The role of early diagnosis of emphysematous cystitis: A case report and literature review.

Emphysematous cystitis (EC) is a rare entity caused by bacteria, which produce gas filled cysts in the bladder wall. We present a case of EC in a 72-year-old woman admitted to Vascular Surgery Department because of diabetic foot syndrome. During the hospital stay, the patient's general condition deteriorated. CT established EC diagnosis. Surgical treatment was inevitable. Salvage cystectomy was performed. Despite macroscopic removal of necrotic tissues, the condition of the patient didn't improve, 75 days past diagnosis of EC she died due to the multi-organ failure. Prompt diagnosis provided by imaging plays a key role in the treatment of EC.

The development of marine biomaterial derived from decellularized squid mantle for potential application as tissue engineered urinary conduit.

Tissue engineering is focusing research effort on search for new biomaterials that might be applied to create artificial urinary conduit. Nevertheless, the demanding biomechanical characteristics necessary for proper conduit function is difficult to be replicated. In this study, we are introducing novel marine biomaterial obtained by decellularization of squid mantle derived from Loligo vulgaris. Squid mantles underwent decellularization according to developed dynamic flow two-staged procedure. Efficacy of the method was confirmed by computational dynamic flow analysis. Subsequently Decellularized Squid Mantle (DSM) underwent extensive histological analysis and mechanical evaluation. Based on gained biomechanical data the computational modelling using finite element method was utilized to simulate behavior of DSM used as a urinary conduit. Taking into account potential application in reconstructive urology, the DSM was then evaluated as a scaffold for urothelial and smooth muscle cells derived from porcine urinary bladder. Conducted analysis showed that DSM created favorable environment for cells growth. In addition, due to polarized structure and natural external polysaccharide layer, it protected seeded cells from urine.

Development of a conductive biocomposite combining graphene and amniotic membrane for replacement of the neuronal network of tissue-engineered urinary bladder.

Tissue engineering allows to combine biomaterials and seeded cells to experimentally replace urinary bladder wall. The normal bladder wall however, includes branched neuronal network propagating signals which regulate urine storage and voiding. In this study we introduced a novel biocomposite built from amniotic membrane (Am) and graphene which created interface between cells and external stimuli replacing neuronal network. Graphene layers were transferred without modifying Am surface. Applied method allowed to preserve the unique bioactive characteristic of Am. Tissue engineered constructs composed from biocomposite seeded with smooth muscle cells (SMC) derived from porcine detrusor and porcine urothelial cells (UC) were used to evaluate properties of developed biomaterial. The presence of graphene layer significantly increased electrical conductivity of biocomposite. UCs and SMCs showed an organized growth pattern on graphene covered surfaces. Electrical filed stimulation (EFS) applied in vitro led additionally to increased SMCs growth and linear arrangement. 3D printed chamber equipped with 3D printed graphene based electrodes was fabricated to deliver EFS and record pressure changes caused by contracting SMCs seeded biocomposite. Observed contractile response indicated on effective SMCs stimulation mediated by graphene layer which constituted efficient cell to biomaterial interface.

Sonography in the instability of the long head of the biceps tendon confronted with histopathologic and arthroscopic findings.

BACKGROUND: Disorders of the long head of the biceps (LHB) tendon are a common source of shoulder pain and disability. This tendon can be well visualised using ultrasonography; however, little is known if such examination allows clinicians to predict pathological changes of the tendon structure. In the study described below, we compare preoperative sonographic findings with the data from shoulder arthroscopy and microscopic examination of the excised tendon fragments in 19 consecutive patients with LHB tendinopathy and clinical suspicion of its instability. MATERIALS AND METHODS: Preoperative ultrasonographic (US) inspection assessed several features of the tendon, whereas its stability was verified arthroscopically. In all cases, tenodesis or tenotomy procedures were performed and excised tendon fragments were harvested for microscopic examination based on the semiquantitative Bonar score. RESULTS: The most common US findings were hypoechoic areas, tendon thickening, an increased power Doppler signal and mechanical instability. Just as shoulder arthroscopy confirmed all mechanical instability cases detected in US, microscopic assessment revealed advanced degeneration in all samples. CONCLUSIONS: Our study indicates that US is a useful tool in identifying cases of advanced instability and LHB tendinopathy, whereas biceps tendon instability is a biomechanically complex, gradually progressing phenomenon, frequently associated with additional shoulder lesions.

