fibrin and Urinary-Bladder-Neoplasms

Urinary proteomics has become one of the most attractive topics in disease biomarker discovery. MS-based proteomic analysis has advanced continuously and emerged as a prominent tool in the field of clinical bioanalysis. However, only few protein biomarkers have made their way to validation and clinical practice. Biomarker discovery is challenged by many clinical and analytical factors including, but not limited to, the complexity of urine and the wide dynamic range of endogenous proteins in the sample. This article highlights promising technologies and strategies in the MS-based biomarker discovery process, including study design, sample preparation, protein quantification, instrumental platforms, and bioinformatics. Different proteomics approaches are discussed, and progresses in maximizing urinary proteome coverage and standardization are emphasized in this review. MS-based urinary proteomics has great potential in the development of noninvasive diagnostic assays in the future, which will require collaborative efforts between analytical scientists, systems biologists, and clinicians.

Topics: Acute Kidney Injury; Antigens, Neoplasm; Antigens, Nuclear; Biomarkers; Biomarkers, Tumor; Cell Cycle Proteins; Fibrin; Humans; Insulin-Like Growth Factor Binding Proteins; Mass Spectrometry; Nuclear Matrix-Associated Proteins; Proteome; Proteomics; Tissue Inhibitor of Metalloproteinase-2; Urinalysis; Urinary Bladder Neoplasms

To determine the impact of fibrin clot inhibitor (FCI) use on oncological outcomes in a large contemporary cohort of patients with non-muscle-invasive bladder cancer (NMIBC) treated with adequate bacille Calmette-Guérin (BCG).. We performed an Institutional Review Board-approved review of patients with NMIBC treated with adequate intravesical BCG, at our institution between 2000 and 2018. FCI use at the time of BCG therapy was recorded for each patient. Patients were stratified according to use of FCI medication. Recurrence- and progression-free survival were analysed using Kaplan-Meier methods and Cox proportional hazard models.. Overall, 226 of 526 patients (43.0%) used a FCI: aspirin (205), clopidogrel (38), warfarin (18) and novel oral anticoagulant (NOAC; seven). The use of FCIs did not adversely affect either recurrence- or progression-free survival (P = 0.385 and P = 0.131, respectively). These results did not change when the impact of aspirin, clopidogrel or warfarin/NOAC use on recurrence and progression was evaluated separately. On multivariate analysis, FCI use was neither associated with tumour recurrence nor progression.. The use of FCIs was not associated with adverse oncological outcomes in a large contemporary cohort of patients receiving adequate intravesical BCG for NMIBC. Based on these results, FCIs may be safely continued during BCG immunotherapy.

Topics: Adjuvants, Immunologic; Administration, Intravesical; Anticoagulants; Aspirin; BCG Vaccine; Clopidogrel; Fibrin; Humans; Neoplasm Recurrence, Local; Thrombosis; Urinary Bladder Neoplasms; Warfarin

Corynebacterium diphtheriae still represents a global medical challenge, particularly due to the significant number of individuals susceptible to diphtheria and the emergence of non-toxigenic strains as the causative agents of invasive infections. In this study, we characterized the clinical and microbiological features of what we believe to be the first case of C. diphtheriae infection of a percutaneous nephrostomy catheter insertion site in an elderly patient with a fatal bladder cancer. Moreover, we demonstrated the potential role of adherence, biofilm formation and fibrin deposition traits in C. diphtheriae from the catheter-related infection. Non-toxigenic C. diphtheriae isolated from the purulent discharge (named strain BR-CAT5003748) was identified by the API Coryne system (code 1 010 324) and a multiplex PCR for detection of dtxR and tox genes. Strain BR-CAT5003748 showed resistance to oxacillin, ceftazidime and ciprofloxacin. In experiments performed in vitro, the catheter isolate was classified as moderately hydrophobic and as moderately adherent to polystyrene surfaces. Glass provided a more effective surface for biofilm formation than polystyrene. Micro-organisms adhered to (>1.5 x 10(6) c.f.u.) and multiplied on surfaces of polyurethane catheters. Microcolony formation (a hallmark of biofilm formation) and amorphous accretions were observed by scanning electron microscopy on both external and luminal catheter surfaces. Micro-organisms yielded simultaneous expression of localized adherence-like and aggregative-like (LAL/AAL) adherence patterns to HEp-2 cells. Interestingly, the coagulase tube test resulted in the formation of a thin layer of fibrin embedded in rabbit plasma by the non-toxigenic BR-CAT5003748 strain. In conclusion, C. diphtheriae should be recognized as a potential cause of catheter-related infections in at-risk populations such as elderly and cancer patients. LAL/AAL strains may be associated with virulence traits that enable C. diphtheriae to effectively produce biofilms on catheter surfaces. Biofilm formation and fibrin deposition could have contributed to the persistence of C. diphtheriae at the infected insertion site and the obstruction of the nephrostomy catheter.

