Expression of estrogen receptor beta correlates with adverse prognosis in resected pancreatic adenocarcinoma.

BACKGROUND: The relevance of estrogen receptor (ER) expression in pancreatic ductal adenocarcinoma (PDAC) is largely unknown. Clinical trials targeting ER with selective estrogen receptor modulators in pancreatic cancer did not show any benefit. Here, we analyze the impact of recently characterized ER isoform beta on survival in a cohort of patients with resected PDAC. METHODS: Eighty-four patients having undergone pancreatic resection for PDAC at a single institution were identified. Tissue microarrays were constructed of archival tumor specimens. The expression of ER beta was determined by immunohistochemistry and quantified by a system established for estrogen receptor expression in breast cancer. ER beta expression was then correlated with clinicopathological parameters, and univariate and multivariate survival analyses were performed. RESULTS: Nuclear expression of ER beta was found in 31% of tumors. No significant correlation was found between ER beta expression and TNM status, tumor grade, age or sex. Univariate analysis revealed nodal metastasis and the expression of ER beta as factors correlating with a shorter overall survival and disease free survival. When comparing ER beta expression in patients surviving more than 24 months with those who died from the tumor within 12 or 24 months, respectively, a significantly lower ER beta expression was found in the long term survivors. In multivariate analysis, ER beta expression was demonstrated to be an independent predictor of shorter overall survival. CONCLUSIONS: In resected PDAC, expression of ER beta seems to correlate with poor prognosis. These data may help to identify patients who may benefit from additional systemic therapy including selective estrogen receptor modulators.

Critical role of platelet glycoprotein ibα in arterial remodeling.

OBJECTIVE: Arteriogenesis is strongly dependent on the recruitment of leukocytes, especially monocytes, into the perivascular space of growing collateral vessels. On the basis of previous findings that platelets are central players in inflammatory processes and mediate the recruitment of leukocytes, the aim of this study was to assess the role of platelets in a model of arterial remodeling. APPROACH AND RESULTS: C57Bl6 wild-type mice, IL4-R/Iba mice lacking the extracellular domain of the glycoprotein Ibα (GPIbα) receptor, and mice treated with antibodies to block GPIbα or deplete circulating platelets were studied in peripheral arteriogenesis. Using a novel model of intravital 2-photon and epifluorescence imaging, we visualized and quantified the interaction of platelets with leukocytes and the vascular endothelium in vivo. We found that transient platelet adhesion to the endothelium of collateral vessels was a major event during arteriogenesis and depended on GPIbα. Furthermore, leukocyte recruitment was obviously affected in animals with defective platelet GPIbα function. In IL4-R/Iba mice, transient and firm leukocyte adhesion to the endothelium of collateral vessels, as well as leukocyte accumulation in the perivascular space, were significantly reduced. Furthermore, we detected platelet-leukocyte aggregates within the circulation, which were significantly reduced in IL4-R/Iba animals. Finally, platelet depletion and loss of GPIbα function resulted in poor reperfusion recovery as determined by laser Doppler imaging. CONCLUSIONS: Thus, GPIbα-mediated interactions between platelets and endothelial cells, as well as leukocytes, support innate immune cell recruitment and promote arteriogenesis-establishing platelets as critical players in this process.

Desmogleins as prognostic biomarkers in resected pancreatic ductal adenocarcinoma.