Tendon - function-related structure, simple healing process and mysterious ageing.

Tendons are connective tissue structures of paramount importance to human ability of locomotion. The understanding of their physiology and pathology is gaining importance as advances in regenerative medicine are being made today. So far, very few studies were conducted to extend the knowledge about pathology, healing response and management of tendon lesions. In this paper we summarise actual knowledge on structure, process of healing and ageing of the tendons. The structure of tendon is optimised for the best performance of the tissue. Despite the simplicity of the healing response, numerous studies showed that the problems with full recovery are common and much more significant than we thought; that is why we discussed the issue of immobilisation and mechanical stimulation during healing process. The phenomenon of tendons' ageing is poorly understood. Although it seems to be a natural and painless process, it is completely different from degeneration in tendinopathy. Recent studies of biological treatment reported faster and optimal healing of the tendons when augmented by growth factors and stem cells. Despite advances in biology of tendons, management of their injuries is still a challenge for physicians; therefore, further studies are required to improve treatment outcomes.

Electron microscope study of the advanced tendinopathy process of the long head of the biceps brachii tendon treated arthroscopically.

BACKGROUND: The ultrastructural alterations related to tendinopathy have not been well described. Most studies on this subject have been conducted many years ago and focused on material from the Achilles tendon. It was demonstrated that various comorbidities can affect ultrastructural alterations in the advanced tendinopathy; however, there is very little data on ultrastructural morphology in tendinopathies related to mechanical overload as in case of the long head of the biceps brachii tendon (LHBT). The aim was to determine intermediate ultrastructural alterations in middle to severe grade the LHBT tendinopathy and to establish if they are different than those reported in the literature for other anatomical locations. MATERIALS AND METHODS: In this study we examined the ultrastructure of a series of the LHBT fragments arthroscopically removed due to tendinopathy and inve-stigated the morphology of tenocytes and collagen fibres in cases of the LHBT tendinopathy. RESULTS: In pathological samples tenocytes were randomly scattered, their shape was round and the shape of nuclei was also disrupted. The presence of apoptotic--like features in tenocytes' nuclei was noted. All samples showed replacement of collagen fibrils by non-collagen extracellular matrix and diffuse collagen disorganisation. CONCLUSIONS: It was demonstrated at ultrastructural level that the LHBT tendino-pathy is not simply a wear and tear phenomenon, since chronic degeneration of the extracellular matrix and tenocytes were present, similarly as in tendinopathies, in other anatomical locations. (Folia Morphol 2018; 77, 2: 371-377).

Process of neovascularisation compared with pain intensity in tendinopathy of the long head of the biceps brachii tendon associated with concomitant shoulder disorders, after arthroscopic treatment. Microscopic evaluation supported by immunohistochemical.

BACKGROUND: Tendinopathy of the long head of the biceps brachii tendon (LHBT) is one of the most common, painful conditions of the anterior part of the shoulder and often coexists with rotator cuff tears. Multifactorial aetiopathology of tendi-nopathy is poorly understood; however, several studies indicated that it is seen predominantly in areas with decreased vascularity of the tissue; the pathology is also characterised by expansive and abundant neovascular in-growth. The aim of the study was to investigate the relationship between the neovascularisation of proximal part of the LHBT and pain along the bicipital groove. MATERIALS AND METHODS: Tissue material was obtained from 28 patients who underwent a shoulder arthroscopy and experienced pain along the bicipital groove measured using Visual-Analog Scale (VAS) score. CD31 and CD34 molecules were visualised by immunohistochemical method to assess biceps tendon neovascula-risation and quantify it based on a Bonar scoring system. RESULTS: Although all patients reported pain prior to arthroscopy (mean VAS score was 7.5), microscopic examination did not reveal neovascularisation in all cases. Immunohistochemical staining for CD31 and CD34 allowed for very precise visualisation and quantification of neovascularisation; however there was also no correlation between vessels in-growth scores and pain. CONCLUSIONS: The obtained data suggest that neovascularisation process in tendino-pathy is not directly related to pain; however, further studies are needed to explain its significance in the LHBT tendinopathy. (Folia Morphol 2018; 77, 2: 378-385).

Doxorubicin-induced F-actin reorganization in cofilin-1 (nonmuscle) down-regulated CHO AA8 cells.

