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Patients and methods : A prospective registration of patients with colorectal cancer and a colonoscopy within the last 10 years. We tried to classify these post-colonoscopy colorectal cancers (PCCRCs) by most reasonable explanation and into subcategories suggested by the World Endoscopy Organization (WEO) and calculated the unadjusted PCCRC rate. Results : 47 PCCRCs were identified. The average age at diagnosis of PCCRC was 73 years. PCCRCs were more located in the right colon with a higher percentage of MSI-positive and B-RAF mutated tumours. The average period between index colonoscopy and diagnosis of PCCRC was 4.2 years. Sixty-eight % of all PCCRCs could be explained by procedural factors. The mean PCCRC-3y of our department was 2.46%. Conclusions : The data of our centre are in line with the data of the literature from which can be concluded that most post-colonoscopy colorectal cancers are preventable. The PCCRC-3y is an important quality measure for screening colonoscopy. Ideally all centres involved in the population screening should measure the PCCRC-3 y annually, with cooperation of the cancer registry and reimbursement data provided by the Intermutualistic Agency (IMA).

Funding Acknowledgements Type of funding sources: None. Introduction Leadless pacemakers (PMs) capable of atrio-ventricular (AV) synchronous pacing have been introduced recently. These devices provide mechanical atrial sensing by detection of the atrial contraction via an accelerometer. Atrial tracking may be disturbed by external influences such as body motions and higher heart rates in real life. To track the amount of AV synchronous pacing, the device statistic classifies all sensed and paced QRS complexes according to presumed AV synchrony. The reliability of this self-diagnostics to estimate the true degree of AV synchrony, however, is insufficiently studied. Purpose To investigate the informative value of the device statistics offered by leadless VDD PMs regarding true AV snychrony. Methods We prospectively included all patients who received a leadless VDD PM between 07/2020 and 05/2021 at our center in this observational study. During the regular outpatient follow-ups, device interrogation was performed, which included evaluation of the PM statistics. True AV synchrony (defined as a QRS complex preceded by a p-wave within 300ms) was analyzed repeatedly during follow-up using Holter ECGs. Results We analysed 34 Holter ECGs from 20 outpatients (816 hours of ECG in total). Patients had a median age of 80 years (interquartile range 76-86 years), 55% were females. For Holter ECG sequences that showed high degree or complete AV-Block (>80% ventricular pacing), the percentage of paced beats that were presumed to be AV synchronous by the device statistic (ratio "AM-VP"/total VP) correlated well with AV synchrony as assessed using Holter-ECGs (Kendall’s τ=0.312, p66.6% presumed AV synchrony (p Conclusion Leadless PMs provide device statistics that show a correlation with true AV synchrony. The clinical setting as well as the device programming (e.g. AV conduction mode switch) significantly influence the predictive value of the different parameters of the device’s statistics.

© 2018, Deutsche Phytomedizinische Gesellschaft. Internal fruit rot in bell pepper is an important fungal disease which results in mycelium growth and/or necrosis on the ovarium and fruit flesh. It is mainly caused by members of the Fusarium lactis species complex and emerged as a major threat for bell pepper production worldwide. Infection already starts during anthesis, but the symptoms are only visible later on in the production chain. An accurate prediction of the disease incidence in the greenhouse based on environmental parameters is an important step towards a sustainable disease control. Based on a large dataset (2011–2016), a binomial, logistic regression model was developed. This model enables an accurate prediction of internal fruit rot occurrence based on simple and robust input parameters such as temperature and relative humidity during anthesis. Spore density was included as a simplified, practical parameter describing the presence or absence of internal fruit rot 1 week earlier. The obtained model was validated with an independent dataset of five different commercial bell pepper greenhouses. The chance of internal fruit rot infection increased with temperature and relative humidity. Once a greenhouse is infected, only lower temperatures can reduce future risks. However, the chance of the disease to occur remains very high. This prediction model offers a strong instrument for growers to optimize greenhouse climate conditions to restrain internal fruit rot incidence. In addition, the model can be used to apply accurate biological or chemical treatments to achieve a more sustainable greenhouse control. A guideline table for climate adjustment is presented. ispartof: Journal of Plant Diseases and Protection vol:125 issue:4 pages:425-432 status: published

