Comparison of tissue biomarkers between non-schistosoma and schistosoma-associated urothelial carcinoma.

BACKGROUND: High-grade urothelial carcinoma either non-Schistosoma (NS-UBC) or Schistosoma (S-UBC)-associated is the tenth cause of death worldwide and represents a serious therapeutic problem. AIM: Evaluation of the immmunohistochemical expression of tumor necrosis factor-alpha (TNFα), epidermal growth factor receptor (EGFR), programmed cell death protein-1 (PDL1), estrogen receptor-alpha (ERα) and UroplakinIII, in the high-grade in NS-UBC and S-UBC as potential prognostic and therapeutic targets analyzed through estimation of area percentage, optical density and international pathological scoring system for each marker. MATERIAL AND METHODS: Sixty high grade urothelial carcinoma cases were enrolled in the study (30 cases of NS-UBC and 30 cases of S-UBC). The cases were immunohistochemically-assessed for TNFα, EGFR, PDL1, ERα and Uroplakin III expression. In S-UBC, parasite load was also evaluated for correlation with the immunohistochemical markers' expression in S-UBC. RESULTS: The area percentage of immune-expression of TNFα and EGFR was higher in S-UBC compared to NS-UBC. On the other hand, the NS-UBC displayed statistically-higher expression of PDL1 and uroplakinIII (p-value <0.001). ERα revealed higher, yet, non-significant expressions in S-UBC compared to NS-UBC (p-value =0.459). PDL1 expression showed the most superior record regarding area percentage (64.6± 34.5). Regarding optical density, TNF-α showed the highest transmittance expression (2.4 ± 0.9). EGFR positively correlated with PDL1 in S-UBC (r= 0.578, p-value =0.001) whereas in NS-UBC, TNFα and PDL1 (r=0.382, p-value=0.037) had positive correlation. Schistosoma eggs in tissues oppose uroplakin III expression and trigger immunomodulation via PDL1. CONCLUSION: Due to lower UroplakinIII expression, S-UBC is supposed to have a poorer prognosis. Hormonal therapy is not hypothesized due to a very minimal ERα expression in both NS-UBC and S-UBC. Regarding immunotherapy, anti-TNF-α is suggested for S-UBC whilst in NS-UBC, blockading PDL1 might be useful. Targeted EGFR therapy seems to carry emphasized outcomes in S-UBC. Correlations encourage combined immune therapy in NS-UBC; nevertheless, in S-UBC, combined anti-EGFR and PDL1 seem to be of benefit.

Differential responses of epithelial cells from urinary and biliary tract to eggs of Schistosoma haematobium and S. mansoni.

Chronic urogenital schistosomiasis can lead to squamous cell carcinoma of the bladder. The International Agency for Research on Cancer classifies the infection with S. haematobium as a group 1 carcinogen, a definitive cause of cancer. By contrast, hepatointestinal schistosomiasis due to the chronic infection with S. mansoni or S. japonicum associated with liver periportal fibrosis, does not apparently lead to malignancy. The effects of culturing human epithelial cells, HCV29, established from normal urothelium, and H69, established from cholangiocytes, in the presence of S. haematobium or S. mansoni eggs were investigated. Cell growth of cells co-cultured with schistosome eggs was monitored in real time, and gene expression analysis of oncogenesis, epithelial to mesenchymal transition and apoptosis pathways was undertaken. Schistosome eggs promoted proliferation of the urothelial cells but inhibited growth of cholangiocytes. In addition, the tumor suppressor P53 pathway was significantly downregulated when exposed to schistosome eggs, and downregulation of estrogen receptor was predicted in urothelial cells exposed only to S. haematobium eggs. Overall, cell proliferative responses were influenced by both the tissue origin of the epithelial cells and the schistosome species.

Computational deconvolution of gene expression by individual host cellular subsets from microarray analyses of complex, parasite-infected whole tissues.

