Effects of opioids on immunologic parameters that are relevant to anti-tumour immune potential in patients with cancer: a systematic literature review.

BACKGROUND: The immune system has a central role in controlling cancer, and factors that influence protective antitumour immunity could therefore have a significant impact on the course of malignant disease. Opioids are essential for the management of cancer pain, and preclinical studies indicate that opioids have the potential to influence these tumour immune surveillance mechanisms. The aim of this systematic literature review is to evaluate the clinical effects of opioids on the immune system of patients with cancer. METHODS: A systematic search of Ovid MEDLINE (PubMed) and Embase, Cochrane database and Web of Knowledge for clinical studies, which evaluated the effects of opioids on the immune system in patients with cancer, was performed. RESULTS: Five human studies, which have assessed the effects of opioids on the immune system in patients with cancer, were identified. Although all of these evaluated the effect of morphine on immunologic end points in patients with cancer, none measured the clinical effects. CONCLUSIONS: Evidence from preclinical, healthy volunteer and surgical models suggests that different opioids variably influence protective anti-tumour immunity; however, actual data derived from cancer populations are inconclusive and definitive recommendations cannot be made. Appropriately designed and powered studies assessing clinical outcomes of opioid use in people with cancer are therefore required to inform oncologists and others involved in cancer care about the rational use of opioids in this patient group.

HAGE (DDX43) is a biomarker for poor prognosis and a predictor of chemotherapy response in breast cancer.

BACKGROUND: HAGE protein is a known immunogenic cancer-specific antigen. METHODS: The biological, prognostic and predictive values of HAGE expression was studied using immunohistochemistry in three cohorts of patients with BC (n=2147): early primary (EP-BC; n=1676); primary oestrogen receptor-negative (PER-BC; n=275) treated with adjuvant anthracycline-combination therapies (Adjuvant-ACT); and primary locally advanced disease (PLA-BC) who received neo-adjuvant anthracycline-combination therapies (Neo-adjuvant-ACT; n=196). The relationship between HAGE expression and the tumour-infiltrating lymphocytes (TILs) in matched prechemotherapy and postchemotherapy samples were investigated. RESULTS: Eight percent of patients with EP-BC exhibited high HAGE expression (HAGE+) and was associated with aggressive clinico-pathological features (Ps<0.01). Furthermore, HAGE+expression was associated with poor prognosis in both univariate and multivariate analysis (Ps<0.001). Patients with HAGE+did not benefit from hormonal therapy in high-risk ER-positive disease. HAGE+and TILs were found to be independent predictors for pathological complete response to neoadjuvant-ACT; P<0.001. A statistically significant loss of HAGE expression following neoadjuvant-ACT was found (P=0.000001), and progression-free survival was worse in those patients who had HAGE+residual disease (P=0.0003). CONCLUSIONS: This is the first report to show HAGE to be a potential prognostic marker and a predictor of response to ACT in patients with BC.

Effect of upper- and lower-limb exercise training on circulating soluble adhesion molecules, hs-CRP and stress proteins in patients with intermittent claudication.

OBJECTIVES: To investigate the effects of exercise training on levels of circulating biomarkers associated with the progression of atherosclerosis and risk of cardiovascular events in patients with intermittent claudication. METHODS: Circulating levels of soluble adhesion molecules (sVCAM-1, sICAM-1, sE-selectin), high sensitivity C-reactive protein (hs-CRP) and stress proteins (Hsp60 and Hsp70) in patients randomised to a 24-week programme of arm- or leg-cranking exercise were compared with those in usual care controls. RESULTS: Arm and leg exercise similarly improved lower-limb aerobic exercise capacity (20% vs 19%, respectively; P<0.001) and maximum walking distance (30% vs 35%, respectively; P<0.001). Improvements in training limb-specific peak oxygen consumption were attenuated for patients in the highest vs lowest quartile for circulating sVCAM-1 levels at baseline (3% vs 25% respectively, P<0.001). Although circulating hs-CRP levels tended to be lower in the arm-cranking group (-1.55 [95% CI: -1.06 to -2.26]mgl(-1)), exercise training had no effect on circulating levels of soluble adhesion molecules or stress proteins. CONCLUSIONS: These findings suggest that high levels of circulating sVCAM-1 are associated with an attenuated exercise training response and that arm-cranking exercise may provide an effective stimulus for evoking systemic anti-inflammatory adaptations in patients with intermittent claudication.