BACKGROUND: Frequent disease relapse and a lack of effective therapies result in a very poor outcome in pancreatic ductal adenocarcinoma (PDAC) patients. Thus, identification of prognostic biomarkers and possible therapeutic targets is essential. Besides their function in cell-cell adhesion, desmogleins may play a role in tumour progression and invasion that has not been investigated in PDAC to date. This study evaluated desmoglein expression as a biomarker in PDAC. METHODS: Using immunohistochemistry, we examined desmoglein 1 (DSG1), desmoglein 2 (DSG2) and desmoglein 3 (DSG3) expression in the tumour tissue of 165 resected PDAC cases. Expression levels were correlated to the patients' clinicopathological parameters and postoperative survival times. We confirmed these results in two independent gene expression data sets. RESULTS: A total of 36% of the tumours showed high DSG3 expression that correlated significantly with shorter patient survival (P=0.011) and poor tumour differentiation (P<0.001), whereas no such association was detected for DSG1 or DSG2. In RNA-Seq data and in microarray expression data, high DSG3 expression correlated significantly with poor survival (P=0.000356 and P=0.00499). CONCLUSIONS: We identify DSG3 as a negative prognostic biomarker in resected PDAC, as high DSG3 expression is associated with poor overall survival and poor tumour-specific survival. These findings suggest DSG3 and its downstream signalling pathways as possible therapeutic targets in DSG3-expressing PDAC.

Heat shock protein 27 as a prognostic and predictive biomarker in pancreatic ductal adenocarcinoma.

A role of heat shock protein 27 (HSP27) as a potential biomarker has been reported in various tumour entities, but comprehensive studies in pancreatic cancer are lacking. Applying tissue microarray (TMA) analysis, we correlated HSP27 protein expression status with clinicopathologic parameters in pancreatic ductal adenocarcinoma specimens from 86 patients. Complementary, we established HSP27 overexpression and RNA-interference models to assess the impact of HSP27 on chemo- and radiosensitivity directly in pancreatic cancer cells. In the TMA study, HSP27 expression was found in 49% of tumour samples. Applying univariate analyses, a significant correlation was found between HSP27 expression and survival. In the multivariate Cox-regression model, HSP27 expression emerged as an independent prognostic factor. HSP27 expression also correlated inversely with nuclear p53 accumulation, indicating either protein interactions between HSP27 and p53 or TP53 mutation-dependent HSP27-regulation in pancreatic cancer. In the sensitivity studies, HSP27 overexpression rendered HSP27 low-expressing PL5 pancreatic cancer cells more susceptible towards treatment with gemcitabine. Vice versa, HSP27 protein depletion in HSP27 high-expressing AsPC-1 cells caused increased gemcitabine resistance. Importantly, HSP27 expression was inducible in pancreatic cancer cell lines as well as primary cells. Taken together, our study suggests a role for HSP27 as a prognostic and predictive marker in pancreatic cancer. Assessment of HSP27 expression could thus facilitate the identification of specific patient subpopulations that might benefit from individualized treatment options. Additional studies need to clarify whether modulation of HSP27 expression could represent an attractive concept to support the incorporation of hyperthermia in clinical treatment protocols for pancreatic cancer.

EFEMP1 expression promotes in vivo tumor growth in human pancreatic adenocarcinoma.

The progression of pancreatic cancer is dependent on local tumor growth, angiogenesis, and metastasis. EFEMP1, a recently discovered member of the fibulin family, was characterized with regard to these key elements of pancreatic cancer progression. Differential gene expression was assessed by mRNA microarray hybridization in FG human pancreatic adenocarcinoma cells and L3.6pl cells, a highly metastatic variant of FG. In vivo orthotopic tumor growth of EFEMP1-transfected FG cells was examined in nude mice. To assess the angiogenic properties of EFEMP1, vascular endothelial growth factor (VEGF) production of tumor cells, endothelial cell proliferation and migration, and tumor microvessel density were analyzed in response to EFEMP1. Further, tumor cell apoptosis, cell cycle progression, and resistance to cytotoxic agents were quantitated by propidium iodide staining and flow cytometry. In microarray hybridization, EFEMP1 was shown to be significantly up-regulated in L3.6pl cells compared with FG cells. Concordantly, EFEMP1 transfection of FG cells stimulated orthotopic and metastatic tumor growth in vivo. EFEMP1 expression resulted in a stimulation of VEGF production by tumor cells and an increased number of CD31-positive microvessels. Endothelial cell proliferation and migration were not altered by EFEMP1, indicating an indirect angiogenic effect. Further, EFEMP1 expression decreased apoptosis and promoted cell cycle progression in response to serum starvation or exposure to gemcitabine, 5-fluorouracil, and irinotecan. EFEMP1 has protumorigenic effects on pancreatic cancer in vivo and in vitro mediated by VEGF-driven angiogenesis and antiapoptotic mechanisms. Hence, EFEMP1 is a promising candidate for assessing prognosis and individualizing therapy in a clinical tumor setting.