Analyses of whole organs from parasite-infected animals can reveal the entirety of the host tissue transcriptome, but conventional approaches make it difficult to dissect out the contributions of individual cellular subsets to observed gene expression. Computational deconvolution of gene expression data may be one solution to this problem. We tested this potential solution by deconvoluting whole bladder gene expression microarray data derived from a model of experimental urogenital schistosomiasis. A supervised technique was used to group B-cell and T-cell related genes based on their cell types, with a semi-supervised technique to calculate the proportions of urothelial cells. We demonstrate that the deconvolution technique was able to group genes into their correct cell types with good accuracy. A clustering-based methodology was also used to improve prediction. However, incorrectly predicted genes could not be discriminated using this methodology. The incorrect predictions were primarily IgH- and IgK-related genes. To our knowledge, this is the first application of computational deconvolution to complex, parasite-infected whole tissues. Other computational techniques such as neural networks may need to be used to improve prediction.

Schistosoma haematobium egg-induced bladder urothelial abnormalities dependent on p53 are modulated by host sex.

INTRODUCTION AND OBJECTIVE: The bladder urothelium changes dramatically during Schistosoma haematobium infection (urogenital schistosomiasis). These alterations include hyperplasia, ulceration, dysplasia, squamous metaplasia and frank carcinogenesis. Defining the pathways underpinning these urothelial responses will contribute to a deeper understanding of how S. haematobium egg expulsion, hematuria, and bladder cancer develop in humans. The tumor suppressor gene p53 is of particular interest, given its role in many cancers, including bladder cancer generally and schistosomal bladder cancer specifically. METHODS: Transgenic mice featuring tamoxifen-inducible Cre recombinase activity in cells expressing the urothelial-specific gene uroplakin-3a (Upk3a-GCE mice) were crossed with either TdTomato-floxed-EGFP reporter or p53-floxed mice. Mice were administered tamoxifen or vehicle control to induce excision of floxed genes. TdTomato-EGFP reporter mice were sacrificed and their bladders harvested, sectioned, and imaged by fluorescence microscopy. p53-floxed mice underwent bladder wall injection with S. haematobium eggs or vehicle controls. Three months later, mice were sacrificed and their bladders subjected to histological analysis (H&E staining). RESULTS: We first confirmed the phenotypic fidelity of Upk3a-GCE mice by crossing them with TdTomato-floxed-EGFP reporter mice and administering tamoxifen to their progeny. As expected, these progeny switched from TdTomato to EGFP expression in their bladder urothelium. Having confirmed the phenotype of Upk3a-GCE mice, we next crossed them to p53-floxed mice. The resulting progeny were given tamoxifen or vehicle control to render them urothelial p53-haploinsufficient or -intact, respectively. Then, we injected S. haematobium eggs or control vehicle into the bladder walls of these mice. Male p53-intact, egg-injected mice exhibited similar histological changes as their p53-haploinsufficient counterparts, including urothelial hyperplasia and ulceration. In contrast, female p53-intact, egg-injected mice featured no urothelial ulceration, whereas their p53-haploinsufficient counterparts often had significant ulceration. CONCLUSIONS: Urothelial p53 signaling indeed seems to affect urothelial homeostasis during S. haematobium infection, albeit in a sex-specific manner. Ongoing work seeks to determine whether p53 mediates associated alterations in urothelial cell cycle status and frank carcinogenesis in the setting of urogenital schistosomiasis.

Schistosomiasis and bladder cancer: similarities and differences from urothelial cancer.

Through the years, schistosoma-associated bladder cancer was believed to be a unique entity of disease, different from urothelial cancer. As carcinogenesis is a highly complex process resulting from the accumulation of many genetic and epigenetic changes leading to alterations in the cell proliferation and regulation process, confirmation of their minute differences or similarities are extremely difficult. In bladder cancer, many of these carcinogenic cascades were not fully documented in spite of the efforts undertaken. The control of schistosomiasis and the subsequent decrease in the intensity of infestation showed feature changes approaching that of urothelial tumors. However, schistosoma-associated bladder cancer still presents in more advanced stages than schistosoma-non-associated urothelial cancer. Furthermore, many data were collected proving that, upon applying the same treatment protocol and management care, stage-by-stage comparison of the treatment end results were found to be similar in bladder cancer patients with the different etiologies.

