Bovine viral diarrhea virus compromises Neutrophil's functions in strain dependent manner.

Bovine viral diarrhea virus (BVDV) infection is a major problem that results in economically important diseases of the cattle industry worldwide. The two major consequences of this disease are persistent infection and immune dysfunction. A number of studies have been done to determine the underline mechanisms of BVDV-induced immune dysfunction, in particular targeting antigen-presenting cells, T- and B- cells and cytokine gene expression. However, little research has focused Eon the effect of BVDV on neutrophils. Neutrophils are one of the predominant leukocytes circulating in blood and are considered the first line of defense in the innate immune system along with macrophages. Neutrophils not only eliminate the invading bacteria but also activate innate as well as adaptive immune responses. Therefore, compromised neutrophil function would affect both arms of immune system and caused immune suppression. In the current study, we used virus strains from both BVDV-1 and BVDV-2 species. Including a highly virulent non-cytopathic type 2a BVDV (ncp BVDV2a-1373), moderately virulent non-cytopathic type 2a (ncp BVDV2a 28508-5), and a pair of non-cytopathic type 1b BVDV (ncp BVDV1b TGAN) and cytopathic type 1b BVDV (cp BVDV1b TGAC) strain isolated from a case of mucosal disease. The highly virulent ncp BVDV2a-1373 significantly increased neutrophil apoptosis. However, none of the other BVDV strains affected neutrophil viability. All BVDV strains used significantly reduced CD18 and L-selectin expression on neutrophils as well as their oxidative burst and neutrophil extracellular traps (NET) activity. Cp BVDV significantly reduced neutrophil's phagocytic activity but ncp BVDV did not have any effect on it. On the other hand, ncp BVDV significantly increased neutrophil's CD14 expression and chemotactic activity while cp BVDV did not show any effect either on neutrophil's CD14 expression or on chemotactic activity. In conclusion, BVDV affected neutrophils variability and functional activity in strain dependent manner. Results of the current study will further help in understanding the pathophysiology of different BVDV strains.

Adjuvants: What a Difference 15 Years Makes!

Vaccination is a critical tool in modern animal production and key to maintaining animal health. Adjuvants affect the immune response by increasing the rate, quantity, or quality of the protective response generated by the target antigens. Although adjuvant technology dates back to the nineteenth century, there was relatively little improvement in adjuvant technology before the late twentieth century. With the discovery of molecular pathways that regulate the timing, quantity, and quality of the immune response, new technologies are focused on bringing safer, more effective, and inexpensive adjuvants to commercial use.

Cell-mediated and humoral immune responses to bovine herpesvirus type 1 and bovine viral diarrhea virus in calves following administration of a killed-virus vaccine and bovine herpesvirus type 1 challenge.

OBJECTIVE To evaluate cell-mediated and humoral immune responses of calves receiving 2 doses of a dual-adjuvanted vaccine containing inactivated bovine herpesvirus type 1 (BHV1) and bovine viral diarrhea virus types 1 (BVDV1) and 2 (BVDV2) before and after exposure to BHV1. ANIMALS 24 Holstein steers negative for anti-BHV1 antibodies and proliferative cell-mediated immune responses against BHV1 and BVDV. PROCEDURES Calves were randomly assigned to 3 groups. The vaccinated group (n = 10) received 2 doses of vaccine on days 0 and 21. Control (n = 10) and seeder (4) groups remained unvaccinated. Calves were commingled during the study except for the 3-day period (days 53 to 55) when seeders were inoculated with BHV1 (1.04 × 107 TCID50, IV) to serve as a source of virus for challenge (days 56 through 84). Rectal temperature and clinical illness scores were monitored, and blood and nasal specimens were obtained for determination of clinicopathologic and immunologic variables. RESULTS After BHV1 challenge, mean rectal temperature and clinical illness scores were lower for vaccinates than controls. In vaccinates, antibody titers against BHV1 and BVDV2, but not BVDV1, increased after challenge as did extracellular and intracellular interferon-γ expression, indicating a T helper 1 memory response. Additional results of cell marker expression were variable, with no significant increase or decrease associated with treatment. CONCLUSIONS AND CLINICAL RELEVANCE Calves administered 2 doses of a killed-virus vaccine developed cell-mediated and humoral immune responses to BHV1 and BVDV, which were protective against disease when those calves were subsequently exposed to BHV1.

Classical natural ovine scrapie prions detected in practical volumes of blood by lamb and transgenic mouse bioassays.

