Interleukin-2-activated killer cell activity in colorectal tumor patients: evaluation of in vitro effects by prothymosin alpha1

The effects of prothymosin alpha1 (Pro alpha1) in combination with interleukin-2 (IL-2) on peripheral blood lymphocytes from 50 colorectal tumor patients at different stages were studied with respect to immunocytotoxicity, adhesion to cultured SW620 colon carcinoma cells, secretion of cytokines and expression of adhesion and surface marker molecules. On average, the patients showed lower natural killer (NK) cell activity than healthy donors, which was associated with a lower adhesion capacity to the tumor target cells. The NK cell activity of the patients was inversely related to the tumor stage. The generation of lymphokine(IL-2)-activated killer (LAK) cell activity was found to be comparable on lymphocytes from healthy individuals and patients and was not correlated to tumor stage. Pro alpha1 stimulated patients' LAK cell activity only, primarily at the early stage (Dukes A/B). The Pro alpha1 effect was associated with an increased adhesion of lymphocytes to tumor target cells and an increased secretion of the deficient IL-2-induced IFN gamma secretion. No significant effects on the low level of TNF alpha secretion was noted. By flow cytometry, Pro alpha1 in combination with IL-2 augmented the expression of the NK cell markers CD56, CD16/56, the subset CD3/16/56 and CD25 on lymphocytes of the patients. In contrast, Pro alpha1 was equally effective by increasing the expression of CD18 and CD11a on lymphocytes from the patients and from normal controls. In conclusion, Pro alpha1, in combination with IL-2, can partially normalize lymphocyte deficiencies of colon cancer patients in vitro. This potential might provide an experimental basis for applying Pro alpha1 or related thymic peptides in selected immunotherapies against colorectal tumors.     

Relative contribution of endothelial cell and polymorphonuclear neutrophil activation in their interactions in systemic inflammatory response syndrome

OBJECTIVE: To examine the relative contribution of polymorphonuclear neutrophil (PMN) vs endothelial cell (EC) activation on the adherence and subsequent killing of ECs by PMNs. DESIGN: In vitro comparative studies of PMN-EC adherence and cytotoxicity. SETTING: Research laboratory and the surgical intensive care unit of a tertiary-level university hospital. PATIENTS: Patients with systemic inflammatory response syndrome admitted to the surgical intensive care unit and hospitalized preoperative noninfected surgical patients. INTERVENTION: None. METHODS: Polymorphonuclear neutrophils were isolated from 21 healthy volunteers, 22 preoperative patients, and 30 patients from the surgical intensive care unit with systemic inflammatory response syndrome. The PMNs were activated with lipopolysaccharide, 100 ng/mL (Escherichia coli 0111:b4), for 40 minutes at 37 degrees C before the adherence and cytotoxicity assays. Human umbilical vein endothelial monolayers were stimulated with tumor necrosis factor alpha, 25 ng/mL, and interleukin 1 beta, 15 U/mL, for 3 hours. The PMNs or EC cells were labeled with sodium chromate Cr 51 and used in a standard adherence or killing assay as required. RESULTS: Control and preoperative patient PMN treatment with lipopolysaccharide produced a modest increase in adherence. The PMNs from patients with systemic inflammatory response syndrome showed moderately increased human umbilical vein endothelial cell adherence, and this could not be augmented further with lipopolysaccharide stimulation. There was a marked increase in PMN adherence to EC after EC activation in all study groups (P < .001). Similar to the adherence data, human umbilical vein endothelial cell cytotoxicity was significantly increased in all groups after human umbilical vein endothelial cell activation (P < .01) but not after PMN stimulation with lipopolysaccharide. CONCLUSION: These data suggest that stimulation of ECs is far more important in producing increased adherence and cytotoxicity of EC than PMN stimulation with lipopolysaccharide in all study groups. Therapeutic efforts in patients with systemic inflammatory response syndrome should be focused on the EC.     