Erythropoietin administration after myocardial infarction in mice attenuates ischemic cardiomyopathy associated with enhanced homing of bone marrow-derived progenitor cells via the CXCR-4/SDF-1 axis.

Mobilization of bone marrow-derived stem cells (BMCs) was shown to have protective effects after myocardial infarction (MI). However, the classical mobilizing agent, granulocyte-colony stimulating factor (G-CSF) relapsed after revealing an impaired homing capacity. In the search for superior cytokines, erythropoietin (EPO) appears to be a promising agent. Therefore, we analyzed in a murine model of surgically induced MI the influence of EPO treatment on survival and functional parameters as well as BMC mobilization, homing, and effect on resident cardiac stem cells (CSCs). Human EPO was injected intraperitoneally after ligation of the left anterior descendens (LAD) for 3 days with a total dose of 5000 IU/kg 6 and 30 days after MI, and pressure volume relationships were investigated in vivo. Cardiac tissues were analyzed by histology. To show the effect on BMCs and CSCs, FACS analyses were performed. Homing factors were analyzed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and ELISA. EPO-treated animals showed a significant improvement of survival post-MI (62 vs. 36%). At days 6 and 30, all hemodynamic parameters associated with attenuated remodeling, enhanced neovascularization, and diminished apoptotic cells in the peri-infarct area were improved. BMC subpopulations (CD31(+), c-kit(+), and Sca-1(+) cells) were mobilized, and homing of Sca-1(+) and CXCR4(+) BMCs toward an SDF-1 gradient into the ischemic myocardium was enhanced. However, there was no beneficial effect on CSCs. We have shown that EPO application after MI shows cardioprotective effects. This may be explained by mobilization of BMCs, which are homing via the CXCR-4/SDF-1 axis. However, EPO has no beneficial effects on resident CSCs. Therefore, new treatment regimes using EPO together with other agents may combine complementary beneficial effects preventing ischemic cardiomyopathy.

Parathyroid hormone effectively induces mobilization of progenitor cells without depletion of bone marrow.

OBJECTIVE: Cytokine-mediated mobilization of hematopoietic stem cells has become an established method in the field of autologous and allogenic stem cell transplantation. Furthermore, it presents a new concept in tissue repair and regenerative medicine. In the present study, we explored the potency of parathyroid hormone (PTH) compared to granulocyte colony-stimulating factor (G-CSF) for mobilization of stem cells and its regenerative capacity on bone marrow. MATERIALS AND METHODS: Healthy mice were either treated with PTH, G-CSF, or saline. Laboratory parameters were analyzed using a hematological cell analyzer. Hematopoietic stem cells characterized by lin(-)/Sca-1(+)/c-kit(+), as well as subpopulations (CD31(+), c-kit(+), Sca-1(+), CXCR4(+)) of CD45(+)/CD34(+) and CD45(+)/CD34(-) cells were measured by flow cytometry. Immunohistology as well as fluorescein-activated cell sorting analyses were utilized to determine the composition and cell-cycle status of bone marrow cells. Serum levels of distinct cytokines (G-CSF, vascular endothelial growth factor [VEGF]) were determined by enzyme-linked immunosorbent assay. Further, circulating cells were measured after PTH treatment in combination with G-CSF or a G-CSF antibody. RESULTS: Stimulation with PTH showed a significant increase of all characterized subpopulations of bone marrow-derived progenitor cells (BMCs) in peripheral blood (1.5- to 9.8-fold) similar to G-CSF. In contrast to G-CSF, PTH treatment resulted in an enhanced cell proliferation with a constant level of lin(-)/Sca-1(+)/c-kit(+) cells and CD45(+)/CD34(+) subpopulations in bone marrow. Interestingly, PTH application was associated with increased serum levels of G-CSF (2.8-fold), whereas VEGF showed no significant changes. Blocking endogenous G-CSF with an antibody significantly reduced the number of circulating cells after PTH treatment. A combination of PTH and G-CSF showed slight additional effects compared to PTH or G-CSF alone. CONCLUSION: PTH induces mobilization of progenitor cells effectively, which can be related to an endogenous release of G-CSF. In contrast to G-CSF treatment, PTH does not result in a depletion of bone marrow, which may be mediated by an activation of PTH receptor on osteoblasts. The novel function of PTH on mobilization and regeneration of BMCs may pave the way for new therapeutic options in bone marrow and stem cell transplantation as well as in the field of ischemic disorders.

