Congenic hematopoietic stem cell transplantation promotes survival of heart allografts in murine models of acute and chronic rejection.

The ability to induce tolerance would be a major advance in the field of solid organ transplantation. Here, we investigated whether autologous (congenic) hematopoietic stem cell transplantation (HSCT) could promote tolerance to heart allografts in mice. In an acute rejection model, fully MHC-mismatched BALB/c hearts were heterotopically transplanted into C57BL/6 (CD45.2) mice. One week later, recipient mice were lethally irradiated and reconstituted with congenic B6 CD45.1 Lin-Sca1+ckit+ cells. Recipient mice received a 14-day course of rapamycin both to prevent rejection and to expand regulatory T cells (Tregs). Heart allografts in both untreated and rapamycin-treated recipients that did not undergo HSCT were rejected within 33 days (median survival time = 8 days for untreated recipients, median survival time = 32 days for rapamycin-treated recipients), whereas allografts in HSCT-treated recipients had a median survival time of 55 days (P < 0.001 vs. both untreated and rapamycin-treated recipients). Enhanced allograft survival following HSCT was associated with increased intragraft Foxp3+ Tregs, reduced intragraft B cells, and reduced serum donor-specific antibodies. In a chronic rejection model, Bm12 hearts were transplanted into C57BL/6 (CD45.2) mice, and congenic HSCT was performed two weeks following heart transplantation. HSCT led to enhanced survival of allografts (median survival time = 70 days vs. median survival time = 28 days in untreated recipients, P < 0.01). Increased allograft survival post-HSCT was associated with prevention of autoantibody development and absence of vasculopathy. These data support the concept that autologous HSCT can promote immune tolerance in the setting of allotransplantation. Further studies to optimize HSCT protocols should be performed before this procedure is adopted clinically.

CD200 mimetic aptamer PEG-M49 markedly increases the therapeutic effects of pegylated liposomal doxorubicin in a mouse model of metastatic breast carcinoma: an effect independent of CD200 receptor 1.

We previously reported that CD200 overexpression in the host decreases progression and metastasis of the highly aggressive metastatic 4THM breast carcinoma. We have explored a possible synergistic interaction between the CD200 mimetic PEG-M49 and pegylated liposomal doxorubicin (Peg-Dox) in wild-type CD200 knockout (CD200-/-) and CD200 Receptor 1 knockout (CD200R1-/-) mice for the first time. A 4THM breast carcinoma model and three groups of BALB/c mice (wild type, CD200-/- and CD200R1-/-) were used. Five days after injection of tumor cells, mice were injected with Peg-Dox (ip, once a week) and PEG-M49 or a control aptamer (iv, every 3 days). Necropsies were performed either 12 (mid-point) or 24 (endpoint) days after injection and the extent of tumor growth, visceral metastasis and changes in the tumor-directed immune response were evaluated. PEG-M49 and Peg-Dox co-treatment induced complete tumor regression and loss of macroscopic lung metastasis in four out of seven WT mice. This synergistic anti-tumoral effect is thought to be due to Peg-M49-induced inhibition of Gr1 + CD11b + cells and Peg-Dox-induced increases in tumor-infiltrating CD8 + and CD8CD4 double-positive cells. Similar changes were observed in CD200R1-/- mice indicating that the primary effects of Peg-M49 are mediated by non-CD200R1 receptors. We also demonstrated for the first time that tumor growth, metastasis, and tumor infiltrating GR1 + CD11b + cells were markedly increased in CD200R1-/- mice, indicating an anti-inflammatory and protective role of CD200. CD200 mimetics might be a safe and effective immunomodulatory treatment in conjunction with classical chemotherapeutics for therapy of aggressive metastatic breast carcinoma.

Importance of B cells to development of regulatory T cells and prolongation of tissue allograft survival in recipients receiving autologous bone marrow transplantation.

