Surface molecules regulating rolling and adhesion to endothelium of neutrophil granulocytes and MDA-MB-468 breast carcinoma cells and their interaction

The extravasation of leukocytes and tumor cells is a multi-step process with the involvement of various adhesion molecules depending on the three steps rolling, adhesion, and diapedesis. We have developed an in vitro model, by which we investigated the rolling and adhesion of neutrophil granulocytes and MDA-MB-468 human breast carcinoma cells to lung endothelial cells under physiological flow-conditions. We found that norepinephrine had an inhibitory function on the fMLP-promoted adhesion of neutrophil granulocytes due to a down-regulation of β2-integrin. Furthermore, neutrophil granulocytes serve as linking cells for the interaction of the MDA-MB-468 cells with the endothelium, which are both β2-integrin negative, but express the β2-integrin ligand ICAM-1. In addition, we show here that N-cadherin is up-regulated on the endothelial cells and on neutrophil granulocytes in response to fMLP. This up-regulation resulted in a significant increase of adherent MDA-MB-468 cells, which are also N-cadherin positive. Received 3 September 2007; received after revision 17 October 2007; accepted 22 October 2007     

Podocalyxin enhances the adherence of cells to platelets

Podocalyxin (PODXL) is a mucin protein of the CD34 family expressed in kidney glomerular podocytes, vascular endothelium, progenitor bone marrow and tumor cells. It is assumed that PODXL plays an anti-adherent role in kidney podocytes. CHO cells stably expressing human PODXL (CHO-PODXL) or human tumor cells (Tera-1) inherently expressing PODXL showed increased adherence to platelets. The adherence of cells was inhibited (70%) by blockers of platelet P-selectin, prevented by the soluble ectodomain of human PODXL (PODXL-Δ) or by the arginine-glycine-aspartate (RGDS) peptide and partially impeded by inhibition of integrin αVβ3/αVβ5, suggesting a coordinated action of P-selectin and integrins. Colocalization of platelet P-selectin and PODXL expressed on CHO cells was demonstrated by confocal immunofluorescence. No adherence to platelets was observed when PODXL was expressed in glycomutant CHO cells deficient in sialic acid. Received 14 August 2007; received after revision 12 September 2007; accepted 13 September 2007     

Developing pharmacological therapies for Alzheimer disease

Alzheimer disease (AD), while chronic and progressive with an average progression of 7 – 10 years, is both multifactorial and heterogeneous. Thus, AD offers a large window of opportunity and a large number of therapeutic targets to inhibit it. The selection of a therapeutic target, however, is one of the biggest challenges in developing a pharmacological treatment of this multifactorial disease. Inhibition of a pivotal downstream event is likely to benefit more patients than inhibition of an upstream event in AD pathogenesis. Neurofibrillary degeneration of abnormally hyperphosphorylated tau offers such a pivotal therapeutic target. Abnormal hyperphosphorylation of tau and not its aggregation into filaments appears to be the most deleterious step in neurofibrillary degeneration. Tau can be abnormally hyperphosphorylated by downregulation of protein phosphatase-2A activity or by upregulation of more than one tau kinase. Restoration of the phosphatase activity which is downregulated in AD brain or inhibition of GSK-3β and cdk5, which are required for AD-type abnormal hyperphosphorylation of tau, are among the most promising therapeutic strategies.     

Alcohol dehydrogenase 2 is a major hepatic enzyme for human retinol metabolism

The metabolism of all-trans- and 9-cis-retinol/ retinaldehyde has been investigated with focus on the activities of human, mouse and rat alcohol dehydrogenase 2 (ADH2), an intriguing enzyme with apparently different functions in human and rodents. Kinetic constants were determined with an HPLC method and a structural approach was implemented by in silico substrate dockings. For human ADH2, the determined K_m values ranged from 0.05 to 0.3 μM and k_cat values from 2.3 to 17.6 min⁻¹, while the catalytic efficiency for 9-cis-retinol showed the highest value for any substrate. In contrast, poor activities were detected for the rodent enzymes. A mouse ADH2 mutant (ADH2Pro47His) was studied that resembles the human ADH2 setup. This mutation increased the retinoid activity up to 100-fold. The K_m values of human ADH2 are the lowest among all known human retinol dehydrogenases, which clearly support a role in hepatic retinol oxidation at physiological concentrations. Received 12 October 2006; received after revision 6 December 2006; accepted 8 January 2007     

The cold case: Are rhinoviruses perfectly adapted pathogens?

Rhinoviruses, which cause common cold, belong to the Picornaviridae family, small non-enveloped viruses (diameter 15-30 nm) containing a single-stranded RNA genome (about 7 kb). Over 100 different rhinoviral serotypes have been identified thus far, establishing rhinoviruses as the most diverse group of Picornaviridae. Based on receptor binding properties, rhinoviruses are divided into two classes: the major group binding to intracellular adhesion molecule-1 and the minor group binding to the very low density lipoprotein receptors. Interactions between virus and the receptor molecules cause a conformational change in the capsid, which is a prerequisite for viral uptake. Rhinoviruses trigger a chemokine response upon infection that may lead to exacerbation of the symptoms of common cold, i.e. asthma and inflammation. The following review aims to summarize the knowledge about rhinoviral infections and discusses therapeutical approaches against this almost perfectly adapted pathogen.     

