C. W. Ramsden

Increased intestinal permeability and altered mucosal immunity in cholestatic jaundice.

OBJECTIVE To examine the effects of cholestatic jaundice on gut barrier function. SUMMARY BACKGROUND DATA Gut barrier failure occurs in animal models of jaundice. In humans, the presence of endotoxemia indirectly implicates failure of this host defense, but this has not previously been investigated in jaundiced patients. METHODS Twenty-seven patients with extrahepatic obstructive jaundice and 27 nonicteric subjects were studied. Intestinal permeability was measured using the lactulose-mannitol test. Small intestinal morphology and the presence of mucosal immunologic activation were examined in endoscopic biopsies of the second part of the duodenum. Systemic antiendotoxin core IgG antibodies and serum interleukin-6 and C-reactive protein were also quantified. Intestinal permeability was remeasured in 9 patients 5 weeks after internal biliary drainage. RESULTS The median lactulose-mannitol ratio was significantly increased in the jaundiced patients. This was accompanied by upregulation of HLA-DR expression on enterocytes and gut-associated lymphoid tissue, suggesting immune activation. A significant increase in the acute phase response and circulating antiendotoxin core antibodies was also observed in the jaundiced patients. After internal biliary drainage, intestinal permeability returned toward normal levels. CONCLUSIONS A reversible impairment in gut barrier function occurs in patients with cholestatic jaundice. Increased intestinal permeability is associated with local immune cell and enterocyte activation. In view of the role of gut defenses in the modern paradigm of sepsis, these data may directly identify an important underlying mechanism contributing to the high risk of sepsis in jaundiced patients.

Pharmacological concentrations of lipid emulsions inhibit interleukin-2-dependent lymphocyte responses in vitro.

Most immunological functions are accomplished by means of interactions between mediator molecules (cytokines or lymphokines) and their specific receptors on the lymphocyte surface. One particular lymphokine, Interleukin-2 (IL-2) is central to the generation of most immune responses including those with antitumor activity. Prompted by two clinical trials which have suggested distinct but apparently opposite effects of lipid emulsions on the production of and lymphocyte responses to IL-2 we have examined the effects of pharmacological concentrations of three lipid emulsions currently in clinical use on IL-2 related interactions in vitro. Mitogen-stimulated and IL-2 activated human lymphocyte proliferation were both inhibited in a dose-dependent manner in the presence of all three lipid emulsions although the effects were less marked with the solution in which 50% of the calories are present as medium-chain triglycerides (MCT) rather than long-chain triglycerides (LCT). Similarly the LCT, but less so the MCT-containing solutions inhibited the generation of cytotoxic lymphokine-activated killer cells. These solutions did not inhibit the proliferation of cell lines which are not growth-factor dependent but did inhibit the growth of an IL-2-dependent cell line. We conclude that lipid emulsions can upset IL-2-dependent lymphocyte responses. These observations may lead to parenteral feeding regimens which are less immunocompromising for the tumor-bearing patient.

Inhibition of lymphokine-activated killer cell generation by cultured tumor cell lines in vitro

SummaryThe co-culture of human peripheral blood mononuclear cells (PBMC) with high concentrations of interleukin 2 normally generates lymphokine-activated killer (LAK) cells capable of indiscriminate lysis of tumor targets. However, the addition of certain cell-line-derived tumor cells to the LAK generation cultures within the first 48 h of culture initiation resulted in the suppression of the LAK cytotoxicity measured after 3–4 days of culture. Suppression could be achieved with tumor cell:PBMC ratios as low as 1:50 when tumor cells were derived from melanoma and colorectal cancer (G361, COLO320, HT-29), but suppression was not observed with cells from the breast cancer cell line SKBr3. No suppression of LAK generation was observed with normal epithelial cells from colon or breast, with autologous or allogeneic lymphoblasts, or with allogeneic vascular endothelial cells. Suppression was independent of the removal of adherent cells from PBMC, could not be prevented by indomethacin and was not attributable to interleukin 2 absorption/adsorption by tumor cells. The suppressive activity of some tumor cells could be augmented by preculture in recombinant gamma interferon. Serum-free supernatants from G361, COLO320 and HT-29 (but not SKBr3 or endothelial cells) were also highly suppressive towards the generation of LAK cells. The elaboration by tumor cells of fractors capable of inhibiting LAK generation may partially explain the failure of LAK/interleukin 2 therapy in some experimental and clinical protocols.

CHANGES IN HUMAN NATURAL KILLER ACTIVITY EARLY AND LATE AFTER RENAL TRANSPLANTATION USING CONVENTIONAL IMMUNOSUPPRESSION

The natural killer (NK) cell activity of human peripheral blood lymphocytes falls following major surgical procedures including renal transplantation but in non-immunosuppressed individuals returns to normal levels within the first 72 hr after operation. In renal allograft recipients, if this early postoperative fall is excluded from the analysis, NK cell function appears to follow changes in allograft function, suggesting that in vivo, as has been reported in vitro, NK activity is generated during activation of the alloreactive process. In an additional group of patients whose grafts were functioning for between 3 and 102 months after cadaveric renal transplantation using conventional immunosuppression, NK function was depressed in comparison with that of control subjects. However, some patients who were more than 48 months post-transplant had normal NK cell activity. Collectively, these results suggest that NK cell function may recover despite the continued administration of conventional immunosuppressive agents.