We previously showed that congenic bone marrow transplantation (BMTx) post myeloablation augmented tissue allograft survival in association with increased regulatory T (Treg) cells of both host and bone marrow donor origin. Regulatory B (Breg) cells can also modulate T-cell immunity and B cells may be implicated in the development of Treg cells. Accordingly, we explored the effect of B-cell depletion in vivo on augmented graft survival post BMTx. C57BL/6 mice received BALB/c skin allografts followed 7 days later by myeloablation using cyclophosphamide and busulphan. Mice then received T-cell-depleted bone marrow from CD45.1 congenic donors, and ongoing immunosuppression with rapamycin (to day 28 after BMTx). Control mice received cyclophosphamide and busulphan followed by rapamycin, but not congenic bone marrow. At different times post BMTx, mice received B-cell-depleting antibody treatment, and the effect on both skin graft survival, and induction of Treg cells was assessed. BMTx resulted in significantly prolonged skin graft survival versus control mice, in association with attenuated donor-specific alloreactivity relative to controls, increased splenic Treg cells and significantly diminished anti-donor IgG. In mice receiving infusion of B-depleting antibodies for 12 days from day 15 post BMTx, both graft survival and Treg cell activity were diminished, particularly for functional Treg cells of donor origin. Adoptive transfer of Breg cells from mice harvested at 15 days post BMTx prolonged survival in naive transplanted mice and increased Treg cell levels. Thus, autologous BMTx augmentation of graft survival is dependent in part upon a population of Breg cells that can modulate the function of donor-derived Treg cells.

Effect of infusion of monoclonal antibodies to tumour necrosis factor-receptor super family 25 on graft rejection in allo-immune mice receiving autologous marrow transplantation.

Significant barriers to transplantation exist for individuals who are pre-sensitized to donor antigen and have high titres of donor-reactive antibody. We report the effect of autologous bone marrow transplantation (BMTx) after myeloablation in pre-sensitized mice along with the use of monoclonal antibodies (mAbs) to tumour necrosis factor-receptor super family 25 (TNFRSF25), expressed on regulatory T (Treg) cells. C57BL/6 mice, which had been sensitized earlier with BALB/c skin allografts, received secondary BALB/c grafts after the primary grafts had been rejected. Subsequently, recipient mice underwent myeloablation with cyclophosphamide and busulphan and were injected with T-cell-depleted bone marrow from CD45.1 congenic donors (BMTx). Recipient mice underwent immunosuppressive treatment with rapamycin. A subgroup of mice was also treated with mAbs to TNFRSF25. Control mice were pre-sensitized mice that received cyclophosphamide and busulphan followed by rapamycin. BMTx-treated mice had significantly prolonged skin graft survival versus control mice. These mice also showed attenuated donor-specific mixed lymphocyte co-culture responses relative to controls, increased splenic Treg cells and markedly diminished serum anti-donor IgG. Infusion of anti-TNFRSF25 mAbs further augmented graft survival and increased graft-infiltrating Treg cells. These mAbs also expanded murine and human Treg cells in vitro with the capacity to attenuate mixed lymphocyte co-cultures using fresh peripheral blood mononuclear cells. Overall, this study delineates the roles of autologous BMTx and anti-TNFRSF25 mAbs in expanding Treg cells and attenuating alloimmune responses in pre-sensitized mice.

A comparison of serum miRNAs influencing metastatic growth of EMT6 vs 4THM tumor cells in wild-type and CD200R1KO mice.

PURPOSE: We investigated whether miRNAs in exosomes from EMT6 or 4THM tumor-bearing mice played a role in regulating inflammatory cytokine expression and/or metastasis in WT mice injected with EMT6 and/or 4THM tumor cells. METHODS: EMT6 tumors in BALB/c CD200R1KO mice resolve following surgical resection of localized tumor and immunization with irradiated EMT6 cells along with CpG as adjuvant. Wild-type (WT) animals treated in the same fashion develop pulmonary and liver metastases within 20 days of surgery. DLNs from CD200R1KO mice contain no tumor cells detectable at limiting dilution. In contrast, 4THM tumor cells injected into CD200R1KO show increased metastasis compared with WT mice. Transfer of serum exosomes from 4THM tumor-bearing mice to WT animals increased metastasis of EMT6 tumors, an effect attenuated by anti-IL-6 antibody. We compared miRNA expression in exosomes from the serum of 4THM/EMT6 WT or CD200R1KO tumor-bearing mice, and the effects of antagomirs to miRNAs on tumor growth. RESULTS: Complex changes in miRNA expression were observed in the isolated exosomes. Some miRNAs, including miR155, have been reported to potentiate inflammatory responses and augment inflammatory cytokine expression. Expression of miR155 increased in exosomes from 4THM relative to EMT6 tumor bearers, and antagomirs to miR155 attenuated tumor growth and metastasis, and improved survival, following infusion into WT mice. Antagomirs to the miR205 family were thought to affect metastasis by targeting epithelial-to-mesenchymal transition (EMT), increased growth and metastasis in both 4THM and EMT6 tumor-bearing mice, and decreased survival, with some modulation of inflammatory cytokine production. CONCLUSIONS: Multiple pathways are implicated in differential metastasis of EMT6/4THM, and targeting these may have clinical utility in human breast cancer.