Survivors of brain tumors are faced with a high risk for a wide range of cognitive problems and learning difficulties. These problems are caused by the lesion itself and its surgical removal as well as by the treatments to follow (chemo- and/or radiation therapy). A few recent studies have indicated that children with brain tumors (BT) might exhibit cognitive problems already at diagnosis, i.e. before the start of any medical treatment. The aim of the present study was to investigate the "baseline" neuropsychological profile in children with BT in comparison to children with an oncological diagnosis not involving the central nervous system (CNS). 20 children with BT and 27 children with an oncological disease without involvement of the CNS (age range: 6.1 to 16.9 years) were evaluated with an extensive battery of neuropsychological tests tailored to the patient's age. Furthermore, the child and its parents completed self-report questionnaires about emotional functioning and quality of life. In both groups, tests were administered before any therapeutic intervention such as surgery, chemotherapy or irradiation. Groups were comparable regarding age, gender and social economic status. Compared to the CG, patients with BTs performed significantly worse in tests of working memory, verbal memory and attention. In contrast the areas of perceptual reasoning, processing speed and verbal comprehension were preserved at this time. Younger children with BT were especially disadvantaged. Compared to aged matched children with malignancies not involving the CNS and older BT patients the young BT patients showed deficits in attention, working memory and verbal memory measures. Our results highlight the need for cognitive assessments and interventions early in the treatment process in order to minimize or even prevent academic difficulties as patients return to school.

Background and purpose Pseudoprogression is a frequent phenomenon observed since the introduction of postoperative therapy with radiotherapy and temozolomide (RT/TMZ) in glioblastoma multiforme (GBM) patients. However, the criteria defining pseudoprogression, its incidence, the time of occurrence and its impact on therapy and outcome remain poorly defined. Methods The objective of this study is to compare two sets of criteria (liberal and stringent), defining pseudoprogression, in a cohort of patients treated before and after the introduction of RT/TMZ in the standard postoperative treatment. This retrospective review includes 136 unselected and consecutively treated patients with pathologically diagnosed GBM. Results Pseudoprogression was observed in 10 (12%) cases applying the stringent criteria, and in 18 (23%) patients when using the liberal criteria, in the cohort treated with RT/TMZ. Pseudoprogression was observed in only one patient treated with RT alone. The median time to pseudoprogression was 4 weeks after the end of RT. Patients with pseudoprogression had a median survival time of 28 months, compared with 12 months for patients without pseudoprogression. Conclusions The incidence of pseudoprogression after RT/TMZ strongly depends on the applied criteria. However, regardless of the stringency of the criteria, the impact on survival remains the same.

ObjectiveTo assess the long-term effect of prepubertal high-dose GH treatment on growth in children with idiopathic short stature (ISS).Design and methodsForty children with no signs of puberty, age at start 4–8 years (girls) or 4–10 years (boys), height SDS −2.0 SDS, were randomly allocated to receive GH at a dose of 2 mg/m2 per day (equivalent to 75 μg/kg per day at start and 64 μg/kg per day at stop) until the onset of puberty for at least 2 years (preceded by two 3-month periods of treatment with low or intermediate doses of GH separated by two washout periods of 3 months) or no treatment. In 28 cases, adult height (AH) was assessed at a mean (s.d.) age of 20.4 (2.3) years.ResultsGH-treated children (mean treatment period on high-dose GH 2.3 years (range 1.2–5.0 years)) showed an increased mean height SDS at discontinuation of the treatment compared with the controls (−1.3 (0.8) SDS versus −2.6 (0.8) SDS respectively). However, bone maturation was significantly accelerated in the GH-treated group compared with the controls (1.6 (0.4) versus 1.0 (0.2) years per year, respectively), and pubertal onset tended to advance. After an untreated interval of 3–12 years, AH was −2.1 (0.7) and −1.9 (0.6) in the GH-treated and control groups respectively. Age was a positive predictor of adult height gain.ConclusionHigh-dose GH treatment restricted to the prepubertal period in young ISS children augments height gain during treatment, but accelerates bone maturation, resulting in a similar adult height compared with the untreated controls.

