Immune restoration with interleukin-2 in patients with squamous cell carcinoma of the head and neck

Patients with head and neck squamous cell carcinoma commonly have depressed cell-mediated immunity which is known to correlate with ultimate prognosis. Selective immune studies were conducted in 27 head and neck cancer patients to determine the potential of interleukin-2 as an immune restorative agent. Patients showed the expected depression of lymphocyte proliferation to phytohemagglutinin and had borderline depressed natural killer cell activity and relatively normal interleukin-2 production. Addition of interleukin-2 at 100 units/ml markedly enhanced natural killer cell activity to normal levels. Serum from head and neck patients was also immune-suppressive. Heat-inactivated serum depressed lymphocyte proliferation and natural killer cell activity of control leukocytes. Lymphocyte incubation with interleukin-2 significantly counteracted immune suppressive serum effects and restored depressed lymphocyte function to normal levels. The effective in vitro interleukin-2 dose is potentially achievable by infusion at approximate doses of 3 X 10(6) units/M2.     

Intravenous fentanyl increases natural killer cell cytotoxicity and circulating CD16(+) lymphocytes in humans.

Opioids, including fentanyl, are often administered to patients who may be at risk for the consequences of impaired immune function. We performed a clinical study to test the effects of the synthetic opioid fentanyl on human immune function. Participants received an IV fentanyl initial dose of 3 microg/kg followed by a 2-h IV infusion of 1.2 microg x kg(-1) x h(-1). Peripheral blood was drawn before and after fentanyl administration to test for neutrophil phagocytic function, neutrophil antibody-dependent cell cytotoxicity, natural killer cell cytotoxicity, percentage of lymphocyte populations, T-lymphocyte proliferative response, and in vivo antibody response to a pneumococcal vaccine inoculation given at the end of the fentanyl infusion. Fentanyl exposure under the conditions of this study caused a rapid and significant increase in natural killer cell cytotoxicity, which was coincident with an increase in the percentage of CD16(+) and CD8(+) cells in peripheral blood. Fentanyl did not significantly affect any of the other immune measurements. IMPLICATIONS: Many previous studies have suggested that opioid drugs can impair immune resistance in patients who may be at risk for infection. This study suggests that the opioid fentanyl, when given to healthy humans without coexisting diseases, does not suppress immune resistance. On the basis of these results, the use of fentanyl should not be restricted because of concerns that it may suppress immune function.     

Natural killer cells and metastases from pharyngeal carcinoma

Natural killer cell activity was assessed in 100 previously untreated pharyngeal carcinoma patients. Diminished natural killer cell function in these patients was associated with an increased risk of death from uncontrolled regional and distant metastases. During the assessment, the cell line MDA686-Ln was established from a metastatic pharyngeal carcinoma of a patient with low natural killer cell cytotoxicity. The initially cytotoxicity-resistant cell line could be lysed when natural killer cell cytotoxicity was enhanced in vitro either through enrichment of a Leu 19+ natural killer cell population by fluorescent-activated cell sorting or by interleukin-2 activation. Additionally, increased circulating immune complexes were identified in these patients, subsequently isolated, and found to block natural killer cell reactivity against MDA686-Ln. In light of this negative interaction, 38 patients were randomly evaluated for both circulating immune complex levels and natural killer cell function. Both parameters examined together were complementary in defining the risk of death with disease; four of five deaths occurred in patients with both high circulating immune complex levels and low natural killer cell function. Results support the biologic modification of natural killer cell activity for controlling metastatic pharyngeal carcinoma and point to the potential confounding influence of circulating immune complex.     