The actin cytoskeleton plays an important role in many cellular processes, including cell mortality, mitosis, cytokinesis, intracellular transport, endocytosis and secretion but also is involved in gene transcription. The dynamics of the actin cytoskeleton is controlled by different classes of actin-binding proteins (ABPs) which regulate the polymerization of actin filaments. In this report we used siRNA against cofilin-1 (nonmuscle) to demonstrate the effect of cofilin on the nuclear and cytoplasmic actin pools in CHO AA8 cells after exposition to various concentrations of doxorubicin. The immunofluorescence studies showed doxorubicin dose dependent tendency to formation the multinucleated giant cells, but also the increase of fluorescence intensity of cofilin in nuclei of untransfected cells. Induction of cell death with doxorubicin treatment in untransfected cells revealed both mitotic catastrophe (in both lower and higher doxorubicin doses) and apoptosis (mostly in higher doxorubicin doses), whereas among cofilin-1 down-regulated cells we observed only mitotic catastrophe. The results suggest that cofilin has apoptosis-inducing ability and that mitotic catastrophe is independent from F-actin content in cell nucleus. In this point of view we conclude that different mechanisms of chromatin reorganization are involved in these two processes. Moreover, we suppose that apoptosis and mitotic catastrophe are independent from each other.

Actin reorganization in CHO AA8 cells undergoing mitotic catastrophe and apoptosis induced by doxorubicin.

Doxorubicin (DOX) is a drug widely used in cancer chemotherapy. Although it has been proven that DOX kills tumor cells, the triggered modes of cell death are not fully understood. There is some evidence that, depending on the dose of DOX, the treated cells undergo senescence, mitotic catastrophe, apoptosis or necrosis. The aim of this study was to assess the type of CHO AA8 cell death induced with different DOX doses. In this context, we also assessed organization and distribution of F-actin, which integrity was suggested to be indispensable for apoptosis. Following treatment with 0.5 and 1 microM DOX, the giant multinucleated cells with extended network of fine microfilaments appeared. Notably, in the nuclei of the enlarged cells microscopy and cytometric analysis showed the presence of F-actin. DOX (2.5 microM) caused the appearance of the giant cells and with apoptotic features and signs of autophagy vacuolization. Flow cytometric studies indicated a dose-dependent increase in the number of TUNEL-positive cells and cells stained with both Annexin V and PI. Cell cycle analysis revealed the increase in the hyperploid DNA content. Our results suggest that treatment of CHO AA8 cells with different DOX doses caused mitotic catastrophe that was followed by apoptosis with signs of autophagy. The increase in F-actin content in the nuclei of the dying cells was evident. We hypothesize that in CHO AA8 cells F-actin may be involved in chromatin reorganization undergoing cell death.

Expression of cyclin A in intestinal biopsies from children with celiac disease.

The aim of this study was to determine the expression of cyclin A and describe its distribution in biopsy samples taken from children with suspected and confirmed celiac disease as well as in control samples. Investigated material consisted of 37 intestinal biopsies: 19 taken from patients with confirmed celiac disease, 9 from patients with its suspicion and 9 from healthy patients, who served as control. Immunohistochemical and immunogold methods were used to estimate cyclin A expression. In celiac disease samples morphological changes in epithelial cells, typical for disease, were shown. We observed weaker cyclin A expression, however there were also some cells with strong labeling in cytoplasm, near the nucleus. In control and suspected celiac disease groups cyclin A was present in the brush border, nucleus and whole cytoplasm, especially in proximity to the nucleus. In conclusion, these studies enabled us visualized pattern of distribution of cyclin A but let us also to presume that observed decrease of expression and its distribution might function as additional factor which could be taken under consideration to establish terminal diagnosis. We are aware of the fact that these are very first observations and that this subject needs to be further investigated with the use of additional methods and samples.

Hyperthermia-induced reorganization of microtubules and microfilaments and cell killing in CHO AA8 cell line.

The aim of this study was to elucidate the effects of hyperthermic treatment on cell morphology and the cytoskeleton in CHO AA8 cell line. The effects of exposure to elevated temperature were analyzed in CHO AA8 cell line by fluorescence microscopy and flow cytometry. The 30 min, at 44.5 degrees C heat shock treatment resulted in the collapse of microtubules (MTs) and microfilaments (MFs) around the nucleus followed by their recovery 24 h after heating. The initial collapse of these cytoskeletal systems, observed 15 min after treatment, was accompanied by the appearance of cells with reduction of volume, shrunken cytoplasm and condensed chromatin. 24 h afterwards, there was the increase in the number of cells with restored and extended MT and MF cytoskeletons. Most of them were larger in size compared to the control cells and had multiple nuclei. 48 h after heat shock the highest number of the giant cells with alternation in nuclear morphology was seen. Flow cytometry analysis revealed the increase in the number of cells with externalized phosphatidylserine 24 h and 48 h after hyperthermic treatment. These results suggest that following heat shock, CHO AA8 cells undergo mitotic catastrophe that presumably represents one of the events resulting in apoptosis.