Analysis of purified gp96 preparations from rat and mouse livers using 2-D gel electrophoresis and tandem mass spectrometry.

The stress protein gp96 exhibits a number of immunological activities, the majority of studies into which have used gp96 purified from a variety of tissues. On the basis of 1-D gel electrophoresis, the purity of these preparations has been reported to range between 70% and 99%. This study analyzed gp96 preparations from rat and mouse livers using 2-D gel electrophoresis and liquid chromatography electrospray ionization tandem mass spectrometry (MS-MS). The procedure for purifying gp96 was reproducible, as similar protein profiles were observed in replicate gels of gp96 preparations. The purity of the preparations was typically around 70%, with minor co-purified proteins of varying molecular weights and mobilities being present. Dominant bands at 95-100 kDa in preparations from Wistar rats and C57BL/6 mice were identified as gp96 by ECL Western blotting. Multiple bands having similar, yet distinct molecular weights and differing pI mobility on ECL Western blots were confirmed as being gp96 in preparations from Wistar rats using MS-MS. The most striking feature of the 2-D gel analysis was the presence of additional dominant bands at 55 kDa in preparations from Wistar rats, and at 75-90 kDa in preparations from C57BL/6 mice. These were identified as gp96 by ECL Western blotting and, in the case of preparations from Wistar rats, by MS-MS. Although the lower molecular weight, gp96-related molecules might be partially degraded gp96, their reproducible presence, definition and characteristics suggest that they are alternative, species-specific isoforms of the molecule. A 55 kDa protein which exhibited a lower pI value than gp96 was present in all preparations and this was identified as calreticulin, another putative immunoregulatory molecule. This study confirms the reproducibility of the gp96 purification protocol and reveals the presence of multiple gp96 isoforms, some of which likely result from post-translational modifications such as differential glycosylation and phosphorylation.

Cytoplasmic PML promotes TGF-ß-associated epithelial-mesenchymal transition and invasion in prostate cancer.

Epithelial-mesenchymal transition (EMT) is a key event that is involved in the invasion and dissemination of cancer cells. Although typically considered as having tumour-suppressive properties, transforming growth factor (TGF)-ß signalling is altered during cancer and has been associated with the invasion of cancer cells and metastasis. In this study, we report a previously unknown role for the cytoplasmic promyelocytic leukaemia (cPML) tumour suppressor in TGF-ß signalling-induced regulation of prostate cancer-associated EMT and invasion. We demonstrate that cPML promotes a mesenchymal phenotype and increases the invasiveness of prostate cancer cells. This event is associated with activation of TGF-ß canonical signalling pathway through the induction of Sma and Mad related family 2 and 3 (SMAD2 and SMAD3) phosphorylation. Furthermore, the cytoplasmic localization of promyelocytic leukaemia (PML) is mediated by its nuclear export in a chromosomal maintenance 1 (CRM1)-dependent manner. This was clinically tested in prostate cancer tissue and shown that cytoplasmic PML and CRM1 co-expression correlates with reduced disease-specific survival. In summary, we provide evidence of dysfunctional TGF-ß signalling occurring at an early stage in prostate cancer. We show that this disease pathway is mediated by cPML and CRM1 and results in a more aggressive cancer cell phenotype. We propose that the targeting of this pathway could be therapeutically exploited for clinical benefit.

Circulating heat shock protein 60 is associated with early cardiovascular disease.