Topics: Aged; Bacterial Adhesion; Biofilms; Catheter-Related Infections; Catheters, Indwelling; Cell Line; Corynebacterium diphtheriae; Diphtheria; Fatal Outcome; Fibrin; Humans; Male; Nephrostomy, Percutaneous; Polyurethanes; Urinary Bladder Neoplasms; Virulence

The features of radiation or chemotherapy cystitis mimicking invasive urothelial cancer are not widely known.. A search of the consultation files from our institution between January 1996 and September 2003 identified 20 patients with bladder biopsies showing cystitis mimicking invasive urothelial cancer.. The mean age at diagnosis was 69 years (range, 40-85 years); 80% were males. Complete history was not available in 1 patient. Seventeen patients had a history of pelvic irradiation (15 prostate cancer and 2 endometrial cancer). Two patients had systemic chemotherapy (1 metastatic colon cancer and 1 mixed connective tissue disease). All patients presented with hematuria. The mean time from radiation and/or chemotherapy to presentation was 27 months (range, 0-84 months). All cases showed epithelial proliferation that mimicked invasive cancer within the lamina propria, with marked proliferation seen in 45% of cases. Mild to moderate nuclear pleomorphism was seen in all cases. A characteristic feature was the presence of pseudoinvasive urothelial nests wrapping around the vessels associated with fibrin deposition. Most cases did not show any mitoses (75%). Ulceration was seen in 39% of cases. All cases showed some degree of hemorrhage, fibrin deposition and fibrin thrombi, fibrosis, and acute and chronic inflammation, with hemosiderin identified in 60% of cases. Edema and vascular congestion were common features (95% and 80%, respectively). Thickened vessels and vascular changes associated with radiation injury were identified in 75% of cases. Seventeen patients were followed for a mean of 9 months (range, 0.25-37 months), and none developed bladder cancer.. Radiation or chemotherapy cystitis can show epithelial proliferations that may be confused with invasive urothelial carcinomas. Other findings characteristic of radiation or chemotherapy cystitis, such as hemorrhage, fibrin, and vascular changes, are often seen in association with the epithelial proliferations and are helpful in distinguishing it from invasive cancer. Pathologists must be aware that these changes may be seen with a remote radiation or chemotherapy history, where this information may not be provided or known at the time of the biopsy evaluation.

Topics: Adult; Aged; Aged, 80 and over; Antineoplastic Agents; Colonic Neoplasms; Cystitis; Diagnosis, Differential; Endometrial Neoplasms; Epithelium; Female; Fibrin; Follow-Up Studies; Hemorrhage; Histocytochemistry; Humans; Male; Middle Aged; Mixed Connective Tissue Disease; Prostatic Neoplasms; Radiation Injuries; Radiotherapy; Thrombosis; Time Factors; Urinary Bladder; Urinary Bladder Neoplasms