The influence of hepatocellular function on NK and T cell tumoricidal activity

Recent studies by this group have demonstrated that hepatocellular integrity is important in the preservation of host cellular immune function. This study evaluated the effect of experimental hepatocellular dysfunction (EHD) on host antineoplastic defense mechanisms. In nonspecific immune studies, we examined the effect of EHD on Wistar Furth (WF) natural killer (NK) cell cytotoxicity; in specific immune studies, we assessed WF C3H/HeJ lymphocytic responsiveness to both T cell mitogen and unmodified syngeneic fibrosarcoma. In concurrent studies, we evaluated the effect of EHD on interleukin-2 (IL-2) synthesis, an important NK and T cell trophic factor. WF rats and C3H/HeJ mice were assigned to three groups: EHD induced by bile duct ligation, sham, and normal control (NC). At day 21 serum bilirubin, WF NK cytotoxicity to YAC-1 tumor cells, WF and C3H/HeJ lymphocytic responsiveness to phytohemagglutinin (PHA) and syngeneic MCA-fibrosarcoma (MCA-F), and WF T-helper IL-2 production were determined in respective groups. Serum total bilirubin was elevated in EHD rats and mice with respect to controls (p less than 0.01). Wistar Furth cytotoxicity to the YAC-1 tumor cells was depressed in EHD animals with respect to sham and NC groups at 12.5:1 (p less than 0.01), 25:1 (p less than 0.05), 50:1 (p less than 0.05), and 100:1 (p less than 0.05) effector/target cell ratios. WF T cell responsiveness to PHA was depressed in EHD with respect to controls (p less than 0.01). C3H/HeJ lymphoproliferative response to MCA-F tumor antigen was also depressed in EHD animals when compared with control groups with the addition of 12.5 X 10(3) (p less than 0.05) and 50 X 10(3) (p less than 0.05) MCA-F cells. These impairments in NK and T cell function in EHD could not be attributed to diminished IL-2 production (EHD vs sham and NC: 112,141 +/- 5232 vs 106,691 +/- 1419 and 120,759 +/- 3248 cpm, respectively). These results demonstrate that hepatocellular failure compromises NK and T cell tumoricidal function, an effect not resultant on diminished T helper IL-2 production.     

Warm ischemia induces alteration in lung immune cell functions

Warm ischemia is an important factor in early allograft dysfunction. To elucidate cellular events involved in such lung injury, we examined the effects of warm ischemia on the cytotoxic function of lymphocytes retrieved by bronchoalveolar lavage as compared with peripheral blood lymphocytes. Warm ischemia of the lung was induced in eight dogs by crossclamping left hilar structures for 1 hour. Bronchoalveolar cells from ischemia left and unaffected right lungs, as well as blood lymphocytes, were isolated before operation and 2 hours, 72 hours, and 7 days after operation. Lung and blood lymphocytes were assayed for natural killer and lectin-dependent cell-mediated cytotoxicity. Warm ischemia resulted in a significant impairment of natural killer activity within 2 hours of reperfusion (49% of preoperative control cytolysis, p less than 0.01). There was a significant increase in natural killer activity in bronchoalveolar lavage mononuclear cells 72 hours after reperfusion injury (178.4% of preoperative value, p less than 0.01). Interestingly, these functional alterations were not paralleled with changes seen in the peripheral blood lymphocytes or the opposite nonaffected lungs, where the natural killer activity appeared significantly depressed at 72 hours. Similarly, lectin-dependent cell-mediated cytotoxicity was noted to be increased in the bronchoalveolar lavage from the ischemic lung (179.5%, p less than 0.01) but decreased in the bronchoalveolar lavage from the nonaffected lung and peripheral blood lymphocytes at 72 hours after injury. We conclude that warm ischemia is associated with a functional alteration of the local lung immune cells. Such alteration is not observed in cells from the opposite lung or peripheral blood. The observed increase in nonspecific cytotoxicity of bronchoalveolar lymphocytes can be causative in the early damage seen in poorly preserved lung allografts.     

T gamma/delta lymphocytes in renal transplant recipients.

T gamma/delta lymphocytes are able to perform allospecific cytotoxicity and natural killer cytotoxicity in vitro. However, very little is known about their function in vivo. To investigate the possible involvement of T gamma/delta lymphocytes in the immune response to renal allografts, fine-needle aspiration biopsies and peripheral blood of 15 renal transplant recipients were studied during the first 4 weeks after transplantation. In addition peripheral blood of patients before transplantation, half a year and one year after transplantation was studied. No increase in the percentage of T gamma/delta lymphocytes in the fine-needle aspiration biopsies, including those taken during acute rejection episodes, was found. A significant decrease in the percentage of T gamma/delta lymphocytes was observed in peripheral blood after transplantation. We conclude that T gamma/delta lymphocytes seem to play no major role in the immune response to renal allografts.     

