Delayed-start cyclosporine immunosuppression reduces graft cell migration after small bowel transplantation.
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Animals↗
Biomedical subjects
A G Pockley
Publications and source records attributed to A G Pockley.
Host cell infiltration of the intraepithelial compartment in small bowel transplantation.
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Animals↗
Heparin modulation of cellular proliferative responses: kinetic and concentration dependency.
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Animals↗
Regulatory role of interleukins 5 and 6 on immunoglobulin production in cultured rat salivary glands.
The effects of interleukin-5 and interleukin-6 on immunoglobulin production in rat salivary glands were investigated using an in vitro tissue culture system. Detectable levels of IgA, IgG and IgM were observed in the culture media of unstimulated tissues of rat salivary glands after 7 days in culture. Incubation of the tissues with recombinant murine IL-5 (50 U/ml) enhanced the levels of IgA (205% of the control) and IgG (136% of the control), but had no effect on the levels of IgM. Similarly, incubation with recombinant human IL-6 (50 U/ml) enhanced IgA (224% of the control) and IgG (149% of the control) production, but had no effect on IgM. A combination of both IL-5 and IL-6 had no additional effect on the enhanced IgA levels than that seen with IL-5 or IL-6 alone. These data demonstrate a potential regulatory function of lymphokines in glandular mucosal tissue that differs from that previously noted in experiments with cultured rat lacrimal glands or isolated cells derived from both mucosal and non-mucosal tissues.
Animals↗
The effects of interleukin-5 on mitogenically stimulated rat spleen cells.
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Animals↗
Modulation of T and B cell proliferative responses by factors present in rat salivary glands.
Rat salivary gland culture supernatants (SGSN) were shown to inhibit the proliferation of rat spleen cells induced by the mitogens concanavalin A (ConA), phytohaemagglutinin (PHA), pokeweed mitogen (PWM), lipopolysaccharide (LPS) and S. typhimurium mitogen (STM). The responses of B cells were more markedly inhibited than the responses of T cells. Factors contained in SGSN which had a molecular weight smaller than 3500 inhibited all responses, whereas factors greater than 3500 only inhibited responses induced by PWM, LPS or STM. Factors present in SGSN also inhibited the proliferation of two B cell hybridoma cell lines, as well as the IL-2-responsive cell line CTLL-2 and the IL-4-responsive cell line CT.4S. However, SGSN factors having a molecular weight greater than 3500 did not inhibit CTLL-2 proliferation. These data indicate that rat salivary glands contain factors which differentially regulate T and B cell proliferative responses in vitro and which may modulate localized immune responses in the salivary gland in vivo.
Animals↗
In vivo adjuvant effect of interleukins 5 and 6 on rat tear IgA antibody responses.
Interleukins 5 and 6 have previously been shown to exhibit IgA-enhancing activity in vitro. This study investigated the effect of IL-5 and IL-6 on antigen-specific tear IgA antibody responses in vivo. Rats were given primary and secondary immunizations by the ocular-topical (OT) route with either antigen (DNP-Pn) alone or antigen plus IL-5 and IL-6 (IL-5/6). Antigen was administered for the first 3 days of each stimulation cycle and lymphokines (50 units of each lymphokine/eye/day) or control media for the first 7 days of the primary and secondary immunizations. Co-administration of antigen with IL-5/6 markedly enhanced (approximately three-fold) the secondary tear IgA anti-DNP antibody response in the lymphokine-treated group. Enhanced tear IgA antibody responses were maintained in the lymphokine-treated group following a tertiary immunization in the absence of IL-5/6. There was no apparent effect on the total tear IgA levels in either group of animals. Ocular topical immunization also elicited secondary and tertiary serum IgG antibody responses which were not affected by the administration of IL-5/6. Neither secondary nor tertiary serum IgM antibody responses were detected in either group of animals. These data indicate that IL-5 and IL-6 are able to augment tear IgA antibody responses in vivo. That the enhancement was maintained following a subsequent antigenic challenge in the absence of the lymphokines suggests a potential therapeutic application for lymphokines in augmenting IgA responses at mucosal surfaces.
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The effect of human placental protein 14 (PP14) on the production of interleukin-1 from mitogenically stimulated mononuclear cell cultures.
