and radiative properties of the samples grown on substrates of various types are compared. Stimulated radiation at wavelengths up to 1.24 μm at 300 K is obtained for structures grown on GaAs substrates and at wavelengths up to 1.1 μm at 77 K, for structures grown on Ge/Si substrates.

According to recent reports, shell morphology is unreliable for the identification of oysters because of the high phenotypic plasticity of these bivalves. Using COI DNA barcoding and sperm morphology, we reinvestigated the species validity of wild Pacific oyster Crassostrea gigas habituating the Peter the Great Bay (Sea of Japan). DNA barcoding confirmed the species validity of samples collected. Application of the single sperm pattern was not possible for species identification due to pronounced sperm plasticity being found. Six sperm morphs were discovered in the testes of each oyster collected. The amount of abundant sperm morphs and the type of the most dominant sperm pattern are particular to geographical localities that are individual depending on the environmental factors. Ecological monitoring of marine areas and commercially assigned intraspecific geo-authentification of the Pacific oyster seems possible based on the analysis of this species’ heterogenic sperm. Further work will be needed to test if sperm heterogeneity exists in other Ostreidae species and if heterogenic sperms could be used for interspecific analysis.

Glutaredoxin 2 (GRX2), a mitochondrial glutathione-dependent oxidoreductase, is central to glutathione homeostasis and mitochondrial redox, which is crucial in highly metabolic tissues like the heart. Previous research showed that absence of Grx2, leads to impaired mitochondrial complex I function, hypertension and cardiac hypertrophy in mice but the impact on mitochondrial structure and function in intact cardiomyocytes and in humans has not been explored. We hypothesized that Grx2 controls cardiac mitochondrial dynamics and function in cellular and mouse models, and that low expression is associated with human cardiac dysfunction. Here we show that Grx2 absence impairs mitochondrial fusion, ultrastructure and energetics in primary cardiomyocytes and cardiac tissue. Moreover, provision of the glutathione precursor, N-acetylcysteine (NAC) to Grx2-/- mice did not restore glutathione redox or prevent impairments. Using genetic and histopathological data from the human Genotype-Tissue Expression consortium we demonstrate that low GRX2 is associated with fibrosis, hypertrophy, and infarct in the left ventricle. Altogether, GRX2 is important in the control of cardiac mitochondrial structure and function, and protects against human cardiac pathologies., Graphical abstract fx1

An expression of calpain and caspase-1 as well as the concomitant ultrastructural alterations were investigated during necrosis of the mouse Ehrlich ascites carcinoma. The calpain expression was registered at 0 h and 1 h although caspase-1 did not induce any signals during these time periods. The rise of the cytoplasmic lytic zones contacted by calpain antibodies was identified as a morphologic event corresponding to the expression of calpain. Lytic zone's distribution followed by the appearance of the calpain/caspase-1 clusters assigned for lysis of the Golgi vesicles and ER. Also, the microapocrine secretion of the vesicles containing the calpain/caspase-1 clusters was detected. Further, the lysis of the plasma membrane occurred due to progression of intracellular lysis. Rupture of the plasma membrane resulted in the termination of secretion and dissemination of cell contents. The nuclei still had their normal shape. Nuclear lysis continued to rise with intranuclear lytic zones, of which the progression was accompanied with the presence of calpain/caspase-1 clusters. The data contribute to the concept of the initial role of calpain for tumor cell destruction, provide first evidence of the calpain/caspase-1 pathway in tumor cells, and highlight microapocrine secretion as a possible tumor cell death signalling mechanism.

Conventional techniques are insufficient precisely to describe the internal structure, the heterogeneous cell populations, and the dynamics of biological processes occurring in diseased liver during surgery. There is a need for a rapid and safe method for the successful diagnosis of liver disease in order to plan surgery and to help avoid postoperative liver failure. We analyze the progression of both acute (cholestasis) and chronic (fibrosis) liver pathology using multiphoton microscopy with fluorescence lifetime imaging and second-harmonic generation modes combined with time-of-flight secondary ion mass spectrometry chemical analysis to obtain new data about pathological changes to hepatocytes at the cellular and molecular levels. All of these techniques allow the study of cellular metabolism, lipid composition, and collagen structure without staining the biological materials or the incorporation of fluorescent or other markers, enabling the use of these methods in a clinical situation. The combination of multiphoton microscopy and mass spectrometry provides more complete information about the liver structure and function than could be assessed using either method individually. The data can be used both to obtain new criteria for the identification of hepatic pathology and to develop a rapid technique for liver quality analysis in order to plan surgery and to help avoid postoperative liver failure in clinic.