The effect of propofol on cytotoxicity and apoptosis of lipopolysaccharide-treated mononuclear cells and lymphocytes

IV anesthetics may inhibit proper immune responses and further compromise an already depressed defense system. To assess the possible role of propofol on human immune function in sepsis, we studied cytotoxicity, and apoptosis of mononuclear cells (MNCs). Peripheral blood MNCs were preincubated in 1 microg/mL of lipopolysaccharide (LPS) and then reincubated in different concentrations of propofol (1 microg/mL, 5 microg/mL, 10 microg/mL, or 50 microg/mL). To determine cytotoxicity, lactate dehydrogenase release was assayed by mixing MNCs (4 x 10(5)/100 microL) with K-562 tumor cells as target cells (1 x 10(4)/100 microL)(E: T ratio of 40:1). Apoptosis was determined by measuring the annexin positive cells using flow cytometry. Cytotoxicity and apoptosis of LPS-treated MNCs were unchanged by clinically acceptable concentrations of propofol (1 microg/mL, 5 microg/mL, and 10 microg/mL). However, significant differences were observed in cytotoxicity (P = 0.004) and apoptosis (P = 0.002) with propofol 50 microg/mL. By gating MNCs, we found that lymphocyte apoptosis was significantly increased at 50 microg/mL of propofol, but monocytes were unaffected (P = 0.02). In terms of cytotoxicity and apoptosis, propofol allowed MNCs to retain their cytotoxicity in septic conditions by protecting immune cells from apoptosis. IMPLICATIONS: Propofol at acceptable therapeutic concentrations, and under experimentally contrived septic conditions, did not affect the cytotoxic activity of mononuclear cells or the apoptosis level of mononuclear cells, lymphocytes, and monocytes from peripheral blood.     

Enhancement of natural cytotoxicity in lymphocytes from animals with carcinogen-induced bladder cancer

Splenic lymphocytes from Fischer rats with carcinogen (FANFT)-induced bladder cancer had depressed natural cytotoxicity that could be enhanced in vitro by the addition of mouse leukocyte interferon to the cytotoxicity assay. Such enhancement appeared to reflect an effect directly on lymphocytes rather than a cytotoxic effect on tumor target cells. The possibility that tumor cell prostaglandin production might partially inhibit lymphocyte cytotoxicity and its enhancement prompted separate attempts to enhance cytotoxicity by inhibiting prostaglandin production during cytotoxicity testing. However, addition of indomethacin to the cytotoxicity assays did not enhance cytotoxicity in lymphocytes from either control or tumor-bearing rats and did not add to the enhancement seen with interferon. Addition to the cytotoxicity assay of unstimulated peritoneal monocytes which themselves have been shown to produce prostaglandins, did not effect lymphocyte cytotoxicity. Correspondingly, indomethacin in parallel samples did not alter baseline levels of cytotoxicity seen. Further stimulation of cytotoxicity by addition of interferon to these samples was also not seen. Taken together, in vitro enhancement of depressed lymphocyte cytotoxicity in tumor-bearing animals was possible with exogenous leukocyte interferon, could not be accomplished by inhibition of prostaglandin production in this system, and did not appear to be influenced by the addition or deletion of monocytes during cytotoxicity testing.     

Negative effect of uraemia and cuprophane haemodialysis on natural killer cells

The evidence indicating the important role of natural killer (NK) cells in immune surveillance against tumours and certain infections is accumulating. Uraemic and dialysed patients are known to be at greater risk of infections and malignant diseases. NK cells were analysed in patients with advanced uraemia, and in patients treated with different dialysis techniques. Number of NK cells was morphologically identified as large granular lymphocytes in blood smears. NK activity was determined as mononuclear cell cytotoxicity against K562 cells. In a group of uraemic patients, large granular lymphocyte number was reduced to 39%, and NK activity to 41%-52% of control values. Large granular lymphocyte number and NK activity in patients haemodialysed on cuprophane membranes was significantly reduced, compared to corresponding values in controls and uraemic patients, declining to 17% and 8%-16% of respective control values. In a group of patients treated by CAPD, and in a group haemodialysed on polyacrylonitrile membranes, NK activity was close to values in the uraemic group, but significantly greater than those of cuprophane-haemodialysed patients. Haemodialysis on cuprophane membranes has an additional negative effect on NK cells, which are already seriously depressed by the uraemic state.     