Unter den hochmalignen Gliomen des Kindes- und Jugendalters werden unterschiedliche Tumorentitaten glialen Ursprungs zusammengefasst, die durch einen malignen histologischen Phanotyp oder ungunstigen klinischen Verlauf charakterisiert sind. Die haufigsten Tumorentitaten sind das Glioblastom, das anaplastische Astrozytom und das Ponsgliom. Die aktuelle Therapieempfehlung besteht aus Operation, Bestrahlung und Chemotherapie, Ponsgliome werden nur bestrahlt und chemotherapiert. Die komplette Tumorresektion besitzt die groste prognostische Bedeutung. Das Gesamtuberleben ist fur die meisten Patienten immer noch schlecht. Eine Ausnahme bilden Sauglinge und Kleinkinder mit hochmalignem Gliom, die unter konventioneller Chemotherapie und Operation ein gutes Uberleben zeigen. Fur diese Patienten wird aktuell ein eigenes europaisches Chemotherapieprotokoll entwickelt. Fur altere Patienten werden neue multimodale Therapiekonzepte etabliert, in die innovative tumorspezifische Therapeutika integriert werden sollen.

Patients with relapsed malignant glioma have a poor prognosis. We developed a strategy of vaccination using autologous mature dendritic cells loaded with autologous tumour homogenate. In total, 12 patients with a median age of 36 years (range: 11-78) were treated. All had relapsing malignant glioma. After surgery, vaccines were given at weeks 1 and 3, and later every 4 weeks. A median of 5 (range: 2-7) vaccines was given. There were no serious adverse events except in one patient with gross residual tumour prior to vaccination, who repetitively developed vaccine-related peritumoral oedema. Minor toxicities were recorded in four out of 12 patients. In six patients with postoperative residual tumour, vaccination induced one stable disease during 8 weeks, and one partial response. Two of six patients with complete resection are in CCR for 3 years. Tumour vaccination for patients with relapsed malignant glioma is feasible and likely beneficial for patients with minimal residual tumour burden.

Desmoplastic infantile ganglioglioma is a rare intracranial tumor of early childhood with a usually excellent prognosis despite malignant features both radiologically and histologically. We present the case of a desmoplastic infantile ganglioglioma with histologically highly anaplastic features and both intracerebral and pial metastases. After partial resection the tumor was rapidly progressive and new metastases appeared. A combination of vincristine and carboplatinum was used according to the Low Grade Glioma Protocol of the International Society of Pediatric Oncology, with a temporary good response. When histologically characterized by highly anaplastic features, it seems the biologic behavior of this tumor remains uncertain. The aggressive behavior and the responsiveness to chemotherapy in this case may challenge the belief in the benign nature of these rare tumors.

Pelvic gynecological malignancies account for 6% of all cancers. In the relapsed state, classical treatments are limited. There is an urgent need for new and personalized treatment. Wilms' tumor gene 1 (WT1) is the most important tumor-associated antigen. Although highly present in gynecological tumors, active immunotherapy against it is still underexplored. This review gives an insight into the importance of WT1 in pelvic gynecological malignancies and the first taken steps into the world of WT1 immunotherapy.