This study describes two experimental models for the in vitro reconstitution of the human bladder mucosa (neo-bladder): human urothelial stabilized cell lines were cultured on three-dimensional matrices, collagen or platelet-fibrin gels, containing murine fibroblast 3T3-J2 cells. Low-density seeding (2x10(4) cells/ml) of both normal (TCA-48) and neoplastic cell lines (TCA-47) on collagen matrix gave rise to isolated papillar colonies, while high-density seeding (3.75x10(6) cells/ml) led to the formation of wide pluristratified epithelial sheets, resembling the normal transitional epithelium. In contrast, high-density seeding (5x10(5) cells/ml) on platelet-fibrin matrix did not allow the formation of epithelial sheets: only isolated voluminous colonies of normal TCA-48 cells, and sparse and small colonies of neoplastic TCA-47 could be observed. Growth assays and cytotoxicity reduction tests showed that the growth inhibitory effect of platelet-fibrin gel on urothelial cells was probably due to the aspecific activation of the complement contained in the plasmatic fraction, whose precipitation forms fibrin-glue. Collectively, these findings allow us to draw the following conclusions: i) neobladders obtained by culturing urothelial cells on collagen matrix reproduce normal bladder mucosa and could be utilized in pharmacological studies; and ii) platelet-fibrin gels, that specifically inhibit neoplastic urothelial cell growth, could be used as scaffolds in surgical bladder reconstitution.

Topics: 3T3 Cells; Animals; Blood Platelets; Cell Count; Cell Line; Cell Line, Tumor; Cell Proliferation; Cell Survival; Collagen; Culture Media; Culture Techniques; Extracellular Matrix; Fibrin; Fibroblasts; Gels; Humans; Mice; Mucous Membrane; Time Factors; Toxicity Tests; Urinary Bladder; Urinary Bladder Neoplasms; Urothelium

During cutaneous wound repair the epidermis avoids the fibrin-rich clot; rather it migrates down the collagen-rich dermal wound margin and over fibronectin-rich granulation tissue. The mechanism(s) underlying keratinocyte movement in this precise pathway has not been previously addressed. Here we demonstrate that cultured human keratinocytes do not express functional fibrinogen/fibrin receptors, specifically alpha v beta 3. Biologic modifiers known to induce integrin expression or activation did not induce adhesion to fibrin, fibrinogen, or its fragments. Epidermal explant outgrowth and single epidermal cell migration failed to occur on either fibrin or fibrinogen. Surprisingly, fibrin and fibrinogen mixed at physiologic molar ratios with fibronectin abrogated keratinocyte attachment to fibronectin. Keratinocytes transduced with the beta 3 integrin subunit cDNA, expressed alpha v beta 3 on their surface and attached to and spread on fibrinogen and fibrin. beta-gal cDNA-transduced keratinocytes did not demonstrate this activity. Furthermore, beta 3 cDNA-transduced keratinocyte adhesion to fibrin was inhibited by LM609 monoclonal antibody to alpha v beta 3 in a concentration-dependent fashion. From these data, we conclude that normal human keratinocytes cannot interact with fibrinogen and its derivatives due to the lack of alpha v beta 3. Thus, fibrinogen and fibrin are authentic anti-adhesive for keratinocytes. This may be a fundamental reason why the migrating epidermis dissects the fibrin eschar from wounds.

Topics: Animals; Antigens, CD; Bucladesine; Carcinogens; Cations, Divalent; Cell Adhesion; Cell Communication; Cell Movement; Centrifugation; DNA, Complementary; Epidermal Cells; Epidermal Growth Factor; Epidermis; Extracellular Matrix Proteins; Fibrin; Fibrin Fibrinogen Degradation Products; Fibrinogen; Gene Expression; Humans; Integrin beta3; Keratinocytes; Platelet Membrane Glycoproteins; Receptors, Vitronectin; Tetradecanoylphorbol Acetate; Transfection; Transforming Growth Factor beta; Tumor Cells, Cultured; Urinary Bladder Neoplasms; Wound Healing

Recent studies showed that the administration of active site-inhibited factor VIIa blocked factor VIIa/tissue factor-induced fibrin and thrombus formation in ex vivo and in vivo model systems. These studies suggest that inactivated factor VIIa competes efficiently with plasma factor VII(a) for a limited number of tissue factor sites. In the present study, we compared the interactions of factor VIIa and active site-inhibited factor VIIa with tissue factor. Competition studies of factor VIIa and active site-inhibited factor VIIa in a factor X activation assay showed that the affinity of the latter for relipidated tissue factor was 5-fold higher than that of factor VIIa. Radioligand binding studies with a human bladder carcinoma cell line (J82) and surface plasmon resonance studies using soluble tissue factor demonstrated a faster association and a slower dissociation for the active site-inhibited factor VIIa. Studies of equilibrium binding to cell surface tissue factor showed that the affinity of active site-inhibited VIIa was 5-fold higher than that of factor VIIa to non-functional tissue factor sites, whereas both inactivated factor VIIa and factor VIIa bound to functional tissue factor sites with the same high affinity. Comparison of the CD spectra of factor VIIa and active site-inactivated factor VIIa revealed structural differences in the protease domain. The potential physiological implications of these findings are discussed.