Elevated prostaglandin E2 production by monocytes is responsible for the depressed levels of natural killer and lymphokine-activated killer cell function in patients with breast cancer

BACKGROUND: Human natural killer (NK) cells mediate spontaneous cytotoxicity against tumor cells and represent the main precursors of lymphokine-activated killer (LAK) cell activity. A comparison of some aspects of NK and LAK cell activity was undertaken in 85 preoperative patients with breast cancer and 75 healthy donors. METHODS: NK cell activity (tested in 18-hour cultures of effector peripheral blood mononuclear cells [PBMC] with K562 or MOLT-4 tumor target cells) was significantly diminished in these patients as it was the fully mature LAK cell activity (i.e., interleukin-2 (IL-2)-induced cytotoxicity in PBMC) against NK resistant target cells. Using immunoenzymatic methods we showed that the reduced NK cell activity was due to abnormally high levels of prostaglandin E2 (PGE2) produced by monocytes in culture. RESULTS: PGE2 was found to suppress the production of IL-2 in these cultures. Removal of monocytes from PBMC restored to almost normal levels the deficient NK and LAK cell activity in patients with breast cancer and was also associated with a normalization in the levels of PGE2 and IL-2. Indomethacin and gamma-interferon (IFN-gamma) increased the NK and LAK cell activity in these patients up to the levels of healthy donors. When highly purified CD56+ cells (obtained by an immunomagnetic isolation technique) were used as effector cells, no differences in LAK cell activity could be noticed between healthy donors and patients with cancer. FACS and northern blot analyses demonstrated a PGE2-mediated down-regulation of IL-2 receptor (IL-2R) expression on CD56+ cells that correlated with reduced LAK cell activity. This inhibitory effect of PGE2 was noticeable in long-term LAK cultures and was abrogated in the presence of IFN-gamma or indomethacin. CONCLUSION: This study may have important implications in the potentiation of NK and LAK cell activity for immunotherapeutic protocols in patients with breast cancer.     

Monocytes from Wiskott-Aldrich patients display reduced chemotaxis and lack of cell polarization in response to monocyte chemoattractant protein-1 and formyl-methionyl-leucyl-phenylalanine

Wiskott-Aldrich syndrome (WAS) is an X-linked disorder characterized by trombocytopenia, eczema, and progressive decline of the immune function. In addition, lymphocytes and platelets from WAS patients have morphologic abnormalities. Since chemokines may induce morphologic changes and migration of leukocytes, we investigated the monocyte response to chemoattractants in cells from WAS patients with an identified mutation in the WAS protein gene. Here, we report that monocytes derived from four patients with molecularly defined typical WAS have a severely impaired migration in response to FMLP and to the chemokines monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1alpha compared with normal donors. Conversely, neither MCP-1 binding to monocytes nor induction of the respiratory burst by MCP-1 and FMLP is significantly different between WAS patients and normal donors. Within a few minutes of stimulation, monocytes respond to chemokines with increased expression of adhesion molecules and with morphologic changes such as cell polarization. Although up-regulation of CD11b/CD18 expression following stimulation with FMLP or MCP-1 is preserved in WAS patients, cell polarization is dramatically decreased. Staining of F-actin by FITC-phalloidin in monocytes stimulated with chemoattractants shows F-actin to have a rounded shape in WAS patients, as opposed to the polymorphic distribution of F-actin in the polarized monocytes from healthy donors. These results suggest that WAS protein is involved in the monocyte response to the chemokines MCP-1 and macrophage inflammatory protein-1alpha.     

Release of soluble tumor necrosis factor receptors from polymorphonuclear cells of breast cancer patients

The release of soluble tumor necrosis factor receptors (sTNF-Rs): soluble tumor necrosis factor receptor I (sTNF-RI) and soluble tumor necrosis factor receptor II (sTNF-RII) by polymorphonuclear cells (PMNs) derived from patients with breast cancer were measured. Serum levels of sTNF-RII and sTNF-RII in patients group were also determined. Significantly higher values of sTNF-RI and sTNF-RII released by PMNs of patients were observed than those in the healthy subjects. There was no significant differences in the serum levels of sTNF-RI and sTNF-RII between breast cancer patients and healthy control. Presented study demostrate that the tumor bearing state, involving breast cancer, may induce an altered soluble cytokine receptors release from the peripheral blood PMNs characterized by increased concentrations of sTNF-RI and sTNF-RII in the culture supernatants of these cells in vitro.     