Occupancy of CTLA-4 (cytotoxic T-lymphocyte antigen-4 or CD152) negatively regulates the activation of mouse T lymphocytes, as indicated by the fate of CTLA-4-deficient mice, by the impact of anti-CTLA-4 monoclonal antibodies (mAbs) on mouse T-cell activation in vitro and by the impact of CTLA-4 blockade on the course of experimental tumoral, autoimmune, alloimmune or infectious disease in this animal. The function of human CTLA-4, however, remains less clear. The expression and function of human CTLA-4 were further explored. CTLA-4 was expressed under mitogenic conditions only, its expression being, at least partially, dependent on the secretion of interleukin-2. Memory T cells expressed CTLA-4 with faster kinetics than naive T cells. The functional role of human CTLA-4 was assessed utilizing a panel of four anti-CTLA-4 mAbs that blocked the interaction between CTLA-4 and its ligands. These mAbs, in immobilized form, profoundly inhibited the activation of T cells by immobilized anti-CD3 mAb in the absence of anti-CD28 mAb, but co-stimulated T-cell activation in the presence of anti-CD28 mAb. Finally, and importantly, blockade of the interaction of CTLA-4 with its ligands using soluble anti-CTLA-4 mAbs, in intact form or as Fab fragments, enhanced T-cell activation in several polyclonal or alloantigen-specific CD80- or CD80/CD86-dependent assays, as measured by cytokine production, cellular proliferation or cytotoxic responses. It is concluded that interaction of CTLA-4 with its functional ligands, CD80 or CD86, can down-regulate human T-cell responses, probably by intracellular signalling events and independent of CD28 occupancy.

Ligation of CD28 on T cells with its natural ligands B7-1 (CD80) or B7-2 (CD86) provides a major costimulatory signal for T cells and is of potential importance for tumor rejection. We previously reported a strong expression of B7-1 on Reed-Sternberg cells and anaplastic large cell lymphoma cells. We report here our findings on B7-2 expression by malignant lymphomas (n = 70). B7-2 was present on the neoplastic cells of anaplastic large cell lymphoma in two of three cases studied, and on a subpopulation of the malignant cells in one out of four cases of follicular lymphoma. B7-2 was not expressed by the neoplastic cells of the other non-Hodgkin's lymphomas (n = 32), including T cell-rich B cell lymphoma. In contrast, Reed-Sternberg cells in lymph nodes affected by Hodgkin's disease are strongly positive for B7-2 (n = 31). Evidence for a functional correlate of this expression was obtained by our findings that the combination of anti-B7-1 and anti-B7-2 monoclonal antibodies was more effective than each separately in blocking allogeneic T cell activation (proliferation and cytokine secretion) by Hodgkin's disease-derived cell lines as stimulators. The possible role of B7-1 and B7-2 expression for the course and symptomatology of Hodgkin's disease is discussed.

BACKGROUND Recently, methylxanthines such as pentoxifylline (PTX) were shown to be immunosuppressive in vitro. Unfortunately, when used in transplant patients, PTX was poorly active as an immunosuppressant. Here we report that the new methylxanthine derivative A802715 not only is more active than PTX, it also suppresses the cyclosporine (CsA)-resistant "signal two"-dependent pathway of T cell proliferation, making it an interesting drug to associate with CsA. METHODS "Signal one"- and "signal two"-dependent T cell activation was investigated with purified human T cells stimulated with immobilized anti-CD3 or anti-CD28 monoclonal antibody (mAb) plus phorbol myristate acetate (PMA) or with a 3T6 mouse fibroblast cell line presenting anti-CD3 mAb on transfected human Fcgamma receptors II (FcgammaRII) in the presence or absence of transfected B7-1 (CD80) molecules. RESULTS A802715 was more immunosuppressive in the mixed lymphocyte reaction (MLR) than PTX. A802715 dose-dependently suppressed polyclonal signal one-dependent T cell activation induced by anti-CD3 mAb/PMA. In addition, A802715 also suppressed signal two-dependent T cell proliferation induced by anti-CD28 mAb/PMA. The expression of the interleukin-2 receptor on T cells stimulated by anti-CD3 mAb presented on 3T6/FcgammaRII cells was equally well suppressed by A802715 and PTX. In contrast, interleukin-2 receptor or CD40L (gp39) expression by T cells after stimulation with the same anti-CD3 mAb- 3T6/FcgammaRII cells, but coexpressing transfected B7-1, was only suppressed by A802715. The anticipated synergism between A802715 and CsA was confirmed in MLR assays. Moreover, generation of cytotoxic T lymphocytes during MLR with Epstein-Barr virus-transformed B cells, which strongly express B7-1 and B7-2, was also inhibited by A802715. CONCLUSIONS These in vitro data indicate that the A802715 (1) is a stronger immunosuppressant for T cells than PTX, (2) suppresses T cell activation pathways that are resistant to PTX or CsA, and (3) acts synergistically with CsA.