The phenomenon of the cytoplasmic localisation of mitochondrial ribosomal subunits (12 S mitochondrial rRNA and 16 S mitochondrial rRNA) has been discovered by scientific teams working with spermatogenic cells of mice. Previous reports showed that the release of mitochondrial substance occurs during interaction of mitochondria with the germ plasm granules (GG). To determine if the interplay between the vasa-positive GG and the mitochondria is associated with cytoplasmic localisation of mtrRNAs, we studied the spermatogenic cells of zebrafish, Danio rerio. It was revealed that in type A undifferentiated spermatogonia the GG did not contact mitochondria, and the extra-mitochondrial localisation of the mtrRNAs was not found. In type A differentiated spermatogonia, the amount of GG in contact with mitochondria increased, but the extra-mitochondrial localisation of the mtrRNAs was not found either. In type B late spermatogonia, which are pre-meiotic cells, the GG/mitochondrion complexes were typically found in contact with the nucleus. This stage was associated with the intra-mitochondrial localisation of GG-originated vasa and extra-mitochondrial localisation of 12 S mtrRNA and 16 S mtrRNA. Until the onset of meiosis, which was determined by the observation of synaptonemal complexes in zygotene-pachytene spermatocytes I, the GG/mitochondrion complexes disappeared, but both types of mtrRNAs persisted in the cytoplasm of spermatids and spermatozoa.

It was shown that tobacco leaf treatment with 100 mM H₂0₂ increased their content ofendogenous H₂0₂ and activities of catalase and hydrolases (acid phosphatase, proteases, and RNase) and also caused'various chang- es in the cell structure. In this case, programmed cell death (PCD) occurred in some cells, which was ob- served as chromatin condensation, cytoplasm collapse, etc. In the meantime, many cells displayed organelle activation rather than PCD. It is suggested that cells that undergo H₂0₂-dependent PCD release signaling molecules inducing protective mechanisms against oxidative stress in neighboring cells not exhibiting PCD.

It was shown that tobacco leaf treatment with 100 mM H₂0₂ increased their content ofendogenous H₂0₂ and activities of catalase and hydrolases (acid phosphatase, proteases, and RNase) and also caused'various chang- es in the cell structure. In this case, programmed cell death (PCD) occurred in some cells, which was ob- served as chromatin condensation, cytoplasm collapse, etc. In the meantime, many cells displayed organelle activation rather than PCD. It is suggested that cells that undergo H₂0₂-dependent PCD release signaling molecules inducing protective mechanisms against oxidative stress in neighboring cells not exhibiting PCD.

Current peroxisome proliferator-activated receptor (PPAR)-targeted drugs, such as the PPARγ-directed diabetes drug rosiglitazone, are associated with undesirable side effects due to robust agonist activity in non-target tissues. To find new PPAR ligands with fewer toxic effects, we generated transgenic zebrafish that can be screened in high throughput for new tissue-selective PPAR partial agonists. A structural analog of coenzyme Q10 (idebenone) that elicits spatially restricted partial agonist activity for both PPARα and PPARγ was identified. Coenzyme Q10 was also found to bind and activate both PPARs in a similar fashion, suggesting an endogenous role in relaying the states of mitochondria, peroxisomes and cellular redox to the two receptors. Testing idebenone in a mouse model of type 2 diabetes revealed the ability to reverse fatty liver development. These findings indicate new mechanisms of action for both PPARα and PPARγ, and new potential treatment options for nonalcoholic fatty liver disease (NAFLD) and steatosis. This article has an associated First Person interview with the first author of the paper., Summary: A zebrafish screen identifies a novel PPARα/γ ligand, idebenone, with potential for treatment of fatty liver diseases, as seen by testing it in a mouse model of type 2 diabetes.

Transmission electron microscopy of the spermatozoa of five species from three families of bivalves has shown that each species has a sperm with unique morphology. However, the morphology of the acrosomes of each species is typical of the subclass of bivalve to which they belong. An examination of spermatogenesis in the five species, along with a re-examination of material from six other species of bivalves, has revealed that pre-spermiogenic cells possess flagella. In addition, acrosome formation begins in the spermatocytes with the formation of proacrosomal vesicles in the Golgi body. During spermiogenesis the proacrosomal vesicles coalesce at the presumptive posterior of the spermatid, with a larger vesicle produced by the Golgi body. The single acrosomal vesicle eventually migrates to the anterior of the spermatid where it assumes its mature form. © 1994 Wiley-Liss, Inc.