Crude human decidual tissue extracts containing placental protein 14 (PP14) were shown to inhibit the production of interleukin-1 beta (IL-1) from mitogenically stimulated mononuclear cell cultures. The inhibition was dose-dependent over the range of PP14 concentrations investigated (0-8.0 mg/l) and was effective on both phytohaemagglutinin-(PHA) and lipopolysaccharide- (LPS)-induced IL-1 secretion. Using these culture systems, a PP14 concentration of 1.0 mg/l induced a 34% suppression of IL-1 secretion following LPS stimulation and 22% following PHA stimulation. For PHA stimulation the suppression of IL-1 secretion was effective throughout the culture period investigated (0-89 hr). Individual crude decidual extracts inhibited the incorporation of [3H]thymidine into PHA-stimulated lymphocytes, such inhibition being partially reversed by the addition of exogenous recombinant IL-1 to the cultures. These results suggest that the previously reported immunosuppressive activity of PP14 may be mediated by the suppression of IL-1 secretion.
Cells, Cultured↗
Placental protein 14 (PP14) content and immunosuppressive activity of human cervical mucus.
Extracts of cervical mucus contain measurable levels of placental protein 14 (PP14), a protein known to suppress proliferative responses of human peripheral blood lymphocytes. Such extracts have been shown to exert suppressive activities on allogeneically stimulated peripheral blood lymphocytes in a dose-dependent manner. However, there was no correlation between the PP14 content of cervical mucus samples and their suppressive activities. Such activity could not be removed from the extracts by dialysis, suggesting high molecular weight mediators. These results indicate that there may be an immunosuppressive factor in human cervical mucus which functions to protect inseminated sperm in a potentially immunologically hostile environment in the female reproductive tract.
Cells, Cultured↗
Placental protein 14 (PP14) inhibits the synthesis of interleukin-2 and the release of soluble interleukin-2 receptors from phytohaemagglutinin-stimulated lymphocytes.
Crude human decidual extracts containing up to 15.0 mg/l PP14 were investigated for their effects on the release of interleukin-2 (IL-2) and of IL-2 receptor (IL-2R) from phytohaemagglutinin (PHA) stimulated lymphocytes. The crude decidual extract suppressed IL-2 production over the culture period investigated (0-90 h) with maximal suppression observed at around 66 h of culture. The suppression was dose dependent over the range of PP14 concentrations used (0-7.0 mg/l). Decidual extracts also inhibited the release of soluble IL-2R into the culture supernatants over the same period. In both cases, the specific reduction of the PP14 content of the extracts by a monoclonal anti-PP14 immunoadsorbent reduced the observed suppression. These results suggest that PP14 inhibits the production of IL-2 from mitogenically stimulated lymphocytes, and leads to a reduced IL-2R release. The mechanism for such an effect is as yet unknown; however, it may explain the previously reported immunosuppressive activity of PP14.
Cells, Cultured↗
Suppression of in vitro lymphocyte reactivity to phytohemagglutinin by placental protein 14.
Crude human decidual extracts containing up to 26.6 mg/l of placental protein 14 (PP14) and purified PP14 were assessed for their effects on the in vitro lymphocyte reactivity to phytohemagglutinin (PHA). Both decidual extract and purified PP14 suppressed the mitogenic response to PHA with the suppression being dose-dependent over the range of PP14 concentrations investigated (0-26.6 mg/l). On the specific reduction of the PP14 content by a monoclonal anti-PP14 immunoadsorbant the suppression was reduced. The suppressive activity of PP14 was related to the degree of proliferation of the stimulated lymphocytes. These results suggest that PP14, which is present at peak levels in the first trimester of pregnancy and constitutes up to 10% of the soluble protein content of decidual tissue, may be an immunomodulator important for the survival of the implanting embryo and maintenance of early pregnancy.
Decidua↗
Identification of placental protein 14 as an immunosuppressive factor in human reproduction.
Extracts of human decidual tissue obtained in the first trimester of pregnancy showed potent suppressive activity in mixed lymphocyte cultures. These extracts contained substantial amounts of the decidual protein PP14. Purified PP14 also exhibited in vitro immunosuppressive activity, and such activity in decidual extract and purified PP14 preparations was removed by treatment with a monoclonal anti-PP14 antibody-based immunoadsorbent. PP14 was present in seminal plasma, which also exhibited immunosuppressive activity that could be reduced, but not removed, by the immunoadsorbent. PP14 may be an important immunomodulator in the human reproductive system.
