The aim of this study was to investigate the role of Mesenchymal Stem Cell (MSC) conditioned medium (CM(MSC)) on apoptosis of cultured mouse primary hepatocytes after in vivo carbon tetrachloride (CCl4)-induced acute liver injury. The acute liver injury was induced by injecting CCl4 intraperitoneally in C57/BL6 mice. Hepatocytes were isolated by liver perfusion, cultured in a defined medium to maintain their differentiation and characterized by reverse transcriptase polymerase chain reaction (RT-PCR) using the hepatic cell specific genes albumin, hepatocyte nuclear factor 4 (HNF4) and cytokeratin 18 (CK18). CM(MSC) was generated from cultured bone marrow-derived MSCs (BM-MSCs). BM-MSCs were positive for CD73, CD90, CD44 by flow cytometry and able to differentiate into chondrocytes, adipocytes and osteocytes. Apoptosis was evaluated by both annexin V. CM(MSC) were examined by flow cytometry to detect MSC-derived annexin V- and CD54/CD44-positive microparticles (MPs). In the CCl4-CM(MSC) treated hepatocytes, interleukin-6 (IL-6) was increased on the first day of culture compared to control and CCl4 and was followed by upregulation of fibroblast-like-protein (FGL1) expression after 48 hrs. This was associated with a significant decrease of annexin V positive CCl4-CM(MSC) treated hepatocytes at day 3 post plating. Recombinant IL-6 was induced FGL1 expression in hepatocytes derived from CCl4-treated mice suggesting that CM(MSC), which is enriched also in microparticles, attenuates CCl4-induced early apoptosis in hepatocytes through activation of FGL1.

T cell-T cell Ag presentation is increasingly attracting attention. We previously showed that the in vitro OVA-pulsed dendritic cell (DC(OVA))-activated CD4(+) Th cells acquired OVA peptide/MHC (pMHC) class I and costimulatory molecules such as CD54 and CD80 from DC(OVA) and acted as CD4(+) Th-APC capable of stimulating OVA-specific CD8(+) CTL responses. In this study, we further applied the OVA-specific TCR-transgenic OT I and OT II mice with deficiency of various cytokines or costimulatory molecule genes useful for studying the molecular mechanisms underlying in Th-APC's stimulatory effect. We demonstrated that DC(OVA)-stimulated OT II CD4(+) Th-APC also acquired costimulatory molecules such as CD40, OX40L, and 4-1BBL and the functional pMHC II complexes by DC(OVA) activation. CD4(+) Th-APC with acquired pMHC II and I were capable of stimulating CD4(+) Th1 and central memory CD8(+)44(+)CD62L(high)IL-7R(+) T cell responses leading to antitumor immunity against OVA-expressing mouse B16 melanoma. Their stimulatory effect on CD8(+) CTL responses and antitumor immunity is mediated by IL-2 secretion, CD40L, and CD80 signaling and is specifically targeted to CD8(+) T cells in vivo via acquired pMHC I. In addition, CD4(+) Th-APC expressing OVA-specific TCR, FasL, and perforin were able to kill DC(OVA) and neighboring Th-APC expressing endogenous and acquired pMHC II. Taken together, we show that CD4(+) Th-APC can modulate immune responses by stimulating CD4(+) Th1 and central memory CD8(+) T cell responses and eliminating DC(OVA) and neighboring Th-APC. Therefore, our findings may have great impacts in not only the antitumor immunity, but also the regulatory T cell-dependent immune tolerance in vivo.

The antigenic phenotype of human synovial lining cells in normal and hyperplastic synovium intima was determined with a panel of monoclonal antibodies directed against a large number of well defined myeloid (macrophage/granulocyte associated) antigens. Synovial lining cells express numerous macrophage associated antigens, including CD11b (CR3), CD13, CD14, CD16 (FcRIII), CD18, CD32 (FcRII), CD45 (leucocyte common antigen), CD54 (ICAM-1), CD64 (FcRI), CD68, and CD71 (transferrin receptor). Few synovial lining cells expressed CD11a (LFA-1) and CD11c (p150,95). Subintimal macrophages expressed all the macrophage associated antigens which were present on synovial lining cells and, in addition, expressed CD15a, CD25 (interleukin-2 receptor), CD34, and CD35 (C3b receptor), none of which was present on synovial lining cells. Synovial lining cell expression of a wide range of macrophage antigens argues in favour of their marrow origin and membership of the mononuclear phagocyte system.