The phylogenetically conserved nature of heat shock proteins (Hsp) has led to the proposition that they may provide a link between infection and the inflammatory component to vascular disease. Hypertension is associated with atherosclerosis. Here, we measured circulating heat shock protein and heat shock protein antibody levels in association with borderline hypertension. Seventy-two men with borderline hypertension patients and 75 normotensive control subjects (diastolic blood pressure 85 to 94 and <80 mm Hg, respectively) were selected from a population-screening program. The levels of Hsp60; Hsp70; and anti-human Hsp60, anti-human Hsp70, and anti-mycobacterial Hsp65 antibodies were determined with enzyme immunoassay. The presence of carotid atherosclerosis and the intima-media thickness values were determined with ultrasonography. A major novel observation in this report was the detection of circulating Hsp60, which was present at a significantly enhanced level in patients with borderline hypertension. Furthermore, serum Hsp60 was associated with intima-media thicknesses (P<0.01). Anti-Hsp65 antibody levels were higher in borderline hypertension (P<0.001), whereas Hsp70 and anti-Hsp70 antibody levels did not differ. In contrast to anti-Hsp65 antibody, anti-Hsp60 antibody levels were lower in borderline hypertension (P<0.03), although the difference was quantitatively small. None of the parameters evaluated were associated with atherosclerosis, metabolic factors, or smoking. We identified elevated Hsp60 levels in patients with borderline hypertension and an association between early atherosclerosis and Hsp60 levels. The physiological role of Hsp60 release has yet to be defined, but given the proinflammatory properties, these proteins could be involved in the induction/progression of both hypertension and atherosclerosis, as well as being markers for early cardiovascular disease.

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.

Heat shock proteins in health and disease: therapeutic targets or therapeutic agents?

For many years, heat shock or stress proteins have been regarded as intracellular molecules that have a range of housekeeping and cytoprotective functions, only being released into the extracellular environment in pathological situations such as necrotic cell death. However, evidence is now accumulating to indicate that, under certain circumstances, these proteins can be released from cells in the absence of cellular necrosis, and that extracellular heat shock proteins have a range of immunoregulatory activities. The capacity of heat shock proteins to induce pro-inflammatory responses, together with the phylogenetic similarity between prokaryotic and eukaryotic heat shock proteins, has led to the proposition that these proteins provide a link between infection and autoimmune disease. Indeed, both elevated levels of antibodies to heat shock proteins and an enhanced immune reactivity to heat shock proteins have been noted in a variety of pathogenic disease states. However, further evaluation of heat shock protein reactivity in autoimmune disease and after transplantation has shown that, rather than promoting disease, reactivity to self-heat shock proteins can downregulate the disease process. It might be that self-reactivity to heat shock proteins is a physiological response that regulates the development and progression of pro-inflammatory immunity to these ubiquitously expressed molecules. The evolving evidence that heat shock proteins are present in the extracellular environment, that reactivity to heat shock proteins does not necessarily reflect adverse, pro-inflammatory responses and that the promotion of reactivity to self-heat shock proteins can downregulate pathogenic processes all suggest a potential role for heat shock proteins as therapeutic agents, rather than as therapeutic targets.

Effect of ex vivo storage on human peripheral blood neutrophil expression of CD11b and the stabilizing effects of Cyto-Chex.

