Diverse Gene Expression in Human Regulatory T Cell Subsets Uncovers Connection between Regulatory T Cell Genes and Suppressive Function.

Regulatory T (Treg) cells have a critical role in the control of immunity, and their diverse subpopulations may allow adaptation to different types of immune responses. In this study, we analyzed human Treg cell subpopulations in the peripheral blood by performing genome-wide expression profiling of 40 Treg cell subsets from healthy donors. We found that the human peripheral blood Treg cell population is comprised of five major genomic subgroups, represented by 16 tractable subsets with a particular cell surface phenotype. These subsets possess a range of suppressive function and cytokine secretion and can exert a genomic footprint on target effector T (Teff) cells. Correlation analysis of variability in gene expression in the subsets identified several cell surface molecules associated with Treg suppressive function, and pharmacological interrogation revealed a set of genes having causative effect. The five genomic subgroups of Treg cells imposed a preserved pattern of gene expression on Teff cells, with a varying degree of genes being suppressed or induced. Notably, there was a cluster of genes induced by Treg cells that bolstered an autoinhibitory effect in Teff cells, and this induction appears to be governed by a different set of genes than ones involved in counteracting Teff activation. Our work shows an example of exploiting the diversity within human Treg cell subpopulations to dissect Treg cell biology.

Targeting Th17 Effector Cytokines for the Treatment of Autoimmune Diseases.

Interleukin (IL)-17-producing T cells, especially T helper (Th)17 cells, play a critical role in the pathogenesis of a variety of autoimmune inflammatory diseases. The pathogenic function of Th17 cells results from their production of Th17 effector cytokines, namely IL-17 (or IL-17A), IL-17F, IL-22 and IL-26. The importance of IL-17 has been demonstrated by antibody neutralization studies in both animal models of autoimmune diseases as well as in human clinical trials. This review highlights the current knowledge of the clinical aspects of the Th17 cytokines as well as therapeutic antibodies against IL-17, IL-17F, IL-17 receptor, IL-22, IL-26 and granulocyte macrophage colony-stimulating factor for the future treatment of autoimmune inflammatory diseases.

Targeting Th17 cells in autoimmune diseases.

T helper 17 (Th17) cells have been implicated in the pathogenesis of most common autoimmune diseases, including psoriasis, rheumatoid arthritis (RA), inflammatory bowel disease (IBD), and multiple sclerosis (MS). Although anti-interleukin-17 (IL-17) antibodies show marked clinical efficacy in psoriasis, targeting IL-17 alone is not sufficient to improve clinical end points in other autoimmune conditions, namely RA and Crohn's disease. Given that Th17 cells express IL-17 together with many other proinflammatory cytokines [IL-17F, IL-22, IL-26, and granulocyte-macrophage colony-stimulating factor (GM-CSF)], targeting the Th17 cell lineage may be superior to blocking a single effector cytokine. Here, we discuss the rationale for targeting two checkpoints in the development and inflammatory function of Th17 cells, retinoid-related orphan receptor-γt (RORγt) and IL-23, and we review recent progress in the development of both RORγt small molecule inhibitors and IL-23 neutralizing antibodies.

Internal hernia through a peritoneal defect in the pouch of Douglas: Report of a case.

INTRODUCTION: Internal hernia is a rare entity which can cause intestinal obstruction. The most common type of internal hernia is the paraduodenal hernia which accounts for 53% of cases, and the internal hernia within the pelvis account for 7%. Perineal hernia, which is classified as pelvic hernia, usually occurs due to weakening of the pelvic floor musculature and thus, should be distinguished from the internal hernia caused by peritoneal defects in the pelvic cavity. PRESENTATION OF CASE: We present a case of 28-year-old female who presented intestinal obstruction. Conservative therapies failed and she required emergency laparotomy. The operative findings revealed a peritoneal defect of 2cm in diameter in the pouch of Douglas, through which the ileum was incarcerated and strangulated. The incarcerated bowel was reduced, and the intestinal color quickly returned to normal. Therefore a primary closure of the peritoneal defect was performed and the postoperative course was uneventful. DISCUSSION: A PubMed search for the case of internal hernia through a defect in the pouch of Douglas revealed only three, making this an extremely rare condition. CONCLUSION: Because of rarity of this hernia, the etiology is unknown. However, our patient is a young female with no history of pregnancy, abdominal surgery, or trauma, therefore the cause of the peritoneal defect is considered congenital.