Topics: Amino Acid Chloromethyl Ketones; Binding Sites; Binding, Competitive; Cell Line; Cell Membrane; Circular Dichroism; Factor VIIa; Fibrin; Humans; Kinetics; Protein Conformation; Recombinant Proteins; Thromboplastin; Tumor Cells, Cultured; Urinary Bladder Neoplasms

After transurethral resection (TUR) of superficial bladder tumors, intravesical instillations with BCG are successfully used to lower the recurrence rate. The mechanism of the antitumor activity of BCG is not completely understood. After TUR, a fibrin clot is formed on the damaged urothelium. Fibronectin (FN) is part of the clot. It has been demonstrated that binding of BCG to the bladder wall is mediated by FN, and therefore FN seems necessary for the antitumor activity of BCG. This binding can be inhibited by fibrin clot inhibitors, like aspirin and coumarin. A reduced efficacy of BCG in patients with superficial bladder cancer using these drugs has been described. We studied 183 patients with superficial bladder cancer, treated with one or two 6-week courses of intravesical BCG instillations. Of the 42 patients that used fibrin clot inhibitors, 13 (31%) experienced a recurrent tumor, as compared to 56 (40%) of 141 patients who did not use these drugs (p = 0.28). The mean time to recurrence was the same in both groups. These results did not depend on the BCG strain used (Tice or RIVM, p = 0.92) and were not influenced by the use of antibiotics against urinary tract infections. We conclude that in our study fibrin clot inhibitors have no adverse effect on the efficacy of BCG therapy for superficial bladder cancer.

Topics: Administration, Intravesical; Anti-Inflammatory Agents, Non-Steroidal; Anticoagulants; Aspirin; BCG Vaccine; Carcinoma in Situ; Carcinoma, Transitional Cell; Coumarins; Dipyridamole; Fibrin; Humans; Male; Random Allocation; Urinary Bladder Neoplasms; Warfarin

Certain evidence indicates that tumor cells in the circulation may be enshrouded with a coat of fibrin. It has been suggested that this fibrin coat protects tumor cells from attack by the immune system. This study compared the interaction of lymphokine activated killer (LAK) cells with tumor cells alone and with fibrin coating. LAK cell killing of cultured human bladder tumor cells (T24) was measured by a 4-hour chromium release assay. Tumor cells (3 x 10(6] were incubated with Na51CrO4 for 2 hours at 37 degrees C and 5% CO2 in serum-free medium. After washing, one half of the cells were coated with fibrin by exposure to recalcified platelet-poor plasma. Fibrin coating was confirmed by immunofluorescence with anti-human-fibrinogen-fluorescein-conjugated antibodies. LAK cells were prepared from peripheral blood lymphocytes by incubation with interleukin-2 at a concentration of 1000 units of interleukin-2/1 ml serum-free medium/1 million cells for 5 days at 37 degrees C, 5% CO2. Five thousand tumor cells with or without fibrin were incubated with varying concentrations of either LAK or peripheral blood lymphocytes (10,000 to 100,000 cells). After 4 hours the supernatants were harvested and counted in a gamma counter for 1 minute. Over a range of effector-to-target cell ratios of 10:1 to 100:1 (LAK to T24), no difference was seen in percentage of specific lysis for T24 alone versus fibrin-coated T24 cells. At a ratio of 100:1 (LAK to T24), percentage of specific lysis was 83.3% versus 87.7% for uncoated and coated T24 cells, respectively. This suggests that fibrin coating of tumor cells is insufficient to provide protection from LAK cell killing.