CD11b is the alpha sub-unit of the CD11b/CD18 heterodimeric complex that has a key role in neutrophil-endothelial cell interactions and early events in inflammatory responses. Accurate assessment of CD11b expression by neutrophils can be problematic since the procedures that isolate cells from whole blood can increase antigen expression. This study used whole blood flow cytometry to monitor neutrophil CD11b expression following 0, 1, 2, 3 and 4 h of ex vivo incubation at room temperature and examined the effects of storage at 4 degrees C and the cell stabilization solution, Cyto-Chex, on antigen expression. Cyto-Chex-treated samples were also re-analyzed after 7 days storage at 4 degrees C. Neutrophil CD11b expression was high (> 90%) in five of the seven samples studied and incubation at room temperature induced a progressive upregulation which was significant after 2, 3, and 4 h (p < 0.05). Storage at 4 degrees C tempered this effect, although an increased expression (from baseline) was still observed after 4 h (p < 0.05). Cyto-Chex had no effect on the light scatter characteristics of treated cells and prevented CD11b upregulation in samples stored at room temperature. Furthermore, expression in Cyto-Chex-treated samples after 7 days was not significantly different from that observed in baseline (time 0) samples. In two samples demonstrating low neutrophil CD11b expression, a temperature-independent increase in the proportion of CD11b+ cells was observed over time. However, this increase was attenuated by treatment with Cyto-Chex. These findings indicate that sample storage significantly affects CD11b expression and caution should be exercised when interpreting data. Given that Cyto-Chex had no effect on the light scatter properties of cells and prevents antigen upregulation, this reagent may be useful for studies involving analysis of neutrophil activation antigen expression.

Effect of red cell lysis protocols on the expression of rat peripheral blood lymphocyte subset and activation antigens.

The effect of ammonium chloride and a commercial red cell lysing agent (Erythrolyse) on cell subset (CD4, CD8, B cell common leucocyte antigen) and activation (p55IL-2R [OX39], MHC class II) antigen expression by freshly isolated and in vitro activated rat peripheral blood lymphocytes was compared. Freshly isolated cells treated with the commercial lysing agent Erythrolyse were 42 +/- 12% p55IL-2R+, whereas p55IL-2R expression was not detected following red cell lysis with ammonium chloride. The expression of cell subset and MHC class II antigens was unaffected by either of the treatment protocols. Density gradient separation of freshly isolated cells had variable effects on antigen expression. In some cases up to a two-fold increase in p55IL-2R expression was observed, whilst in others there was a reduced receptor expression. Similarly, MHC class II antigen expression was either increased or reduced following density gradient separation. IL-2R expression on cells activated in vitro with the mitogen concanavalin A (95 +/- 2% IL-2R+) was unaffected by ammonium chloride lysis. Incubation of freshly isolated lymphocytes with p55IL-2R antibody prior to ammonium chloride treatment did not prevent the reduction in receptor expression. These findings suggest that ammonium chloride lysis of freshly isolated rat peripheral blood prior to IL-2R expression analysis should be avoided.

Development of an enzyme immunoassay for rat soluble interleukin-2 receptors.

This study describes the development of a sandwich enzyme immunoassay (ELISA) for rat soluble IL-2 receptors (sIL-2R) using a combination of monoclonal antibodies reactive with different epitopes on the rat IL-2R. Coating plates with NDS61 and NDS64 monoclonal antibodies produced similar dose-response curves when incubated with a standard sIL-2R preparation followed by biotinylated OX39, streptavidin-alkaline phosphatase and the substrate. Although normal rat serum inhibited the assay, the effects were more profound when NDS64 was used as the capture antibody and subsequent development of the assay was performed using NDS61. The intra- and interassay variations were typically less than 5%. This assay will be valuable for monitoring immune activation status in a variety of experimental models.

Heat shock proteins, anti-heat shock protein reactivity and allograft rejection.

Heat shock proteins are families of highly conserved immunodominant molecules, reactivity to which has been implicated in the pathogenesis of a number of autoimmune and vascular disease states. However, heat shock proteins are cytoprotective, and in clinical and experimental arthritis, anti-heat shock protein reactivity can down modulate immune responses via a self-Hsp reactive, Th2-type mechanism. Despite a number of studies associating heat shock protein expression and anti-heat shock protein reactivity with allograft rejection, the balance between protective and damaging effects and the precise influence of these responses on graft outcome is unclear. This article reviews current knowledge surrounding heat shock proteins, autoimmunity, and allograft rejection and presents a perspective on the potential influence of these proteins and the stress response on allograft outcome.