Parathyroid hormone treatment after myocardial infarction promotes cardiac repair by enhanced neovascularization and cell survival.

AIMS: An ongoing concept is that stem cells have the potential to regenerate the injured myocardium. In addition to direct vasorelaxing effects on the vasculature, which are mediated by an increased cAMP production leading to a decreased calcium influx in smooth muscle cells, parathyroid hormone (PTH) was recently shown to facilitate stem cell mobilization. Therefore, we analysed in a murine model of experimental myocardial infarction (MI) the influence of PTH treatment on survival, functional parameters, stem cell migration, and expression of vascular endothelial growth factor A (VEGF-A). METHODS AND RESULTS: Mice (C57BL/6) were treated with PTH (80 microg/kg/d) for up to 14 days after coronary artery ligation. Functional and immunohistochemical analyses were performed at days 6 and 30 after MI. Stem cells and VEGF expression in the myocardium were analysed by FACS and qRT-PCR at day 2 after MI. PTH-treated animals revealed a significant improvement of post-MI survival and myocardial function that was related to a subsequent reduction of left ventricular wall thinning and scar extension. Infarcted hearts of PTH-treated mice revealed increased numbers of CD45(+)/CD34(+) progenitor cells as well as an upregulation of VEGF-A mRNA associated with increased neovascularization and cell survival. CONCLUSIONS: PTH application after MI increases migration of angiogenic CD45(+)/CD34(+) progenitor cells to the ischaemic heart, which may attenuate ischaemic cardiomyopathy. As PTH is already used in patients with osteoporosis, our findings may have a direct impact on the initiation of clinical studies in patients with ischaemic heart disease.

Of rodents and humans: a light microscopic and ultrastructural study on cardiomyocytes in pulmonary veins.

Cardiomyocytes in pulmonary veins (PVs) have been reported in rodents and humans. In humans they were related to atrial arrhythmias, including atrial fibrillation (AF). To investigate histological similarities and differences in PV cardiomyocyte localization and distribution, we performed comparative light and electron microscopic studies on humans, rats and mice, and generated a transgenic mouse strain. Results on mice (C57BL/6 and BALBc) and rats (Wistar) revealed that cardiomyocytes regularly extend from the hilus along venous vessels into the lung tissue surrounding individual intrapulmonary veins of varying diameters (70-250 microm). The cardiomyocytes showed the ultrastructure of a normal working myocardium with intact intercalated discs and tightly packed contractile filaments. In both lung and hilus cardiomyocytes were localized either close to the basal lamina of the endothelium or separated from it by smooth muscle cells and/or collagen fibres. In humans (autopsies, n=20) extrapericardiac cardiomyocytes were only found in 23 out of 78 veins and showed an incomplete sleeve at the lung hilus. In addition, cardiomyocytes occurred significantly more often in right than in left veins, however, never in intrapulmonary veins. We discuss the hypothesis that the variance in distribution of PV cardiomyocytes in humans and rodents might reflect the difference in pathogenesis and development of AF.

Subtyping Of Triple Negative Breast Carcinoma On The Basis Of RTK Expression.