Topics: Cytotoxicity, Immunologic; Fibrin; Fluorescent Antibody Technique; Humans; Interleukin-2; Killer Cells, Lymphokine-Activated; Lymphocytes; Time Factors; Tumor Cells, Cultured; Urinary Bladder Neoplasms

Although intravesical bacillus Calmette-Guerin therapy has proved to be efficacious in the treatment and prophylaxis against tumor recurrence of superficial bladder tumors, its mechanism of action has not been fully elucidated. Previous work has suggested that bacillus Calmette-Guerin organisms attach to the matrix protein, fibronectin, during fibrin clot formation at sites of urothelial disruption and that this attachment was required for the antitumor effect of bacillus Calmette-Guerin to be expressed. Furthermore, drugs inhibiting clot formation were found to abrogate the antitumor effect of intravesical bacillus Calmette-Guerin therapy in a murine bladder tumor model. To examine the effect of inhibitors of fibrin clot formation on the results of intravesical bacillus Calmette-Guerin therapy, a retrospective analysis of 149 evaluable patients receiving intravesical bacillus Calmette-Guerin for superficial bladder tumors was performed. The over-all response rate free of tumor for 29 patients who concomitantly received inhibitors of fibrin clot formation with bacillus Calmette-Guerin therapy was 48%, as compared with 67% for 120 patients who were not receiving these medications (p = 0.0655, chi-square). The most striking difference was noted for patients who failed with recurrent superficial disease. Of the patients who received fibrin clot inhibitors during intravesical bacillus Calmette-Guerin therapy 35% had recurrent superficial tumors compared to only 8% of those who did not receive these drugs during a mean followup of 29.8 plus or minus 11 months (p = 0.005, chi-square). Our study suggests that inhibitors of fibrin clot formation may have an adverse influence on the results of intravesical bacillus Calmette-Guerin therapy for superficial bladder tumors.

Topics: Administration, Intravesical; BCG Vaccine; Carcinoma in Situ; Carcinoma, Transitional Cell; Fibrin; Fibronectins; Humans; Neoplasm Recurrence, Local; Platelet Aggregation Inhibitors; Retrospective Studies; Urinary Bladder Neoplasms; Warfarin

In order to characterize further the previously observed induction of a highly metastatic phenotype in mouse bladder carcinoma cells by Ha-ras transfection, we studied production of plasminogen activator, in vitro invasiveness, and the potential for lung colonization of these cells. The parent carcinoma cells produced predominantly tissue-type plasminogen activator. Out of 13 clones of ras-transfected cells tested, 8 secreted quantitatively elevated levels of plasminogen activator (up to 3.5-fold) as compared to the control transfectants. The plasminogen activator activity in cell lysates was maximally increased 3-fold, the surface-associated activity increased 2.5-fold. The secreted plasminogen activator of cloned ras-transfected cells was characterized to be predominantly of the urokinase type (71.3% compared to 20.5% with the parental BL cells). Thus, in addition to the quantitative augmentation of plasminogen activator production and secretion in a large fraction of the ras-transfected cell population, a significant qualitative shift from tissue-type to urokinase-type has been observed. In addition, ras-transfection augmented the capacity of the cells for invasion into Matrigel in a double-filter in vitro assay as well as their ability to colonize the lungs of syngeneic animals. These malignant properties of the transfected cells might be responsible for their highly metastatic behaviour induced by ras transfection.

Topics: Animals; Fibrin; Genes, ras; Humans; Lung Neoplasms; Mice; Neoplasm Invasiveness; Neoplasm Metastasis; Phenotype; Transfection; Tumor Cells, Cultured; Urinary Bladder Neoplasms; Urokinase-Type Plasminogen Activator

Rapid in vivo growth of cultured human cancer or leukemia cells was achieved by implantation into the subrenal capsule of mice. A solid structure, necessary for accurate implantation and measurement of tumor growth in this model, was provided by stepwise addition of fibrinogen and thrombin to the tumor cells, leading to rapid enzymatic formation of a solid tumor-fibrin matrix. Human leukemia and epithelial cancers increased in volume between 6- and 40-fold when measured 6-10 days after implantation into normal or immunosuppressed mice. Immunosuppression of host CD-1 mice was achieved by cyclosporine given daily after tumor implantation, cyclophosphamide given preimplantation combined with cyclosporine, or whole-body irradiation given preimplantation. Confirming the validity of tumor measurements, tumor histology in the immunosuppressed mice revealed cell proliferation, invasion, and neovascularization. Similarly, no artifactual measurement of tumor growth was observed by nonviable cancer cells, implanted after in vitro exposure to a known cytotoxic concentration of thiotepa. This model provides an economical, short-term technique for the in vivo study of human tumor growth, for the evaluation of new cancer therapies, and for in vitro - in vivo drug activity correlations in specific types of human cancer or leukemia cell lines.