Dendritic cells (DC) are the main stimulators of primary T-cell responses and, thus, probably play a role in the immune reactions after stem cell transplantation. Very little is known about DC in cord blood (CB) and about their potential involvement in the low incidence and severity of acute graft-versus-host disease after CB transplantation. Here, CBDC were identified as a HLA-DR+ cell population, lacking the CD3, CD11b, CD14, CD16, CD19, CD34, CD56, and glycophorin A lineage markers (lin). This lin-/HLA-DR+ population represented 0.3% +/- 0.1% (mean +/- SD; range, 0.1% to 0. 6%; n = 15) of CB mononuclear cells, and CB contained 5.4 +/- 3.2 x 10(3) CBDC/mL (1.8 to 13.0 x 10(3); n = 15). CBDC expressed CD4, CD11a, CD18, CD45RA, CD50, CD54, and CD123, but showed no expression of CD1a, CD11c, CD33, CD40, CD45R0, CD80, CD83, and CD86 and only limited expression of CD58, CD102, and CD116. Despite this immature phenotype, immunomagnetically lin--enriched CBDC were potent stimulators of allogeneic CB T cells. As few as 266 +/- 107 (193 to 530; n = 10) lin-/HLA-DR+ CBDC stimulated a significant response. However, CBDC failed to take up protein or peptide antigens. Thus, in CB there is a prevalence of a DC subpopulation, resembling the CD11c- DC identified in tonsils, the so-called plasmacytoid T cells, which may exert a function distinct from the CD11c+ DC subpopulation.

BACKGROUND

Previous in vitro data indicate that perforin containing drug-specific cytotoxic T cells are involved in cutaneous drug reactions.

OBJECTIVE

The aim of this study was to investigate the in situ expression of perforin and granzyme B together with the nature of the inflammatory infiltrate in acute drug-induced exanthem. Furthermore, expression of interleukin (IL)-12 and interferon (IFN)-gamma, which are known to stimulate cytotoxic T cells, was investigated.

METHODS

Skin biopsy specimens were obtained from 10 patients with a generalized maculopapular exanthem and from nine controls with normal skin. Expression of CD3, CD4, CD8, CD56, CD1a, CD68, CD25, HLA-DR, CD54, perforin, granzyme B, IL-12 and IFNgamma was analysed using immunohistochemistry.

RESULTS

In contrast to the controls, the skin of patients with an exanthem was mainly infiltrated by T cells (CD4>> CD8) and showed a marked enhancement of perforin and granzyme B immunostaining. Double immunostaining revealed that perforin and granzyme B were expressed in both CD4+ and CD8+ cells, which were partly located at the dermoepidermal junction and in the epidermis. In addition, strong immunreactivity for IL-12 and IFNgamma was observed in the mononuclear cells infiltrate, indicating that these cytokines may be important in activation of these cytotoxic T cells.

CONCLUSIONS

The increased numbers of perforin and granzyme B containing T cells infiltrating the dermoepidermal junction may contribute to the damage of epidermal cells, which is frequently observed as a typical feature of interface dermatitis in drug-induced exanthem. Our data provide further evidence that cytotoxic T cells play an essential role in cutaneous drug reactions.

Treating human malignant melanoma cells with tumor necrosis factor-alpha (TNF-alpha) or interferon-gamma (IFN-gamma) causes a dose- and time-dependent increase in surface expression of ICAM-1. Increased ICAM-1 expression corresponds to greater binding of human leukocyte functional antigen-1 (CD11a/CD18)-expressing peripheral blood mononuclear cells (PBMC) to C8161 monolayers, suggesting that cytokine-treated melanoma cells would be more metastatic due to PBMC-tumor cell emboli. The purpose of this study was: (1) to test whether TNF-alpha-treated human melanoma cells are indeed more metastatic than untreated C8161 and (2) to determine whether ICAM-1 plays a role in metastasis of C8161. When surface ICAM-1 expression is upregulated, formation of lung metastases in nude mice increases 1.5- to 4-fold (P < 0.05) for human melanoma cell lines C8161, MeWo, and A375. Treatment of C8161 with ICAM-1 phosphorothioate antisense oligonucleotides using cationic lipids results in>> 90% inhibition of ICAM-1 surface expression as determined by ELISA and flow cytometry. Antisense ICAM-1-treated cells form 41-64% fewer metastases than sham-treated cells. Metastasis does not increase when antisense-treated melanoma cells are exposed to TNF-alpha. However, treatment of C8161 with antisense 5-lipoxygenase (5-LO) oligonucleotides inhibits metastases 39% in Lipofectin-treated cells, but does not inhibit TNF-alpha-induced upregulation of experimental metastases. Similar experiments were performed to measure PBMC adhesion to antisense oligonucleotide-treated C8161 cells; however, TNF-alpha-inducible increase in adhesion was unaffected by ICAM-1 or 5-LO antisense oligonucleotides. These results demonstrate that ICAM-1 is involved in melanoma metastasis, but probably not at the step of PBMC adhesion to C8161 cells.