Background: "Triple-negative breast cancers" (TNBC) comprise a heterogeneous group of about 15% of invasive BCs lacking the expression of estrogen and progesterone receptors (ER, PR) and the expression of HER2 (ERBB2) and are therefore no established candidates for targeted treatment options in BC, i.e., endocrine and anti-HER2 therapy. The aim of the present study was to use gene expression profiling and immunohistochemical (IHC) characterization to identify receptor tyrosine kinase (RTK) profiles that would allow patient stratification for the purposes of target-oriented personalized tumor therapy in TNBC. Methods: Twenty-nine cases of TNBC selected according to routine diagnostic IHC/cytogenetic criteria were examined by reverse transcription polymerase chain reaction (RT-PCR). RTK mRNA expression profiles were generated for a total of 31 tumor-relevant biomarkers, mainly belonging to the IGF- and EGF-receptor families but also including biomarkers related to downstream signaling. Protein expression of selected biomarkers was investigated by IHC. Results: Hierarchical cluster analysis revealed a dichotomous differentiation pattern amongst TNBCs. A significant difference in gene expression was observed for 16 of the 31 RTK-associated tumor relevant biomarkers between the two newly identified TNBC subgroups. The findings were verified at the posttranslational level by the IHC data. The RTKs HER4, IGF-1R and IGF-2R and the hormone receptors ER and PR below the IHC detection limit play a central role in the differentiation of the two TNBC subgroups. Observed survival was reported as Kaplan-Meier estimates and point towards an improved survival of patients with RTK-high with superior three-year survival rate of 100% compared to RTK-low gene signatures with superior three-year survival rate of 60% (log-rank test, p-value = 0.022). Conclusion: Gene-expression and IHC analysis of the EGF and IGF receptor families and biomarkers associated with downstream signaling point to the existence of two distinct TNBC subtypes. The RTKs HER4, IGF-1R, IGF-2R and the hormone receptors ER and PR appear to be of particular importance here. Based on survival analysis the differentiation of TNBC with RTK-high and RTK-low gene signatures seems to be of prognostic relevance. Additionally, correlation analysis of the relationship between RTKs and ER suggests co-regulatory mechanisms that may have potential significance in new therapeutic approaches.

Expression of phosphorylated estrogen receptor beta is an independent negative prognostic factor for pancreatic ductal adenocarcinoma.

PURPOSE: The role of estrogen receptor beta (ER-ß) expression in pancreatic ductal adenocarcinoma (PDAC) is largely unknown. Ligand-independent phosphorylation and activation of ER-ß may play a relevant role in the IL-6/STAT3 signaling pathway and, as a result, in tumor progression. Here, we examined the effect of ER-ß, phosphorylated ER-ß (pER-ß), STAT3, phosphorylated STAT3 (pSTAT3) and IL-6 expression on the overall and recurrence-free survival in a cohort of patients with resected PDAC. METHODS: We identified 175 patients who underwent pancreatic resection for PDAC. Tissue microarrays were constructed from the archival tumor specimens. These were stained with specific antibodies for the above molecules. The expression of the markers was then correlated with clinicopathological parameters and survival analysis was performed. RESULTS: High nuclear expression of ER-ß was found in 61.7% and pER-ß in 80.6% of the tumors. STAT3 was expressed in 54.3% of the tumor samples, pSTAT3 in 68% and IL-6 in 76.6%. The median overall survival for patients with low pER-ß expression was 29 months, whereas for patients with high pER-ß expression was 15.1 months (p = 0.016). Multivariate analysis revealed that pER-ß expression was an independent factor correlating with shorter overall survival (hazard ratio 1.9; p = 0.013) and disease-free survival (hazard ratio 1.9; p = 0.029). CONCLUSIONS: Expression of pER-ß constitutes an independent prognostic marker for PDAC and is correlated with poor prognosis. These data may help in identifying novel drug targets in PDAC and patients who could benefit from additional therapeutic regimens, including selective estrogen receptor modulators.

G-CSF administration after myocardial infarction in mice attenuates late ischemic cardiomyopathy by enhanced arteriogenesis.