Omalizumab (anti-IgE) therapy increases blood glucose levels in severe persistent allergic asthma patients with diabetes mellitus: 18 month follow-up.

BACKGROUND: Therapeutic anti-IgE antibodies (Xolair, omalizumab) able to reduce free IgE levels and to block the binding of IgE to Fcepsilon RI without cross-linking IgE and triggering degranulation of IgE-sensitised cells have been developed. METHODS: We had two male patients of severe persistent allergic asthma with type-2 diabetes mellitus at the ages of 57 and 52 and who had suffered a side-effect of increased blood glucose level that caused a need for an extra insulin injection to control the hyperglycemia. Their asthma was not under control, frequent emergency department admissions lead us to use omalizumab treatment. Assessment of clinical changes and adverse effects were evaluated at each bimonthly patient visit including vital signs, full physical examination, details of any allergy incidents, total and specific IgE levels, serum ECP (eosinophilic cationic peptid) levels, pulmonary function test, exhaled nitric oxide concentrations, and asthma control test. RESULTS: Both patients were on week 42 - 45 of omalizumab treatment with a the dosage of 375 and 300 mg when they had the adverse reaction we reported here; they also had no other complaints. Blood levels of ECP and high sensitive CRP (hs-CRP) were decreased after starting the treatment of anti-IgE. CONCLUSIONS: To our knowledge, this is the first time an association between omalizumab use and hyperglycemia has been documented. Every vial of Xolair (150 mg) contains 145.5 mg sucrose and it might increase the blood levels of glucose in diabetics. As a conclusion the prescribing information might have been revised based on post marketing surveillance data and reported such cases indicating that different side effects may occur beyond 2 hours of the injection.

High hypoxia status in pancreatic cancer is associated with multiple hallmarks of an immunosuppressive tumor microenvironment.

Introduction: Pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer, is a particularly lethal disease that is often diagnosed late and is refractory to most forms of treatment. Tumour hypoxia is a key hallmark of PDAC and is purported to contribute to multiple facets of disease progression such as treatment resistance, increased invasiveness, metabolic reprogramming, and immunosuppression. Methods: We used the Buffa gene signature as a hypoxia score to profile transcriptomics datasets from PDAC cases. We performed cell-type deconvolution and gene expression profiling approaches to compare the immunological phenotypes of cases with low and high hypoxia scores. We further supported our findings by qPCR analyses in PDAC cell lines cultured in hypoxic conditions. Results: First, we demonstrated that this hypoxia score is associated with increased tumour grade and reduced survival suggesting that this score is correlated to disease progression. Subsequently, we compared the immune phenotypes of cases with high versus low hypoxia score expression (HypoxiaHI vs. HypoxiaLOW) to show that high hypoxia is associated with reduced levels of T cells, NK cells and dendritic cells (DC), including the crucial cDC1 subset. Concomitantly, immune-related gene expression profiling revealed that compared to HypoxiaLOW tumours, mRNA levels for multiple immunosuppressive molecules were notably elevated in HypoxiaHI cases. Using a Random Forest machine learning approach for variable selection, we identified LGALS3 (Galectin-3) as the top gene associated with high hypoxia status and confirmed its expression in hypoxic PDAC cell lines. Discussion: In summary, we demonstrated novel associations between hypoxia and multiple immunosuppressive mediators in PDAC, highlighting avenues for improving PDAC immunotherapy by targeting these immune molecules in combination with hypoxia-targeted drugs.