Scrapie is diagnosed antemortem in sheep by detecting misfolded isoforms of prion protein (PrP(Sc)) in lymphoid follicles of the rectal mucosa and nictitating membranes. Assay sensitivity is limited if (a) the biopsy is collected early during disease development, (b) an insufficient number of follicles is collected, or (c) peripheral accumulation of PrP(Sc) is reduced or delayed. A blood test would be convenient for mass live animal scrapie testing. Currently approved techniques, however, have their own detection limits. Novel detection methods may soon offer a non-animal-based, rapid platform with detection sensitivities that rival the prion bioassay. In anticipation, we sought to determine if diseased animals could be routinely identified with a bioassay using B lymphocytes isolated from blood sample volumes commonly collected for diagnostic purposes in small ruminants. Scrapie transmission was detected in five of six recipient lambs intravenously transfused with B lymphocytes isolated from 5~10 mL of blood from a naturally scrapie-infected sheep. Additionally, scrapie transmission was observed in 18 ovinized transgenic Tg338 mice intracerebrally inoculated with B lymphocytes isolated from 5~10 mL of blood from two naturally scrapie-infected sheep. Based on our findings, we anticipate that these blood sample volumes should be of diagnostic value.

Prion formation, but not clearance, is supported by protein misfolding cyclic amplification.

Prion diseases are fatal transmissible neurodegenerative disorders that affect animals including humans. The kinetics of prion infectivity and PrP(Sc) accumulation can differ between prion strains and within a single strain in different tissues. The net accumulation of PrP(Sc) in animals is controlled by the relationship between the rate of PrP(Sc) formation and clearance. Protein misfolding cyclic amplification (PMCA) is a powerful technique that faithfully recapitulates PrP(Sc) formation and prion infectivity in a cell-free system. PMCA has been used as a surrogate for animal bioassay and can model species barriers, host range, strain co-factors and strain interference. In this study we investigated if degradation of PrP(Sc) and/or prion infectivity occurs during PMCA. To accomplish this we performed PMCA under conditions that do not support PrP(Sc) formation and did not observe either a reduction in PrP(Sc) abundance or an extension of prion incubation period, compared to untreated control samples. These results indicate that prion clearance does not occur during PMCA. These data have significant implications for the interpretation of PMCA based experiments such as prion amplification rate, adaptation to new species and strain interference where production and clearance of prions can affect the outcome.

Metformin and berberine prevent olanzapine-induced weight gain in rats.

Olanzapine is a first line medication for the treatment of schizophrenia, but it is also one of the atypical antipsychotics carrying the highest risk of weight gain. Metformin was reported to produce significant attenuation of antipsychotic-induced weight gain in patients, while the study of preventing olanzapine-induced weight gain in an animal model is absent. Berberine, an herbal alkaloid, was shown in our previous studies to prevent fat accumulation in vitro and in vivo. Utilizing a well-replicated rat model of olanzapine-induced weight gain, here we demonstrated that two weeks of metformin or berberine treatment significantly prevented the olanzapine-induced weight gain and white fat accumulation. Neither metformin nor berberine treatment demonstrated a significant inhibition of olanzapine-increased food intake. But interestingly, a significant loss of brown adipose tissue caused by olanzapine treatment was prevented by the addition of metformin or berberine. Our gene expression analysis also demonstrated that the weight gain prevention efficacy of metformin or berberine treatment was associated with changes in the expression of multiple key genes controlling energy expenditure. This study not only demonstrates a significant preventive efficacy of metformin and berberine treatment on olanzapine-induced weight gain in rats, but also suggests a potential mechanism of action for preventing olanzapine-reduced energy expenditure.

The effect of bovine viral diarrhea virus (BVDV) strains on bovine monocyte-derived dendritic cells (Mo-DC) phenotype and capacity to produce BVDV.

BACKGROUND: Dendritic cells (DC) are important antigen presentation cells that monitor, process, and present antigen to T cells. Viruses that infect DC can have a devastating impact on the immune system. In this study, the ability of bovine viral diarrhea virus (BVDV) to replicate and produce infectious virus in monocyte-derived dendritic cells (Mo-DC) and monocytes was studied. The study also examined the effect of BVDV infection on Mo-DC expression of cell surface markers, including MHCI, MHCII, and CD86, which are critical for DC function in immune response. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from bovine blood through gradient centrifugation. The adherent monocytes were isolated from PBMCs and differentiated into Mo-DC using bovine recombinant interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GMCSF). To determine the effect of BVDV on Mo-DC, four strains of BVDV were used including the severe acute non-cytopathic (ncp) BVDV2a-1373; moderate acute ncp BVDV2a 28508-5; and a homologous virus pair [i.e., cytopathic (cp) BVDV1b TGAC and ncp BVDV1b TGAN]. The Cooper strain of bovine herpesvirus 1 (BHV1) was used as the control virus. Mo-DC were infected with one of the BVDV strains or BHV-1 and were subsequently examined for virus replication, virus production, and the effect on MHCI, MHCII, and CD86 expression. RESULTS: The ability of monocytes to produce infectious virus reduced as monocytes differentiated to Mo-DC, and was completely lost at 120 hours of maturation. Interestingly, viral RNA increased throughout the course of infection in Mo-DC, and the viral non-structural (NS5A) and envelope (E2) proteins were expressed. The ncp strains of BVDV down-regulated while cp strain up-regulated the expression of the MHCI, MHCII, and CD86 on Mo-DC. CONCLUSIONS: The study revealed that the ability of Mo-DC to produce infectious virus was reduced with its differentiation from monocytes to Mo-DC. The inability to produce infectious virus may be due to a hindrance of virus packaging or release mechanisms. Additionally, the study demonstrated that ncp BVDV down-regulated and cp BVDV up-regulated the expression of Mo-DC cell surface markers MHCI, MHCII, and CD86, which are important in the mounting of immune responses.

