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Biomedical subjects

Alan J Young

Publications and source records attributed to Alan J Young.

11 recordsLinked to original sources
PubMed2005

Chemokine receptor CCR7 required for T lymphocyte exit from peripheral tissues.

Lymphocytes travel throughout the body to carry out immune surveillance and participate in inflammatory reactions. Their path takes them from blood through tissues into lymph and back to blood. Molecules that control lymphocyte recruitment into extralymphoid tissues are well characterized, but exit is assumed to be random. Here, we showed that lymphocyte emigration from the skin was regulated and was sensitive to pertussis toxin. CD4(+) lymphocytes emigrated more efficiently than CD8(+) or B lymphocytes. T lymphocytes in the afferent lymph expressed functional chemokine receptor CCR7, and CCR7 was required for T lymphocyte exit from the skin. The regulated expression of CCR7 by tissue T lymphocytes may control their exit, acting with recruitment mechanisms to regulate lymphocyte transit and accumulation during immune surveillance and inflammation.

Animals↗
PubMed2005

A novel mechanism of immune regulation: interferon-gamma regulates retention of CD4 T cells during delayed type hypersensitivity.

The local immune response is characterized by an increase in the rate of entry of lymphocytes from the blood into regional lymph nodes and changes in the output of cells in lymph. While significant data are available regarding the role of inflammation-induced vascular adhesion processes in regulating lymphocyte entry into inflamed tissues and lymph nodes, relatively little is known about the molecular processes governing lymphocyte exit into efferent lymph. We have defined a novel role for lymphatic endothelial cells in the regulation of lymphocyte exit during a delayed type hypersensitivity (DTH) response to mycobacterial purified protein derivative (PPD). Soluble, pro-adhesive factors were identified in efferent lymph concomitant with reduced lymphocyte output in lymph, which significantly increased lymphocyte binding to lymphatic endothelial cells. While all lymphocyte subsets were retained, CD4+ T cells appeared less susceptible than others. Among a panel of cytokines in inflammatory lymph plasma, interferon (IFN)-gamma alone appeared responsible for this retention. In vitro adhesion assays using physiological levels of IFN-gamma confirmed the interaction between recirculating lymphocytes and lymphatic endothelium. These data demonstrate a new level of immune regulation, whereby the exit of recirculating lymphocytes from lymph nodes is selectively and sequentially regulated by cytokines in a manner equally as complex as lymphocyte recruitment.

Animals↗
PubMed2003

Microangiectasias: structural regulators of lymphocyte transmigration.

The migration of lymphocytes into inflammatory tissue requires the migrating cell to overcome mechanical forces produced by blood flow. A generally accepted hypothesis is that these forces are overcome by a multistep sequence of adhesive interactions between lymphocytes and endothelial cells. This hypothesis has been recently challenged by results demonstrating wall shear stress on the order of 20 dyn/cm(2) in vivo and infrequent lymphocyte-endothelial adhesion at wall shear stress >1-2 dyn/cm(2) in vitro. Here, we show that lymphocyte slowing and transmigration in the skin is associated with microangiectasias, i.e., focal structural dilatations of microvessel segments. Microangiectasias are inducible within 4 days of the onset of inflammation and lead to a greater than 10-fold local reduction in wall shear stress. These findings support the hypothesis that a preparatory step to lymphocyte transmigration involves structural adaptations in the inflammatory microcirculation.

Animals↗
PubMed2003

Dynamic deformation of migratory efferent lymph-derived cells "trapped" in the inflammatory microcirculation.

The cellular immune response depends on the delivery of lymphocytes from the lymph node to the peripheral site of antigenic challenge. During their passage through the inflammatory microcirculaton, the migratory cells can become transiently immobilized or "trapped" in small caliber vessels. In this report, we used intravital microscopy and temporal area mapping to define the dynamic deformation of efferent lymph-derived mononuclear cells trapped in the systemic inflammatory microcirculation. Mononuclear cells obtained from the efferent lymph draining the oxazolone-stimulated microcirculation were labeled with fluorescent dye and reinjected into the feeding arterial circulation. Intravital video microscopy observed thousands of cells passing through the microcirculation; 35 cells were "trapped" in the oxazolone-stimulated microcirculation. Temporal area maps of the trapped cells demonstrated dramatic slowing and deformation. The cells were trapped in the microcirculation for a median of 8.90 sec (range 5-17 sec) prior to returning to the flow stream. During this period, the cells showed sustained movement associated with both antegrade locomotion (mean cell velocity = 7.92 microm/sec; range 1.16-14.23 microm/sec) and dynamic elongation (median cell length = 73.8 microm; range 58-144 microm). In contrast, efferent lymph-derived cells passing unimpeded through the microcirculation demonstrated rapid velocity (median velocity = 216 microm/sec) and spherical geometry (median diameter = 14.6 microm). Further, the membrane surface area of the "trapped" cells, calculated based on digital image morphometry and corrosion cast scanning electron microscopy, suggested that the fractional excess membrane of the cells in the efferent lymph was significantly greater than previous estimates of membrane excess. These data indicate that transient immobilization of efferent lymph-derived mononuclear cells in the systemic inflammatory microcirculation is rare. When "trapping" does occur, the shape changes and sustained cell movement facilitated by excess cell membrane may contribute to the return of the "trapped cells" into the flow stream.

Adjuvants, Immunologic↗
PubMed2003

Engraftment of human cord blood-derived stem cells in preimmune ovine fetuses after ultrasound-guided in utero transplantation.