Introduction and Aim: Glioblastoma Multiforme is a WHO grade IV neoplasm and is the most frequent primary brain tumor in adults. In spite of current multimodal treatment the prognosis remains poor, with a median overall survival of 14.6 months and a mortality rate of 88% within 3 years. At time of relapse (which is universal), the prognosis is particularly dismal with reports of 100% mortality within 1.5 years. In view of these discouraging results, (pre)clinical research is focused on long-term and tumor-specific treatments, such as oncolytic virotherapy. Not only do oncolytic viruses have the potential to lower tumor volume and viability considerably, they might also alter the local microenvironment within the tumor, to subsequently induce a better immune control through immunotherapeutic approaches. Here we investigate the efficacy of the oncolytic Reovirus Type 3 Dearing, Parvovirus H-1 and Newcastle disease virus (NDV) Hitchner B1 strains in the orthotopic murine GL261 glioma model. Materials and Methods: monolayers of GL261 cells and healthy astrocytes were inoculated with Reovirus, Parvovirus or NDV at different multiplicities of infection, ranging from 0,05 to 10 plague forming units/cells. 72 hours later, cell death was measured via MTT assays. C57/BL6 and Rag2-/- mice were challenged with GL261 cells orthotopically and treated intratumorally with NDV on day 3 or day 7 after tumor induction. Animals were followed for symptoms and survival. Long-term surviving animals were rechallenged with GL261 cells after 100 days. Results: We demonstrated in vitro that Reovirus, Parvovirus and NDV exhibit strong cytotoxicity towards the GL261 cell line. In vivo we showed that local injection of NDV was feasible and significantly prolonged survival of treated animals versus untreated controls. Furthermore, 45% of treated animals survived long-term versus none in the control group. Similar survival data were obtained whether NDV was injected at time of minimal disease or at time of bulky tumor. Tumor rechallenge in long-term surviving animals resulted in 80% survival, suggesting immunization in most mice after NDV therapy. In immunodeficient Rag2-/- animals, NDV failed to prolong survival after glioma challenge, further substantiating the notion that the anticancer effects of NDV can be tracked not only to direct viral oncolysis, but also to an additional immune component. Conclusion: For the first time, we describe that NDV has therapeutic activity against GL261 tumor cells, evidenced in an orthotopic mouse model. NDV therapy significantly improves median survival and induces an antitumor memory response in treated animals. This is a highly innovative finding and ongoing work is focused on further elucidating the immune component in successful oncolytic virotherapy. Furthermore, this model will allow us to study the potential synergism between NDV therapy and dendritic cell mediated immunotherapy, which thus far has not been studied in the field of glioma research

Objective The development of osteoarthritis (OA) may be caused by activation of hypertrophic differentiation of articular chondrocytes. Healthy articular cartilage is highly resistant to hypertrophic differentiation, in contrast to other hyaline cartilage subtypes, such as growth plate cartilage. The purpose of this study was to elucidate the molecular mechanism responsible for the difference in the propensity of human articular cartilage and growth plate cartilage to undergo hypertrophic differentiation. Methods Whole-genome gene-expression microarray analysis of healthy human growth plate and articular cartilage derived from the same adolescent donors was performed. Candidate genes, which were enriched in the articular cartilage, were validated at the messenger RNA (mRNA) and protein levels and examined for their potential to inhibit hypertrophic differentiation in two models. In addition, we studied a possible genetic association with OA. Results Pathway analysis demonstrated decreased Wnt signaling in articular cartilage as compared to growth plate cartilage. This was at least partly due to increased expression of the bone morphogenetic protein and Wnt antagonists Gremlin 1, Frizzled-related protein (FRP), and Dkk-1 at the mRNA and protein levels in articular cartilage. Supplementation of these proteins diminished terminal hypertrophic differentiation without affecting chondrogenesis in long-bone explant cultures and chondrogenically differentiating human mesenchymal stem cells. Additionally, we found that single-nucleotide polymorphism rs12593365, which is located in a genomic control region of GREM1, was significantly associated with a 20% reduced risk of radiographic hip OA in 2 population-based cohorts. Conclusion Taken together, our study identified Gremlin 1, FRP, and Dkk-1 as natural brakes on hypertrophic differentiation in articular cartilage. As hypertrophic differentiation of articular cartilage may contribute to the development of OA, our findings may open new avenues for therapeutic intervention.