Recombinant Streptococcus gordonii expressing on the surface the C-fragment of tetanus toxin was tested as an Ag delivery system for human monocyte-derived dendritic cells (DCs). DCs incubated with recombinant S. gordonii were much more efficient than DCs pulsed with soluble C-fragment of tetanus toxin at stimulating specific CD4+ T cells as determined by cell proliferation and IFN-gamma release. Compared with DCs treated with soluble Ag, DCs fed with recombinant bacteria required 102- to 103-fold less Ag and were at least 102 times more effective on a per-cell basis for activating specific T cells. S. gordonii was internalized in DCs by conventional phagocytosis, and cytochalasin D inhibited presentation of bacteria-associated Ag, but not of soluble Ag, suggesting that phagocytosis was required for proper delivery of recombinant Ag. Bacteria were also very potent inducers of DC maturation, although they enhanced the capacity of DCs to activate specific CD4+ T cells at concentrations that did not stimulate DC maturation. In particular, S. gordonii dose-dependently up-regulated expression of membrane molecules (MHC I and II, CD80, CD86, CD54, CD40, CD83) and reduced both phagocytic and endocytic activities. Furthermore, bacteria promoted in a dose-dependent manner DC release of cytokines (IL-6, TNF-alpha, IL-1beta, IL-12, TGF-beta, and IL-10) and of the chemokines IL-8, RANTES, IFN-gamma-inducible protein-10, and monokine induced by IFN-gamma. Thus, recombinant Gram-positive bacteria appear a powerful tool for vaccine design due to their extremely high capacity to deliver Ags into DCs, as well as induce DC maturation and secretion of T cell chemoattractans.

BACKGROUND

We hypothesized that the aggressive LNCaP-derived androgen-independent cell line, CL1, might differ from LNCaP in their repertoire of cell surface markers and that these differences might typify changes that occur during clinical prostate cancer progression.

METHODS

The cell surface marker expression profiles of CL1 and LNCaP were examined using flow cytometry. Markedly differential gene expression was confirmed using RT-PCR and further examined using immunohistochemistry among the prostate cancer cell lines LAPC-4, LNCaP, CL1, CL2, DU145, and PC-3. The expression of the most markedly differentially expressed surface marker, CD10, was further explored in a tissue microarray containing radical prostatectomy samples from 219 hormone naïve prostate cancer patients.

RESULTS

There were marked differences in the expression of CD10, CD13, CD26, CD33, CD44, CD54, CD55, and CD104 between CL1 and LNCaP. Results from both the RT-PCR and immunohistochemistry confirmed the differential expression and found that CD10 demonstrated a pattern of expression in hormone sensitive but not hormone refractory cell lines. When CD10 expression was examined in a tissue microarray, CD10 expression was below the 25th percentile of matched normal prostate tissue in 68% of prostate cancers, below the median expression of matched normal prostate tissue in 86% of cancers, and completely absent in 34% of cancers. Samples of prostatic intraepithelial neoplasia demonstrated CD10 expression that was intermediate between normal prostatic tissue and prostate cancer. Among prostate cancer patients, CD10 expression did not correlate with Gleason score, pathological stage, or biochemical recurrence following radical prostatectomy.

CONCLUSIONS

These findings demonstrate that loss or decreased expression of CD10 is an early and frequent event in human prostate cancer and implicates CD10 as a potential therapeutic target for early stage hormone sensitive prostate cancer.

OBJECTIVE

To study distribution and toxicity of cytolytic T lymphocytes (CTLs) against a single melanoma epitope.

METHODS

CD8(+) T cells obtained by leukapheresis from 10 patients with disseminated HLA-A2.1(+), tyrosinase-positive melanomas were immunized in vitro against tyrosinase(369-377) (YMNGTMSQV). Drosophila cells transduced with HLA-A2.1, CD80, and CD54 (intracellular adhesion molecule-1) were used for priming, followed by two rounds of immunization with mononuclear cells as antigen-presenting cells. 1 x 10(8) CTL were infused intravenously (IV) on day 1. CTL frequency was measured by limiting dilutions in five patients. (111)In labeling and scintigraphy measured distribution of CTL in next five. Five days later, 1 x 10(8) CTLs were infused on 4 successive days to both groups. Immunohistology of response was judged by biopsies.