Role of endothelin-1 in astrocyte responses after acute brain damage

The mucosal injury of active ulcerative colitis (UC) could involve enhanced migration and activation of neutrophils (PMNs). Because, in vitro, PMNs from patients with UC appear normal and are not therefore a likely cause for the enhancements, we hypothesized an abnormal colonic milieu. We previously found that factors in the UC colonic milieu markedly increase production of reactive oxygen species (ROS) by control PMNs. We now hypothesize that these factors also regulate PMN surface integrins, that regulation of UC PMNs is different than that of control PMNs, and that the integrin regulation is consistent with the ROS regulation. Using rectal dialysis, we sampled the colonic milieu in patients with active UC, in patients with inactive UC, and in control subjects. We monitored a key PMN adhesion molecule, CD11b. When control PMNs were tested, active UC rectal dialysate was almost as effective (+115%) as N-formyl-methionyl-leucyl-phenylalanine (+132%) in up-regulating CD11b. When inactive UC PMNs were tested, baseline CD11b was 50% higher than that for control PMNs. In contrast, rectal dialysates failed to up-regulate CD11b of inactive UC PMNs and in fact down-regulated CD11b. Preincubating control PMNs with UC rectal dialysates converted their CD11b response to PMN activators from up-regulation to down-regulation, mimicking inactive UC PMNs. Changes in intracellular calcium levels paralleled these changes in CD11b. Rectal dialysate-induced changes in both CD11b and calcium paralleled our previous findings on rectal dialysate-induced changes in ROS production. Thus the net overall effect of factors in the colonic milieu is a consistent and predictable regulation of PMN function--proinflammatory in UC, anti-inflammatory in control subjects. These factors may be a critical part of the pathophysiology of UC.     

Naturally processed class II epitope from the tumor antigen MUC1 primes human CD4+ T cells

Epithelial cell mucin MUC1 is expressed on adenocarcinomas in an underglycosylated form that serves as a tumor antigen in breast, pancreatic, ovarian, and other tumors. Two predominant MUC1-specific immune responses are found in patients: CD8+ CTLs, which recognize tandemly repeated epitopes on the MUC1 protein core, and IgM antibodies. There have been no reports to date of MUC1-specific CD4+ T-helper cells in cancer patients. We show here that MUC1-specific CD4+ T cells are neither deleted nor tolerized and that CD4+ T cell responses can be generated when an appropriate soluble form of MUC1 is used. Naive CD4+ T cells from healthy donors were primed in vitro to a synthetic MUC1 peptide of 100 amino acids, representing five unglycosylated tandem repeats, presented by dendritic cells. They produced IFN-gamma and had moderate cytolytic activity. We identified one core peptide sequence, PGSTAPPAHGVT, that elicits this response when it is presented by HLA-DR3.     

Carney complex, Peutz-Jeghers syndrome, Cowden disease, and Bannayan-Zonana syndrome share cutaneous and endocrine manifestations, but not genetic loci

Carney complex (CC), Peutz-Jeghers syndrome (PJS), Cowden disease (CD), and Bannayan-Zonana syndrome (BZS) share clinical features, such as mucocutaneous lentigines and multiple tumors (thyroid, breast, ovarian, and testicular neoplasms), and autosomal dominant inheritance. A genetic locus has been identified for CC on chromosome 2 (2p16), and the genes for PJS, CD, and BZS were recently identified; genetic heterogeneity appears likely in both CC and PJS. The genes for PJS and CD/BZS, STK11/LKB1 and PTEN, respectively, may act as tumor suppressors, because loss of heterozygosity (LOH) of the PJS and CD/BZS loci has been demonstrated in tumors excised from patients with these disorders. We studied 2 families with CC in whom the disease could not be shown to segregate with polymorphic markers from the 2p16 locus. Their members presented with lesions frequently seen in PJS and the other lentiginosis syndromes. We also tested 16 tumors and cell lines established from patients with CC for LOH involving the PJS and CD/BZS loci. DNA was extracted from peripheral blood, tumor cell lines, and tissues and subjected to PCR amplification with primers from microsatellite sequences flanking the STK11/LKB1 and PTEN genes on 19p13 and 10q23, respectively, and a putative PJS locus on 19q13. All loci were excluded as candidates in both families with LOD scores less than 2 and/or by haplotype analysis. LOH for these loci was not present in any of the tumors that were histologically identical to those seen in PJS. The overall rate of LOH for the PJS and CD/BZS loci in tumors from patients with CC was less than 10%. We conclude that despite substantial clinical overlap among CC, PJS, CD, and BZS, LOH for the STK11 and PTEN loci is an infrequent event in CC-related tumors. Linkage analysis excluded the PJS and CD/BZS loci on chromosomes 19 (19p13 and 19q13) and 10 (10q23) from harboring the gene defect(s) responsible for the phenotype in these 2 families.     