Granulocyte-colony stimulating factor (G-CSF) has been shown to improve cardiac function after myocardial infarction (MI) by bone marrow cell mobilization and by protecting cardiomyocytes from apoptotic cell death. However, its role in collateral artery growth (arteriogenesis) has not been elucidated. Here, we investigated the effect of G-CSF on arteriolar growth and cardiac function in a murine MI model. Mice were treated with G-CSF (100 microg/kg/day) directly after MI for 5 consecutive days. G-CSF application resulted in a significant increase of circulating mononuclear cells expressing stem cell markers. Arterioles in the border zone of infarcted myocardium showed an increased expression of ICAM-1 accompanied by an accumulation of bone marrow derived cells and a pronounced proliferation of endothelial and smooth muscle cells. Histology of G-CSF treated mice revealed a lower amount of granulation tissue (67.8 vs. 84.4%) associated with a subsequent reduction in free LV wall thinning and scar extension (23.1 vs. 30.8% of LV). Furthermore, G-CSF treated animals showed a significant improvement of post-MI survival (68.8 vs. 46.2%). Pressure-volume relations revealed a partially restored myocardial function at day 30 (EF: 32.5 vs. 17.2%). Our results demonstrate that G-CSF administration after MI stimulates arteriogenesis and attenuates ischemic cardiomyopathy after MI.

Comparison of color-Doppler and qualitative and quantitative strain-elastography for differentiation of thyroid nodules in daily practice.

OBJECTIVE: Strain-elastography provides a new ultrasound-based method that can offer information about the stiffness of thyroid nodules as an indicator of malignancy. The aim of our study was to compare the utility of color-Doppler and strain-elastography in differentiating between benign and malignant nodules. DESIGN AND METHODS: 77 thyroid nodules (70 benign and 7 malignant) from 70 unselected patients (48 female/22 male, mean age 49.7±14.3 years) were evaluated with color-Doppler and elastography based on a five-scale elastogram score for qualitative elastography and strain ratio for quantitative elastography. As reference tissue we chose normal thyroid tissue [strain ratio a (SR a)] and cervical muscles [strain ratio b (SR b)]. The cytological or histological results were used as a reference standard. Diagnostic performances of qualitative and quantitative elastography were compared using ROC curves. RESULTS: Vascularization score 3 or 4 was associated with malignancy (p=0.024) as well as elastogram score 4 or 5 (p=0.070, n.s.s.). SR a was indicatively higher and SR b lower in the group of malignant nodules (p=0.065 and p=0.246, n.s.s.). The best cut-off points predicting malignancy were 3.32 for SR a (66.7% sensitivity, 83.3% specificity) and 0.10 for SR b (71.4% sensitivity, 67.1% specificity). CONCLUSION: In our study, the accuracy of elastography did not surpass other sonographic parameters in differentiating thyroid nodules. The technique can play a role as a supplementary parameter in assessment of malignancy to improve diagnostic efficacy. The best parameter is SR a, but SR b can serve as an alternative if SR a is not assessable.

Perivascular Mast Cells Govern Shear Stress-Induced Arteriogenesis by Orchestrating Leukocyte Function.

The body has the capacity to compensate for an occluded artery by creating a natural bypass upon increased fluid shear stress. How this mechanical force is translated into collateral artery growth (arteriogenesis) is unresolved. We show that extravasation of neutrophils mediated by the platelet receptor GPIbα and uPA results in Nox2-derived reactive oxygen radicals, which activate perivascular mast cells. These c-kit(+)/CXCR-4(+) cells stimulate arteriogenesis by recruiting additional neutrophils as well as growth-promoting monocytes and T cells. Additionally, mast cells may directly contribute to vascular remodeling and vascular cell proliferation through increased MMP activity and by supplying growth-promoting factors. Boosting mast cell recruitment and activation effectively promotes arteriogenesis, thereby protecting tissue from severe ischemic damage. We thus find that perivascular mast cells are central regulators of shear stress-induced arteriogenesis by orchestrating leukocyte function and growth factor/cytokine release, thus providing a therapeutic target for treatment of vascular occlusive diseases.