Effects of FK409 on intestinal ischemia-reperfusion injury and ischemia-induced changes in the rat mucosal villus microcirculation.

BACKGROUND: The small intestine is extremely sensitive to ischemia-reperfusion (I/R) injury and a range of microcirculatory disturbances contribute to tissue damage. Nitric oxide (NO) seems to be involved in tissue protection after I/R injury. This study therefore assessed the effects of the NO donor, FK409, on intestinal I/R injury and changes induced in intestinal microcirculation. METHODS: PVG rats were subjected to 30-min intestinal ischemia with a subgroup of animals receiving FK409 (10 mg/kg i.v.) 30 min before ischemia and 30 min postreperfusion. Controls underwent sham surgery. The mucosal surface was visualized via an incision made in an exteriorized ileal segment and FITC-BSA or acridine orange was used to quantitate macromolecular leak (MML) and leukocyte adhesion, respectively. MML from, and numbers of adherent leukocytes within, individual villi were determined every 15 min for 2 hr after removal of the vessel clamp. Heart rate and mean blood pressure (mBP) were monitored throughout the experiment. RESULTS: Eleven of 12 untreated animals subjected to intestinal I/R injury failed to survive the 2 hr reperfusion period, whereas all 12 FK409-treated animals survived. MML and leukocyte adhesion were increased in untreated animals (P<0.001), and blood flow stasis eventually ensued. Although FK409 decreased mBP (P<0.001), MML and leukocyte adhesion were significantly (P<0.001) reduced, and villus blood flow was maintained throughout the observation period. CONCLUSIONS: FK409 prevented mortality after intestinal I/R, significantly reduced leukocyte adhesion, and maintained blood flow after intestinal ischemia and may therefore be of value in reducing tissue damage and improving outcome after small bowel transplantation.

Induction of antigraft and antirecipient antibody responses after fully allogeneic and semiallogeneic rat small bowel transplantation.

BACKGROUND: Given the potential influence of alloantibodies on organ graft outcome, this study investigated the induction of antigraft and antirecipient antibodies after allogeneic and semiallogeneic rat small bowel transplantation. METHODS: Fully allogeneic, unidirectional rejection and unidirectional graft-versus-host disease (GvHD) heterotopic small bowel transplantation was performed using DA, PVG, and (PVGxDA)F1 donor-recipient combinations. Serum was obtained before and at time points after transplantation and incubated with blood from untransplanted DA and PVG rats. Antibody binding to T cells was detected by whole blood flow cytometry using FITC-conjugated anti-rat IgM murine monoclonal antibody. Antibody levels were determined by reference to a standard curve of fluorescent intensity generated using a serum sample with known anti-target cell IgM activity. Data are presented as arbitrary units/ml (AU/ml). RESULTS: In the PVG-->DA combination, five of six DA recipients had detectable anti-graft (PVG) antibodies by day 4 after transplantation (mean 72 AU/ml) and all animals were positive by day 6 (976 AU/ml). Antirecipient (DA) antibodies were also induced, however, they were only apparent after 6 days in five of eight animals (90 AU/ml). Antigraft (DA) antibody responses were also induced in the DA-->PVG combination (day 6-218 AU/ml), however no antirecipient (PVG) response was apparent. Transplantation induced antirecipient (DA) antibodies in the unidirectional GvHD model (day 6-90 AU/ml) and an anti-graft (PVG) response in the unidirectional rejection model (day 6-60 AU/ml). However, the latter was quantitatively lower than that generated in the PVG-->DA combination (day 6-976 AU/ml). CONCLUSIONS: Antigraft and antirecipient antibody responses are simultaneously induced after fully allogeneic small bowel transplantation, despite rejection being the predominant clinical feature. Further studies are required to elucidate their influence on graft outcome.

Antimurine immunoglobulin antibody responses after the administration of murine monoclonal antibodies to rats are altered by small bowel allograft rejection.