Topics: Animals; Colonic Neoplasms; Female; Fibrin; Humans; Immunosuppression Therapy; Karyotyping; Kidney; Leukemia, Myeloid, Acute; Melanoma; Methods; Mice; Mice, Nude; Neoplasm Transplantation; Transplantation, Heterologous; Urinary Bladder Neoplasms; Vulvar Neoplasms

Topics: Adult; Aged; Blood Coagulation; Carcinoma, Transitional Cell; Female; Fibrin; Fibrin Fibrinogen Degradation Products; Fibrinogen; Fibrinolysis; Histocytochemistry; Humans; Male; Middle Aged; Urinary Bladder Neoplasms

An enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody was developed to determine the clinical value of urinary fibrinogen/fibrin degradation product levels for the identification and management of patients with bladder cancer. Assays were performed on 286 serial urine specimens from 56 bladder carcinoma patients. Specimens were grouped according to whether the patient had an evident tumor at the time of specimen collection (134 specimens, 41 patients) or was clinically disease-free following treatment (152 specimens, 38 patients). Many patients contributed specimens to both groups as determined by their clinical status at the time of collection. In addition, 45 specimens from 33 patients with inflammation of the urogenital tract and 81 specimens from 19 patients with renal or prostatic cancer were assayed for urinary fibrin degradation products. The ELISA, using a high-sensitivity procedure, identified 83% of the specimens from bladder cancer-positive patients with an overall accuracy with all specimens of 78% and a false-negative rate of 5% for all specimens tested. The high-sensitivity ELISA appeared most appropriate for monitoring bladder cancer patients for recurrence of tumor after surgery. The ELISA using a high-specificity procedure appeared most appropriate for screening. The high-specificity ELISA accurately identified 96% of urine specimens from non-bladder cancer patients with a false-positive rate of only 5%. These results demonstrate that the ELISA is an efficient, reliable, quantitative, and noninvasive immunoassay that can be useful both for the identification of bladder cancer patients and for monitoring the course of the disease.

Topics: Aged; Antibodies, Monoclonal; Cell Line; Cell Membrane; Enzyme-Linked Immunosorbent Assay; Female; Fibrin; Fibrinogen; Humans; Male; Prognosis; Urinary Bladder Neoplasms

Topics: Adult; Aged; Bleomycin; Female; Fibrin; Humans; Male; Middle Aged; Solutions; Urinary Bladder Neoplasms

Topics: Adult; Aged; Cytodiagnosis; Female; Fibrin; Fibrin Fibrinogen Degradation Products; Humans; Male; Plasminogen Activators; Urinary Bladder Neoplasms; Urine

Topics: Aminocaproates; Blood Coagulation; Breast Neoplasms; Colonic Neoplasms; Female; Fibrin; Fibrinogen; Fibrinolysis; Gastrointestinal Neoplasms; Humans; Immunoelectrophoresis; Intestinal Neoplasms; Kidney Neoplasms; Lung Neoplasms; Lymphatic Metastasis; Male; Neoplasm Metastasis; Neoplasm Recurrence, Local; Pancreatic Neoplasms; Prostatic Neoplasms; Rectal Neoplasms; Thrombin; Urinary Bladder Neoplasms; Urogenital Neoplasms

Topics: Aged; Anuria; Cystitis; Diverticulum; Duodenal Ulcer; Female; Fibrin; Hernia, Inguinal; Humans; Ileum; Laparotomy; Male; Necrosis; Peritonitis; Rupture, Spontaneous; Tuberculosis, Urogenital; Urinary Bladder Calculi; Urinary Bladder Diseases; Urinary Bladder Neoplasms; Urination Disorders; Urography