Clinical aspects of transplantability of human gastric and colorectal carcinomas in nude mice

Transplantability of human tumors into nude mice might represent higher grade of malignancy. In our institution, surgically resected tumors have been subjected to transplantation into nude mice since 1975. We retrospectively investigated clinical outcomes of donor patients with gastric and colorectal cancer who underwent operation and analyzed the relationship between the transplantability and clinical characteristics of the patients. Thirty five out of 119 gastric tumors were successfully transplanted into nude mice with a 29.4% take rate. Results of transplantation among 92 patients' tumor in stage 2, 3 and 4 revealed 35 takes and 57 non-takes. There were only 3 ten-year survivors in the 'take' group while there were 17 ten-year survivors in the 'non-take' group. Patients with tumors which are transplantable into nude mice appeared to have poor prognosis (p < 0.05). The take rate with fifty colorectal tumors was 50%. Among 46 patient's tumors in stage 2, 3 and 4, there were 25 takes and 21 non-takes. Ten-year survivors included 11 'take' donors and 7 'non-take' donors. As opposed to gastric cancers, no statistically significant difference in survival was observed between the two groups.     

Human CD8+ TCR-alpha beta(+) and TCR-gamma delta(+) cells modulate autologous autoreactive neuroantigen-specific CD4+ T-cells by different mechanisms

To investigate the regulatory interactions among autologous T-cells during the course of multiple sclerosis (MS), proteolipid protein peptide-specific CD4+ T-cell clones (TCCs) were irradiated and used as immunogens to stimulate purified populations of autologous CD8+ TCR-alpha beta+ and TCR-gamma delta+ T-cells isolated from the peripheral blood of MS patients, patients with other non-inflammatory neurological diseases, and healthy blood donors. The resulting blasts were expanded in the presence of hIL-2 and then cloned by limiting dilution. Two different groups of CD8+ TCCs were revealed. A first group of CD8+ TCCs recognized autologous CD4+ T-cells based in their TCRV beta structures (anti-idiotypic responsiveness). A second group of CD8+ TCCs recognized Ag activated autologous CD4+ TCCs irrespective of their Ag specificity or TCRV beta expression (anti-ergotypic responsiveness). Both groups showed MHC class I restricted cytotoxicity against CD4+ T-cells and were able to secrete IFN-gamma, TNF alpha/beta and TGF-beta. TCR-gamma delta+ TCCs isolated in response to stimulation with autologous peptide-specific CD4+ TCCs showed only anti-ergotypic cytotoxicity, which was not inhibited by anti-MHC class Ia monoclonal antibodies. Moreover, they were able to secrete IFN-gamma and TNF alpha/beta, but not TGF-beta. These data demonstrate that regulatory mechanisms among human autologous T-cells can be mediated by cytolytic interactions or by the release of specific cytokines. Furthermore, they provide evidence that CD8+ TCR-alpha beta+ and TCR-gamma delta+ cells differ in their patterns of recognition and in their abilities to modulate the immune response mediated by autologous autoreactive CD4+ T-cells.     