RESULTS

Infusions were nontoxic. CTLs were undetectable in the blood, going to lungs within 5 minutes. At 4, 24, and 72 hours, they were found in liver and spleen. Lesions were visualized by scintiscans in one responding patient where two subcutaneous nodules were noted at 4 and 24 hours. A second patient had a partial response and remains alive with disease 2 years later. CD8(+) T cells were found in lesions of responders, associated with the presence of HLA-A2 molecules and tyrosinase. Two nonresponders without tyrosinase and HLA-A2 molecules had a paucity of CD8(+) T cells in their lesions. Whether the CD8(+) T cells in lesions of responders were those we had reinfused is uncertain.

CONCLUSIONS

CTLs immunized against a single melanoma epitope were nontoxic but did not specifically localize to tumor sites. Nevertheless, two patients had disease regression. Additional therapeutic studies with specifically immunized CTL seem justified.

OBJECTIVE

Previous work suggested the potential utility of therapy with a monoclonal antibody (Mab) to intercellular adhesion molecule-1 (ICAM-1; CD54) in patients with longstanding rheumatoid arthritis (RA). Immunomodulatory interventions, including adhesion receptor directed therapies, might be expected to have greater efficacy in patients with less established or less aggressive disease. Therefore, we assessed the efficacy and safety of an anti-ICAM-1 Mab in patients with early RA.

METHODS

An open label study of a 5 day infusion of an anti-ICAM-1 Mab in 10 patients with early or indolent RA was conducted. These patients were defined as having previously used < or = 1 disease modifying antirheumatic drug.

RESULTS

Based on composite criteria, 7/10 patients had a marked or moderate response to therapy at one month of followup. Clinical benefit was sustained through 2 months for 5/10 patients and 3/10 had extended benefit (11, 8, and>> 7 months). Clinical benefit was more likely to be obtained in patients with subacute onset of disease than in those with a fulminant onset.

CONCLUSIONS

A single course of therapy with an anti-ICAM-1 Mab was associated with clinical improvement in a group of patients with early or indolent RA to an extent apparently greater than previously observed in patients with longstanding, aggressive RA.

Human uterine fibroblasts (HuF) isolated from the maternal part (decidua parietalis) of a term placenta provide a useful model of in vitro cell differentiation into decidual cells (decidualization, a critical process for successful pregnancy). After isolation, the cells adhere to plastic and have either a small round or spindle-shaped morphology that later changes into a flattened pattern in culture. HuF robustly proliferate in culture until passage 20 and form colonies when plated at low densities. The cells express the mesenchymal cell markers fibronectin, integrin-beta1, ICAM-1 (CD54), and collagen I. Flow cytometry of HuF has detected the presence of CD34, a marker of the hematopoietic stem cell lineage, and an absence of CD10, CD11b/Mac, CD14, CD45, and HLA type II. Furthermore, they also express the pluripotency markers SSEA-1, SSEA-4, Oct-4, Stro-1, and TRA-1-81 as detected by confocal microscopy. Treatment for 14-21 days with differentiation-inducing media leads to the differentiation of HuF into osteoblasts, adipocytes, and chondrocytes. The presence of alpha-smooth muscle actin, calponin, and myosin light-chain kinase in cultured HuF implies their similarity to myofibroblasts. Treatment of the HuF with dimethyl sufoxide causes reversion to the spindle-shaped morphology and a loss of myofibroblast characteristics, suggesting a switch into a less differentiated phenotype. The unique abilities of HuF to exhibit multipotency, even with myofibroblast characteristics, and their ready availability and low maintenance requirements make them an interesting cell model for further exploration as a possible tool for regenerative medicine.