BACKGROUND: This study monitored the induction of antimurine immunoglobulin antibody responses after the administration of anti-CD4 (OX38) and anti-LFA-1 (WT.1) monoclonal antibodies to DA rats. METHODS: Monoclonal antibody was administered i.v. on 3 consecutive days to untransplanted DA rats, and DA recipients of PVG small bowel allografts. Control animals received no monoclonal antibody. Antimurine immunoglobulin antibody levels in serum samples were determined by enzyme immunoassay. RESULTS: No antimurine immunoglobulin antibody was detected in untransplanted animals receiving OX38 alone. Reactivity was apparent in WT.1-treated animals, but this response was totally abrogated by the co-administration of OX38. A combination of OX38 and WT.1 had no effect on allograft recipient survival and antimurine immunoglobulin antibody responses were detected in all allograft recipients, irrespective of the treatment regimen. CONCLUSIONS: Although OX38 inhibited the antibody response both to itself and to WT.1 in untransplanted animals, the immune reaction induced by small bowel allograft rejection overcame this inhibitory capacity.

CD44 expression in rejecting rat small bowel allografts.

CD44 is a widely expressed cell surface protein that recognizes multiple ligands and is involved in extra- and intercellular adhesion. The precise role of CD44 in immune interactions is currently unknown, but it is believed to be a homing receptor involved in lymphocyte trafficking and inflammatory responses. This study investigated CD44 expression in intestinal tissue after heterotopic rat small bowel transplantation and assessed the effect of transplantation on intestinal epithelial cell proliferation using an antibody to the nuclear activation Ag Ki67. Lamina propria and intestinal epithelial cell expression of CD44 was graded blindly by five observers, and villus epithelial cells were noted as being positive or negative for Ki67 staining. CD44 expression was high in the lamina propria of both allografted and isografted animals; however, there was no significant difference between the two groups. In contrast, the expression of CD44 on the villus epithelium was greater in allografted animals and progressed toward the villus tips as rejection developed, declining thereafter because of loss of villus integrity. Ki67 positivity was also greater in allografted animals but did not progress toward the villus tip. This is the first reported observation of CD44 expression on intestinal epithelium that is not restricted to the crypts. The findings indicate the involvement of CD44 in the rejection process and demonstrate changes in the proliferative profile of rejecting small intestinal epithelium. Further studies into adhesion molecules, such as CD44, may help to improve understanding of graft failure and promote the development of new therapeutic approaches for controlling and preventing graft rejection.

Differential expression of adhesion molecules during rat small bowel allograft rejection.

The adhesion molecules intercellular adhesion molecule 1 (ICAM-1), lymphocyte function-associated antigen-1 (LFA-1) (alpha and beta chains), and very late activation antigen-4 (VLA-4) have an essential role in cell-cell interactions and the initiation of immune responses. This study used an indirect immunoperoxidase technique to investigate the expression of these molecules in the lamina propria of allografts and isografts after heterotopic rat small bowel transplantation. Normal untransplanted small bowel served as additional controls. Overall, ICAM-1 and LFA-1 alpha expression was significantly higher in allografts, although there was variable expression of these molecules in isografted animals. There were temporal differences in the expression of ICAM-1 and LFA-1 alpha in that increased ICAM-1 expression was more pronounced in the the early posttransplant period, whereas there was a progressive increase in LFA-1 alpha as rejection developed. In contrast, there was no difference between allograft and isograft expression of LFA-1 beta and VLA-4. This study has demonstrated a preferential increase in adhesion molecule expression with developing rat small bowel allograft rejection and suggests that adhesion molecules are involved in the development and progression of allograft rejection. Although the observed differences in antigen expression are not as marked as those previously reported in other organ transplants, appropriate adhesion molecules may present suitable targets for immunotherapeutic protocols after small bowel transplantation.