We performed conventional cytogenetic analysis and fluorescence in situ hybridization in short-term cultures of normal and neoplastic kidney tissues. Cell populations carrying an extra chromosome 7 or an extra chromosome 10 as the only chromosome change could be identified in kidney tumors, mostly renal cell carcinomas, and in the surrounding kidney tissue, but not in nonneoplastic kidneys. To identify the type of cells displaying these aneuploidies, we performed in situ hybridization (ISH) with probes specific for the centromeric region of chromosomes 7 and 10 on frozen kidney tissue sections. Trisomy 7 and trisomy 10 were restricted to infiltrating inflammatory cells in the tumor as well as in the surrounding tissue. Trisomy 7 and trisomy 10 were also found in subpopulations of peripheral blood T cells of cancer patients and of normal individuals, as well as in the thymus of five normal fetuses (21-29 weeks), but not in noninvaded reactive lymph node sections of patients without malignancy. When lymphocytes were enriched from kidney tumors and surrounding tissue by either Ficoll/Hypaque density gradient or immunomagnetic selection with anti-CD3, anti-CD4, or anti-CD8 monoclonal antibodies, it was confirmed that they contained a high percentage of trisomy 7 and trisomy 10 cells. Further proof for T-lymphocyte origin of the trisomy 7 and trisomy 10 cells was obtained by simultaneous staining of lymphocytes isolated from tumor tissue with anti-CD3, anti-CD4, and anti-CD8 monoclonal antibodies and ISH. We conclude that trisomy 7 and trisomy 10, found in renal carcinomas and surrounding kidney tissue, characterize subpopulations of tumor-infiltrating lymphocytes. The biologic significance of this phenomenon is unknown and requires further investigation.

ispartof: Histopathology vol:54 issue:3 pages:384-387 ispartof: location:England status: published

The value of bioluminescence imaging (BLI) for experimental cancer models has become firmly established. We applied BLI to the GL261 glioma model in the context of dendritic cell (DC) immunotherapy. Initial validation revealed robust linear correlations between in vivo, ex vivo and in vitro luciferase activity measurements. Ex vivo BLI demonstrated midline crossing and leakage of tumor cells. Orthotopically challenged mice followed with BLI showed an initial adaptation phase, after which imaging data correlated linearly with stereologically determined tumor dimensions. Transition from healthy to moribund state corresponded with an increasing in vivo flux but the onset of neurological deficit was clearly delayed compared to the onset of in vivo flux increase. BLI was implemented in prophylactic immunotherapy and imaging data were prognostic for therapy outcome. Three distinct response patterns were detected. Our data underscore the feasibility of in vivo BLI in an experimental immunotherapeutic setting in the GL261 glioma model.

A 69-year-old man presented with progressive blurred vision and corneal clouding (Fig. 1A, B). No other ocular abnormalities were present. There was no history of cardiovascular problems. On slit-lamp examination, corneal clouding was seen (Fig. 1C: left eye). Lipid profile showed total cholesterol 186 mg/dl (normal e190), low-density lipoproteins 132 mg/dl (e115), high-density lipoproteins 9 mg/dl (Q40) and triglycerides 236 mg/dl (e180). Esterified cholesterol fraction in plasma of the patient (0.59) was decreased compared to controls (0.73), with the most marked decline observed in HDL particles (0.54 versus 0.81). The diagnosis of fish-eye disease (FED, OMIM #136120), caused by a mutation in the lecithin:cholesterol acyltransferase (LCAT) gene (Carlson and Holmquist 1985) was made, based on the presentation, the lipid profile and homozygosity for a known point mutation (440C>T, J Inherit Metab Dis DOI 10.1007/s10545-008-0853-6