ICAM-1 (CD54) plays an important role in the cell-cell interaction and migration of leukocytes. Previous studies have shown that ICAM-1 is involved in inflammatory reactions and that a defect in ICAM-1 gene inhibits allergic contact hypersensitivity. This study indicates that the migration of hapten presenting Langerhans cells into the regional lymph nodes was significantly reduced in ICAM-1-deficient mice compared to wild-type C57BL/6 mice. The reduced number of dendritic cells in regional lymph nodes did not result from abnormal migration of Langerhans cells into the skin of ICAM-1-deficient mice. The concentration and distribution of Langerhans cells in the naïve skin of ICAM-1-deficient mice was equal to that of wild-type mice. Following hapten sensitization, Langerhans cell migration out of the skin and recruitment of fresh Langerhans cells back to the epidermis was not affected in ICAM-1-deficient mice. Further experiments demonstrated that ICAM-1 deficiency on lymphatic endothelium rather than on dendritic cells was responsible for the reduced migration of Langerhans cells into draining lymph nodes. This study indicates that ICAM-1 regulates the migration of dendritic cells into regional lymph nodes but not into or out of the skin.

Endothelial adhesion molecules are directly involved in the localization and migration of leukocytes from the circulation into tissues at sites of inflammation. We have compared the expression of PECAM-1 (CD31), ELAM-1, ICAM-1 (CD54), and VCAM-1 in pretransplant (n = 20) and needle-core biopsies from renal transplants obtained during different clinical circumstances (n = 42). PECAM-1 was consistently expressed on all endothelium in both pretransplant and transplant biopsies. In contrast, there was variation in endothelial expression of ELAM-1 and in proximal tubular expression of ICAM-1 and VCAM-1 between pretransplant biopsies. After transplantation induced expression of endothelial ELAM-1 and VCAM-1 and tubular induction of ICAM-1 and VCAM-1 was detected. Induced adhesion molecule expression was frequently associated with focal leukocyte infiltration, and there was a significantly higher level of CD45 and CD25 positive cell infiltration in biopsies with induced adhesion molecule expression. The induction of adhesion molecule expression is evidence of endothelial activation in these transplant biopsies. Comparison of adhesion molecule expression and HLA-class II antigen expression revealed that induced tubular class II antigens may be detected in the absence of induced adhesion molecule expression.

The majority of T cells at the site of an inflammatory lesion do not appear to be Ag-specific, but they still contribute to the inflammatory response. Herein, we report that sCD23 activates monocytes to participate in the stimulation of resting T cells in the absence of TCR engagement. First, sCD23 selectively triggers monokine release by purified monocytes in the absence of costimulus. It induces TNF-alpha, IL-1 beta, IL-8, granulocyte-macrophage-CSF, and prostaglandin E2 but no IL-10, IL-12, TGF-beta, or leukotriene B4. The sCD23-induced TNF-alpha production is significantly inhibited by IL-4 and IL-10 but not by TGF-beta. Also, monocytes activated by sCD23 express decreased levels of HLA-DR and increased levels of CD14, CD54, CD40, and B7 Ag. Next, we show that, in the presence of monocytes, sCD23 is a potent costimulator of IL-2 or IL-12-induced IFN-gamma production by resting T cells in the absence of exogenous Ag and that this effect is partially reduced by anti-TNF-alpha mAb. B cells cannot substitute for monocytes, and CD4+ and CD8+ T cells are equal responders. The data further indicate that monocyte-T cell contact, more particularly CD40-CD40L interactions, is required for IFN-gamma production in response to IL-2 plus sCD23, and the response to IL-12 plus sCD23 is CD40- and B7-independent but is still partially contact-dependent. It is proposed that sCD23, when produced locally at a site of immune response, may trigger an inflammatory process via monokine release and may further amplify it via the stimulation of bystander non-Ag-specific T cells.

Human mesenchymal stem cells (hMSCs) have been paid a great deal of attention because of their unprecedented therapeutic merits endowed by powerful ex vivo expansion and multilineage differentiation potential. Umbilical cord blood (UCB) is a convenient but not fully proven source for hMSCs, and hence, greater experience is required to establish UCB as a reliable source of hMSCs. To this end, we attempted to isolate hMSC-like adherent cells from human UCB. The isolated cells were highly proliferative and exhibited an immunophenotype of CD13+ CD14- CD29+ CD31- CD34- CD44+ CD45- CD49e+ CD54+ CD90+ CD106- ASMA+ SH2+ SH3+ HLA-ABC+ HLA-DR-. More importantly, these cells, under appropriate conditions, could differentiate into a variety of mesenchymal lineage cells such as osteoblasts, chondrocytes, adipocytes, and skeletal myoblasts. This mesengenic potential assures that the UCB-derived cells are multipotent hMSCs and further implicates that UCB can be a legitimate source of hMSCs.

