Urinary tract infections are among the most common bacterial infections in humans. Moreover, they are highly recurrent and increasingly often resistant to antibiotics. The antimicrobial properties of the amniotic membrane (AM), the innermost layer of fetal membranes, have been briefly reported in the literature, however, the results of published studies are often inconsistent and unclear; moreover, its effect on uropathogenic bacteria has not yet been investigated. Further, there is no data in the literature about the effect of AM preparation and storage on its antimicrobial properties. To examine the impact of several preparation procedures on the antimicrobial properties of AM, we prepared patches and homogenates of fresh (fAM) and cryopreserved (cAM) human AM and tested them on 14 selected Gram-positive and Gram-negative uropathogenic bacteria. By employing novel antimicrobial efficiency assays we showed that fAM and cAM homogenates have broad-spectrum antimicrobial activity against all here tested uropathogenic bacteria, except for Serratia marcescens. Moreover, they had a potent effect also on the multiple-resistant clinical strains of uropathogenic Escherichia coli. Interestingly, the patches of fAM and cAM had no antimicrobial effect on any of the tested strains. We therefore prepared and stored AM patches according to the standard procedure for clinical use in ophthalmology, which includes the cryopreservation of antibiotic-treated AM, and performed antimicrobial...
COBISS.SI-ID: 34747609
Tunneling membrane nanotubes (TnTs) are membrane protrusions connecting nearby or distant cells in vitro and in vivo. Functions of TnTs in cellular processes are various and rely on TnT structure, which also depends on cytoskeletal composition. In the present study, we focused on the organization of microtubules (MTs) and intermediate filaments (IFs) in TnTs of urothelial cells. We analysed TnTs of normal porcine urothelial cells, which morphologically and physiologically closely resemble normal human urothelial cells, and of cancer cells derived from invasive human urothelial neoplasm. Wide-field fluorescence, confocal and super-resolution microscopy techniques, together with image analyses and 3D reconstructions enlightened specific MT-IF organization in TnTs, and for the first time revealed that MTs and IFs co-occur in the majority of normal and cancer urothelial cell TnTs. Our findings show that in the initiation segment of TnTs, MTs are cross-linked with each other into filamentous network, however in the middle and the attaching segment of TnT, MTs can helically enwrap IFs, the phenomenon that has not been shown before within the TnTs. In this study, we assess MT-IF co-occurrence in TnTs and present evidence that such helical organization of MTs enwrapping IFs is only occurring in a minority of the TnTs. We also discuss the possible cell-biological and physiological reasons for helical organization of MTs in TnTs.
COBISS.SI-ID: 34052057
The apical surface of the terminally differentiated mammalian urothelial um-brella cell is mechanically stable and highly impermeable, in part due to its coverage by urothelial plaques consisting of 2D crystals of uroplakin particles. The mechanism for regulating the uroplakin/plaque level is unclear. We found that genetic ablation of the highly tissue-specific sorting nexin Snx31, which localizes to plaques lining the multivesicular bodies (MVBs) in urothelial umbrella cells, abolishes MVBs suggesting that Snx31 plays a role in stabilizing the MVB-associated plaques by allowing them to achieve a greater curvature. Strikingly, Snx31 ablation also induces a massive accumulation of uroplakin-containing mito-chondria-derived lipid droplets (LDs), which mediate uroplakin degradation via autophagy/lipophagy, leading to the loss of apical and fusiform vesicle plaques. These results suggest that MVBs play an active role in suppressing the excessive/wasteful endocytic degradation of uroplakins. Failure of this suppression mechanism triggers the formation of mitochondrial LDs so that excessive uroplakin membranes can be sequestered and degraded. Because mitochondrial LD formation, which occurs at a low level in normal urothelium, can also be induced by disturbance in uroplakin polymerization due to individual uroplakin knockout and by arsenite, a bladder carcinogen, this pathway may represent an inducible, versatile urothelial detoxification mechanism.
COBISS.SI-ID: 15006211
Induced desquamation of urinary bladder epithelial cells, also called urothelial cells, is frequently used in studies of bladder epithelial regeneration and also in treating recurrent bacterial cystitis. Positively charged polymer chitosan is known to cause large-scale desquamation of terminally differentiated urothelial cells called umbrella cells. Aiming to compare the desquamation ability of another polycation poly-L-lysine, we studied the effect of this polymer on the functional and structural integrity of the urothelium in ex vivo and in vivo experiments. The urothelium was analyzed by measuring transepithelial electrical resistance, and the structural changes of its luminal surface were analyzed with scanning electron microscopy. The results revealed a selective and concentration-dependent desquamation effect of poly-L-lysine on superficial urothelial cells followed by quick regeneration of the urothelium, which functionally and structurally recovers in 2 to 3 h after poly-L-lysine-induced injury. Poly-L-lysine was thus proven to be a promising polymer to be used when desquamation of urothelial cells is required in basic and potentially clinical studies.
COBISS.SI-ID: 4789617
Desmosomal cadherins, desmocollins, and desmogleins are cholesterol-dependent entities responsible for the stable adhesion of desmosomes in epithelial cells. Here, we investigated the influence of cellular cholesterol depletion on the dynamic properties of the desmosomal cadherin desmocollin, particularly the lateral mobility and distribution of desmocollin 2 (Dsc2-YFP) in the plasma membrane, and how these properties influence the adhesion strength of desmosomes. Depletion of cellular cholesterol decreased the lateral mobility of Dsc2-YFP and caused dispersion of Dsc2-YFP in the plasma membrane of epithelial MDCK cells. As a consequence of the altered Dsc2-YFP dynamics, the adhesive strength of desmosomes was weakened. Moreover, our study is the first to show and quantify the co-association of desmosomes with cholesterol/ sphingomyelin-enriched membrane domains at the ultrastructural level. Taken together, our data emphasize a critical role for the cellular cholesterol content in regulating the lateral mobility and distribution of Dsc2 and show that cholesterol depletion reduces the strength of desmosomal adhesions.
COBISS.SI-ID: 34355417