Dysregulated expression of CD66a (BGP, C-CAM), an adhesion molecule of the CEA family, in endometrial cancer

CD66a (BGP, C-CAM) is an adhesion molecule of the carcinoembryonic antigen family that has been shown to be down-regulated in colorectal, prostate, and breast cancers. The purpose of the present study was to determine its expression pattern in the normal human endometrium and in endometrial neoplasia. For this purpose, we performed immunohistochemistry using the 4D1/C2 monoclonal antibody on a series of 24 normal endometrial samples and 47 endometrial carcinomas. Strong CD66a expression was observed in glandular and luminal epithelial cells of the normal endometrium with a consistent localization at the apical poles of these cells throughout the cycle. In late secretory (premenstrual) phase, loss of cellular polarity resulted in a membranous expression pattern in some glandular cells. In the analyzed tumor samples increasing areas with a complete loss of expression were observed with increasing malignancy grade. The apical expression pattern of the normal epithelium was changed to a membranous all-around pattern in 55% of the tumors, mostly in solid areas. This change correlated with malignancy grade and could be observed in 3 of 15 G1 tumors, 4 of 12 G2 tumors, 11 of 12 G3 tumors, and 8 of 8 serous-papillary carcinomas. Areas with membranous expression pattern could be observed along with areas with a normal apical expression pattern in lower grade carcinomas and with areas with complete loss of expression in high grade tumors. Northern blot analysis showed a loss of mRNA expression in tumor samples and HEC-1B endometrial adenocarcinoma cells. Loss of protein expression in the tumor samples was also observed by Western blot. In conclusion, CD66a protein expression is dysregulated in endometrial carcinomas, showing reduction or loss of expression with increasing malignancy grade and a change from the apical to a membranous localization.     

Constitutive up-regulation of integrin-mediated adhesion of tumor-infiltrating lymphocytes to osteoblasts and bone marrow-derived stromal cells

Tumor-reactive T cells, known as tumor-infiltrating lymphocyte(TIL)s are known to infiltrate various tumors. Although TILs exert cytotoxic activities against tumor cells, only a small percentage of tumors usually contain TILs that specifically react to tumor antigens. Because the exact role of these lymphocytes is unclear, we investigated the mechanisms of migration and adhesion of TILs to bone metastatic tumors, particularly to osteoblasts and bone marrow-derived stromal cell(BMSC)s. Histopathological examination showed that most TILs in secondary bone metastatic tumors (from primary tumors in the lung or breast) were found in the supporting tissue stroma between the bone and tumor mass. Cultured TILs (obtained from breast tumors) adhered spontaneously to osteoblasts and BMSCs (obtained from patients with osteoarthritis) without exogenous stimulation. Adhesion was further enhanced by chemokines macrophage inflammatory protein (MIP)-1alpha and MIP-1beta. TILs highly expressed activation antigens CD25 and CD69. A spontaneous activation of an integrin, lymphocyte function-associated antigen-1 (LFA-1), was also detected on TILs. TILs produced high concentrations of MIP-1alpha and MIP-1beta and spontaneous polymerization of cytoskeletal F-actin was observed in these cells. Adhesion of TILs to osteoblasts and BMSCs via LFA-1 and very late antigen-4 was associated with the production of osteoclastogen interleukin 6 by the latter cells. Our results indicate that integrins on TILs are activated in an autocrine manner by MIP-1alpha and MIP-1beta, and that treatment with the chemokines increases the binding of TILs on osteoblasts and stromal cells via a mechanism involving intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 as targets for the integrin. Our data also indicated that interactions between TILs and osteoblasts/stromal cells lead to the secretion by the latter of the osteoclastogenic cytokine interleukin 6.     

Neutrophil transfusions: kinetics and functions of neutrophils mobilized with granulocyte-colony-stimulating factor and dexamethasone

BACKGROUND: The collection of adequate numbers of neutrophils (polymorphonuclear leukocytes, PMNs) from normal donors has long hampered the development of neutrophil transfusion therapy. The stimulation of donors with granulocyte-colony-stimulating factor (G-CSF) plus dexamethasone is a promising way of improving PMN collections. STUDY DESIGN AND METHODS: Sixteen normal subjects received G-CSF (600 micrograms subcutaneously) and dexamethasone (8 mg by mouth) 12 hours before leukapheresis. Measurements included PMN morphology, immunophenotype analysis, chemiluminescence, bactericidal activity, in vivo kinetics, and adverse effects. RESULTS: A mean of 77.4 +/- 6.4 x 10(9) PMNs was collected with each leukapheresis; 14 percent were bands. PMNs had increased surface expression of CD11b, CD18, CD14, CD32, and CD64. Bactericidal capacity against Staphylococcus aureus was normal. Inducible respiratory burst was maintained, although the responses to some agonists were diminished. Returned leukapheresis cells labeled with 3H-diisopropylfluorophosphate had a modestly decreased percentage of recovery and circulated with a prolonged half-life. Migration of these cells to skin chambers was approximately equal to that of the subjects' own blood PMNs. Adverse effects included transient bone pain, headache, hunger, and insomnia. CONCLUSIONS: Precollection treatment of leukapheresis donors with G-CSF plus dexamethasone is an effective way to enhance the collection of PMNs with normal or near-normal functional properties for PMN transfusion therapy.     