Animals↗
The effects of dietary omega-3 polyunsaturated fatty acids on erythrocyte membrane phospholipids, erythrocyte deformability and blood viscosity in healthy volunteers.
We have examined, in normal subjects, the effects of a daily dietary supplement of fish oil concentrate ('maxEPA'), providing 3 g of omega-3 fatty acids, on erythrocyte membrane phospholipids, erythrocyte deformability and blood viscosity. After 3 weeks, incorporation of C20:5 omega 3 into erythrocyte phosphatidyl choline (PC) was greater compared to phosphatidyl ethanolamine (PE) and phosphatidyl serine (PS). After 6 weeks, there was no further increase in total erythrocyte C20:5 omega 3, but its distribution amongst phospholipid subclasses had changed. C20:5 omega 3 had increased further in PE and PS, but decreased in PC. Incorporation of C20:5 omega 3 also occurred into PC, PE and PS. omega-3 Fatty acids were incorporated almost entirely at the expense of C18:2 omega 6, but total unsaturation of phospholipids was increased. This is consistent with increased lipid fluidity, which may be an important determinant of erythrocyte deformability. The same dosage of maxEPA also resulted in a significant increase in erythrocyte deformability and a concomitant reduction in whole blood viscosity. Since plasma viscosity and haematocrit were unchanged it seems likely that the effects on blood rheology were mediated by changes in erythrocyte lipid fluidity. Modification of blood rheology by dietary omega-3 fatty acids is of potential value in the treatment of vascular disease.
Blood Viscosity↗
A rapid microplate-based fluorometric assay for phagocytosis.
A highly sensitive quantitative fluorometric assay for phagocytosis, previously measured using fluorescence spectrophotometry or flow cytometry, has been adapted for use with a 96-well fluorescence plate reader. The technique allows rapid analysis of large numbers of samples, and requires only a small sample volume. Comparison of plate types demonstrated that opaque white 96-well luminostrips produced a 100 fold greater fluorescent output, and were more sensitive than black fluoroplates. Intraplate variability was also significantly lower using white luminostrips. For the phagocytic assay, fluorescein conjugated polystyrene beads were incubated with macrophage monolayers in white luminostrips. After incubation, cells were washed, lysed and phagocytosis quantified by determining the fluorescent intensity using a fluorescence plate reader. The number of beads phagocytized was determined from a standard curve of bead number versus fluorescent output. The phagocytic activity of resident and thioglycollate-elicited peritoneal macrophages was compared using this technique.
Animals↗
Identification of migratory graft and host cell populations after allogeneic rat small bowel transplantation.
This study used flow cytometry to identify graft cells in the recipient peripheral blood and spleen and host cells infiltrating the graft mesenteric lymph node and Peyer's patches after heterotopic rat small bowel transplantation. Transplantation had little effect on the overall cell subset composition of these compartments and no changes appeared characteristic or indicative of developing rejection, suggesting that physiological control of cell migration remained unaltered. A small and transient population of graft cells was detected in the peripheral blood and spleen of the recipient which disappeared after 5 and 3 days respectively. Graft-derived cells in the peripheral blood comprised predominantly CD4+ cells on day 1 with B cells predominating on day 5. Graft cells infiltrating the spleen were predominantly B cells. Host cells infiltrated the graft mesenteric lymph nodes and Peyer's patches to a lesser extent than previously reported using immunohistochemical analysis. For both tissues, infiltrating host-derived cells initially comprised mainly CD4+ cells. On day 4 approximately equal proportions of CD4+ and B cells were present in the mesenteric lymph node, whereas B cells were predominant in the host cell infiltrate of the graft Peyer's patches. In summary, these findings indicate that the cell subset composition of recipient and graft lymphoid compartments does not change after small bowel transplantation, even in the presence of a substantial recipient cell infiltration. The reasons for the apparent discrepancies in the degree of host cell infiltration when assessed using immunohistochemical and flow cytometric techniques are currently uncertain, but may result from the localised release of soluble MHC class I in graft tissues as a consequence of infiltrating host cell activation or localised cell destruction.