The capacity to migrate from peripheral tissues, where antigen is encountered, to lymphoid organs, where the primary immune response is initiated, is crucial to the immunogenic function of dendritic cells (DC). The skin is a suitable tissue to study migration. DC were observed to gather in distinct nonrandom arrays ("cords") in the dermis upon culture of murine whole skin explants. It is assumed that cords represent lymphatic vessels. Using a similar organ culture model with human split-thickness skin explants, we investigated migration pathways in human skin. We made the following observations. 1) Spontaneous emigration of Langerhans cells took place in skin cultured for 1-3 d. Nonrandom distribution patterns of strongly major histocompatibility complex class II-expressing DC (cords) occurred in cultured dermis. A variable, yet high (>50%) percentage of these DC coexpressed the Birbeck granule-associated antigen "Lag." Ultrastructurally, the cells corresponded to mature DC. 2) Electron microscopy proved that the dermal structures harboring the accumulations of DC (i.e., cords) were typical lymph vessels. Moreover, markers for blood endothelia (monoclonal antibody PAL-E, Factor VIII-related antigen) and markers for cords (strong major histocompatibility complex class II expression on nonrandomly arranged, hairy-appearing cells) were expressed in a mutually exclusive pattern. 3) On epidermal sheets we failed to detect gross changes in the levels of expression of adhesion molecules (CD44, CD54/ ICAM-1, E-cadherin) on keratinocytes in the course of the culture period. The reactivity of a part of the DC in the dermal cords with Birbeck granule-specific monoclonal antibody "Lag" suggests that the migratory population is composed of both epidermal Langerhans cells and dermal DC. We conclude that this organ culture model may prove helpful in resolving pathways and mechanisms of DC migration.

Dendritic cell (DC) precursors and immature DC reside in epithelium where they encounter pathogens and cytokines, which stimulate their differentiation. We hypothesized that type-I interferons (IFN-alpha and -beta), cytokines that are produced early in the innate immune response against viruses and some bacteria, may influence DC differentiation and function. To examine this possibility, we used an in vitro model of DC differentiation in which initial culture of human CD14(+) monocytes with granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4 generates immature DC, and subsequent culture with tumor necrosis factor (TNF)-alpha drives the final development into mature DC. We found in this model that IFN-alpha/beta, added from the initiation of the culture on, significantly reduced the survival and altered the morphology and differentiation of DC. TNF-alpha-dependent maturation of IFN-beta-treated immature DC led to cells with reduced expression of CD1a, CD40, CD54, and CD80 when compared with mature DC controls. IFN-alpha/beta-treated DC further had a reduced capacity to induce naive Th-cell proliferation through allostimulation or anti-CD3 monoclonal antibody stimulation. In addition, IFN-alpha/beta-treated DC secreted less IL-12 upon stimulation with Staphylococcus aureus Cowan strain or with CD4(+) T cells, and this decrease correlated directly with their inability to support CD4(+) T-cell secretion of IFN-gamma, even though T-cell lymphotoxin production was unaffected. These findings indicate that type-I IFNs can influence the generation of acquired immune responses by modifying T-helper cell differentiation through the regulation of DC differentiation and function.

The Revised European-American Lymphoma classification gives Burkitt-like lymphoma (BLL) provisional status, leaving unresolved the differential diagnosis with Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL). This study compared the biologic features of adult BLL and DLBCL. The phenotypic distinction between BLL and DLBCL was determined by immunohistochemical staining of frozen tissue from 13 patients with BLL and 55 patients with DLBCL by using an extensive antibody panel including Ki-67, CD10, CD11a/lymphocyte function-associated antigen 1alpha (LFA-1alpha), CD18/LFA-1beta, CD58/LFA-3, and CD54/intercellular adhesion molecule, CD8 for tumor-infiltrating cytotoxic T cells (T-TILs), CD44 homing receptor, and p53 and Bcl-2 oncogenic proteins. Compared with DLBCL, BLL had a higher proliferative rate (mean Ki-67, 88% versus 53%), greater expression of CD10 and p53 antigens, and decreased expression of Bcl-2. BLL cases had a consistent absence of one or more cell adhesion molecules (92% versus 27%), low T-TIL numbers, and absence of CD44 homing receptor (92% versus 14%). The t(8;14) translocation was identified in 80% of BLL cases, but no patients with BLL had the t(14;18) translocation. In a 10-year analysis, median survival of patients with BLL was 1.2 years, and that of patients with DLBCL was 2.5 years. Although the proportion of patients cured was similar in the 2 groups, BLL patients had an increased risk of early death. We conclude that BLL can be recognized by its combined morphologic and phenotypic features and that it represents a high-grade lymphoma much closer to BL than DLBCL. Retention of the BLL category or inclusion of BLL as a variant of BL is biologically and clinically more appropriate than absorbing the category of BLL into DLBCL. (Blood. 2001;97:3713-3720)