CD44s and CD44v6 in diagnosis and prognosis of human bladder cancer.

UNLABELLED: The cell adhesion molecules (CAMs) CD44 standard (CD44s) and its variant 6 (CD44v6) are involved in the progression and invasion of human malignancies. However, discrepancies in the prognostic value of CD44s and CD44v6 expression need to be addressed. AIMS: To investigate the expression of CD44s and CD44v6 in bladder carcinomas and relate the results to the established prognostic factors. MATERIALS AND METHODS: 50 bladder carcinoma specimens, 30 cases with transitional cell carcinoma (TCC: 6 bilharzial and 24 nonbilharzial) and 20 cases with squamous cell carcinoma (SCC: 8 bilharzial and 12 nonbilharzial), were included. Immunohistochemical analysis for CD44s and CD44v6 was carried out using avidin-biotin peroxidase method. RESULTS: The level of both CD44s and CD44v6 in TCC was significantly higher in invasive than in preinvasive tumors and normal urothelium (p < .05). A direct association between the percentage of expression of both markers and the grade of TCC (p < .05) was observed. An inverse correlation between CD44s and SCC was seen, where metaplastic urothelium showed higher expression than invasive carcinomas. No association was observed between the expressions of both CD44s and CD44v6 and bilharzial ova, sex and age of the patient, or size of the tumor. CONCLUSIONS: The authors report statistically significant correlation between CD44s and CD44v6 expression and increasing grade and stage of TCC. No such correlation with SCC and with bilharzial cystitis, sex and age of the patient, or size of the tumor was documented.

Confocal laser scanning microscopy, a new in vivo diagnostic tool for schistosomiasis.

BACKGROUND: The gold standard for the diagnosis of schistosomiasis is the detection of the parasite's characteristic eggs in urine, stool, or rectal and bladder biopsy specimens. Direct detection of eggs is difficult and not always possible in patients with low egg-shedding rates. Confocal laser scanning microscopy (CLSM) permits non-invasive cell imaging in vivo and is an established way of obtaining high-resolution images and 3-dimensional reconstructions. Recently, CLSM was shown to be a suitable method to visualize Schistosoma mansoni eggs within the mucosa of dissected mouse gut. In this case, we evaluated the suitability of CLSM to detect eggs of Schistosoma haematobium in a patient with urinary schistosomiasis and low egg-shedding rates. METHODOLOGY/PRINCIPAL FINDINGS: The confocal laser scanning microscope used in this study was based on a scanning laser system for imaging the retina of a living eye, the Heidelberg Retina Tomograph II, in combination with a lens system (image modality). Standard light cystoscopy was performed using a rigid cystoscope under general anaesthesia. The CLSM endoscope was then passed through the working channel of the rigid cystoscope. The mucosal tissue of the bladder was scanned using CLSM. Schistoma haematobium eggs appeared as bright structures, with the characteristic egg shape and typical terminal spine. CONCLUSION/SIGNIFICANCE: We were able to detect schistosomal eggs in the urothelium of a patient with urinary schistosomiasis. Thus, CLSM may be a suitable tool for the diagnosis of schistosomiasis in humans, especially in cases where standard diagnostic tools are not suitable.

Microscopic investigations in a diabetic rat urinary bladder infected with Trichosomoides crassicauda.