Ultraviolet radiation (UV) induces reorganization of actin cytoskeleton in CHOAA8 cells.

Effect of UV radiation on actin cytoskeleton was studied in CHOAA8 cells by fluorescence and electron microscopy. UV irradiated cells showed impaired adherence, disruption of the actin filaments and stronger F-actin labeling in the center of the cell. Attached cells, especially enlarged ones showed rather weak labeling of stress fibers and bundles of F-actin in the cytoplasm, but some cells with intensive labeling of these structures were also observed. Detached cells were rounded, showed strong F-actin labeling and often had buds. At the ultrastructural level UV-irradiated cells showed segmented nuclei, bodies resembling micronuclei, dilatation of endoplasmic reticulum, swollen and disturb mitochondria. Immunogold labeling of actin at the ultrastructural level was observed in non-radiated and UV irradiated cells. Actin labeling was seen in nuclei and cytoplasm. In nuclei gold particles were localized in the area of condense chromatin. Labeling for actin was not found after control incubation. Our observations show that UV radiation promotes changes in the distribution of actin in CHOAA8 cells. The results also suggest that not only reorganization of actin but changes in organelles are involved in the process of apoptosis initiated by UV radiation.

Reorganization of actin in K-562 and HL-60 cells treated with taxol.

Influence of taxol, microtubular poison, has been studied on distribution of actin. K-562 and HL-60 cells were treated with taxol in a range of concentrations 0.02-10 microM for 72 hours. The reorganization of F-actin was dependent on its dose. Phalloidin conjugated to TRITC was used to evaluate actin distribution by classical fluorescence and confocal microscopy. Actin was visualized at ultrastructural level by using a postembedding streptavidin-gold method. The treatment of K-562 and HL-60 cells with 2-10 microM of taxol resulted in an increase of F- actin in the cytoplasm, with intense labeling as a ring close to surface of the cell. In HL-60 cells a concentration of F- actin at the site of apoptotic bodies was often observed. Immunogold labeling of actin was localized in the nuclei and cytoplasm in control cells and cells treated with all doses of taxol. At higher doses, compaction of chromatin in the nucleus with strong actin labeling was observed. These observations at the ultrastructural level suggest actin involvement in chromatin reorganization during the process of apoptosis. The present study demonstrated a dose dependent reorganization of actin after treatment with taxol.

Estimation of taxol influence on changes in tubulin and vimentin systems in K-562 and HL-60 cell lines by immunofluorescence microscopy.

The cytoskeletal system may be considered as an additional pathway involved in process of apoptosis and can be promising target for development of new chemotherapies. The study describes alterations in the distribution of vimentin and tubulin in taxol treated K-562 and HL-60 cells in relation to apoptotic changes. K-562 and HL-60 cells were treated with taxol in a range of concentrations (0.02-10 microM) for 72 hours. Significant changes in distribution of studied proteins occurred in the range 2-10 microM of taxol. K-562 cells showed thin network of vimentin distributed throughout cells or collapsed on nucleus. There were also cells with bright aggregates remembering apoptotic bodies. HL-60 cells showed strong labeling of vimentin in the cytoplasm as well as at the site of apoptotic bodies. Vimentin collapsed on the nucleus, labeling at poles and along the major axis of the cell were also seen. K-562 and HL- 60 cells showed radial labeling of tubulin from the centre, aggregates at the surface and bundled microtubules. These findings indicate that alterations in expression of studied cytoskeletal proteins after treatment with taxol were dose-dependent and related with characteristic features of apoptosis.

Effect of doxorubicin on actin reorganization in Chinese hamster ovary cells.

