Deciphering von Hippel-Lindau (VHL/Vhl)-Associated Pancreatic Manifestations by Inactivating Vhl in Specific Pancreatic Cell Populations

The von Hippel-Lindau (VHL) syndrome is a pleomorphic familial disease characterized by the development of highly vascularized tumors, such as hemangioblastomas of the central nervous system, pheochromocytomas, renal cell carcinomas, cysts and neuroendocrine tumors of the pancreas. Up to 75% of VHL patients are affected by VHL-associated pancreatic lesions; however, very few reports in the published literature have described the cellular origins and biological roles of VHL in the pancreas. Since homozygous loss of Vhl in mice resulted in embryonic lethality, this study aimed to characterize the functional significance of VHL in the pancreas by conditionally inactivating Vhl utilizing the Cre/LoxP system. Specifically, Vhl was inactivated in different pancreatic cell populations distinguished by their roles during embryonic organ development and their endocrine lineage commitment. With Cre recombinase expression directed by a glucagon promoter in α-cells or an insulin promoter in β-cells, we showed that deletion of Vhl is dispensable for normal functions of the endocrine pancreas. In addition, deficiency of VHL protein (pVHL) in terminally differentiated α-cells or β-cells is insufficient to induce pancreatic neuroendocrine tumorigenesis. Most significantly, we presented the first mouse model of VHL-associated pancreatic disease in mice lacking pVHL utilizing Pdx1-Cre transgenic mice to inactivate Vhl in pancreatic progenitor cells. The highly vascularized microcystic adenomas and hyperplastic islets that developed in Pdx1-Cre;Vhl f/f homozygous mice exhibited clinical features similar to VHL patients. Establishment of three different, cell-specific Vhl knockouts in the pancreas have allowed us to provide evidence suggesting that VHL is functionally important for postnatal ductal and exocrine pancreas, and that VHL-associated pancreatic lesions are likely to originate from progenitor cells, not mature endocrine cells. The novel model systems reported here will provide the basis for further functional and genetic studies to define molecular mechanisms involved in VHL-associated pancreatic diseases

Tissue microarray: A simple technology that has revolutionized research in pathology

Tissue microarray (TMA) technology is a high-throughput research tool, which has greatly facilitated and accelerated tissue analyses by in-situ technologies. TMAs are amenable to every research method that can be applied on the standard whole sections at enhanced speed. It plays a central role in target verification of results from cDNA arrays, expression profiling of tumors and tissues, and is proving to be a powerful platform for proteomic research. In this review article, primarily meant for students of pathology and oncology, we briefly discuss its basic methodology, applications and merits and limitations

Tissue microarray: A simple technology that has revolutionized research in pathology

Tissue microarray (TMA) technology is a high-throughput research tool, which has greatly facilitated and accelerated tissue analyses by in-situ technologies. TMAs are amenable to every research method that can be applied on the standard whole sections at enhanced speed. It plays a central role in target verification of results from cDNA arrays, expression profiling of tumors and tissues, and is proving to be a powerful platform for proteomic research. In this review article, primarily meant for students of pathology and oncology, we briefly discuss its basic methodology, applications and merits and limitations

Supplementary Material for: Prognostic Significance of AMP-Dependent Kinase Alpha Expression in Cervical Cancer

<b><i>Objectives:</i></b> Cervical cancer is one of the most common gynecological malignancies worldwide, and its association with the AMP-activated protein kinase (AMPK) is still unknown. We aimed to investigate the clinical correlation between AMPK expression and cervical cancer. <b><i>Methods:</i></b> The expression of AMPKα1, AMPKα2 and phosphorylated AMPKα (p-AMPKα) was determined immunohistochemically in 524 formalin-fixed, paraffin-embedded malignant and premalignant cervical tissues. Subsequently, associations with clinicopathological characteristics and patient survival were assessed. <b><i>Results:</i></b> AMPKα2 expression was observed in the cytoplasm and nucleus, while expression of AMPKα1 and p-AMPKα was mainly observed in the cytoplasm. p-AMPKα expression increased during the normal-to-tumor transition of cervical carcinoma (p < 0.001), but, once cancer developed, the expression of AMPKα2 and p-AMPKα decreased in large-sized tumors when compared to smaller tumors (36 vs. 68%, p = 0.004 and 39 vs. 64%, p = 0.029, respectively). Notably, AMPKα2 expression was significantly associated with better disease-free survival (HR 0.29, 95% CI 0.10-0.86, p = 0.026). <b><i>Conclusion:</i></b> The AMPKα2 isoform showed potential as a favorable prognostic marker in cervical cancer. Therefore, additional studies are necessary to further clarify the complex contribution of AMPK isoforms and of phosphorylation status to cervical cancer progression and prognosis

Multiple Endocrine Neoplasia Type 1 Deletion in Pancreatic α-Cells Leads to Development of Insulinomas in Mice

The pancreatic α- and β-cells are critical components in regulating blood glucose homeostasis via secretion of glucagon and insulin, respectively. Both cell types are typically localized in the islets of Langerhans. However, little is known about the roles of paracrine interactions that contribute to their physiological functions. The lack of suitable cell lines to study α- and β-cells interactions have led us to develop an α-cell-specific Cre-expressing transgenic line utilizing a glucagon promoter sequence, the Glu-Cre transgenic mouse. Here, we demonstrate that the Glu-Cre could specifically and efficiently excise floxed target genes in adult islet α-cells. We further showed that deletion of the tumor suppressor gene, multiple endocrine neoplasia type 1 (Men1), in α-cells led to tumorigenesis. However, to our surprise, the lack of Men1 in α-cells did not result in glucagonomas but rather β-cell insulinomas. Because deletion of the Men1 alleles was only present in α-cells, our data suggested that cross communication between α- and β-cells contributes to tumorigenesis in the absence of Men1. Together, we believed that the new model systems described here will allow future studies to decipher cellular interactions between islet α- and β-cells in a physiological context

The 19q12 bladder cancer GWAS signal: association with cyclin E function and aggressive disease

A genome-wide association study (GWAS) of bladder cancer identified a genetic marker rs8102137 within the 19q12 region as a novel susceptibility variant. This marker is located upstream of the CCNE1 gene, which encodes cyclin E, a cell-cycle protein. We performed genetic fine-mapping analysis of the CCNE1 region using data from two bladder cancer GWAS (5,942 cases and 10,857 controls). We found that the original GWAS marker rs8102137 represents a group of 47 linked SNPs (with r(2) >/= 0.7) associated with increased bladder cancer risk. From this group, we selected a functional promoter variant rs7257330, which showed strong allele-specific binding of nuclear proteins in several cell lines. In both GWASs, rs7257330 was associated only with aggressive bladder cancer, with a combined per-allele OR = 1.18 [95% confidence interval (CI), 1.09-1.27, P = 4.67 x 10(-5)] versus OR = 1.01 (95% CI, 0.93-1.10, P = 0.79) for nonaggressive disease, with P = 0.0015 for case-only analysis. Cyclin E protein expression analyzed in 265 bladder tumors was increased in aggressive tumors (P = 0.013) and, independently, with each rs7257330-A risk allele (P(trend) = 0.024). Overexpression of recombinant cyclin E in cell lines caused significant acceleration of cell cycle. In conclusion, we defined the 19q12 signal as the first GWAS signal specific for aggressive bladder cancer. Molecular mechanisms of this genetic association may be related to cyclin E overexpression and alteration of cell cycle in carriers of CCNE1 risk variants. In combination with established bladder cancer risk factors and other somatic and germline genetic markers, the CCNE1 variants could be useful for inclusion into bladder cancer risk prediction models