Expression patterns of microRNAs are altered in hypoxic human neuroblastoma cells.

BACKGROUND/PURPOSE: There was a report that microRNA (miRNA) controls multiple genes. In addition, there are some reports that the presence of neoplastic cells that are hypoxic because of rapid tumor development is related to prognosis. As a step toward identifying the role of miRNA in hypoxic tumor cells, the present study was designed to determine which miRNAs have increased expression and which have decreased expression in hypoxic neuroblastoma cells. METHODS: For this study, we used seven neuroblastoma cell lines. In four with MYCN was amplified; in the other three MYCN was non-amplified. Neuroblastoma cells were cultured under hypoxic conditions. The expression levels of 662 kinds of miRNA in the hypoxic cells were quantified by gene array. RESULTS: We found that the expression of 85 kinds of miRNA was increased. Expression of six of these mRNAs was increased in two or more cell lines. Hsa-miR-143, -145, and -210 were each expressed in four of the seven cell lines. In addition, expression of 48 kinds of miRNA was decreased. Expression of five was decreased in two cell lines. There was no relation between the expression of miRNA and the amplification of MYCN. CONCLUSION: Our results thus suggest a possible causal relation between these three miRNAs and the malignancy of neuroblastoma in hypoxic conditions.

Gain-of-function of mutated C-CBL tumour suppressor in myeloid neoplasms.

Acquired uniparental disomy (aUPD) is a common feature of cancer genomes, leading to loss of heterozygosity. aUPD is associated not only with loss-of-function mutations of tumour suppressor genes, but also with gain-of-function mutations of proto-oncogenes. Here we show unique gain-of-function mutations of the C-CBL (also known as CBL) tumour suppressor that are tightly associated with aUPD of the 11q arm in myeloid neoplasms showing myeloproliferative features. The C-CBL proto-oncogene, a cellular homologue of v-Cbl, encodes an E3 ubiquitin ligase and negatively regulates signal transduction of tyrosine kinases. Homozygous C-CBL mutations were found in most 11q-aUPD-positive myeloid malignancies. Although the C-CBL mutations were oncogenic in NIH3T3 cells, c-Cbl was shown to functionally and genetically act as a tumour suppressor. C-CBL mutants did not have E3 ubiquitin ligase activity, but inhibited that of wild-type C-CBL and CBL-B (also known as CBLB), leading to prolonged activation of tyrosine kinases after cytokine stimulation. c-Cbl(-/-) haematopoietic stem/progenitor cells (HSPCs) showed enhanced sensitivity to a variety of cytokines compared to c-Cbl(+/+) HSPCs, and transduction of C-CBL mutants into c-Cbl(-/-) HSPCs further augmented their sensitivities to a broader spectrum of cytokines, including stem-cell factor (SCF, also known as KITLG), thrombopoietin (TPO, also known as THPO), IL3 and FLT3 ligand (FLT3LG), indicating the presence of a gain-of-function that could not be attributed to a simple loss-of-function. The gain-of-function effects of C-CBL mutants on cytokine sensitivity of HSPCs largely disappeared in a c-Cbl(+/+) background or by co-transduction of wild-type C-CBL, which suggests the pathogenic importance of loss of wild-type C-CBL alleles found in most cases of C-CBL-mutated myeloid neoplasms. Our findings provide a new insight into a role of gain-of-function mutations of a tumour suppressor associated with aUPD in the pathogenesis of some myeloid cancer subsets.