ALK expression is absent in pancreatic ductal adenocarcinoma.

PURPOSE: It has not yet been clearly defined whether anaplastic lymphoma kinase (ALK) expression can be detected in pancreatic ductal adenocarcinoma (PDAC). METHODS: Within a retrospective study, archival PDAC surgical specimens were screened for ALK expression in tumor and normal tissue by immunohistochemistry (IHC) with the use of a specific ALK detection kit on a tissue microarray (TMA). RESULTS: PDAC tumor tissue was available from 99 resected cases: fifty-eight out of 99 patients (59 %) had nodal-positive disease, and 80 patients (81 %) had pT3 tumors. Forty-nine patients underwent R0 resection, and in 48 cases, resection status was classified R1. Regarding ALK expression, five cases showed faint immunoreactivity on TMA, which was negative on whole mount sections. All other 94 cases showed no ALK expression. CONCLUSION: In 99 PDAC cases, no ALK expression was detected by IHC; ALK thus may not serve as a relevant drug target in PDAC.

Antidiabetic gliptins in combination with G-CSF enhances myocardial function and survival after acute myocardial infarction.

BACKGROUND: Medical stimulation of endogenous progenitor cell circulation may serve as a new therapeutic tool for treatment of acute myocardial infarction. We analyzed the effects of antidiabetic gliptins plus GCSF (granulocyte colony stimulating factor) on myocardial regeneration after myocardial infarction in a mouse model. METHODS AND RESULTS: After surgical LAD-ligation (left anterior descending artery), Sitagliptin/Vildagliptin was applied yielding sufficient blood levels verified by mass spectrometry and significantly reducing activity of dipeptidyl peptidase (DPP) IV. GCSF or saline was administered intraperitoneally for 6 days. We assessed stem cell mobilization and homing (flow cytometry), infarct size (histology), neovascularization and cellular proliferation (immunohistology), heart function (Millar tip catheterization) and survival (Kaplan-Meier-curves). Gliptins±GCSF administration increased mobilization and cardiac homing of bone-marrow derived stem cells by stabilization of cardiac SDF1 (stromal cell-derived factor). For Sitagliptin, it could be shown that resident cardiac stem cells were stimulated, neovascularization was enhanced and cardiac remodeling was reduced. These effects finally improved myocardial function and increased survival for both gliptins. Although gliptins as a mono therapy lead to remarkable effects in a dose dependent manner and were superior to G-CSF mono-therapy, dual application of GCSF and gliptins revealed the best results. Since both gliptins yielded comparable effects concerning stem cell homing, cardiac function and survival, we suggest a class-effect of DPP-IV-inhibitors. CONCLUSIONS: Thus, gliptins+GCSF and in high concentrations even as mono therapy have beneficial effects on cardiac regeneration after myocardial infarction beyond its anti-diabetic potential.

Cancer of unknown primary originating from oropharyngeal carcinomas are strongly correlated to HPV positivity.

Human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma (OSCC) has been identified as a distinct entity within squamous cell carcinoma of the head and neck. It is associated with special characteristics and is preponderantly restricted to palatial tonsils and base of tongue. These primary locations have for long been associated with the clinical situation of cancer of unknown primary (CUP). In order to investigate the putative relationship between CUP and HPV, we investigated 26 patients who initially presented as CUP and were finally diagnosed with carcinomas of these two locations. Twenty-one cases proved to be positive for high-risk HPV. Primary carcinomas were small and frequently presented in a submucosal location. HPV-positive carcinomas, presented more often in women, showed atypical basaloid differentiation and correlated to cystic lymph node metastases. This study demonstrates an over-representation of HPV-associated OSCC in patients who were initially diagnosed with CUP. This finding indicates a strong relationship between HPV-association and CUP in OSCC. The frequent manifestation as CUP is presumably caused by the unusual predisposition for small size and submucosal location combined with early lymphatic metastization. In order not to miss clinically occult carcinomas, consequent interdisciplinary cooperation in combination with meticulous histological workup is mandatory.

