This Month in Investigative Urology

Article Outline

• The Genetic Landscape of Bladder Development in Mice
• Neutrophil Gelatinase Associated Lipocalin and Urinary Tract Infection
• RAS Gene Activation in Urothelial Cancer
• Tamm-Horsfall Protein Protects Against Urinary Tract Infection
• Matrix Metalloproteinase Genetic Polymorphisms and Prostate Cancer
• Copyright

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The Genetic Landscape of Bladder Development in Mice 

Knowledge of the biology of normal bladder development is necessary to understand the pathobiology of human congenital bladder abnormalities such as posterior urethral valves and persistent cloaca, and disorders associated with dedifferentiation such as bladder cancer. Price et al (page 2366) from London, United Kingdom performed microarray analysis focusing on determining the gene expression profile at the initiation of bladder development. They extracted RNA from embryonic day 13 and 18 mouse bladders (anatomically equivalent to 7 and 13 weeks of human gestation), and evaluated gene expression using microarrays. Alterations in select genes of biological interest were confirmed using real-time quantitative polymerase chain reaction (PCR) and localization was determined by immunohistochemistry. They found that the genetic profile in the initiating mouse bladder at embryonic day 13 was dominated by transcription factors, retinoic acid signaling genes, Eph/ephrin bidirectional signaling molecules, and genes associated with regulating cell cycle and differentiation. At embryonic day 18 genes associated with smooth muscle, innervation and epithelial differentiation were up-regulated. These data provide novel insights into the molecules that orchestrate bladder development and highlight genes that may be involved in diseases associated with abnormal differentiation.

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Neutrophil Gelatinase Associated Lipocalin and Urinary Tract Infection 

It is well documented that 30% to 50% of urinary tract infections in children develop in the presence of vesicoureteral reflux. Recurrent upper urinary tract infections often lead to irreversible renal scarring. Ichino et al (page 2326) from Toyoake, Japan investigated the product of an up-regulated gene product, neutrophil gelatinase-associated lipocalin (NGAL), in an animal model to evaluate its potential usefulness as a biomarker of renal scarring. Global gene expression profiling of the kidney during the onset of renal scarring was performed, and renal NGAL mRNA and protein levels were examined by real-time reverse transcriptase PCR, Western blot and immunohistochemistry. Urinary NGAL levels were monitored by direct enzyme-linked immunosorbent assay. The authors found that NGAL mRNA and protein levels increased soon after bacterial injection, then decreased rapidly but subsequently persisted at high levels until 6 weeks after injection. Histological analysis revealed that the NGAL protein was overproduced in macrophages and renal tubular cells 2 weeks after injection. However, renal tubular cells continued to produce NGAL 6 weeks after injection, whereas this expression was lost in infiltrating cells. Urinary NGAL levels were also markedly increased at the early stages of infection and persisted at high levels throughout the latter stages of the experiment. These findings suggest the potential usefulness of NGAL as a marker of renal scarring in patients with vesicoureteral reflux in the absence of an active urinary tract infection. In addition, the authors suggest that determination of the urinary levels of this protein could be routinely performed in a clinical setting, potentially replacing renal scanning methods.

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RAS Gene Activation in Urothelial Cancer 

The most frequently detected alterations in oncogenes in animal and tumor models of human cancer are mutations in the RAS family of oncogenes. Most of these mutations are point mutations in codons 12, 13 or 61 that convert the RAS gene into a transforming oncogene. Boulalas et al (page 2312) from Athens, Greece determined RAS codon 12 and 13 point mutations and evaluated mRNA expression levels in bladder cancer, the fifth most common malignancy in men in Western society. They analyzed samples from 30 human bladder cancers and 30 normal tissues using PCR/restriction fragment length polymorphism and direct sequencing to determine the occurrence of mutations in codons 12 and 13 of RAS family genes. Real-time reverse transcriptase PCR was used to evaluate the expression profile of RAS genes in bladder cancer specimens compared to that in adjacent normal tissues. H-RAS mutations in codon 12 were observed in 9 tumor samples (30%). Of the 9 patients 2 (22%) had invasive bladder cancer and 7 (77%) had noninvasive bladder cancer. One H-RAS mutation was homozygous (11%) and the remaining 89% were heterozygous. All samples were WT for the K and N-RAS oncogenes. Moreover, 23 of 30 samples (77%) exhibited over expression in at least 1 RAS family gene compared to adjacent normal tissue. K and N-RAS presented the highest levels of over expression in bladder cancer specimens (50%), whereas 27% of transitional cell carcinomas exhibited H-RAS over expression relative to paired normal tissues. These results emphasize the importance of H-RAS activation in human bladder cancer by codon 12 mutations, and provide evidence that the increased expression of all 3 RAS genes is a common event in bladder cancer that is associated with disease development.

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Tamm-Horsfall Protein Protects Against Urinary Tract Infection 

Proteus mirabilis is a common cause of urinary tract infection, which is the most common nosocomial infection in North America occurring at a rate of approximately 1 million episodes per year. Tamm-Horsfall protein (THP) has been identified as a host defense factor against urinary tract infection by Escherichia coli, Staphylococcus saprophyticus and Klebsiella pneumoniae. Raffi et al (page 2332) from Oklahoma City, Oklahoma determined the role of THP as a host defense factor against cystitis and pyelonephritis caused by P. mirabilis. They generated THP gene knockout mice using homologous recombination, and introduced P. mirabilis transurethrally into the bladder of THP deficient (THP^−/−) and genetically similar WT (THP^+/+) mice. Cultured urine was used to quantitate the degree of bacteriuria. Bladders and kidneys were examined grossly and histomorphometrically to determine the intensity of inflammation. The THP^−/− mice had more severe bacteriuria and cystitis than the THP^+/+ mice. In addition, the THP^−/− mice had more pyelonephritic abscesses than the THP^+/+ mice. Thus, the authors concluded that THP acts as a host defense factor against P. mirabilis induced urinary tract infection.

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Matrix Metalloproteinase Genetic Polymorphisms and Prostate Cancer 

Prostate cancer is a common disease with a multifactorial and complex etiology. It is the most common male malignancy and the second leading cause of death in many countries. Molecular markers that can predict prostate cancer predisposition and progression are needed. Single nucleotide polymorphisms have been demonstrated to exist in the promoter regions of matrix metalloproteinase (MMP) genes, and are associated with the development and progression of some cancers. dos Reis et al (page 2320) from Sao Paulo, Brazil investigated the correlation between MMP1, 2, 7 and 9 polymorphisms with susceptibility to prostate cancer and classic prognostic parameters of prostate cancer. DNA genomic was extracted using conventional protocols and the DNA sequence containing the polymorphic site was amplified by real-time PCR using TaqMan® fluorescent probes. For the MMP1 gene the polymorphic allele was more common in the control group than in the prostate cancer group (p <0.001). For the MMP9 gene the incidence of the polymorphic homozygote genotype was higher in the prostate cancer group (p <0.001). For higher stage tumors (pT3) a polymorphic allele in the MMP2 gene was more common (p = 0.026). When considering Gleason score the polymorphic homozygote genotype of MMP9 was more common in Gleason 6 or less tumors (p = 0.003) while a polymorphic allele in the MMP2 gene was more common in Gleason 7 or greater tumors (p = 0.042). The authors concluded that MMP1 and MMP2 may protect against prostate cancer development, and that MMP9 may be related to higher risk. In contrast, the MMP9 polymorphism was associated with a lower Gleason score and the MMP2 polymorphism was associated with nonorgan confined disease.