Motility-related protein-1 (MRP-1/CD9) reduction as a factor of poor prognosis in breast cancer

The application of reliable markers is of major importance for predicting the prognosis of and instituting the appropriate postsurgical treatment of patients with breast cancer. Previously we showed that motility-related protein-1 (MRP-1), which is identical to CD9, regulates cell motility, and that cultured tumor cells transfected with MRP-1/CD9 cDNA have low motility and low metastatic potential. In addition, MRP-1/CD9 immunoblotting and immunohistochemical study with breast cancer revealed that MRP-1/CD9 expression diminished as the clinical stage of a given breast cancer advanced and that the MRP-1/CD9 gene and protein expression in the metastatic lymph nodes was strikingly lower than in the primary breast cancers. In this study, we also investigated the expression of MRP-1/CD9 by immunoblotting and immunohistochemical analysis in 143 freshly resected invasive ductal carcinomas of the breast: 52 tumors were stage I, 61 were stage II, and 30 were stage III. Tumors were classified as MRP-1/CD9 positive when a band intensity of >30% compared with positive control cells, ZR-75-30 were evaluated with the antibody M31-15, and those with intensities <30% as negative. Moreover, these results were ascertained by immunostaining. Tumor specimens classified as MRP-1/CD9 positive using Western blotting had >50% of the cancer cells immunostained with M31-15, and those classified as MRP-1/CD9 reduced had <50% of the cancer cells immunostained with M31-15. There were 97 patients with MRP-1/CD9 positive tumors and 46 patients whose tumors had reduced MRP-1/CD9 levels. The disease-free rate of the former group of patients was strikingly higher than that of the latter (84.7% versus 51.4%, P<0.001). Similarly, the overall survival rate was also significantly different between the two groups (93.6% versus 69.6%, P=0.004). Multivariate analysis with the Cox regression model indicated that MRP-1/CD9 positively correlated better with disease-free survival (P<0.001) than estrogen receptor, tumor, and lymph node status. Our data suggest that low MRP-1/CD9 expression by tumors of the breast may be associated with poor prognosis. It is conceivable that testing for MRP-1/CD9 may identify node-negative breast cancer patients who are at high risk for early disease recurrence.     

High-titer HER-2/neu protein-specific antibody can be detected in patients with early-stage breast cancer

PURPOSE: To evaluate HER-2/neu-specific antibody immunity in patients with breast cancer, to determine the rate of occurrence of serum antibodies to HER-2/neu in patients with breast cancer, and to relate the presence of specific immunity to overexpression of HER-2/neu protein in primary tumor. METHODS: The antibody response to HER-2/neu protein was analyzed in 107 newly diagnosed breast cancer patients. Sera was analyzed for the presence of HER-2/neu-specific antibodies with a capture enzyme-linked immunosorbent assay (ELISA) and verified by Western blot. Sera from 200 volunteer blood donors was used as a control population. RESULTS: The presence of antibodies to HER-2/neu correlated with the presence of breast cancer. HER-2/neu antibodies at titers of > or = 1:100 were detected in 12 of 107 (11%) breast cancer patients versus none of 200 (0%) normal controls (P < .01). The presence of antibodies to HER-2/neu also correlated to overexpression of HER-2/neu protein in the patient's primary tumor. Nine of 44 (20%) patients with HER-2/neu-positive tumors had HER-2/neu-specific antibodies, whereas three of 63 (5%) patients with HER-2/neu-negative tumors had antibodies (P = .03). The antibody responses could be substantial. Titers of greater than 1:5,000 were detected in five of 107 (5%). CONCLUSION: The presence of HER-2/neu antibodies in breast cancer patients and the correlation with HER-2/neu-positive cancer implies that immunity to HER-2/neu develops as a result of exposure of patients to HER-2/neu protein expressed by their own cancer. These findings should stimulate further studies to develop the detection of immunity to oncogenic proteins as tumor markers, as well as the development and testing of vaccine strategies to induce and augment immunity to HER-2/neu for the treatment of breast cancer or prevention of recurrent disease.     