[Mediastinal large B-cell lymphoma associated with systemic sclerosis].

Malignant lymphoma (ML) is frequently associated with several forms of collagen diseases such as Sjören syndrome, systemic lupus erythematodes, and rheumatoid arthritis. However, the occurrence of ML in systemic sclerosis (SSc) patients has rarely been reported. Here we report an SSc patient who developed mediastinal (thymic) large B-cell lymphoma (MLBCL). A 31-year-old woman was diagnosed as having SSc in August 2007. The patient was treated with low-dose prednisolone (10 mg/day) without any effect. One year after the diagnosis, chest computed tomography-scan demonstrated thymic tumor in the anterior mediastinum. Thymectomy was performed, and a pathohistological diagnosis of MLBCL was established. Immunohistochemical analysis demonstrated that the tumor cells were positive for CD45 and CD20, but negative for CD30 and EBV-encoded RNA. The patient was treated with 6 courses of CHOP regimen, resulting in complete remission of lymphoma. This report describes the first SSc patient associated with MLBCL. SSc patients occasionally develop ML after a relatively short interval. Our case suggests that intensive monitoring for the development of ML is needed in newly diagnosed SSc patients.

Arterial infusion chemotherapy in patient with repeated recurrent tumor of cecal cancer: report of a case.

PURPOSE: We report a patient with a repeated recurrent tumor after Right-hemicolectomy for advanced cecal cancer who was treated by intra-arterial infusions of 5-fluorouracil (5-FU). METHODS: A computed tomography scan revealed a pelvic mass involving the psoas major muscle and quadratos lumborum muscle, in contact with the widely projecting toward L2-S2. The fluorodeoxyglucose-positron emission tomography (FDG-PET) revealed an accumulation spot in the same place. This case was deemed in operable, and one-shot bolus of 5-FU was administered through the tumor feeding arteries: the left 3rd, 4th lumbar, and ilio -- lumbar arteries at a dosage of 250 mg/body from each artery. RESULTS: A partial regression of the tumor was observed by computed tomography. The serum level of carbohydrate antigen 19-9 returned normal in 8 months. During chemotherapy, the side effect and complications were tolerable, and she experienced only grade-1 nausea caused by 5-fluorouracil. CONCLUSION: A long-time, intra-arterial 5-fluorouracil infusion could control effectively and safely.

Role of the RUNX1-EVI1 fusion gene in leukemogenesis.

RUNX1-EVI1 is a chimeric gene generated by t(3;21)(q26;q22) observed in patients with aggressive transformation of myelodysplastic syndrome or chronic myelogenous leukemia. RUNX1-EVI1 has oncogenic potentials through dominant-negative effect over wild-type RUNX1, inhibition of Jun kinase (JNK) pathway, stimulation of cell growth via AP-1, suppression of TGF-beta-mediated growth inhibition and repression of C/EBPalpha. Runx1-EVI1 heterozygous knock-in mice die in uteri due to central nervous system (CNS) hemorrhage and severe defects in definitive hematopoiesis as Runx1-/- mice do, indicating that RUNX1-EVI1 dominantly suppresses functions of wild-type RUNX1 in vivo. Acute myelogenous leukemia is induced in mice transplanted with bone marrow cells expressing RUNX1-EVI1, and a Runx1-EVI1 knock-in chimera mouse developed acute megakaryoblastic leukemia. These results suggest that RUNX1-EVI1 plays indispensable roles in leukemogenesis of t(3;21)-positive leukemia. Major leukemogenic effect of RUNX1-EVI1 is mainly through histone deacetyltransferase recruitment via C-terminal binding protein. Histone deacetyltransferase could be a target in molecular therapy of RUNX1-EVI1-expressing leukemia.

Poorly differentiated adenocarcinoma of the colon and rectum: clinical characteristics.