Cure of metastatic growth of EMT6 tumor cells in mice following manipulation of CD200:CD200R signaling.

In previous studies, we observed that regulation of expression of CD200, both on cells of a transplantable breast cancer, EMT6, and of the host, as well as of the receptor, CD200R in host mice, regulated local tumor growth and metastasis in immunocompetent animals. This in turn led to an improved ability to document immunity to EMT6 in CD200R1KO mice. In the current study, we have explored the ability to cure BALB/c CD200KO or CD200R1KO mice of tumors ≤1 cm(3) in size by surgical resection of localized tumor, followed by immunization with irradiated EMT6 cells along with CpG as adjuvant. While control animals treated in this fashion developed significant pulmonary and liver metastases within 30 days of surgery, significant protection was seen in both CD200KO or CD200R1KO mice, with no macroscopic lung/liver metastases observed in CD200R1KO mice on sacrifice at day 300. Following surgical resection and immunization, draining lymph nodes from control mice contained tumor cells cloned at limiting dilution in vitro even before pulmonary and hepatic metastasis was seen. In contrast, within the limits of detection of the assay used (sensitivity ~1 in 10(7) cells), no tumor cells were detected at limiting dilution in similarly treated CD200R1KO mice, and significant reductions were seen in CD200KO mice. Infusion of anti-CD4, but less so anti-CD8, mAb into surgically treated and immunized CD200R1KO mice attenuated protection from both macroscopic (liver/lung) and microscopic (assayed by limiting dilution of DLN) metastasis. Adoptive transfer of lymphocytes from treated CD200R1KO mice to surgically treated control mice also attenuated metastatic growth of tumor, which was abolished by pretreatment of transferred cells with anti-CD4 mAb. Our data suggest that CD200:CD200R attenuates a potentially tumor-protective CD4 host response to breast cancer.

Clinical course and side effects of anti-IgE monoclonal antibody in patients with severe persistent asthma.

BACKGROUND: Omalizumab, a recombinant humanized monoclonal antibody to IgE, is recommended as a new option for the treatment of severe persistent allergic asthma. The purpose of this study is to assess the effects omalizumab treatment on life quality and its side effects in severe persistent asthma patients. METHODS: In this study, we evaluated 19 severe persistent asthma patients who received therapy with omalizumab for 8 months. Omalizumab was administered every 2 weeks at doses between 150 to 375 mg. Symptoms and severity of allergic reactions were recorded before and after being on omalizumab. IgE levels, mean platelet volume (MPV), platelet levels, pulmonary function test, and asthma control test were evaluated in all patients before and 8 months after the treatment. Local and systemic side effects of omalizumab were evaluated. Stool parasites were examined in the 4th and 8th month after initiation of treatment to investigate any parasitosis. RESULTS: The patients had severe persistent asthma for periods ranging from 3 to 8 years, and they were diagnosed with allergic asthma for 7 - 28 years. Thrombocytopenia developed in a male patient after the 22nd dose of the drug was given. When the platelet count fell down to 55000, the omalizumab treatment was suspended. During the therapy period, one patient had parasitosis (giardiasis), one patient had severe side effects, one patient had dyspnea two hours after the injection, and one patient had a dyspnea attack 2 hours after the injection. The changes in MPV levels were not statistically significant. There was also a significant decrease in IgE levels after the treatment. CONCLUSIONS: Monitoring of complete blood cell count is very important when using this drug. Though we did not see anaphylaxis in any patients, we believe that the patients should be monitored at least for 3 hours after the omalizumab injection.

A case of toxic epidermal necrolysis with diverse etiologies: successful treatment with intravenous immunoglobulin and pulse prednisolone and effects on sTRAIL and sCD200 levels.