Role of interleukin 8 in the genesis of acute respiratory distress syndrome through an effect on neutrophil apoptosis

OBJECTIVE: To evaluate the role of interleukin 8 (IL-8) in the regulation of neutrophil (PMN) apoptosis in normal plasma and plasma from patients with early, fulminant acute respiratory distress syndrome (ARDS). DESIGN: Experimental study using cultured human PMNs. SETTING: University hospital, level I trauma center. PARTICIPANTS: Plasma was obtained from 6 patients with early, fulminant posttraumatic ARDS (mean Injury Severity Score, 26). All samples were drawn within 24 hours after injury. Plasma was also taken from 13 healthy control subjects. These controls were also used as sources of PMNs. MAIN OUTCOME MEASURES: Effect of early, fulminant ARDS and normal plasma on spontaneous apoptosis, CD16, and CD11-b expression in PMNs in vitro; levels of IL-8 in plasma; correlation of extracellular IL-8 concentration with rate of PMN apoptosis; and effect of IL-8 blockade on PMN apoptosis, CD16, and CD11-b expression in ARDS and normal plasma. RESULTS: Plasma from patients with early, fulminant ARDS inhibited spontaneous PMN apoptosis at 24 hours (35%+/-5% vs 54%+/-5%; P=.01). Neither CD16 nor CD1l-b differed significantly between the 2 groups. The mean plasma level of IL-8 in patients with early, fulminant ARDS was 359+/-161 pg/mL vs 3.0+/-0.4 pg/mL in healthy controls (P<.05). Interleukin 8 inhibited apoptosis in plasma-free medium at low doses (1-50 pg/mL) but had no significant effect at higher doses (100-5000 pg/mL) (P<.05). Interleukin 8 blockade with monoclonal antibody suppressed apoptosis in normal plasma (28%+/-5% with monoclonal antibody vs 51%+/-5% without monoclonal antibody; P=.008) but not in plasma from patients with early, fulminant ARDS (29%+/-5% with monoclonal antibody vs 34%+/-6% without monoclonal antibody; P=.67). It had no effect on CD16 or CD11-b expression in either plasma. CONCLUSIONS: Plasma from patients with early, fulminant ARDS contains soluble factors that inhibit PMN apoptosis in vitro. Low levels of IL-8 inhibit PMN apoptosis in normal plasma. Although plasma levels of IL-8 are markedly elevated in early, fulminant ARDS, IL-8 is not directly responsible for the antiapoptotic effect of plasma from patients with early, fulminant ARDS.     

Decreased expression of signal-transducing zeta chain in peripheral T cells and natural killer cells in patients with cervical cancer

An impaired immune response is frequently observed in patients and experimental animals with advanced cancer. We and others have shown alterations in CD3-associated signal-transducing zeta molecules in tumor-infiltrating T cells and peripheral blood lymphocytes (PBLs) of patients with advanced cancer. By using flow cytometric analysis of permeabilized cells with a monoclonal antibody (TIA-2) that reacts with the cytoplasmic domain of the zeta chain, here we demonstrate a marked decrease (P < 0.01) in the expression of the signal-transducing CD3 zeta chain of PBLs in patients with cervical cancer (n = 22) as compared to PBLs from healthy donors (n = 21). In addition, PBLs isolated from patients (n = 23) with cervical intraepithelial neoplasia (CIN), to a lesser but significant (P < 0. 01) extent, expressed reduced CD3 zeta levels as compared to those from healthy donors. This decreased expression of zeta chains was also observed on CD16(+) natural killer cells in PBLs from patients with cervical cancer. Surface expression of CD3 epsilon on PBLs was also decreased in cervical cancer patients as compared to healthy donors, but not on PBLs from patients with CIN. CD3 zeta chain expression significantly (r = 0.53, P < 0.01) correlated with the ability of the PBLs to produce tumor necrosis factor in response to anti-CD3 stimulation. These findings suggest that alterations of signal-transducing zeta molecules commonly occur in patients with cervical cancer and to a lesser extent with CIN, and that they are associated with reduced cellular functions such as production of tumor necrosis factor.     

