[Regulatory dysfunctions in nasal polyposis]. / Regulatorische Dysfunktionen bei der Polyposis nasi.

Chronic rhinosinusitis is a heterogeneous group of inflammatory diseases with significant annual costs for the health care system. To date, there is no distinct signaling pathway known that explains the entire process from the beginning to tissue transformation. Due to the diversity of chronic rhinosinusitis, no uniform treatment has yet been developed. With a focus on chronic rhinosinusitis with nasal polyps (CRSwNP), molecular biologic gene expression studies have been performed to identify specific characteristics of nasal polyps that might allow the development of new therapeutic procedures. Microarray analysis revealed alterations in cell adhesion and differentiation as well as blood vessels. Further examinations identified two mechanisms that could play an important role in the pathogenesis of nasal polyps. In the context of the underlying disease, i.e., Th2-mediated chronic inflammation with predominantly eosinophilic cell infiltration, these findings might explain the pathogenesis of nasal polyps and allow development of new therapeutic strategies.

[Chemotherapy with paclitaxel leads to microRNA release]. / Chemotherapie mit Paclitaxel fuhrt zu microRNA-Freisetzung

BACKGROUND: During recent years, microRNAs (Greek: micros = small; miRNA) have become more important. miRNAs are highly conserved, noncoding, single-stranded RNA molecules 17­28 nucleotide in length. Secreted by tumor cells, miRNAs regulate many biological processes and are also involved in chemoresistance. Classical forms of cancer treatment lead to miRNA release. Which miRNAs are correlated to head and neck squamous cell carcinomas (HNSCC) and their chemoresistance to paclitaxel remains unknown. OBJECTIVES: Identification of miRNAs expressed in HNSCC and elucidation of those involved in conferring chemoresistance to paclitaxel. MATERIALS AND METHODS: To identify changes in gene expression, HNSCC cell lines were treated with 10 µM paclitaxel for 48 h and analyzed by microarray analysis. Thereafter, changed in expression of single miRNAs (miR221*, miR222 and miR222*) following paclitaxel treatment were analyzed using a quantitative real-time polymerase chain reaction (qRT-PCR). RESULTS: Under treatment with paclitaxel, miRNAs were released. The dominant change is upregulation of MIR222 gene expression. Regulation of miR222* expression under paclitaxel treatment seems to be different in human papillomavirus (HPV)-negative and HPV-positive HNSCC cell lines. CONCLUSION: Expression of mirR221/222 is correlated to cell cycle regulation, carcinogenesis, and chemoresistance. Detailed knowledge of the molecular mechanisms and effects ofmiRNAs is important for identifying miRNAs as cancermarkers, as well as for increasing the efficiency of cancer therapeutics.

Decline of lactate in tumor tissue after ketogenic diet: in vivo microdialysis study in patients with head and neck cancer.

In head and neck squamous cell carcinoma (HNSCC) aerobic glycolysis is the key feature for energy supply of the tumor. Quantitative microdialysis (µD) offers an online method to measure parameters of the carbohydrate metabolism in vivo. The aim was to standardize a quantitative µD-study in patients with HNSCC and to prove if a ketogenic diet would differently influence the carbohydrate metabolism of the tumor tissue. Commercially available 100 kDa-CMA71-µD- catheters were implanted in tumor-free and in tumor tissue in patients with HNSCC for simultaneous measurements up to 5 days. The metabolic pattern and circadian rhythm of urea, glucose, lactate, and pyruvate was monitored during 24 h of western diet and subsequent up to 4 days of ketogenic diet. After 3 days of ketogenic diet the mean lactate concentration declines to a greater extent in the tumor tissue than in the tumor-free mucosa, whereas the mean glucose and pyruvate concentrations rise. The in vivo glucose metabolism of the tumor tissue is clearly influenced by nutrition. The decline of mean lactate concentration in the tumor tissue after ketogenic diet supports the hypothesis that HNSCC tumor cells might use lactate as fuel for oxidative glucose metabolism.

Pancreatic carcinoma cell lines reflect frequency and variability of cancer stem cell markers in clinical tissue.

BACKGROUND: Pancreatic cancer is one of the most deadly malignancies with insufficient therapeutic options and poor outcome. Cancer stem cells (CSCs) are thought to be responsible for progression and therapy resistance. We investigated the potential of pancreatic cell lines for CSC research by analyzing to what extent they contain CSC populations and how representative these are compared to clinical tissue. METHODS: Six pancreatic cancer cell lines were analyzed by flow cytometry for CD326, CD133, CD44, CD24, CXCR4 and ABCG2. Subsequently, 70 primary pancreatic tissues were evaluated for CD326, CD133 and CD44 by immunohistochemistry. RESULTS: All the cell lines but one showed a stable expression pattern throughout biological replicates. Marker expression in clinical tissue of CD44 distinguished normal patients from pancreatic carcinoma patients with a sensitivity of 50% at 80% specificity and metastasized from nonmetastasized carcinomas with 69% sensitivity at 100% specificity. CONCLUSIONS: Our results indicate a link between elevated CD44 expression, malignancy and metastasis of pancreatic tissue. Furthermore, individual pancreatic cell lines show a substantial amount of cells with CSC properties which is comparable with interpatient variability detected in primary tissue. These pancreatic cancer cell lines could thus serve for urgently needed pharmacological CSC in vitro research.

Human Th17 cells can be induced through head and neck cancer and have a functional impact on HNSCC development.