Malignant gliomas are highly proliferative and invasive tumors with poor prognosis. We investigated the influence of Interferon-gamma (IFN-gamma) on the human malignant glioma cell line A172, measuring cell viability (MTT-test), proliferation (3H-thymidine-uptake), cell death (FACS) adhesion to hyaluronic acid (HA, adhesion-assay) and migration (Boyden-chamber). IFN-gamma significantly decreased cell viability and proliferation. Measured by FACS, an up-regulation of CD95 expression has been shown in combination with an increased rate of cell death, first seen after 96 hours IFN-gamma treatment. Adhesion to HA was decreased after pre-treatment with IFN-gamma. This was not mediated by down-regulation of the main HA-receptor CD44, since IFN-gamma did not change CD44 expression. IFN-gamma-treated cells showed a significantly diminished migration rate through a native or HA-coated 8-microm polycarbonate membrane. To summarise, IFN-gamma influences both the main characteristics of malignancy: it decreases cell proliferation and induces cell death, further it diminishes migration of A172 human glioblastoma cells.

Various immunotherapeutical approaches are presently evaluated for their efficacy to eradicate glioma cells. Complicating the concepts, these tumors secrete cytokines which modulate the immune response.We analyzed the influence of interferon gamma (IFN-gamma) on the cytokine production and IFN-gamma receptor expression in T98G and U87-MG human glioma cells.The IFN-gamma receptors were own-regulated after IFN-gamma treatment. Secretion of interleukin-6 (IL-6) protein was elevated by factors of 6.4 in T98G cells and 5.2 in U87-MG. Other cytokines were increased as well, but less constantly: IL-8, and VEGF were elevated significantly only in T98G, but not in U87-MG. Increases of IL-1 beta, IL-1 alpha and TGF beta-2 were only detectable at the mRNA level. TNF was not detectable in any of the cell lines, and TGF-beta, alpha FGF and HG were not influenced by IFN-gamma.IL-6 produced by glioma cells in response to IFN-gamma might support immune eradication of glioma cells.

gamma-Butyrobetaine hydroxylase catalyse the last step in carnitine biosynthesis, the formation of L-carnitine from gamma-butyrobetaine, a reaction dependent on Fe2+, alpha-ketoglutarate, ascorbate and oxygen. Initial attempts to purify the protein from rat liver showed that gamma-butyrobetaine hydroxylase is unstable. We, therefore, determined the influence of various compounds on the stability of gamma-butyrobetaine hydroxylase at different storage temperatures. The enzyme activity was best conserved by storing the protein at 4 degrees C in the presence of 200 g/l glycerol and 10 mM DTT. We subsequently purified the enzyme from rat liver to apparent homogeneity by liquid chromatography.

gamma-Butyrobetaine hydroxylase (EC 1.14.11.1) is the last enzyme in the biosynthetic pathway of L-carnitine and catalyzes the formation of L-carnitine from gamma-butyrobetaine, a reaction dependent on alpha-ketoglutarate, Fe2+, and oxygen. We report the purification of the protein from rat liver to apparent homogeneity, which allowed N-terminal sequencing using Edman degradation. The obtained amino acid sequence was used to screen the expressed sequence tag database and led to the identification of a human cDNA containing an open reading frame of 1161 base pairs encoding a polypeptide of 387 amino acids with a predicted molecular weight of 44.7 kDa. Heterologous expression of the open reading frame in the yeast Saccharomyces cerevisiae confirmed that the cDNA encodes the human gamma-butyrobetaine hydroxylase. Northern blot analysis showed gamma-butyrobetaine hydroxylase expression in kidney (high), liver (moderate), and brain (very low), while no expression could be detected in the other investigated tissues.