Stress responses in graft and native intestine after rat heterotopic small bowel transplantation.

BACKGROUND: Cardiac transplantation has been shown to induce heat shock protein expression, and reactivity to these stress proteins has been implicated in acute and chronic allograft rejection. This study assessed Hsp60 and Hsp70 expression in graft and native small intestine after rat small bowel transplantation. METHODS: Heterotopic small bowel transplantation was performed between PVG donor and DA recipient rats, a subgroup of which received tacrolimus immunosuppression (1 mg x kg(-1) x day(-1)). Untransplanted and isografted (PVG-->PVG) animals served as controls. Paraffin sections of graft and native intestine on day 5 after transplantation were stained by immunohistochemistry, and heat shock protein expression was graded blindly by three observers. RESULTS: Villus epithelial cell expression of Hsp60, but not Hsp70, was increased in allografts. The induction of Hsp60 in the villus epithelium was not controlled by tacrolimus. Hsp60 and Hsp70 expression was induced in the lamina propria of isografts and allografts. This response was more pronounced in allografts and was significantly reduced, but not totally abrogated, by tacrolimus. Interestingly, heat shock protein expression was also induced in the native intestine lamina propria and epithelium of allograft recipients, suggesting the induction of stress responses at sites other than the transplanted organ. CONCLUSIONS: Small bowel transplantation induces a stress response in both the graft and native intestine. The early and prolonged expression of these proteins may influence the induction of anti-heat shock protein reactivity and have an adverse effect on graft outcome after small bowel transplantation.

Evidence that orthotopic transposition following rat heterotopic small bowel transplantation corrects overgrowth of potentially pathogenic bacteria.

An overgrowth of pathogenic organisms occurs following rat heterotopic small bowel transplantation. This study assessed whether the bacterial microflora return to normal following subsequent orthotopic transposition of the graft. After 14 days the heterotopic graft was placed into continuity following resection of 15 cm of the host midintestinal loop. Quantitative and qualitative analyses of the intraluminal bacteria were performed studying the resected host intestine, the heterotopic graft at 14 days, and the graft 14 days after transposition. A group of normal rats were used as controls. An overgrowth of Staphylococcus epidermidis evident in the heterotopic graft at 14 days returned to a more normal bacterial profile following orthotopic transposition. These findings suggest that early interposition of a small bowel graft into an orthotopic position may prevent an alteration in the small bowel ecology toward potentially pathogenic organisms capable of translocation.

Activation antigen expression on peripheral blood neutrophils following rat small bowel transplantation. NKR-P1 is a novel antigen preferentially expressed during allograft rejection.

This study used flow cytometric analyses to monitor activation antigen expression (MHC class II; interleukin-2 receptor, p55IL-2R and 3.2.3/NKR-P1 antigen) on peripheral blood neutrophils following rat small bowel transplantation. The rat 3.2.3 antigen is a member of the NKR-P1 family of natural killer (NK) cell-associated molecules, which are expressed at high levels on NK cells and lymphokine-activated killer cells, and low levels on at least one T cell subset. Peripheral blood neutrophils in normal animals express very low or undetectable levels of NKR-P1. Detectable levels of NKR-P1 were induced as early as day 1 following small bowel transplantation in all allografted animals, whereas expression was only rarely detected in isografted animals. In addition, NKR-P1 density was significantly higher in allografted animals and was maintained as rejection developed. MHC class II and p55IL-2R expression was also induced following transplantation. The mechanisms of induction and functional relevance of NKR-P1 expression on neutrophils remain to be defined. However, the concomitant increased expression of MHC class II and p55IL-2R suggest NKR-P1 to be a neutrophil activation marker and implicate a potential role for NKR-P1+ neutrophils in small bowel allograft rejection. This hypothesis is further supported by the loss of detectable peripheral blood neutrophils only with developing rejection. Flow cytometric analysis of neutrophil activation antigen expression may be useful for monitoring human small bowel transplant recipients.