BACKGROUND: Here, we report the first case of patient with intracranial tumors (ICT) who developed a cutaneous adverse drug reaction during lansoprazole and prophylactic anticonvulsant treatment. SCORTEN is a scoring system used to predict mortality in TEN patients. If SCORTEN index is 5 or more, mortality rate is more than 90%. SCORTEN of our patient was calculated as 5. METHODS: Our patient is a 64 year-old white female, who had glioma and had been on post-op prophylactic anticonvulsant therapy. On the 3rd day post operation, lansoprazole was added to the therapy. After the first lansoprazole dose, erythematous dusky red macules occurred on extremities and trunk and on the following day confluent purpuric lesions tended to run together in 95% of the whole body including scalp, oral and genital mucosa. Nikolsky's Sign was positive on the skin. Physical examination; body temperature was 38.4 degrees C with a heart rate of 146 beats/minute and 80/50 mm Hg arterial blood pressure, Glascow Coma Scale was E1 M1e, pupillary light reflex was 2/2 +/+ and she was confused. Her biopsy resulted as toxic epidermal necrolysis. Moreover, sTRAIL and sCD200 levels of serum and blister fluid were investigated as an apoptotic marker and a negative marker for inflammation. RESULTS AND CONCLUSIONS: sTRAIL and sCD200 were evaluated both in the sera and blister fluid. sTRAIL level was lower than for healthy individuals with high levels in blister fluid; and sCD200 level was depressed by up to 10% of the normal values of healthy individuals but with high levels in the blister fluid during the active phase of the disease. After our successful treatment with human albumin, prednisolone pulse therapy, and IVIG at a dose of 400 mg/kg, she was discharged from the hospital on the 23rd day and followed up after 2 months. The increase in sTRAIL (up to two-fold) and sCD200 (up to six-fold) levels may provide useful information in understanding disease pathogenesis and monitoring treatment efficacy.

Pollen aero allergens and the climate in Mediterranean region and allergen sensitivity in allergic rhinoconjunctivitis and allergic asthma patients.

BACKGROUND: We evaluated the profiles of allergic rhino-conjunctivitis and asthma patients annually in Antalya, a Mediterranean coastal city in Turkey. MATERIAL AND METHODS: We evaluated patients' allergic clinical status, and recorded the climate and pollens in the city center air, investigating any correlation between pollination, climatic conditions and allergic disorders. The meteorological conditions and the pollen count/cm2 during every month of the year and the concordance of this with the patient's clinical status were evaluated. RESULTS: SPT positivity for plantago lanceolata, aspergillus fumigatus and d. pteronyssinus was significant in patients younger than 40 years old. Pollination levels are consistent from March 2010 to February 2011. In Antalya, high levels occur mostly from April to June, thus we performed skin prick tests mostly in May/June (~30%). During these months meteorological conditions of the city were windy with low humidity, without rain, and lukewarm temperatures, all of which contribute to high-risk conditions for seasonal allergies. CONCLUSIONS: The major allergen between April and June was derived from Graminea; between February and March was Cupressus spp; and between March and June was Pinus spp. These results suggest that the pollination is correlated with allergic conditions and thus SPT might be best performed according to the pollen count.

Further evidence for a role of tumor CD200 expression in breast cancer metastasis: decreased metastasis in CD200R1KO mice or using CD200-silenced EMT6.

Previous studies reported that CD200 expression on cells of the transplantable EMT6 mouse breast cancer line was increased during growth in immunocompetent mice. Low levels of expression persisted in NOD-SCID.IL-2(γr-/-) mice or mice with generalized over-expression of a CD200 transgene (CD200(tg) mice), despite the faster tumor growth in both of these latter strains. We also showed that CD200 expression (by the host and/or tumor cells) led to increased seeding of tumor cells to DLN in immunocompromised (CD200(tg) or NOD-SCID.IL-2(γr-/-)) vs immunocompetent mice, using limiting dilution cloning of tumor cells from DLN (vs contralateral lymph nodes, CLN). Evidence for an important role for CD200 expression in this increased metastasis came from the observation that neutralization of CD200 by anti-CD200mAbs decreased tumor metastasis and increased levels of cytotoxic anti-tumor immune cells in DLN. In the current studies, we have extended these observations by exploring tumor growth/metastasis in CD200R1 KO mice in which we have previously shown, in a transplant model, that expression of CD200 fails to deliver an immunosuppressive signal. In addition, we have studied local and metastatic growth in healthy control mice of EMT6 tumor cells stably transduced with shRNA able to silence CD200 expression. In both scenarios, decreased metastasis was observed, with increased immunity to EMT6 detected by cytotoxicity assays. In addition, adoptive transfer of DLN to control mice attenuated EMT6 metastases implying a potential therapeutic benefit from neutralizing CD200 expression in breast cancer.

