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MitoBlue is a fluorescent bisamidine that can be used to easily monitor the changes in mitochondrial degradation processes in different cells and cellular conditions. MitoBlue staining pattern is exceptional among mitochondrial dyes and recombinant fluorescent probes, allowing the dynamic study of mitochondrial recycling in a variety of situations in living cells. MitoBlue is a unique tool for the study of these processes that will allow the detailed characterization of communication between mitochondria and lysosomes

[Abstract] Aiming to address a stable chondrogenesis derived from mesenchymal stromal cells (MSCs) to be applied in cartilage repair strategies at the onset of osteoarthritis (OA), we analyzed the effect of arginine–glycine–aspartate (RGD) density on cell condensation that occurs during the initial phase of chondrogenesis. For this, we seeded MSC-derived from OA and healthy (H) donors in RGD-dendrimer-poly(L-lactic) acid (PLLA) nanopatterned substrates (RGD concentrations of 4 × 10−9, 10−8, 2.5 × 10−8, and 10−2 w/w), during three days and compared to a cell pellet conventional three-dimensional culture system. Molecular gene expression (collagens type-I and II–COL1A1 and COL2A1, tenascin-TNC, sex determining region Y-box9-SOX9, and gap junction protein alpha 1–GJA1) was determined as well as the cell aggregates and pellet size, collagen type-II and connexin 43 proteins synthesis. This study showed that RGD-tailored first generation dendrimer (RGD-Cys-D1) PLLA nanopatterned substrates supported the formation of pre-chondrogenic condensates from OA- and H-derived human bone marrow-MSCs with enhanced chondrogenesis regarding the cell pellet conventional system (presence of collagen type-II and connexin 43, both at the gene and protein level). A RGD-density dependent trend was observed for aggregates size, in concordance with previous studies. Moreover, the nanopatterns’ had a higher effect on OA-derived MSC morphology, leading to the formation of bigger and more compact aggregates with improved expression of early chondrogenic markers.

Allergic reactions towards b-lactam antibiotics pose an important clinical problem. The ability of small molecules, such as a b-lactams, to bind covalently to proteins, in a process known as haptenation, is considered necessary for induction of a specific immunological response. Identification of the proteins modified by b-lactams and elucidation of the relevance of this process in allergic reactions requires sensitive tools. Here we describe the preparation and characterization of a biotinylated amoxicillin analog (AX-B) as a tool for the study of protein haptenation by amoxicillin (AX). AX-B, obtained by the inclusion of a biotin moiety at the lateral chain of AX, showed a chemical reactivity identical to AX. Covalent modification of proteins by AX-B was reduced by excess AX and vice versa, suggesting competition for binding to the same targets. From an immunological point of view, AX and AX-B behaved similarly in RAST inhibition studies with sera of patients with non-selective allergy towards b-lactams, whereas, as expected, competition by AX-B was poorer with sera of AX-selective patients, which recognize AX lateral chain. Use of AX-B followed by biotin detection allowed the observation of human serum albumin (HSA) modification by concentrations 100-fold lower that when using AX followed by immunological detection. Incubation of human serum with AX-B led to the haptenation of all of the previously identified major AX targets.In addition, some new targets could be detected. Interestingly, AX-B allowed the detection of intracellular protein adducts, which showed a cell type-specific pattern. This opens the possibility of following the formation and fate of AX-B adducts in cells. Thus, AX-B may constitute a valuable tool for the identification of AX targets with high sensitivity as well as for the elucidation of the mechanisms involved in allergy towards b-lactams.