OBJECTIVE: The fetal sheep in utero transplantation model has developed into an important tool to study the efficacy of human in utero stem cell transplantation and gene therapy because of similarities in both the scale and development of immunocompetence relative to gestational age. The aim of this study was to determine whether human stem cells can be successfully transplanted to the first-trimester ovine fetus by use of a newly developed minimally invasive technique. STUDY DESIGN: Human cord blood-derived, CD34(+)-enriched stem cells were injected into the peritoneal cavity of 45- to 60-day-old ovine fetuses by using ultrasound-guided transabdominal percutaneous needle puncture. Engraftment was determined 1 to 3 months after birth by flow cytometry with use of human-specific anti-CD45 antibodies. RESULTS: In contrast to previous studies that used surgical techniques, we observed a fetal loss rate of 24%, significantly below previous values and only marginally higher than natural loss. Successful human cell engraftment was achieved in 18% of lambs available for analysis. Engraftment levels of human cells in bone marrow of the recipient were up to 0.8% of total nucleated cells. CONCLUSION: Ultrasound-guided percutaneous transplantation of stem cells to fetal sheep in the first trimester is feasible. Although we were unable to observe a significant improvement in the level of engraftment of human cells in sheep, the decreased fetal loss rate associated with this technique allows greater use for further studies that use this model of in utero transplantation.

Animals↗
PubMed2003

Tracking dendritic cells: use of an in situ method to label all blood leukocytes.

Here we describe an in situ procedure with a labeling index (percent of labeled blood leukocytes) >98%, which is high enough to permit the direct tracking of dendritic cell (DC) precursors from blood into lymphoid tissues, while circumventing the pitfalls associated with in vitro labeling. DC and lymphocytes have similar blood to afferent lymph migratory capabilities. This method has additional applications in tracking other rare cell populations in both normal and pathological states.

Animals↗
PubMed2002

Biochemical changes in the efferent lymph plasma after oxazolone stimulation.

The lymph node is a specialized microenvironment for the regulation of immune responses. The reticular network of the lymph node provides a structure that facilitates not only intercellular interactions, but the intranodal flow of lymph fluid. To assess biochemical changes in the nodal lymph plasma after antigen stimulation, prescapular lymph nodes in sheep were stimulated with the epicutaneous antigen oxazolone. The efferent lymph from both antigen-stimulated and contralateral control prescapular lymph nodes was continuously monitored for more than 120 h. The oxazolone-stimulated lymph plasma was associated with a selective increase in cholesterol content during the 'recruitment' phase of lymph node enlargement. The peak in cholesterol was followed closely by a significant increase in lymph plasma LDH concentration. In contrast, there was no significant difference between oxazolone-stimulated and control lymph plasma in the concentration of triglycerides, albumin, alkaline phosphatase, and alanine transferase. These selective biochemical changes in the efferent lymph appeared to reflect the dynamics of lymphocyte activation within the lymph node as well as provide a practical measure of the lymph node response to antigen stimulation.

Animals↗
PubMed2002

Stimulation of regional lymphatic and blood flow by epicutaneous oxazolone.

The application of the epicutaneous antigen oxazolone results in persistent induration and erythema; however, the relative changes in lymph and blood flow in the inflammatory skin are largely unknown. To define the contribution of lymph and blood flow to the clinical appearance of cutaneous inflammation, we studied the sheep ear after the application of oxazolone. As a model for the study of these changes, the sheep ear had several experimental advantages: 1) a simplified superficial vascular network, 2) defined lymphatic drainage, and 3) an avascular and alymphatic cartilaginous barrier. Lymph flow was continuously monitored by cannulation of the prescapular efferent lymph duct. Blood flow, as reflected by cutaneous erythema, was noninvasively measured by use of a visible-spectrum spectrophotometer. The application of the epicutaneous oxazolone resulted in increased ear thickness for >7 days. The lymph flow from the oxazolone-stimulated ear peaked between 24 and 48 h after oxazolone stimulation. Spectrophotometric evaluation indicated that the cutaneous erythema peaked 72-96 h after application of oxazolone. Corrosion casting and scanning electron microscopy of the microcirculation at 96 h after antigen stimulation demonstrated significant dilatation of the superficial vascular network. These results suggest a biphasic response to oxazolone stimulation: 1) an early increase in vascular permeability associated with increased lymph flow and 2) a subsequent increase in relative blood flow associated with a dilated inflammatory microcirculation.

Adjuvants, Immunologic↗
PubMed2002

Using a neural network with flow cytometry histograms to recognize cell surface protein binding patterns.

Flow cytometric systems are being used increasingly in all branches of biological science including medicine. To develop analytic tools for identifying unknown molecules such as the antibodies that recognize different structure in the identical antigens, we explored use of a neural network in flow cytometry data comparison. Peak locations were extracted from flow cytometry histograms and we used the Marquardt backpropagation neural networks to recognize identical or similar binding patterns between antibodies and antigens based on the peak locations. The neural network showed 93.8% to 99.6% correct classification rates for identical or similar molecules. This suggests that the neural network technique can be useful in flow cytometry histogram data analysis.

Antibodies, Monoclonal↗
PubMed2002

Matching of flow-cytometry histograms using information theory in feature space.

Flow cytometry is a widely available technique for analyzing cell-surface protein expression. Data obtained from flow cytometry is frequently used to produce fluorescence intensity histograms. Comparison of histograms can be useful in the identification of unknown molecules and in the analysis of protein expression. In this study, we examined the combination of a new smoothing technique called SiZer with information theory to measure the difference between cytometry histograms. SiZer provides cross-bandwidth smoothing and allowed analysis in feature space. The new methods were tested on a panel of monoclonal antibodies raised against proteins expressed on peripheral blood lymphocytes and compared with previous methods. The findings suggest that comparing information content of histograms in feature space is effective and efficient for identifying antibodies with similar cell-surface binding patterns.

Animals↗