We analyzed the stimulatory effect of oncostatin M (OSM), leukemia inhibitory factor (LIF), interleukin 6 (IL-6), IL-11, and the inhibitory effect of anti-IL-6 antibody (Ab), anti-IL-6 receptor monoclonal antibody (mAb), and anti-gp130 mAb on the growth of human plasmacytoma cells freshly isolated from a patient with multiple myeloma. The purified cells showed a plasmacytoid morphology and expressed CD38, CD54, and CD56 antigens but no CD3, CD5, CD10, CD19, CD20, or very late antigen 5. IL-6 receptor (IL-6R) and its signal transducer, gp130, were expressed on their cell surface at a low level. Dose-dependent proliferation of the cells in response to OSM, LIF, and IL-6, but not to IL-11, was observed using [3H]TdR incorporation in vitro. Both anti-IL-6 Ab and anti-IL-6R mAb inhibited the growth of the cells in the presence or absence of exogenous IL-6. These cells release IL-6 but not OSM or LIF into the culture supernatant during short-term culture. Therefore, an autocrine growth mechanism mediated by IL-6, but not by OSM or LIF, was confirmed. Furthermore, anti-gp130 mAb completely inhibited the proliferation of the cells induced by OSM, LIF, as well as IL-6. These data indicate that OSM, LIF, and IL-6 can act as growth factors of human plasmacytoma cells through a common signal transducer, gp130, on their cell surface, and also suggest the potential therapeutic application of anti-gp130 mAb, as well as anti-IL-6R mAb against myeloma/plasmacytomas.

NK cells constitute the first line of defense against pathogens and transformed cells. They mature in secondary lymphoid organs, including tonsils, where common pathogens, such as EBV, enter the host and potentially imprint differentiating cells, which then patrol the body via the blood stream. Therefore, we set out to characterize a distinct human NK cell population in tonsils that produces high amounts of the immunomodulatory and antiviral cytokine IFN-γ. We found that the tonsilar IFN-γ(high) NK cell subset is CD56(bright)NKG2A(+)CD94(+)CD54(+)CD62L(-), is present in tonsils ex vivo and is more mature than other CD56(bright) NK cells in tonsils and less mature than other NK cells in blood, shows very low plasticity even after prolonged cytokine stimulation, accumulates in tonsils of EBV carriers, and is able to potently restrict EBV-induced transformation of B cells. Thus, we characterized a distinct and stable IFN-γ(high) NK cell subpopulation that can specifically restrict malignant transformation of EBV-infected B cells. This subset should be exploited for future development of cell-based therapeutic approaches in EBV-associated malignancies.

BACKGROUND

Mature circulating endothelial cells (CEC) are surrogate markers of endothelial damage. CEC measured in patients with advanced cancer are thought not only to derive from damaged normal vasculature (n-CEC), but also from damaged (t-CEC). Therefore, assays that allow the discrimination between these two putative types of CEC are thought to improve the specificity of the enumeration of CEC in cancer.

METHODS

Identification of tumour-associated endothelial markers (TEM) by comparing antigen expression on normal vs t-CEC and assess the presence of t-CEC in peripheral blood of cancer patients by incorporating TEM in our novel flow cytometry-based CEC detection assay.

RESULTS

No difference in antigen expression between normal and malignant endothelial cells (ECs) was found for CD54, CD109, CD137, CD141, CD144 and CXCR7. In contrast, overexpression for CD105, CD146, CD276 and CD309 was observed in tumour ECs compared with normal ECs. CD276 was most differentially expressed and chosen as a marker for further investigation. CD276-expressing CEC were significantly higher in 15 patients with advanced colorectal cancer (median 9 (range 1-293 cell per 4 ml); P<0.005), in 83 patients with a glioblastoma multiforme (median 10 (range 0-804); P<0.0001) and in 14 patients with advanced breast cancer (median 14 (range 0-390) P<0.05) as compared with 24 healthy individuals (median 3 (range 0-11)). Of all patients with malignancies, 58% had CD276(+) CEC counts above the ULN (8 cell per 4 ml).