BACKGROUND: The T helper 17 (Th17) cells recently identified as distinct T helper cell lineage are characterised by their production of the proinflammatory cytokine interleukin 17. Although much effort has been made in understanding the function of Th17 cells in the pathogenesis of different diseases, their influence in carcinogenesis remain largely unknown. METHODS: We studied the prevalence and induction of Th17 cells in head and neck squamous cell carcinoma (HNSCC) patients by flow cytometry. To determine the migration mechanism of Th17 cells into primary tumours and metastasis of HNSCC, we performed chemotaxis assays. We analysed the proliferation and the angiogenesis-related proteins of HNSCCs in the presence of Th17 cells with MTT-based proliferation assay and an angiogenesis protein array. RESULTS: In this study, we showed that the prevalence of Th17 cells is elevated in peripheral blood of HNSCC patients. In addition, tumour tissue and tumour-draining lymph nodes are infiltrated by a huge number of Th17 cells representing an important fraction of the tumour-infiltrating lymphocytes (TILs). We further showed that Th17 cells can be induced and expanded in tumour microenvironment through cytokines produced by tumour cells and TILs, and in addition can be recruited to the tumour milieu through a CCR6/CCL20-dependent mechanism. Furthermore, we showed that the proliferation and angiogenesis of HNSCC are impaired in the presence of Th17 cells. CONCLUSION: We conclude that Th17 cells have a substantial impact on the carcinogenesis of HNSCCs and on their metastasis and could serve as a potential therapeutic target to modulate anti-tumour response in HNSCC.

[Constitutive expression of the potential stem cell marker CD44 in permanent HNSCC cell lines]. / Konstitutive Expression des potentiellen Stammzellmarkers CD44 in permanenten HNSCC-Zelllinien.

BACKGROUND: Despite significant advances and the use of new diagnosic and therapy methods to treat head and neck squamous cell carcinoma (HNSCC), prognosis has improved only marginally in the last decades. Thus, there is an enormous need for novel immunotherapeutic approaches. It is becoming more and more obvious that stem cells play an important role in tumor development and progression. The identity of these cells and the underlying cellular and molecular mechanisms remain mostly unknown in HNSCC. MATERIAL AND METHODS: Solid tumors as well as cells from the peripheral blood of patients with HNSCC were analyzed by flow cytometry concerning the expression of different putative stem cell marker proteins. RESULTS: Distinct populations of CD44-positive (CD44+), lin-negative (lin-) potential stem cells could be identified in solid tumors of HNSCC patients with strong individual variations. Surprisingly, the potential stem cell marker CD44 was found to be constitutively expressed on the surface of all permanent HNSCC cell lines analyzed. CONCLUSION: These data strongly suggest that CD44+ tumor stem cells may play a key role in establishment of permanent HNSCC cell lines, selecting especially robust cell entities that in real life might drive progression and metastasis of HNSCC. Individual analysis of tumor stem cell markers will be an important tool for innovative therapies and prognosis of patients with HNSCC. Additional stem cell markers will be investigated concerning their expression and cellular function.

HARO7 encodes chorismate mutase of the methylotrophic yeast Hansenula polymorpha and is derepressed upon methanol utilization.

The HARO7 gene of the methylotrophic, thermotolerant yeast Hansenula polymorpha was cloned by functional complementation. HARO7 encodes a monofunctional 280-amino-acid protein with chorismate mutase (EC 5.4. 99.5) activity that catalyzes the conversion of chorismate to prephenate, a key step in the biosynthesis of aromatic amino acids. The HARO7 gene product shows strong similarities to primary sequences of known eukaryotic chorismate mutase enzymes. After homologous overexpression and purification of the 32-kDa protein, its kinetic parameters (k(cat) = 319.1 s(-1), n(H) = 1.56, [S](0.5) = 16.7 mM) as well as its allosteric regulatory properties were determined. Tryptophan acts as heterotropic positive effector; tyrosine is a negative-acting, heterotropic feedback inhibitor of enzyme activity. The influence of temperature on catalytic turnover and the thermal stability of the enzyme were determined and compared to features of the chorismate mutase enzyme of Saccharomyces cerevisiae. Using the Cre-loxP recombination system, we constructed mutant strains carrying a disrupted HARO7 gene that showed tyrosine auxotrophy and severe growth defects. The amount of the 0.9-kb HARO7 mRNA is independent of amino acid starvation conditions but increases twofold in the presence of methanol as the sole carbon source, implying a catabolite repression system acting on HARO7 expression.

Nuclear import of yeast Gcn4p requires karyopherins Srp1p and Kap95p.

The yeast transcription factor Gcn4p contains two stretches of amino acid residues, NLS1 and NLS2, which are independently able to relocate the cytoplasmic protein chorismate mutase into the nucleus. Only NLS2 is conserved among fungi. A truncated version of CPCA (the counterpart of Gcn4p in Aspergillus nidulans), which lacks the conserved NLS, accumulates in the cytoplasm instead of the nucleus. Nuclear uptake mediated by the NLS1 of Gcn4p is impaired by defects in genes for several different karyopherins, whereas NLS2-dependent nuclear import specifically requires the alpha-importin Srp1p and the beta-importin Kap95p. Yeast strains that are defective in either of these two karyopherins are unable to respond to amino acid starvation. We have thus identified Gcn4p as a substrate for the Srp1p/Kap95p transport complex. Our data suggest that NLS2 is the essential and specific nuclear transport signal; NLS1 may play only an unspecific or accessory role.