Cellular internalization of proinsulin C-peptide

Proinsulin C-peptide is known to bind specifically to cell membranes and to exert intracellular effects, but whether it is internalized in target cells is unknown. In this study, using confocal microscopy and immunostained or rhodamine-labeled peptide, we show that C-peptide is internalized and localized to the cytosol of Swiss 3T3 and HEK-293 cells. In addition, transport into nuclei was found using the labeled peptide. The internalization was followed at 37°C for up to 1 h, and was reduced at 4°C and after preincubation with pertussis toxin. Hence, it is concluded to occur via an energy-dependent, pertussis toxin-sensitive mechanism and without detectable degradation within the experimental time course. Surface plasmon resonance measurements demonstrated binding of HEK-293 cell extract components to C-peptide, and subsequent elution of bound material revealed the components to be intracellular proteins. The identification of C-peptide cellular internalization, intracellular binding proteins, absence of rapid subsequent C-peptide degradation and apparent nuclear internalization support a maintained activity similar to that of an intracrine peptide hormone. Hence, the data suggest the possibility of one further C-peptide site of action. Received 31 October 2006; received after revision 27 December 2006; accepted 30 December 2006     

SMAD4-deficient intestinal tumors recruit CCR1+ myeloid cells that promote invasion

Inactivation of TGF-beta family signaling is implicated in colorectal tumor progression. Using cis-Apc(+/Delta716) Smad4(+/-) mutant mice (referred to as cis-Apc/Smad4), a model of invasive colorectal cancer in which TGF-beta family signaling is blocked, we show here that a new type of immature myeloid cell (iMC) is recruited from the bone marrow to the tumor invasion front. These CD34(+) iMCs express the matrix metalloproteinases MMP9 and MMP2 and the CC-chemokine receptor 1 (CCR1) and migrate toward the CCR1 ligand CCL9. In adenocarcinomas, expression of CCL9 is increased in the tumor epithelium. By deleting Ccr1 in the background of the cis-Apc/Smad4 mutant, we further show that lack of CCR1 prevents accumulation of CD34(+) iMCs at the invasion front and suppresses tumor invasion. These results indicate that loss of transforming growth factor-beta family signaling in tumor epithelium causes accumulation of iMCs that promote tumor invasion.     

Population-based resequencing of ANGPTL4 uncovers variations that reduce triglycerides and increase HDL

Resequencing genes provides the opportunity to assess the full spectrum of variants that influence complex traits. Here we report the first application of resequencing to a large population (n = 3,551) to examine the role of the adipokine ANGPTL4 in lipid metabolism. Nonsynonymous variants in ANGPTL4 were more prevalent in individuals with triglyceride levels in the lowest quartile than in individuals with levels in the highest quartile (P = 0.016). One variant (E40K), present in approximately 3% of European Americans, was associated with significantly lower plasma levels of triglyceride and higher levels of high-density lipoprotein cholesterol in European Americans from the Atherosclerosis Risk in Communities Study and in Danes from the Copenhagen City Heart Study. The ratio of nonsynonymous to synonymous variants was higher in European Americans than in African Americans (4:1 versus 1.3:1), suggesting population-specific relaxation of purifying selection. Thus, resequencing of ANGPTL4 in a multiethnic population allowed analysis of the phenotypic effects of both rare and common variants while taking advantage of genetic variation arising from ethnic differences in population history.     

Stepwise replication identifies a low-producing lymphotoxin-alpha allele as a major risk factor for early-onset leprosy

Host genetics has an important role in leprosy, and variants in the shared promoter region of PARK2 and PACRG were the first major susceptibility factors identified by positional cloning. Here we report the linkage disequilibrium mapping of the second linkage peak of our previous genome-wide scan, located close to the HLA complex. In both a Vietnamese familial sample and an Indian case-control sample, the low-producing lymphotoxin-alpha (LTA)+80 A allele was significantly associated with an increase in leprosy risk (P = 0.007 and P = 0.01, respectively). Analysis of an additional case-control sample from Brazil and an additional familial sample from Vietnam showed that the LTA+80 effect was much stronger in young individuals. In the combined sample of 298 Vietnamese familial trios, the odds ratio of leprosy for LTA+80 AA/AC versus CC subjects was 2.11 (P = 0.000024), which increased to 5.63 (P = 0.0000004) in the subsample of 121 trios of affected individuals diagnosed before 16 years of age. In addition to identifying LTA as a major gene associated with early-onset leprosy, our study highlights the critical role of case- and population-specific factors in the dissection of susceptibility variants in complex diseases.