BACKGROUND/AIMS: The aims of this study were to assess the prognosis and histopathological factors of poorly differentiated colorectal adenocarcinoma, and the clinical relevance of the proposed histopathological sub classifications. METHODOLOGY: Fifty eight patients with poorly differentiated adenocarcinoma were enrolled in this study. According to the lymphatic canal spread in tumor tissue, they were classified into lymphangitic type (tumor spread beyond the intra mucosal tumor space through the lymphatic canal widely) and non-lymphangitic type (tumor spread within that space). Next, they were sub classified into medullary, intermediate and scirrhous types according to the amount of fibrous stroma. In addition, immunohistological examinations were performed on the expression of an intercellular adhesion molecule (E-cadherin). RESULTS: In 33 cases (57%) Lymphangitic type was present and in 25 cases non-lymphangitic type (43%) was present. In the lymphangitic types, 5 cases were medullary type (15%), 17 cases were intermediate type (52%) and 11 cases were scirrhous type (33%) that included 2 cases of signet ring cell carcinoma. In the non-lymphangitic types, medullary type was dominant (20 cases, 80%) while intermediate type and scirrhous type were 3 cases (12%) and 2 cases (8%), respectively. The survival rates were calculated for both types and a large difference was found in terms of 5-year survival rate; 0% for lymphangitic type and 72% for non-lymphangitic type (p<0.05). There was no correlation found between the expression of cadherin and the subclassification. CONCLUSIONS: In conclusion, a wide tumor infiltration and growth in lymphatic vessels appears to be an important prognostic factor for poorly differentiated adenocarcinoma compared to the metastasis patterns.

Histone deacetylase inhibitors trichostatin A and valproic acid circumvent apoptosis in human leukemic cells expressing the RUNX1 chimera.

Disturbance of the normal functions of wild-type RUNX1 resulting from chromosomal translocations or gene mutations is one of the major molecular mechanisms in human leukemogenesis. RUNX1-related chimeras generated by the chromosomal translocations repress transcriptional activity of wild-type RUNX1 by recruiting the co-repressor/histone deacetylase complex. Thus, histone deacetylase inhibitors are expected to restore normal functions of wild-type RUNX1 and thereby affect the growth and differentiation ability of leukemic cells expressing the chimera. We investigated the in vitro effects of histone deacetylase inhibitors, trichostatin A and valproic acid, on human leukemic cell lines such as SKNO-1 and Kasumi-1 expressing RUNX1/ETO, Reh expressing TEL/RUNX1 and SKH-1 co-expressing RUNX1/EVI1 and BCR/ABL. We also employed K562 cells expressing BCR/ABL without such a chimera as a control. Treatment with each inhibitor increased acetylated histone 4 in all of these cell lines. Interestingly, proliferation of SKNO-1, Kasumi-1, SKH-1 and Reh cells was significantly suppressed after 3-day culture with trichostatin A or valproic acid, when compared to that of K562 cells. We observed cell cycle arrest and apoptotic induction in the RUNX1 chimera-expressing cells by the propidium iodide staining. Up- and downregulation of cell cycle regulator genes appeared to be the molecular basis for the former, and activation of both extrinsic and intrinsic apoptotic caspases for the latter. We propose histone deacetylase inhibitors to be an attractive choice in the molecular targeting therapy of RUNX1-related leukemia.

Molecular profiling reveals distinct functional attributes of CD1d-restricted natural killer (NK) T cell subsets.

CD1d-restricted natural killer T (NKT) cells can have multiple effects on an immune response, including the activation, regulation and attraction of innate immune cells, and modulation of adaptive immunity. Recent studies reveal that there are distinct subsets of NKT cells which selectively perform some of the functions attributed to CD1d-restricted cells, but the mechanisms underlying these functional differences have not been resolved. Our aim in this study was to identify novel NKT cell associated traits that would provide important insight into NKT cell activation and function. To this end, we have performed gene expression profiling of two separate subsets of NKT cells, analyzing genes differentially expressed in these cells compared to conventional CD4(+)NK1.1(-) T cells. We identify different sets of genes over expressed in each of the two NKT cell types, as well as genes that are common to the two CD1d-restricted NKT cell populations analyzed. A large number of these genes are highly relevant for NKT cell development, activation and function. Each NKT subtype displayed a unique set of chemokine receptors, integrins and molecules related to effector function, supporting the notion that distinct NKT cells can be selectively engaged and have diverse functions in different types of immune reactions.