CONCLUSIONS

The present study shows that CD276 can be used to discriminate ECs from malignant tissue from ECs from normal tissue. In addition, CD276(+) CEC do occur in higher frequencies in patients with advanced cancer.

BACKGROUND

Probiotic bacteria are proposed to alleviate atopic dermatitis (AD) in infants. There are few indications about the effect of probiotics on AD in adults.

OBJECTIVE

The purpose of this study was to elucidate the influence of a probiotic drink containing a combination of the probiotics Lactobacillus paracasei Lpc-37, Lactobacillus acidophilus 74-2 and Bifidobacterium animalis subsp. lactis DGCC 420 (B. lactis 420) in healthy volunteers and in patients with AD on clinical and immunological parameters and their detection in feces.

METHODS

A double-blind, placebo-controlled, randomized cross-over study was conducted in 15 healthy adults and 15 patients with AD. The probiotic product or placebo was given over 8 weeks. A 2-week washout period was interconnected before the intervention was crossed. At the end of each period, blood and stool samples were collected. In patients, the severity of AD was evaluated using the Scoring of Atopic Dermatitis (SCORAD).

RESULTS

L. paracasei and B. lactis were recovered in high numbers in feces after supplementation, whereas L. acidophilus marginally increased. In patients, the SCORAD tended to decrease by 15.5% (P=0.081). Major lymphocyte subsets were not affected by the probiotic intervention. However, CD57(+) increased significantly (P=0.034) in healthy subjects after probiotic intake and was not changed in patients, whereas CD4(+)CD54(+) decreased significantly (P=0.031) in patients with AD and remained uninfluenced in healthy subjects. The expression of CD4(+)CD25(+) T cells was similar in healthy subjects and AD patients. The phagocytic activity of monocytes and granulocytes was significantly increased in healthy subjects after probiotic intervention (P=0.014).

CONCLUSIONS

L. paracasei Lpc-37 and B. lactis 420 are able to colonize the intestine transiently. This study reveals that the probiotics differently modulate peripheral immune parameters in healthy subjects and patients with AD.

Asthma is characterized by airway inflammation rich in eosinophils. Airway eosinophilia is associated with exacerbations and has been suggested to play a role in airway remodelling. Recruitment of eosinophils from the circulation requires that blood eosinophils become activated, leading to their arrest on the endothelium and extravasation. Circulating eosinophils can be envisioned as potentially being in different activation states, including non-activated, pre-activated or 'primed', or fully activated. In addition, the circulation can potentially be deficient of pre-activated or activated eosinophils, because such cells have marginated on activated endothelium or extravasated into the tissue. A number of eosinophil surface proteins, including CD69, L-selectin, intercellular adhesion molecule-1 (ICAM-1, CD54), CD44, P-selectin glycoprotein ligand-1 (PSGL-1, CD162), cytokine receptors, Fc receptors, integrins including αM integrin (CD11b), and activated conformations of Fc receptors and integrins, have been proposed to report cell activation. Variation in eosinophil activation states may be associated with asthma activity. Eosinophil surface proteins proposed to be activation markers, with a particular focus on integrins, and evidence for associations between activation states of blood eosinophils and features of asthma are reviewed here. Partial activation of β1 and β2 integrins on blood eosinophils, reported by monoclonal antibodies (mAbs) N29 and KIM-127, is associated with impaired pulmonary function and airway eosinophilia, respectively, in non-severe asthma. The association with lung function does not occur in severe asthma, presumably due to greater eosinophil extravasation, specifically of activated or pre-activated cells, in severe disease.

The immune, neural, and endocrine systems do not act autonomously; rather, multiple communicative pathways exist among the nervous, endocrine, and immune systems that facilitate physiological immunoregulation. Patients with spinal cord injury (SCI) have decreased natural and adaptive immune responses by 2 weeks after injury. In patients with SCI, adrenocorticotropic hormone (ACTH) and urine-free cortisol levels were increased while zinc and albumin levels were decreased, respectively. In addition, the surface markers alpha 2, alpha 3, alpha 4, CD11a, CD11b, CD18, CD54, and CD8 found on lymphocytes and alpha 3, alpha 4, CD11a, CD18, and CD8 surface markers found on granulocytes were also decreased in the patient population. Finally, the adhesion molecules binding ability in the SCI group was also decreased when compared with a control group. Overall, the investigation showed that patients with SCI have a decreased immune function, especially succeeding the SCI injury, an impaired nutrition status, and a decreased number of adhesion molecules, all of which contribute to delayed wound healing.