Rats are widely used laboratory animals and have several parasites. One of these are helminths, known not only to cause serious effects on the experimental results in healthy subjects, but also in subjects with heavy infections. One of the relatively pathologic helminth is Trichosomoides crassicauda, which lives in the nodules of the urinary bladder. It is known that diabetics are more prone to infections with several microorganisms. Observations in a diabetic rat bladder showed T. crassicauda eggs inside the transitional epithelium, and structural changes in the bladder epithelium were evident. Urinary-bladder tissues taken from streptozotocin-injected diabetic subjects and citrate buffer-injected control subjects were fixed, embedded in araldite and investigated under a light microscope. Distinct changes in the histological structure of a diabetic urinary bladder transitional epithelium were observed after T. crassicauda infection. Many papillomas were formed and the epithelial tissues were completely degenerated. In addition, electron microscopic examinations also revealed degeneration of the subepithelial tissues.

Clinical utility of squamous and transitional nuclear structure alterations induced by Schistosoma haematobium in chronically infected adults with bladder damage verified by ultrasound in Ghana.

OBJECTIVE: To evaluate the clinical utility of quantitative nuclear morphometry--i.e., alteration in nuclear size/shape, DNA content and chromatin structure-of intact cells obtained from the sediment of urine specimens collected from people living in an area highly endemic for Schistosoma haematobium in Ghana. STUDY DESIGN: Digital images of Feulgen-DNA-stained squamous cell (SC) and transitional cell (TC) urothelial nuclei were captured using the AutoCyte imaging system, and nuclear morphometric descriptors (NMDs) were calculated. A total of 3,495 and 4,523 SC and TC nuclei from normal bladder ultrasound subjects (n =21) and 3,465 and 3,064 SC and TC nuclei from severely abnormal bladder ultrasound subjects (n = 20) were captured. RESULTS: Univariate logistic regression analyses of pooled SC and TC nuclei training sets showed that 27/40 NMDs and 24/40 NMDs were univariately significant for differentiating between SCs and TCs of subjects with normal and severely abnormal bladder ultrasound. Multivariate models constructed using NMDs with > or = 50% inclusion frequency yielded AUC-ROCs of 75.23% and 74.42% in the SC training and validation, and 69.90% and 66.70% for TC training and validation. Further, a squamous cell patient-specific model predicted severe bladder damage with an AUC-ROC of 86.90%, yielding the sensitivity, specificity and accuracy of 85.00%, 76.19% and 80.49%, respectively. CONCLUSION: Quantitative nuclear structure alterations can be used to make a noninvasive assessment of cytologic changes observed in both SC and TC bladder epithelia due to S haematobium infection.

Rhinosporidiosis presenting as an urethral polyp.

Rhinosporidiosis is an inflammatory disease caused by Rhinosporidium seeberi, a protoctistan mesomycetozoa, member of a group of novel aquatic parasites, characterized by hyperplastic polypoid lesions of the nasal cavity and rarely other mucous membranes. We report an unusual presentation of rhinosporidiosis as an urethral polyp, which is only the second case of rhinosporidiosis reported from Pakistan.

Analysis of fibronectin expression in the bilharzial granulomas and of laminin in the transformed urothelium in schistosoma haematobium infested patients.

UNLABELLED: The bilharzial granulomas and urothelial transformation are common findings in Schistosoma haematobium infested patients. We hypothesize that the distribution of extrinsic (fibronectin, FN) and intrinsic basement membrane (BM) proteins (laminin, LN) is altered during the evolution of these lesions. METHODS: To test this hypothesis, 70 cystectomy specimens, entailing variable associations of normal and dysplastic urothelium (all cases), and bilharzial granulomas were examined for FN and LN protein expression. RESULTS: The biharzial granulomas were formed of admixture of CD3+T cells, CD68+histiocytes and CD220B cells. The CD3+T cells and and CD68+histiocytes were the predominant cell populations. Increased deposition of FN occurred with the evolution from cellular (loose fibrillary network, 20 cases) to fibrocellualr (dense fibrillary network, 30 cases) to fibrotic (tight conglomerates, 20 cases) granulomas. Alternatively, BM staining for LN was linear and continuous underlying normal and metaplastic urothelium. In dysplastic urothelium (20 cases), it showed breaks in continuity. CONCLUSIONS: Alterations of FN and LN occur during the development of the bilharzial granuloma and urothelial transformation.