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Identification of localized anti-host responses in the graft mesenteric lymph node and Peyer's patches after rat small bowel transplantation.
This study determined the activation status of recipient and donor lymphocyte populations in the graft mesenteric lymph node (MLN) and Peyer's patches (PP) after allogeneic, heterotopic rat small bowel transplantation without immunosuppression. Untransplanted and isografted animals served as controls. The activation status of lymphocyte subsets was determined by flow cytometric evaluation of lymphoblastoid transformation (forward light scatter; FSc). The proportion of activated lymphocytes in the MLN and PP of allografted animals progressively increased. There was also an early transient activation of MLN lymphocytes in isografted animals which probably resulted from surgery-related inflammation. Activated alpha/beta TCR+ and CD4+ cells were detected in the MLN as early as day 3, whereas there was little activation of CD8+ cells. Interestingly, donor lymphocytes became more activated than recipient lymphocytes. Allografting also led to activation of graft-derived PP alpha/beta TCR+ and CD8+ cells, yet there was no detectable activation of recipient-derived lymphocytes. In summary, this study has identified activated donor lymphocytes in the graft MLN and PP after allogeneic small bowel transplantation. Although rejection predominates without immunosuppression, the presence of an underlying anti-recipient response within the small bowel allograft may contribute to graft damage via the localized release of cytokines and inflammatory mediators.
Animals↗
Modulation of leukocyte phagocytic and oxidative burst responses by human seminal plasma.
The typical absence of immune responses to spermatozoa in the female reproductive tract at the time of insemination, despite the presence of a marked leukocytic infiltrate into the cervical mucus is intriguing. It may be that localised immunoregulatory mechanisms exist and this study used whole blood flow cytometry to determine the effects of human seminal plasma on neutrophil and monocyte function. Seminal plasma inhibited the proportion of neutrophils and monocytes phagocytosing E. coli, and the intensity of neutrophil phagocytosis, but enhanced the magnitude of the phagocytic response of those monocytes that escaped inhibition relative to PBS treated controls. Oxidative burst responses to E. coli were also inhibited and this effect was mediated by low molecular weight species, as dialysis totally abrogated the inhibitory activity. Seminal plasma had no effect on the neutrophil burst response to fMLP when compared to the controls, however there was a significant difference between the responses of undialysed and dialysed seminal plasma treated samples. Undialysed seminal plasma significantly inhibited the proportion of monocytes undergoing the burst response to fMLP and there were significant differences between the proportion of cells responding and their intensity in undialysed and dialysed seminal plasma treated samples. In summary, this study reports differential modification of neutrophil and monocyte function by human seminal plasma. The residual capacity of these cells to undergo phagocytosis and generate oxidative burst responses suggests that localised innate immune function remains intact and is possibly enhanced in the female reproductive tract at the time of insemination. Other mechanisms must protect inseminated sperm at this time.
Dialysis↗
Identification of lacrimal gland associated immunomodulatory activities having differential effects on T and B cell proliferative responses.
The supernatants from overnight cultures of rat lacrimal glands (LGSN) were found to suppress the proliferative response of rat splenic mononuclear cells following mitogenic stimulation. LGSN inhibited LPS and STM responses to a greater degree (55% inhibition at 1.56% v/v) than ConA and PHA responses (9 and 28% inhibition, respectively, at 1.56% v/v). PWM-induced responses were inhibited at an intermediate level (32% inhibition at 1.56% v/v). The inhibitory activity was not mediated by a suppression of lymphokine production, as the activity was not reversed by the addition of exogenous recombinant IL-2 (25 U/ml) or conditioned medium from allogeneic mixed lymphocyte reactions, nor was the suppressive activity mediated by a nonspecific cytotoxic effect. Dialysis of LGSN indicated that species less than 3500 Da were able to inhibit all mitogenic responses whereas factors greater than 3500 Da were only able to inhibit LPS and STM responses. LGSN were shown to contain prostaglandin E2, however, at levels which were insufficient to explain the low molecular weight inhibitory activity. In addition to the inhibition of mitogenic responses, LGSN also inhibited B cell hybridoma and growth factor (IL-2, IL-4) dependent T cell proliferation, an inhibition which was abrogated following dialysis. These findings suggest the presence of multiple factors in rat lacrimal glands which are able to modulate T and B cell proliferative responses.
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