Interaction of allergic disease and (immune) response to cancer in patient with stage-4 adenocarcinoma of the lung.

A 50 year old male patient who was watched because of penicillin and venom allergy in the allergy unit had been admitted to the medical oncology polyclinic with cough, expectoration and hemoptysis 10 years ago. His symptoms had first begun 1 month prior to that admission, and he was then diagnosed with stage IV adeno carcinoma. The first allergic reaction occurred at 1 hour following the last transfusion, while the second transfusion reaction occurred within the first 10 minutes of the transfusion and was associated with respiratory arrest. Also, he reported both penicillin and venom allergy history, and, subsequent to diagnosis and treatment, developed an anaphylactic reaction to a European Cornet sting on two occasions. Our aim in publishing this case is that after diagnosis of metastatic lung adeno cancer, our patient developed anaphylactic reactions associated with bee stings and on 2 occasions suffered transfusion associated anaphylactic reactions. His subsequent survival (9 years to date with stable disease) is extraordinary.

Erratum to "IL8 and serum soluble TRAIL levels following anti-VEGF monoclonal antibody treatment in patients with metastatic colon cancer" [Clin. Lab. 2012; 58:501-505].

BACKGROUND: Colorectal cancer is the third most common human cancer and the third leading cause of cancer related death. BevacizumAb is a humanized monoclonal antibody developed against vascular endothelial growth factor (VEGF) for the treatment of metastatic cancers. Our goal was to evaluate the possibility of using serum sTRAIL and IL8 as markers of treatment efficacy and prognosis in patients with metastatic colon cancer. METHODS: The study was conducted in Denizli between November 10, 2009 and September 20, 2010. 25 patients (6 female, 19 male) with metastatic colon cancer whose mean age was 58.7 years, were selected and included in the study. All patients received therapy with BevacizumAb. All patients were followed in the Oncology Clinic of Denizli State Hospital and were evaluated by clinical status. sTRAIL and IL8 levels were measured by ELISA in the sera of 25 BevacizumAb treated metastatic colon cancer patients and 20 healthy age-gender matched controls. Measurements were taken before and after treatment. RESULTS: The serum sTRAIL concentrations in patients before therapy were similar to those of healthy age-gender matched controls, namely 1.23 +/- 0.06 ng/mL and 1.21 +/- 0.04 ng/mL, respectively. After BevacizumAb treatment, sTRAIL ratios were increased significantly in 11 of 25 patients. 14 patients showed progressive disease with median overall survival of only 8.1 +/- 0.4 months. These 14 patients were the same ones who showed no increase in sTRAIL levels after BevacizumAb treatment. We explored evidence for a correlation between sTRAIL levels and overall survival rates and observed that elevated sTRAIL levels after the BevacizumAb treatment were significantly associated with increased median overall survival up to 22.6 months. Serum IL8 levels were decreased in all patients who received BevacizumAb therapy. CONCLUSIONS: In BevacizumAb therapy, serum IL8 levels were decreased in all patients, and thus, changes in such levels were not correlated with disease outcome. Our data suggest measurement of changes in sTRAIL following BevacizumAb treatment may have prognostic value in metastatic colon cancer patients.

CD40, CD45 CTLA-4 levels are elevated in healthy older adults.