Catheter-related infection: an update on diagnosis, treatment, and prevention

PURPOSE: Neoadjuvant chemotherapy is becoming the standard of care for locally advanced breast cancer. This study was performed to determine whether pathologic primary tumor response to neoadjuvant chemotherapy might predict axillary lymph node status and so be used to identify patients in whom surgery could be effectively limited to biopsy of the previous primary tumor site without axillary dissection. PATIENTS AND METHODS: Between 1992 and 1996, 170 consecutive patients with locally advanced breast cancer were treated in a prospective trial with four preoperative cycles of 5-fluorouracil, doxorubicin, and cyclophosphamide. Disease was staged before initiation of preoperative chemotherapy and before surgery. Segmental resection with axillary lymph node dissection or modified radical mastectomy was performed first, followed by postoperative chemotherapy and radiation therapy of the breast (or chest wall) and regional lymphatics. Patient and tumor characteristics associated with complete versus incomplete pathologic primary tumor response to neoadjuvant chemotherapy and correlation between primary breast tumor pathologic response and axillary lymph node status found at surgery were analyzed. RESULTS: Of 156 evaluable patients, 30 patients (19%) had primary breast tumors that were completely eliminated after induction chemotherapy based on histologic assessment. Nineteen of those 30 patients (63%) had negative axillary lymph nodes at dissection, compared with 13 patients (33%) of the 40 who had a near-complete pathologic primary tumor response (< or = 1 cm3 remaining) and only 15 patients (17%) of the 86 who had > 1 cm3 tumor remaining in the pathology specimen of the breast primary. Of the 22 patients with a complete pathologic response in the breast and a clinically negative axilla after induction chemotherapy, axillary dissection revealed positive lymph nodes in four. These four patients had only one or two positive lymph nodes. DISCUSSION: Because initial clinical regression of primary tumor with neoadjuvant chemotherapy is considered an excellent prognostic indicator and because patients with locally advanced breast cancer routinely receive local and regional radiation treatment followed by additional chemotherapy, the role of breast and axillary surgery has been questioned. In this study, a complete pathologic response of the primary tumor to induction chemotherapy is highly predictive of negative axillary lymph node status. Therefore, axillary lymph node dissection may be omitted in certain subsets of patients who have a biopsy-proven complete pathologic response in the primary tumor and a clinical negative axillary examination. Further prospective, randomized investigation is needed to confirm this finding.     

Neurotrophin mRNA expression in the developing tooth suggests multiple roles in innervation and organogenesis

Dendritic cells (DCs), which are antigen presenting cells of potential use in human antitumor vaccination trials, are presently the subject of intense investigation. Many recent studies have reported the possibility of generating ex vivo large numbers of DCs with high antigen presenting capacity by the culture of bone marrow or blood progenitors. In this study, we examined the differentiation into DCs of CD34+ progenitors isolated from the G-CSF mobilized blood of 3 healthy donors and 5 patients with breast cancer and cultured in the presence of GM-CSF + IL-13. The characteristics of the cells were compared to those of cells obtained in the presence of GM-CSF + TNF alpha. By day 15, one third of the bulk cells cultured with IL-13 were CD1a+/CD14- and strongly expressed CD1c, CD40, CD80 and HLA-DR. In contrast, cells obtained with TNF alpha expressed CD1a on one in three cells but with a considerably lower fluorescence intensity than on IL-13-cultured cells and strongly expressed CD14 on more than 50% of cells. CD1a+/CD14- cells emerged in IL-13 cultures at day 5, while in TNF alpha cultures CD14+ cells appeared before CD1a+ cells. Cells grown in the presence of IL-13 had an increased capacity to present antigens to autologous lymphocytes and to stimulate allogeneic T-lymphocytes. This effect was greater than that of cells grown in the presence of TNF alpha. These cells should therefore have greater effector potential in any therapeutic applications in humans.