Comparison of in vitro glioma cell cytotoxicity of LAK cells from glioma patients and healthy subjects

Peripheral blood mononuclear cells from 11 glioma patients and 11 healthy control subjects were cultured in medium containing recombinant interleukin-2 for a period of 5 days. The cytotoxicity of these lymphokine-activated killer (LAK) cells was tested on chromium-51-labeled freshly prepared allogeneic glioblastoma cells, and on the cell lines K562 (natural killer cell (NK)-sensitive) and Daudi (NK-resistant). Peripheral blood mononuclear cells from all subjects showed high levels of cytotoxicity against these targets. There was no significant difference between the patients and the control group when LAK cytotoxicity was compared. Thus, although glioma patients are known to have depressed immunological reactivity, the cytotoxic capacity of LAK cells derived from glioma patients is similar to that of LAK cells from healthy control subjects. However, the glioma patients had significantly reduced numbers of mononuclear cells in their peripheral blood, possibly due to steroid treatment. Therefore, the volume of blood required to generate the same number of LAK cells was approximately three times larger from the glioma patients than from control subjects.     

Functional analysis of interleukin-2 in immune surveillance against brain tumors

We studied the capacity of peripheral blood lymphocytes (PBLs) from patients with malignant brain tumors to produce interleukin-2 (IL-2) and to respond to IL-2. The role of IL-2 in the generation of T cells cytotoxic against tumor cells was also studied. PBLs from the patients with malignant brain tumors tended to produce a level of IL-2 lower than that in normal controls because of the decreased number of IL-2-producing T cells. Phytohemagglutinin activated PBLs from normal controls and the patients, however, responded equally well to IL-2. This indicates that the expression of IL-2 receptors is abundant in PBLs of these patients, although IL-2 production may be depressed. Furthermore, after incubation with IL-2, PBLs from the patients with malignant glioma exhibited higher natural killer activity and strong cytotoxicity against glioma cells. This increased cytotoxicity was evident by Day 3 of culture in IL-2 and remained effective for at least 2 days. These observations of antitumor cytotoxicity make IL-2 a likely candidate for use in adoptive immunotherapy.     

Functional properties of natural killer cells in carcinoma of the prostate

Functional properties of natural killer cells were analyzed in a 51chromium-release assay by modulating their activity with human beta-interferon using 1) peripheral mononuclear cells of 16 patients with carcinoma of the prostate and 7 healthy male donors, and 2) mononuclear cells from the periprostatic lymph nodes of 6 patients with stage pT2N0M0 prostatic cancer and 5 without malignancy. Cell lines EB 33, PC 3, DU 145 and CaKi 1 were used as target cells. Spontaneous cell-mediated cytotoxicity of peripheral mononuclear cells was depressed in patients with advanced prostatic cancer; however, the natural killer cells responded as those of the healthy controls to beta-interferon. The beta-interferon effect was time and dose dependent. In mononuclear periprostatic lymph node cells virtually no spontaneous cell-mediated cytotoxicity was detected. The reactivity of mononuclear periprostatic lymph node cells from patients with prostatic cancer was significantly less improved by beta-interferon stimulation than the mononuclear periprostatic lymph node cells of healthy subjects. The stimulation of mononuclear periprostatic lymph node cells by beta-interferon was reduced significantly compared to simultaneously tested autologous peripheral mononuclear cells.     

The effects of tramadol and morphine on immune responses and pain after surgery in cancer patients

There has been growing interest in determining the possible immune consequences of opioid administration for the management of postoperative pain. We studied the effects of morphine and tramadol on pain and immune function during the postoperative period in 30 patients undergoing abdominal surgery for uterine carcinoma. Phytohemoagglutinin-induced T lymphocyte proliferation and natural killer cell activity were evaluated immediately before and after surgery, and 2 h after the acute administration of either 10 mg of morphine IM or 100 mg tramadol IM for pain. In all patients, phytohemagglutinin-induced lymphoproliferation was significantly depressed by surgical stress. However, in the morphine-treated group, proliferative values remained lower than basal levels for 2 h after treatment, whereas in tramadol-administered patients proliferative values returned to basal levels. Natural killer cell activity was not significantly affected by surgery nor by morphine administration, whereas tramadol significantly enhanced the activity of natural killer cells. Both drugs produced a comparable reduction in postoperative pain. We conclude that, as previously observed in the experimental animal, tramadol and morphine, when administered in analgesic doses, induce different immune effects. Implications: Recent studies suggest that opioids can have an adverse impact on the immune system. Because surgical stress also induces immune dysfunction, the search for analgesic drugs devoid of immunosuppressive effects is of import. This study compared the effects on immune responses of morphine and of the atypical opioid analgesic, tramadol, given for postoperative pain to gynecological cancer patients. Tramadol and morphine showed comparable analgesic activity; however, tramadol, in contrast to morphine, induced an improvement of postoperative immunosuppression and, therefore, may be preferred to morphine for the treatment of postoperative pain.     