ß-Catenin mutations in 2 nested stromal epithelial tumors of the liver--a neoplasia with defective mesenchymal-epithelial transition.

Nested stromal epithelial tumor of the liver is a rare neoplasm of early childhood and adolescence with a characteristic nested morphology of spindle and epithelioid cells. Histogenesis and pathogenesis of this neoplasm are, however, still unclear. Because the characteristic nested morphology with spindle mesenchymal and epithelioid cells is suggestive of altered mesenchymal-epithelial transition and ß-catenin mutations are rather common in other liver tumors such as hepatoblastomas, we investigated the ß-catenin gene in 2 nested stromal epithelial tumors of the liver and analyzed additional factors involved in mesenchymal-epithelial transition, such as E-cadherin, vimentin, c-Met, TWIST, SNAIL, and SLUG by molecular genetic and immunohistochemical methods. Mutation analysis of both cases revealed large deletions in exon 3 of the ß-catenin gene (155 and 228 base pairs), resulting in an accumulation of ß-catenin in the cytoplasm and nuclei of tumor cells, as evidenced by immunohistochemistry. The expression of the mesenchymal-epithelial transition factors SNAIL, SLUG, TWIST, c-Met, vimentin, and ß-catenin was generally increased, whereas E-cadherin was decreased. Morphological and immunohistochemical analysis, however, showed a variable expression pattern of various epithelial and mesenchymal markers both in the spindle and epithelioid cell compartments of the tumors, thus illustrating the transitional status of the tumor cells. In conclusion, our data clearly identify protein stabilizing mutations of the ß-catenin gene as a common feature of nested stromal epithelial tumors of the liver, similarly as in hepatoblastomas. Therefore, nested stromal epithelial tumors of the liver may be regarded as a variant of hepatoblastoma, despite differing from it in clinical and morphological aspects. The characteristic epithelioid-spindle morphology along with the incomplete epithelial differentiation proposes impaired mesenchymal-epithelial transition as a possible pathogenetic mechanism of this rare tumor. However, because only 2 cases were studied, this hypothesis awaits further validation.

The cardioprotective effects of parathyroid hormone are independent of endogenous granulocyte-colony stimulating factor release.

AIMS: Parathyroid hormone (PTH) administration after myocardial infarction (MI) is known to attenuate ischaemic cardiomyopathy. This effect mainly resulted from an increase in mobilization and homing of CD34+/CD45+ cells into the ischaemic myocardium. PTH-related stem cell mobilization was shown to be related to endogenous granulocyte-colony stimulating factor (G-CSF) release. The aim of our study is to determine the role of G-CSF on the cardioprotective effects of PTH. METHODS AND RESULTS: G-CSF +/+ (C57BL/6) and G-CSF -/- mice were treated with PTH for 6 days after inducing a MI. The myocardial homing factor stromal cell-derived factor-1 (SDF-1) was analysed on day 2 with enzyme-linked immunosorbent assay. Stem cell populations in peripheral blood and hearts were examined by FACS on days 6 and 2, respectively. Cardiac function and immunohistochemistry were investigated on day 6 and day 30. PTH treatment resulted in a significant increase in CD45+/CD34+ cells in peripheral blood in G-CSF +/+ but not in G-CSF -/- mice. However, a significant increase in SDF-1 and enhanced migration of CD45+/CD34+ cells into the ischaemic myocardium was revealed after PTH administration in both G-CSF +/+ and G-CSF -/- mice. Enhanced stem cell homing was associated with improved cardiac function and post-MI survival after PTH treatment. Furthermore, infarct size, wall thickness, and neovascularization showed a significant improvement in both groups 30 days after MI. CONCLUSION: The cardioprotective effects of PTH were shown to be independent of endogenous G-CSF release and therefore from stem cell mobilization. This puts more emphasis on the role of stem cell homing into ischaemic myocardium.