Recently, methylxanthines such as pentoxifylline (PTX) were shown to be immunosuppressive in vitro. Unfortunately, when used in transplant patients, PTX was poorly active as an immunosuppressant. Here we report that the new methylxanthine derivative A802715 not only is more active than PTX, it also suppresses the cyclosporine (CsA)-resistant "signal two"-dependent pathway of T cell proliferation, making it an interesting drug to associate with CsA."Signal one"- and "signal two"-dependent T cell activation was investigated with purified human T cells stimulated with immobilized anti-CD3 or anti-CD28 monoclonal antibody (mAb) plus phorbol myristate acetate (PMA) or with a 3T6 mouse fibroblast cell line presenting anti-CD3 mAb on transfected human Fcgamma receptors II (FcgammaRII) in the presence or absence of transfected B7-1 (CD80) molecules.A802715 was more immunosuppressive in the mixed lymphocyte reaction (MLR) than PTX. A802715 dose-dependently suppressed polyclonal signal one-dependent T cell activation induced by anti-CD3 mAb/PMA. In addition, A802715 also suppressed signal two-dependent T cell proliferation induced by anti-CD28 mAb/PMA. The expression of the interleukin-2 receptor on T cells stimulated by anti-CD3 mAb presented on 3T6/FcgammaRII cells was equally well suppressed by A802715 and PTX. In contrast, interleukin-2 receptor or CD40L (gp39) expression by T cells after stimulation with the same anti-CD3 mAb- 3T6/FcgammaRII cells, but coexpressing transfected B7-1, was only suppressed by A802715. The anticipated synergism between A802715 and CsA was confirmed in MLR assays. Moreover, generation of cytotoxic T lymphocytes during MLR with Epstein-Barr virus-transformed B cells, which strongly express B7-1 and B7-2, was also inhibited by A802715.These in vitro data indicate that the A802715 (1) is a stronger immunosuppressant for T cells than PTX, (2) suppresses T cell activation pathways that are resistant to PTX or CsA, and (3) acts synergistically with CsA.

The interaction of CD28 with one of the B7 molecules (CD80 and CD86) on professional antigen-presenting cells (APC) is generally considered as the most important co-stimulatory signal for T cell activation. APC in a resting condition express either no or only low levels of B7 molecules. These are up-regulated as a result of interactions with activated T cells, thus suggesting that B7-CD28 interaction is not required at initiation of T cell activation. To study this issue, we blocked B7-CD28 interaction at various time points after in vitro stimulation of peripheral blood T cells with allogeneic monocytes. Epstein-Barr virus-transformed B cells or soluble antigens. We observed that T cell proliferation and IL-2 production were inhibited by B7-blocking agents (CTLA-4-Ig or anti-B7 mAb) almost to the same degree when added either at initiation of culture or 24 h later. B7-blocking agents still resulted in significant inhibition of allogeneic T cell activation when added after 48 h. Furthermore, when CTLA-4-Ig was added at the start of an allogeneic T cell stimulation, addition of anti-CD28 mAb after 24 h of culture nearly fully restored T cell proliferation to control levels. Finally, we demonstrate that delayed addition of B7-blocking agents together with cyclosporin A 1 day after the onset of culture of T cells with allogeneic B cells is highly efficient to induce energy as evaluated by lack of proliferation, cytotoxic T lymphocyte reactivity and IFN-gamma or IL-5 production upon alloantigen rechallenge. Taken together, our data can explain why B7 expression on APC is not required at the time of initial APC-T cell contact, and suggest that the effect of the CD28 signal indeed consists in prolonging IL-2 production and amplifying T cell responses, rather than in providing a critical co-stimulatory signal at the time of initial TCR triggering.

Polyclonal intravenous immunoglobulins (IVIg) are increasingly used in clinical practice, not only as substitutive agents but also in the treatment of immunologically mediated diseases.1,2 How IVIg modulate the immune system is not yet clear, but several immunoregulatory mechanisms have been postulated.2 We studied the production of the immunosuppressive cytokine interleukin 10 (IL10)3 as a potential explanation for the beneficiary effects of IVIg in autoimmune diseases. IL10 concentrations were measured in six patients: one patient (aged 19) with X-linked agammaglobulinaemia, two patients (aged 63 and 64) with common variable immunodeficiency, one patient (aged 15) with hyper IgM syndrome, two patients (aged 4) with IgG2 subclass deficiency. All patients or their parents gave informed consent. Blood was collected before and immediately after infusion of IVIg (Sandoglobulin; Novartis, Brussels, Belgium), and IL10 levels were measured with a sandwich enzyme linked …