胃癌围手术期T细胞亚群和NK细胞活性检测的临床意义

目的 探讨胃癌围手术期细胞免疫功能的变化及与肿瘤分期的关系。方法 采用流式细胞仪测定40例胃癌患者围手术期外周血T淋巴细胞亚群和NK细胞活性。结果 胃癌患者术前外周血CD_3~+细胞、CD_4~+细胞、CD_4~+/CD_8~+比值和NK细胞活性明显低于正常对照组,CD_8~+细胞显著升高,并与病理分期相关,分期越晚改变越大。手术后以上指标可显著改善。结论 胃癌患者细胞免疫功能低下,病理分期越晚细胞免疫功能越低,肿瘤负荷是其主要原因。     

Impairment of helper T-cell function and lymphokine-activated killer cytotoxicity following severe head injury.

Infection is a major complication of severe head injury, occurring in 50% to 75% of patients who survive to hospitalization. Previous investigations of immune activity following head injury have demonstrated suppression of helper T-cell activation. In this study, the in vitro production of interferon-gamma (INF-gamma), interleukin-1 (IL-1), and interleukin-2 (IL-2) was determined in 25 head-injured patients following incubation of peripheral blood lymphocytes (PBL's) with the lymphocyte mitogen phytohemagglutin (PHA). In order to elucidate the functional status of cellular cytotoxicity, lymphokine-activated killer (LAK) cell cytotoxicity assays were performed both prior to and following incubation of PBL's with IL-2 in five patients with severe head injury. The production of INF-gamma and IL-2 by PHA-stimulated PBL's was maximally depressed within 24 hours of injury (p less than 0.001 for INF-gamma, p = 0.035 for IL-2) and partially normalized within 21 days of injury. There was no change in the production of IL-1. When comparing the in vitro LAK cell cytotoxicity of PBL's from head-injured patients and normal subjects, there was a significant depression in LAK cell cytotoxicity both prior to (p = 0.010) and following (p less than 0.001) incubation of PBL's with IL-2. The results of this study indicate that IL-2 and INF-gamma production, normally required for inducing cell-mediated immunity, is suppressed following severe head injury. The failure of IL-2 to enhance LAK cell cytotoxicity suggest that factors other than decreased IL-2 production, such as inhibitory soluble mediators or suppressor lymphocytes, may be responsible for the reduction in cellular immune activity following severe head injury. These findings may have significant implications in designing clinical studies aimed at reducing the incidence of infection following severe head injury.     

阻塞性黄疸患者手术前后红细胞免疫功能和T淋巴细胞亚群的变化

为探讨阻塞性黄疸患者免疫功能的变化，采用红细胞Ｃ３ｂ受体花环率、红细胞免疫复合物花环率、肿瘤红细胞花环率及ＣＤ＋３、ＣＤ＋４、ＣＤ＋８细胞作为观测指标，测定了３５例阻塞性黄疸患者免疫功能，并与３０例正常人比较。结果显示：疾病组患者术前红细胞及Ｔ淋巴细胞免疫功能低于正常人（Ｐ＜０．００１），术后５天变化不大，术后１４天较术前１天明显增高（Ｐ＜０．０５），但尚未恢复正常。提示解除胆道梗阻及切除肿瘤有助于阻塞性黄疸患者免疫功能的恢复。     

Immunologic characteristics of fibrillary glomerulonephritis.

