Modulation of endothelial cell surface glycoconjugate expression by organ-derived biomatrices

Cell surface molecules play an important role in cellular communication, migration, and adherence. Here, we show the effect of organ-derived biomatrices on endothelial cell surface glycosylation. Five different lectins (with and without neuraminidase treatment) have been used as probes in an enzyme-linked lectin assay to quantitatively detect glycoconjugates on endothelial cells (BAEC) grown on tissue culture plastic or biomatrices isolated from bovine lung, liver, and kidney. BAEC generally exhibit strong binding of concanavalin A (Con A), Ricinus communis agglutinin I (RCA-I), wheat germ agglutinin (WGA), and soybean agglutinin, and peanut agglutinin after neuraminidase pretreatment of cells (Neu-SBA and Neu-PNA), while SBA and PNA consistently bind weakly to BAEC. BAEC grown on organ-derived biomatrices exhibit significantly altered binding intensities of Con A, RCA-I, WGA, and Neu-PNA: BAEC cultured on lung- or kidney-derived biomatrices express significantly stronger binding affinities for Con A and RCA-I than BAEC grown on liver-derived biomatrix or tissue culture plastic. In contrast, BAEC binding of WGA and PNA (after treatment of cells with neuraminidase) is significantly reduced when BAEC are grown on liver- or kidney-derived biomatrix. Quantitative lectin immunogold electron microscopy reveals consistently stronger lectin binding over nuclear regions compared to junctional regions between neighboring cells. These results indicate that extracellular matrix components regulate endothelial cell surface glycoconjugate expression, which determines cellular functions, e.g., preferential adhesion of lymphocytes or metastatic tumor cells.     

Eph receptor and ephrin ligand-mediated interactions during angiogenesis and tumor progression

Eph receptors comprise the largest family of receptor tyrosine kinases. They are classified into an A family and a B family on the basis of the characteristic properties of the corresponding ephrin ligands which are either GPI-anchored peripheral membrane molecules (A class ephrins) or transmembrane molecules (B class ephrins). Eph receptors and ephrin ligands were originally identified as neuronal pathfinding molecules. Yet, gene targeting experiments in mice have identified the EphB/ephrinB system as critical and rate-limiting determinant of arterio-venous differentiation during embryonic vascular development. Identification of vascular EphB/ephrinB functions has in the last few years stimulated two emerging fields of vascular biology research, namely (1) the molecular analysis of the structural and functional mechanisms of arterio-venous differentiation, and (2) the molecular study of the commonalities between vascular and neuronal guidance and patterning mechanisms. This review summarizes the current understanding of vascular Eph receptor and ephrin ligand functions and provides an overview of emerging roles of the Eph/ephrin system in controlling tumor and vascular functions during tumorigenesis and tumor progression.     

Integration of Endothelial Cells in Multicellular Spheroids Prevents Apoptosis and Induces Differentiation

Single endothelial cells (EC) seeded in suspension culture rapidly undergo apoptosis. Addition of survival factors, such as VEGF and FGF-2, does not prevent apoptosis of suspended EC. However, when cells are allowed to establish cell–cell contacts, they become responsive to the activities of survival factors. These observations have led to the development of a three-dimensional spheroid model of EC differentiation. EC spheroids remodel over time to establish a differentiated surface layer of EC and a center of unorganized EC that subsequently undergo apoptosis. Surface EC become quiescent, establish firm cell–cell contacts, and can be induced to express differentiation antigens (e.g., induction of CD34 expression by VEGF). In contrast, the unorganized center spheroid cells undergo apoptosis if they are not rescued by survival factors. The responsiveness to the survival factor activities of VEGF and FGF-2 was not dependent on cell shape changes since it was retained after cytochalasin D treatment. Taken together, these findings characterize survival factor requirements of unorganized EC and indicate that polarized surface EC differentiate to become independent of exogenous survival factors. Furthermore, they demonstrate that spheroid cell culture systems are useful not just for the study of tumor cells and embryonic stem cells but also for the analysis of differentiated functions of nontransformed cells.     

Quantitative mathematical evaluation of growth studies from the viewpoint of exogenously or genetically influenced variability

Firstly, the ideas are sketched which serve as the basis for the phenomenologic-mathematical kind of modeling of the body length growth process of man. For proving the biological relevance of the spurts analyzed by numerical procedures one has to consider the social and the biological circumstances in which the growth process takes place. An analysis of longitudinal data series of the body length of (monozygotic) twins will give further hints to the possible meaning of the growth spurts by way of separation of exogenous and of genetic determined endogenous agents on the growth process. The data available for our examinations cover the time interval of the praepubertal and the postpuberal development as well as the time interval in which the puberal growth spurt takes place. The way to proceed in evaluation these time series will be presented.     