Role of phosphatidylinositol 3-kinase (PI3K) and Akt1 kinase in porcine reproductive and respiratory syndrome virus (PRRSV) replication.

We have previously reported that inhibition of phosphatidylinositol 3-kinase (PI3K) reduces porcine reproductive and respiratory syndrome (PRRSV) replication. Here, we further investigate the mechanism by which PI3K inhibition affects virus replication and the role of Akt1 kinase in virus replication. We found that PI3K inhibition reduced viral gene transcription by approximately 1.5-fold. Accordingly, viral protein synthesis was significantly reduced by PI3K inhibition. Interestingly, cells overexpressing the dominant negative mutant Akt1 exhibited a significant reduction in viral gene transcription compared to cells overexpressing the constitutively active Akt1. Viral protein synthesis was also enhanced in cells overexpressing the constitutively active Akt1. Overall, our data show that both PI3K and Akt1 play a role in viral gene expression, leading to an increase in virus replication.

The skin, a novel niche for recirculating B cells.

B cells infiltrate the skin in many chronic inflammatory diseases caused by autoimmunity or infection. Despite potential contribution to disease, skin-associated B cells remain poorly characterized. Using an ovine model of granulomatous skin inflammation, we demonstrate that B cells increase in the skin and skin-draining afferent lymph during inflammation. Surprisingly, skin B cells are a heterogeneous population that is distinct from lymph node B cells, with more large lymphocytes as well as B-1-like B cells that coexpress high levels of IgM and CD11b. Skin B cells have increased MHC class II, CD1, and CD80/86 expression compared with lymph node B cells, suggesting that they are well-suited for T cell activation at the site of inflammation. Furthermore, we show that skin accumulation of B cells and Ab-secreting cells during inflammation increases local Ab titers, which could augment host defense and autoimmunity. Although skin B cells express typical skin-homing receptors, such as E-selectin ligand and α-4 and ß-1 integrins, they are unresponsive to ligands for chemokine receptors associated with T cell homing into skin. Instead, skin B cells migrate toward the cutaneously expressed CCR6 ligand CCL20. Our data support a model in which B cells use CCR6-CCL20 to recirculate through the skin, fulfilling a novel role in skin immunity and inflammation.

Salivary prions in sheep and deer.

Scrapie of sheep and chronic wasting disease (CWD) of cervids are transmissible prion diseases. Milk and placenta have been identified as sources of scrapie prions but do not explain horizontal transmission. In contrast, CWD prions have been reported in saliva, urine and feces, which are thought to be responsible for horizontal transmission. While the titers of CWD prions have been measured in feces, levels in saliva or urine are unknown. Because sheep produce ~17 L/day of saliva, and scrapie prions are present in tongue and salivary glands of infected sheep, we asked if scrapie prions are shed in saliva. We inoculated transgenic (Tg) mice expressing ovine prion protein, Tg(OvPrP) mice, with saliva from seven Cheviot sheep with scrapie. Six of seven samples transmitted prions to Tg(OvPrP) mice with titers of -0.5 to 1.7 log ID50 U/ml. Similarly, inoculation of saliva samples from two mule deer with CWD transmitted prions to Tg(ElkPrP) mice with titers of -1.1 to -0.4 log ID50 U/ml. Assuming similar shedding kinetics for salivary prions as those for fecal prions of deer, we estimated the secreted salivary prion dose over a 10-mo period to be as high as 8.4 log ID50 units for sheep and 7.0 log ID50 units for deer. These estimates are similar to 7.9 log ID50 units of fecal CWD prions for deer. Because saliva is mostly swallowed, salivary prions may reinfect tissues of the gastrointestinal tract and contribute to fecal prion shedding. Salivary prions shed into the environment provide an additional mechanism for horizontal prion transmission.

Classical scrapie prions in ovine blood are associated with B lymphocytes and platelet-rich plasma.