Exposure of Chinese hamster ovary (CHO AA8) cells to doxorubicin in doses of 75 microM resulted in reorganization of F-actin filaments and characteristic feature of apoptosis. Even increase in size of CHOAA8 cells was observed. Attached cells become more flattered and elongated than control ones. Intense labeling of F-actin around the nucleus and disrupted filaments in cytoplasm as well as stress fibres and bundles of F-actin were seen. Cells detached were rather rounded and there were not stress fibres present. In these cells the network of F-actin was weak and rather disrupted and cells with buds labelled for F-actin were observed. In section from confocal microscopy labeling of F-actin in nucleus was confirmed. Electron microscopy showed cells with multisegmented nuclei. Cells with intracellular area with small and large vacuoles and containing also electron-dense material were seen. Other cellular organelles were rather well preserve. There were margination and condensation of chromatin in nucleus. Immunogold labelling of actin was observed in cells whether or not treated with doxorubicin. Positivity for actin was localized in the nuclei and cytoplasm. In the nucleus gold particles were localized predominantly in the area of condense chromatin. Positive labeling for actin was not found after control incubation. These findings show that doxorubicin promotes changes in the distribution of actin in CHOAA8 cells and that reorganization of actin in these cells is involved in process of apoptosis.

Cytoskeletal reorganization during process of apoptosis induced by cytostatic drugs in K-562 and HL-60 leukemia cell lines.

The aim of the present study was to investigate the reorganization of F-actin, vimentin and tubulin in K-562 and HL-60 cell lines during apoptosis induced by etoposide, doxorubicin and taxol. The distribution of cytoskeletal proteins was analyzed by fluorescence microscopy. Actin was also studied by confocal microscopy and at the ultrastructural level. Changes in the distribution of cytoskeletal proteins were found to be dose-dependent and appeared to be more intense in HL-60 cells. Etoposide- and doxorubicin-treated cells showed similar changes in the distribution of F-actin, vimentin and tubulin. The reorganization of cytoskeletal proteins seemed to be consistent with features of apoptosis. An increase in bright staining of F-actin, vimentin and tubulin at the site of apoptotic bodies formation was observed. Immunogold labeling of actin in HL-60 cells was associated with features typical for apoptosis, i.e. compaction and margination of nuclear chromatin. K-562 cells showed cytoplasmic actin-positivity in the cytoplasm. Significant changes in morphology of HL-60 cells were found in the following concentrations: etoposide 20, 200 microM; doxorubicin 5, 10 microM and taxol 2-10 microM. The investigated proteins seemed to be involved in the above-reported apoptotic changes. Bright staining of F-actin, vimentin and tubulin, concentrated at the site of apoptotic bodies formation might suggested importance of these proteins for this process. Moreover, the increase in actin labeling in areas of chromatin compaction and margination of nuclear chromatin especially in HL-60 cells, which are more susceptible to apoptosis might implicate that actin might be involved in the chromatin remodeling during apoptosis.

The expression of proliferating cell nuclear antigen (PCNA) in leukemia cell lines HL-60 and K-562 at the light and electron microscope level.

PCNA antigen was localized at the light and electron microscopes level in two human leukemia cell lines HL-60 and K-562. PCNA expression was used to discriminate cycling from non-cycling cells. PCNA protein at the level of the light microscope was present in 70% of the cell in HL-60 cell line and in 65% of the cells in K-562 line. Streptavidin immunogold method was used for localization of PCNA expression at the ultrastructural level. Positive staining for this protein was seen as granular pattern in the nucleus and in the cytoplasm. In the nucleus the gold particles were seen to be associated with heterochromatin and euchromatin of the leukemia cells. In cytoplasm it was found on the endoplasmic reticulum and associated with ribosomes. Controls of the leukemia cells incubation with normal mouse serum showed no labelling at the light and electron microscope level.

Immunoelectron microscopical identification of the c-erbB-2 oncoprotein in patients with laryngeal squamous cell carcinoma.

The peroxidase-antiperoxidase (PAP) method was used for localization of the c-erbB-2 oncoprotein at the electron microscopical level in 15 patients with laryngeal squamous cell carcinoma. It was found that c-erbB-2 oncoprotein was present in 7 of 15 samples. Electron microscopical examination revealed expression of the c-erbB-2 oncoprotein both on the membrane of individual cells and in the cytoplasm. In 5 of the 7 cases of positive labeling, it was observed only on the plasma membrane of cells whereas in 2 cases, there was also cytoplasmic staining. Reaction product was associated with endoplasmic reticulum, and the nuclear envelope and was scattered throughout the cytoplasm on ribosomes. Control incubations using normal rabbit serum instead of the primary antibody showed no labeling. A significant correlation between c-erbB-2 oncoprotein and pathological characteristics such as nodal status and histological grade was not found.