Phenomenologic-mathematical modeling of the human-body-length growth through the analysis of growth periods and their quantitative-analytical registration

The body height growth (of masculine beings) was modelled in a phenomenologic-mathematical manner by partitioning the time course of measured growth curve in parts every of which corresponds to a separated growth period. This partitioning was reached in a natural way so that a superposition of the single spurts yields the whole measured course. Every growth batch will be described in its time course by one term of inverse tangent function. The biological meaning and an explanation of the succession of the growth spurt as an effect of control circuits need further exploratory work. For detailed statements on acceleration phenomena concerning the body height growth this analysis gives possibilities for comparing the single growth spurts of the mean growth process of two populations in question. For measured values given by BROCK (1954) and SALZLER (1967) there are five growth periods in the time intervall reaching from time of conception until the end of the first year. Comparing the mathematical functions of the corresponding growth spurts for these two groups one can conclude that the second spurt (fetal spurt) is responsible for an increase of birth body height and the fourth for an increase of body height in the suckling age of the latter group against the former one.     

Multiple angiopoietin recombinant proteins activate the Tie1 receptor tyrosine kinase and promote its interaction with Tie2

The Tie1 receptor tyrosine kinase was isolated over a decade ago, but so far no ligand has been found to activate this receptor. Here, we have examined the potential of angiopoietins, ligands for the related Tie2 receptor, to mediate Tie1 activation. We show that a soluble Ang1 chimeric protein, COMP-Ang1, stimulates Tie1 phosphorylation in endothelial cells with similar kinetics and angiopoietin dose dependence when compared with Tie2. The phosphorylation of overexpressed Tie1 was weakly induced by COMP-Ang1 also in transfected cells that do not express Tie2. When cotransfected, Tie2 formed heteromeric complexes with Tie1, enhanced Tie1 activation, and induced phosphorylation of a kinase-inactive Tie1 in a ligand-dependent manner. Tie1 phosphorylation was also induced by native Ang1 and Ang4, although less efficiently than with COMP-Ang1. In conclusion, we show that Tie1 phosphorylation is induced by multiple angiopoietin proteins and that the activation is amplified via Tie2. These results should be important in dissecting the signal transduction pathways and biological functions of Tie1.     

Recruitment of human cord blood-derived endothelial colony-forming cells to sites of tumor angiogenesis

Background aimsEndothelial progenitor cells (EPCs) specifically home to sites of malignant growth, rendering them attractive for anti-cancer therapies. Data are conflicting on the phenotype and quantitative contribution toward tumor angiogenesis based on differing culture assays to outgrow EPCs. To evaluate the origin and early phenotype of EPCs and to define a population with enhanced tumor-targeting capacity, we evaluated a hierarchy of cord blood-derived EPCs modeling the multi-step nature of tumor homing.MethodsCD34⁺ mononuclear cells were isolated from fresh cord blood and cultured to derive endothelial colony-forming cells (ECFCs). Human umbilical vein endothelial cells (HUVECs) served as control. Using intra-vital microscopy, the recruitment was analyzed in mice bearing C6 xenografts. Adhesion, migration, transmigration and differentiation were further addressed.ResultsWithin the primary passage, ECFCs underwent a rapid maturation from a CD45⁺ and CD31⁺ phenotype to a CD45^? and endothelial marker positive phenotype. Assessing in vivo tumor recruitment, ECFCs had the highest activity in all steps analyzed. In vitro, ECFCs demonstrated significantly higher adhesion under static and flow conditions. Similarly, ECFCs exhibited highest migratory and trans-migratory activity toward tumor-conditioned medium. On subcutaneous implantation, only ECFCs formed blood vessels covered with perivascular cells, similar to HUVECs.ConclusionsOur study indicates that ECFCs emerge from a CD45⁺ and CD31⁺ progenitor and rapidly mature in culture. ECFCs have a significantly higher potential for tumor targeting than non-cultured CD34⁺ cells and HUVECs. They are ideal candidates for future cell-based anti-cancer therapies.     

Evaluation of Uric Acid as a Prognostic Blood-Based Marker in a Large Cohort of Pancreatic Cancer Patients

Background

Recently, chemical blood parameters gain more attraction as potential prognostic parameters in pancreatic cancer (PC). In the present study we investigated the prognostic relevance of the uric acid (UA) level in blood plasma at the time of diagnosis for overall survival (OS) in a large cohort of patients with PC.

Patients and Methods

Data from 466 consecutive patients with ductal adenocarcinoma of the pancreas were evaluated retrospectively. Overall survival (OS) was analysed using the Kaplan-Meier method. To further evaluate the prognostic significance of the UA level, univariate and multivariate Cox regression models were calculated.

Results

None of the clinicopathological parameters (tumour grade, clinical stage, age, CA19-9 level, Karnofski Index (KI) or surgical resection) except gender was associated with UA level. In univariate analysis we observed the elevated UA level (<5.1 versus ≥5.1 mg/dl, p = 0.017) as poor prognostic factor for OS. In the multivariate analysis that included age, gender, tumour grade, tumour stage, surgical resection, CA19-9 level, the KI and UA level we confirmed the UA level as independent prognostic factor for OS (HR = 1.373%; CI = 1.077–1.751; p = 0.011).

Conclusion

In conclusion, we identified the UA level at time of diagnosis as an independent prognostic factor in PC patients. Our results indicate that the UA level might represent a novel and useful marker for patient stratification in PC management.