BACKGROUND: The immune system changes with age. In this study we characterized immune changes by performing immunologic screening profiles on ageing individuals. METHODS: This study was performed at Akdeniz University, in the Faculty of Medicine, Department of Immunology. Healthy volunteers consisted of a younger group (22 donors) and an older group (45 individuals). All subjects had no serious health problems (i.e. chronic heart, lung, liver or immunological diseases) and were taking no prescribed medications. Flow cytometry analysis was used to evaluate CD3, CD4, CD8, CD16, CD19, CD28, CD40, CD45, CD56, CD80, CD86, CTLA-4 and ELISA for IL-1 beta, IL-2, IL-6, IL-10, IFN-gamma, TNF-alpha expression In addition, NK activity and induced cytokine expression (by bioassay and ELISA, respectively) were evaluated. RESULTS: No statistical differences were observed between the two groups in expression of CD3, CD8, CD19, CD80, CD86, CD16, CD 56, or CD28. A higher frequency of expression of CD4, CTLA-4, CD40, and CD45 was seen in older subjects by comparison with younger subjects. Cytokine profiles expressed by stimulated monocytes and lymphocytes from the two groups showed no difference in IL-1 beta, IL-2, IL-6, IL-10, TNF-alpha, and IFN-gamma production levels. CONCLUSIONS: We found increased expression levels of CD40 and CD45 levels in healthy older (age: 59.42 +/- 5.89) versus younger individuals (age: 30.32 +/- 2.29). CTLA-4 expression levels were also higher in older subjects, with no difference in CD28 expression levels between younger/older individuals.

IL8 and serum soluble TRAIL levels following anti-VEGF monoclonal antibody treatment in patients with metastatic colon cancer.

BACKGROUND: Colorectal cancer is the third most common human cancer and the third leading cause of cancer related death. BevacizumAb is a humanized monoclonal antibody developed against vascular endothelial growth factor (VEGF) for the treatment of metastatic cancers. Our goal was to evaluate the possibility of using serum sTRAIL and IL8 as markers of treatment efficacy and prognosis in patients with metastatic colon cancer. METHODS: The study was conducted in Denizli between November 10, 2009 and September 20, 2010. 25 patients (6 female, 19 male) with metastatic colon cancer whose mean age was 58.7 years, were selected and included in the study. All patients received therapy with BevacizumAb. All patients were followed in the Oncology Clinic of Denizli State Hospital and were evaluated by clinical status, sTRAIL and IL8 levels were measured by ELISA in the sera of 25 BevacizumAb treated metastatic colon cancer patients and 20 healthy age-gender matched controls. Measurements were taken before and after treatment. RESULTS: The serum sTRAIL concentrations in patients before therapy were similar to those of healthy age-gender matched controls, namely 1.23 +/- 0.06 ng/mL and 1.21 +/- 0.04 ng/mL, respectively. After BevacizumAb treatment, sTRAIL ratios were increased significantly in 11 of 25 patients. 14 patients showed progressive disease with median overall survival of only 8.1 +/- 0.4 months. These 14 patients were the same ones who showed no increase in sTRAIL levels after BevacizumAb treatment. We explored evidence for a correlation between sTRAIL levels and overall survival rates and observed that elevated sTRAIL levels after the BevacizumAb treatment were significantly associated with increased median overall survival up to 22.6 months. Serum IL8 levels were decreased in all patients who received BevacizumAb therapy. CONCLUSIONS: In BevacizumAb therapy, serum IL8 levels were decreased in all patients, and thus, changes in such levels were not correlated with disease outcome. Our data suggest measurement of changes in sTRAIL following BevacizumAb treatment may have prognostic value in metastatic colon cancer patients.

Total antioxidant capacity, hydrogen peroxide, malondialdehyde and total nitric oxide concentrations in patients with severe persistent allergic asthma: its relation to omalizumab treatment.