IgG and IgA immune complexes, mononuclear phagocytic system function, interleukin-2 (IL-2) production by peripheral blood lymphocytes (PBL), serum-soluble IL-2 receptors, tumor necrosis factor, beta 2-microglobulin and IL-1 beta, HLA-DNA polymorphisms, immuno-isoelectrofocusing, phenotype of PBL, lymphocyte cytotoxicity, activation of lymphokine-activated killer cells and natural killer cell activity were evaluated in 8 patients with tubular/fibrillary glomerulonephritis (GN). No common serologic, immunologic or immunogenetic features suggestive of plasma cell dyscrasias were found. No elements to state whether these GNs represent a new entity or just atypical forms of known GN were found.     

Keyhole limpet hemocyanine can enhance the natural killer activity of patients with transitional cell carcinoma of the bladder

It has been shown that natural killer (NK) cells are involved in the immunosurveillance of tumors. In patients with transitional cell carcinoma (TCC) of the bladder, there is a negative correlation between the levels of NK activity in peripheral blood mononuclear cells (PBMNCs) and both the clinical evolution and the pathological stages of the disease. We have investigated the immunoregulatory effect of keyhole limpet hemocyanin (KLH) on the NK activity of patients with TCC of the bladder. We found that KLH enhances the cytotoxic activity of PBMNC from patients with superficial TCC against NK-sensitive target cells in a time-dependent manner. This KLH-enhanced cytotoxicity is not directed against NK-resistant target cells. However, KLH fails to normalize the depressed NK activity of PBMNCs from patients with infiltrative TCC. The present in vitro studies suggest that this immunoregulatory effect of KLH on NK cytotoxicity may be implicated in its therapeutic effect on superficial TCC of the bladder.     

Foxp3 is critical for human natural CD4+CD25+ regulatory T cells to suppress alloimmune response

Naturally occurring CD4+CD25+ regulatory T cells (nTregs) that express high level of Foxp3 actively suppress pathological and physiological immune responses, contributing to the maintenance of immunological self-tolerance and immune homeostasis. Although Foxp3 is required for nTreg development and appears to be necessary for mature murine Treg function, the precise role of Foxp3 in regulating natural human Treg function in alloimmune response is unclear. In this study, we used siRNA-mediated gene silencing to knockdown Foxp3 expression in natural human Tregs and investigated the importance of Foxp3 in maintaining human nTreg suppressive function. We showed that Foxp3 knockdown resulted in impaired phenotype and nonresponsiveness, downregulated expression of function molecules, and reduced production of suppressive cytokines in nTregs. These changes correlated with diminished nTreg activity in suppressing proliferation of effector CD4+CD25- T cells, their cytotoxicity against allogeneic target cells and production of effector cytokines in response to allogeneic stimulation. Thus, this study shows that ongoing Foxp3 expression is required for natural human Tregs to maintain their phenotype and suppressive function in the alloimmune response.     

Immunoregulatory cell function in peripheral blood leukocytes of patients with intracranial gliomas

Levels of indomethacin-sensitive, glass-adherent, preculture-sensitive, and lymphocyte-mediated immunoregulatory activity were measured in peripheral blood mononuclear cells from 12 patients with intracranial astrocytomas. The levels of regulatory cell function were determined in assays of T cell immunocompetence as judged by phytohemagglutinin (PHA)-induced in vitro lymphocyte DNA synthesis. Of 6 patients studied before surgical exploration and diagnosis, 6 exhibited significantly depressed levels of PHA responsiveness in association with significantly increased levels of regulatory cell function by glass-adherent, preculture-sensitive and/or indomethacin-sensitive cells. Six patients were studied after diagnosis and treatment. Of those, 2 in remission had normal levels of immune function in association with normal levels of regulatory cell activity, whereas 2 of 4 patients who had recurrent disease had significantly depressed T cell function in association with increased glass-adherent, preculture-sensitive and/or indomethacin-sensitive regulatory cell activity. Although some alterations in lymphocyte-medicated suppressor cell activity were seen in 2 patients, those changes could not be correlated with impaired T cell function as measured in the PHA stimulation assay. The changes in regulatory cell function could not be correlated with levels of immune complexes in patient sera. These data suggest that increased levels of regulatory cell function by glass-adherent, preculture-sensitive and/or indomethacin-sensitive cells alone or in conjunction with lymphocyte and T cell depletion are a primary determinant of impaired immunocompetence in glioma patients.