Before firm adhesion, leukocytes roll slowly along the walls of small venules at velocities ranging from 0.7 to 36% of mean blood flow velocity. To investigate the nature of the adhesive process underlying leukocyte rolling, synthetic (dextran sulfate) and naturally occurring sulfated polysaccharides (heparin, chondroitin sulfates, keratan sulfate, and heparan sulfate) were infused via glass micropipettes into the lumen of small venules (20-60 microns diam) of the rabbit mesentery. Leukocyte rolling was observed and quantified using both transmitted light and incident fluorescence intravital microscopy. Rolling leukocytes accounted for 27-80% of total leukocyte flux, exhibiting a wide range of individual velocities (0.01-0.84 mm/s) with a mean value of 4% of centerline velocity. Dextran sulfate (Mr 500,000) inhibited leukocyte rolling very effectively [half-effective concentration (ED50) approximately 10 micrograms/ml] and was able to almost completely abolish rolling at 500 micrograms/ml. Heparin (ED50 approximately 50 micrograms/ml), chondroitin 6-sulfate C (ED50 approximately 500 micrograms/ml), and heparan sulfate (ED50 approximately 5 mg/ml) also reduced leukocyte rolling. At 5 mg/ml, chondroitin 4-sulfate B (dermatan sulfate) was marginally effective, but chondroitin 4-sulfate A and keratan sulfate were ineffective. The present data suggest that an adhesion receptor-ligand system distinct from the leukocyte integrins may be underlying transient leukocyte adhesion (rolling). Endothelial glycoproteins or proteoglycans containing sulfated side chains may be involved in mediating this adhesive process.
Leukocyte distribution was studied in 58 arterioles and capillaries constituting eight networks in the fascia adjacent to the rabbit tenuissimus muscle. Fluorescence video microscopy (acridine red) and digital image processing were used to visualize the leukocytes. Leukocyte concentration in the transverse arterioles leaving the muscle proper to irrigate the fascia was significantly elevated to 143 +/- 13% (mean +/- SE) of the systemic count. Leukocytes were found to further accumulate along the arteriolar tree, reaching 244 +/- 16% of systemic in the most remote branches. Leukocyte accumulation in the connective tissue can be calculated to lead to a decrease of leukocyte concentration in muscle capillaries to 89% of the systemic value. This shunting effect may be important in preventing leukocyte-induced capillary obstruction. At individual arteriolar bifurcations, leukocytes were found to preferentially enter the downstream branch with higher flow rate (regression coefficient 1.11 +/- 0.04, n = 100). This preferential distribution is quantitatively sufficient to account for the observed leukocyte distribution in the network.
Background: Cohesive visualization and interpretation of hyperdimensional, large-scale -omics data is an ongoing challenge, particularly for biologists and clinicians involved in current highly complex sequencing studies. Multivariate studies are often better suited towards non-linear network analysis than differential expression testing. Here, we present PRESTO, a 'PREdictive Stochastic neighbor embedding Tool for Omics', which allows unsupervised dimensionality reduction of multivariate data matrices with thousands of subjects or conditions. PRESTO is intuitively integrated into an interactive user interface that helps to visualize the multidimensional patterns in genome-wide transcriptomic data from basic science and clinical studies.Results: PRESTO was tested with multiple input omics' platforms, including microarray and proteomics from both mouse and human clinical datasets. PRESTO can analyze up to tens of thousands of genes and shows no increase in processing time with a large number of samples or patients. In complex datasets, such as those with multiple time points, several patient groups, or diverse mouse strains, PRESTO outperformed conventional methods. Core co-expressed gene networks were intuitively grouped in clusters, or gates, after dimensionality reduction and remained consistent across users. Networks were identified and assigned to physiological and pathological functions that cannot be gleaned from conventional bioinformatics analyses. PRESTO detected gene networks from the natural variations among mouse macrophages and human blood leukocytes. We applied PRESTO to clinical transcriptomic and proteomic data from large patient cohorts and detected disease-defining signatures in antibody-mediated kidney transplant rejection, renal cell carcinoma, and relapsing acute myeloid leukemia (AML). In AML, PRESTO confirmed a previously described gene signature and found a new signature of 10 genes that is highly predictive of patient outcome. Conclusions: PRESTO offers an important integration of powerful bioinformatics tools with an interactive userinterface that increases data analysis accessibility beyond bioinformaticians and 'coders'. Here, we show that PRESTO out performs conventional methods, such as DE analysis, in multi-dimensional datasets and can identify biologically relevant co-expression gene networks. In paired samples or time points, co-expression networks could be compared for insight into longitudinal regulatory mechanisms. Additionally, PRESTO identified disease-specific signatures in clinical datasets with highly significant diagnostic and prognostic potential.
The regulation of integrin expression and function controls interactions of immune cells and targets their trafficking locally and systemically. We show here that the tyrosine phosphatase SHP-1 is required for lymphocyte surface expression of the intestinal immune response-associated integrin β 7 , but not for β 1 or β 2 integrins. Viable motheaten mice deficient for SHP-1 have less β 7 on T cells and lack β 7 on B cells. SHP-1 function is targeted in B cells by the B cell specific lectin CD22 (Siglec-2), suggesting a potential role for CD22 in β 7 expression. CD22-deficiency on B cells phenocopies the effects of SHP-1 haplodeficiency. Mechanistically, we show that SHP-1 suppresses β 7 endocytosis: internalization of β 7 but not β 1 integrin is accelerated in SHP-1 +/and CD22 -/-B cells. Moreover, mutations in CD22 cytoplasmic SHP1-binding ITIM sequences reduce α 4 β 7 comparably, and loss of CD22 lectin activity has an intermediate effect suggesting a model in which the CD22 ITIM sequences recruit SHP-1 to control β 7 expression. Integrin α 4 β 7 selectively contributes to cell interactions in intestinal immunity. Consistent with this, CD22 deficient and SHP-1 +/-B cells display reduced β 7 -dependent homing to gut associated Peyer's patches (PP); and CD22-deficiency impairs intestinal but not systemic antibody responses and delays clearance of the gut pathogen rotavirus. The results define a novel role for SHP-1 in the differential control of leukocyte integrins and an unexpected integrin β 7 -specific role for CD22-SHP-1 interplay in mucosal immunity. 1470 1887 3759 WT CD22 R130E
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