BACKGROUND: Classical scrapie is a naturally occurring transmissible spongiform encephalopathy of sheep and goats characterized by cellular accumulation of abnormal isoforms of prion protein (PrPSc) in the central nervous system and the follicles of peripheral lymphoid tissues. Previous studies have shown that the whole blood and buffy coat blood fraction of scrapie infected sheep harbor prion infectivity. Although PrPSc has been detected in peripheral blood mononuclear cells (PBMCs), plasma, and more recently within a subpopulation of B lymphocytes, the infectivity status of these cells and plasma in sheep remains unknown. Therefore, the objective of this study was to determine whether circulating PBMCs, B lymphocytes and platelets from classical scrapie infected sheep harbor prion infectivity using a sheep bioassay. RESULTS: Serial rectal mucosal biopsy and immunohistochemistry were used to detect preclinical infection in lambs transfused with whole blood or blood cell fractions from preclinical or clinical scrapie infected sheep. PrPSc immunolabeling was detected in antemortem rectal and postmortem lymphoid tissues from recipient lambs receiving PBMCs (15/15), CD72+ B lymphocytes (3/3), CD21+ B lymphocytes (3/3) or platelet-rich plasma (2/3) fractions. As expected, whole blood (11/13) and buffy coat (5/5) recipients showed positive PrPSc labeling in lymphoid follicles. However, at 549 days post-transfusion, PrPSc was not detected in rectal or other lymphoid tissues in three sheep receiving platelet-poor plasma fraction. CONCLUSIONS: Prion infectivity was detected in circulating PBMCs, CD72+ pan B lymphocytes, the CD21+ subpopulation of B lymphocytes and platelet-rich plasma of classical scrapie infected sheep using a sheep bioassay. Combining platelets with B lymphocytes might enhance PrPSc detection levels in blood samples.

Enhancement of immunohistochemical staining of scrapie proteins and immune cells within lymph nodes of early scrapie-infected sheep.

Transmissible spongiform encephalopathies (TSE) are a group of fatal neurodegenerative diseases that affect animals as well as humans. The oldest of these diseases is Scrapie seen in sheep. Scrapie is caused by an altered form (PrP(sc)), capable of inducing "self-replication" of the normal host prion protein (PrP(c)). There is currently no universal standard for antigen retrieval when using immunohistochemistry to simultaneously stain the PrP(c) protein and other cellular markers. The use of formalin-fixed tissue creates a challenge by concealing the antigenic sites where an antibody would bind, and lengthy antigen retrieval methods must be applied in order to facilitate staining. Further complicating sheep tissue immunohistochemistry is a significant lack of commercial antibodies to sheep cell markers available in research. Here we developed a novel immunohistochemical technique using trypsin, formic acid, and hydrated autoclaving using citraconic anhydride buffer to increase sensitivity of staining for scrapie proteins and immune cell subsets. This allowed us to stain and identify cells within lymphoid tissue associated with early lymphoid pathogenesis in scrapie.

B cells and platelets harbor prion infectivity in the blood of deer infected with chronic wasting disease.

Substantial evidence for prion transmission via blood transfusion exists for many transmissible spongiform encephalopathy (TSE) diseases. Determining which cell phenotype(s) is responsible for trafficking infectivity has important implications for our understanding of the dissemination of prions, as well as their detection and elimination from blood products. We used bioassay studies of native white-tailed deer and transgenic cervidized mice to determine (i) if chronic wasting disease (CWD) blood infectivity is associated with the cellular versus the cell-free/plasma fraction of blood and (ii) in particular if B-cell (MAb 2-104(+)), platelet (CD41/61(+)), or CD14(+) monocyte blood cell phenotypes harbor infectious prions. All four deer transfused with the blood mononuclear cell fraction from CWD(+) donor deer became PrP(CWD) positive by 19 months postinoculation, whereas none of the four deer inoculated with cell-free plasma from the same source developed prion infection. All four of the deer injected with B cells and three of four deer receiving platelets from CWD(+) donor deer became PrP(CWD) positive in as little as 6 months postinoculation, whereas none of the four deer receiving blood CD14(+) monocytes developed evidence of CWD infection (immunohistochemistry and Western blot analysis) after 19 months of observation. Results of the Tg(CerPrP) mouse bioassays mirrored those of the native cervid host. These results indicate that CWD blood infectivity is cell associated and suggest a significant role for B cells and platelets in trafficking CWD infectivity in vivo and support earlier tissue-based studies associating putative follicular B cells with PrP(CWD). Localization of CWD infectivity with leukocyte subpopulations may aid in enhancing the sensitivity of blood-based diagnostic assays for CWD and other TSEs.

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.

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.

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.

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.

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.

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.