BACKGROUND: Asthma is a chronic inflammatory disease and omalizumab is indicated for moderate-to-severe persistent asthma. The results of many studies have shown that oxidative stress is involved in asthma pathogenesis. However, there is no data available to evaluate the alterations in total antioxidant capacity, hydrogen peroxide, malondialdehyde, and total nitric oxide concentrations. OBJECTIVES: The objective was to determine whether treatment with omalizumab in severe allergic asthma influences systemic levels of oxidative stress markers. METHODS: The first group of 14 patients included 6 male and 8 female subjects with severe persistent asthma, having a mean age of 42.4 years. The second group included 14 newly diagnosed allergic asthma patients with a mean age of 43.8 years. All patients were followed in the Immunology and Allergy Clinic of the Antalya Education and Training Hospital and were evaluated by clinical status. A third group of 14 age-sex matched healthy controls were also included. Serum samples were collected and stored at -70 degrees C until use for the determination of total antioxidant capacity, hydrogen peroxide, malondialdehyde, and total nitric oxide concentrations. Serum IgE levels, ANA (antinuclear antibody), RF (rheumatoid factor), hepatitis markers, C3, C4, and eosinophil levels were evaluated in all patients. All assays were carried out in duplicate. RESULTS: The mean IgE levels were as follow: Group I: 459.785 IU/mL; Group II: 124.8 IU/mL, and Group III: 39.88 IU/mL. Total antioxidant capacity levels of Group IB, group II, and group III were lower than the IA group. Total antioxidant capacity levels of groups II and III were higher than in group IB. Hydrogen peroxide concentrations in group IB were lower than in group IA, while concentrations in group II were higher than in group IB. The malondialdehyde concentration of group IB was lower than in all other groups. The malondialdehyde concentration of group III was higher than all other groups. The malondialdehyde concentration of group II was lower than in group III. The total nitric oxide level of group IB was lower than all other groups. The total nitric oxide level of group III was higher than all other groups, while that of group II was higher than for both groups IA/IB. CONCLUSIONS: To monitor the omalizumab treatment efficacy in severe allergic asthma patients, total antioxidant capacity, hydrogen peroxide, malondialdehyde, and total nitric oxide concentrations might be new markers.

Serum-soluble TRAIL levels in patients with severe persistent allergic asthma: its relation to omalizumab treatment.

BACKGROUND: In this study we compare the Omalizumab treatment modality in the dynamics of cell apoptosis regulating molecules in both severe persistent asthma patients who had no other any allergic disease, newly diagnosed patients with allergic asthma, and healthy volunteers. MATERIAL/METHODS: Severe persistent allergic asthma patients were subjected to measurement of serum soluble TRAIL (TNF-related apoptosis-inducing ligand) levels during the active disease phase and the stable phase which occurred 4 months after Omalizumab treatment. Serum sTRAIL concentrations were measured by a solid phase sandwich enzyme-linked immunosorbent assay. Concentration levels were compared with those of age- and sex-matched newly diagnosed patients with allergic asthma, and healthy controls. All assays were carried out in duplicate. Total serum IgE levels, antinuclear antibody (ANA), rheumatoid factor (RF), hepatitis markers, C3, C4 and eosinophil levels were evaluated in all patients. RESULTS: ANA, RF, hepatitis markers were negative in all patients. Complement 3 and 4 levels were normal in all patients. Prick tests in all patients were detected in mite and grass allergy. These results correlated with specific IgE. There were no differences between the healthy controls, newly diagnosed allergic asthma patients, and non-treated severe persistent allergic asthma patients during the active phase. Interestingly, the levels in variances of the patients who had the effective omalizumab treatment were significantly lower than the healthy controls, while the mean values were not statistically significant. CONCLUSIONS: Our study gives a different perspective on severe persistent allergic asthma and omalizumab treatment efficacy at the cell apoptosis-linked step by the serum sTRAIL levels.

Alternative splicing of CD200 is regulated by an exonic splicing enhancer and SF2/ASF.

CD200, a type I membrane glycoprotein, plays an important role in prevention of inflammatory disorders, graft rejection, autoimmune diseases and spontaneous fetal loss. It also regulates tumor immunity. A truncated CD200 (CD200(tr)) resulting from alternative splicing has been identified and characterized as a functional antagonist to full-length CD200. Thus, it is important to explore the mechanism(s) controlling alternative splicing of CD200. In this study, we identified an exonic splicing enhancer (ESE) located in exon 2, which is a putative binding site for a splicing regulatory protein SF2/ASF. Deletion or mutation of the ESE site decreased expression of the full-length CD200. Direct binding of SF2/ASF to the ESE site was confirmed by RNA electrophoretic mobility shift assay (EMSA). Knockdown of expression of SF2/ASF resulted in the same splicing pattern as seen after deletion or mutation of the ESE, whereas overexpression of SF2/ASF increased expression of the full-length CD200. In vivo studies showed that viral infection reversed the alternative splicing pattern of CD200 with increased expression of SF2/ASF and the full-length CD200. Taken together, our data suggest for the first time that SF2/ASF regulates the function of CD200 by controlling CD200 alternative splicing, through direct binding to an ESE located in exon 2 of CD200.