Peri‐operative modulation of cellular immunity in patients with colorectal cancer

The peri‐operalive cellular immune response is depressed in patients with gastrointestinal cancer, a factor which may facilitate maligtiant dissemination. We have investigated the effects of perioperative rIL‐2 and a combination of rlL‐2 and interferon‐alpha (IFN‐α) on both peripheral blood lymphocyte function and number in patients undergoing surgical resection for colorectal cancer. Fifty‐two patients were randomly allocated to either control, rIL‐2 or rIL‐2 with IFN‐α treatment arms. In vitro studies were performed pre‐operatively and on post‐operative days I, 4, 7 and 10. Natural ikller (NK) and lymphokine‐activated killer (LAK) cell function were profoundly depressed in control patients (P < 0.001; P < 0.001), an effect abrogated in both treatment groups; indeed NK function was augmented in the rIL‐2 and IFN‐α group on the first post‐operative day in association with an increase in the percentage of cells expressing CD16 and CD56 (P < 0.01). Flow cytometric analysis of lymphocyte subsets in the control group was unremarkable, except for an early post‐operative fall in numbers of lymphocytes. Treatment with either rIL‐2 or rIL‐2 and IFN‐α produced an initial profound reduction in T lymphocyte numbers, followed by a ‘rebound’ lymphocytosis of activated CD3+ T cells, as demonstrated by a significant increase in co‐expression of CD25, CD38, and CD45RO. No significant differences were observed between either of the treatment groups. Adjuvant immunotherapy affects peri‐operative anti‐tumour immune responses, and this may influence long term outcome in patients undergoing surgery for gastrointestinal cancer.

Reversible Impairment in Monocyte Major Histocompatibility Complex Class II Expression in Malnourished Surgical Patients

BACKGROUND Upregulation of major histocompatibility complex (MHC) class II antigen in response to the T-cell lymphokine interferon-gamma (IFN-gamma) is central to T cell-macrophage cooperation and immune homeostasis. We evaluated this property in malnourished surgical patients and assessed the impact of nutrition repletion with total parenteral nutrition (TPN). METHODS Sixty-two patients were studied: 37 malnourished and 25 controls. Whole blood was cultured with or without IFN-gamma (100 U mL-1), dual-labeled with anti-CD14 (monocyte) and anti-human leukocyte antigen-DR antibodies and analyzed by flow cytometry. Phagocytosis was measured by flow cytometry. In a second study, 10 severely malnourished patients received 5 days of TPN and MHC class II expression was measured at the end of this period. RESULTS The magnitude of the increase in monocyte MHC class II expression in response to IFN-gamma was significantly increased in the control group compared with the malnourished group (107% vs 53%; p < .05). This impairment directly correlated with severity of malnutrition, but did not correlate with age or disease type. The number of bacteria phagocytozed per cell was significantly decreased (p < .05) in the malnourished group. In study 2, there was a significant increase in MHC class II induction with IFN-gamma after short-term TPN (58% before vs 173% after, p < .001). CONCLUSIONS MHC class II induction in response to IFN-gamma is significantly impaired in malnourished patients, correlating with the severity of malnutrition. This defect is reversed by short-term TPN. These data identify the reversible loss of a key mechanism, fundamental to host defense, that may enhance the risk of infection in malnourished patients.

Natural Killer-Cell Activity, Interferon-Alpha2 Production, and Interleukin-2 Production in Cyclosporine-Treated and Conventionally Immunosuppressed Human Allograft Recipients

Natural killer (NK) activity, interferon (IFN)-alpha production, and interleukin-2 (IL-2) production were measured in renal transplant recipients undergoing immunosuppression with either azathioprine and steroids (Az + P) or cyclosporine (CyA). Overall, both IFN-alpha production and IL-2 production were impaired in these two groups compared with identical studies in healthy individuals. However, on the basis of control data these two patient groups were divided into those with “normal” NK activity and those with “low” NK activity. In the CyA group those with a low NK reaction produced less IL-2 and IFN-alpha than those with normal NK activity. No such relationship between cytokine production and NK activity was discerned in the Az + P group. These data conflict within vitro studies, which have failed to demonstrate any effect of CyA on IFN-alpha production. In addition, they suggest that whereas cyclosporine influences NK activityin vivo by inhibiting cytokine production, other factors may play a role in impairing NK activity in conventionally immunosuppressed patients.

The response to interferon of NK and K cells from conventionally immunosuppressed and cyclosporine-treated renal allograft recipients

Interferon is a potent stimulator of natural killer (NK) and killer (K) cell activity in human beings, both these cytotoxic functions representing host defense mechanisms against viral infections and lymphoid malignancy. Both NK and K cell functions are markedly impaired in conventionally immunosuppressed allograft recipients but coincubation of lymphocytes from these patients with purified human lymphoblastoid interferon considerably augments both these activities. Cyclosporine immunosuppression causes only a moderate, but significant, impairment of NK activity—but K cell activity appears to be normal. Again IFN increases NK activity of the lymphocytes of these patients but produces a fall or only moderate increases in K cell activity. We conclude that these data support the functional distinction between NK and K cells and suggest that immunosuppressive agents act at the pre-NK/K cell stage of maturation, though possibly via different mechanisms.