The same genomic region conditions clonal deletion and clonal deviation to the CD8alphaalpha and regulatory T cell lineages in NOD versus C57BL/6 mice.

Clonal deviation is a mechanism by which immature thymocytes expressing a self-reactive T cell antigen receptor (TCR) are rescued from clonal deletion by adopting an alternative differentiation pathway resistant to apoptosis. Here, we confirm and generalize previous indications that genetic alleles in NOD mice condition ineffective clonal deviation toward the CD8alphaalpha lineage, a peculiar population of TCRalphabeta lymphocytes that electively colonizes the intraepithelial lymphocyte pool in the gut. Thymic selection of CD8alphaalpha cells was very age-dependent, occurring almost exclusively in the postnatal period. Fewer CD8alphaalpha cells were found in the thymus and intraepithelial lymphocytes of BDC2.5 TCR transgenic mice on the NOD than on the C57BL/6 (B6) background; this paucity extended to standard NOD mice, albeit to a lesser extent. CD8alphaalpha cells resided in the BDC2.5 pancreatic infiltrate, and they were more abundant on the B6 than the NOD background, correlating with aggressivity of the lesion. A (B6(g7) x NOD)F(2) intercross in agonist-challenged BDC2.5 fetal thymic organ cultures demonstrated the existence of a major quantitative trait locus on chromosome 3, coincident with an interval associated with resistance to clonal deletion. A replicate linkage confirmed these positions and showed that the same region also controls clonal deviation toward the CD4(+)FoxP3(+) regulatory T cell lineage. That clonal deviation toward the CD8alphaalpha and regulatory T cell pathways share genetic control further highlights the similarities between these two "rescue lineages," consistent with an immunoregulatory role for CD8alphaalpha cells.

[Multiple hepatic metastases of colon cancer responding to combined therapy with TS-1 and CPT-11--a case report].

We treated a patient with multiple liver metastases arising from colon cancer in whom the metastatic tumors were responsive to treatment with the combination of TS-1 and CPT-11. The patient was a 71-year-old woman with cancer of the ascending colon and metastatic hepatic tumors. She had undergone surgery on July 28, 2004, and abdominal contrast CT scans obtained after discharge from hospital revealed numerous LDA (low-density areas) in both lobes of the liver. The patient was given ambulatory chemotherapy with TS-1 (120 mg/day on days 1-14) and CPT-11 (100 mg/day on days 1 and 8). After completion of 2 courses of chemotherapy, abdominal contrast CT scans revealed that most of the LDAs in both lobes of the liver had disappeared, and the patient was judged to have achieved PR. No adverse reactions were observed except for a slight decrease of WBC, and her chemotherapy is being continued at present. This case suggests that the combination of TS-1 and CPT-11 may be an effective form of chemotherapy for the treatment of colon cancer with multiple hepatic metastases.

TEL/ETV6 induces apoptosis in 32D cells through p53-dependent pathways.

TEL is an ETS family transcription factor that is critical for maintaining hematopoietic stem cells in adult bone marrow. To investigate the roles of TEL in myeloid proliferation and differentiation, we introduced TEL cDNA into mouse myeloid 32Dcl3 cells. Overexpression of TEL repressed interleukin-3-dependent proliferation through blocking cell cycle progression. Also, the presence of TEL triggered apoptosis through the mitochondrial intrinsic pathway on exposure to granulocyte colony-stimulating factor. We found an increase in p53 protein and its DNA binding in the TEL-overexpressing cells. Forced expression of TEL stimulated transcription via the p53-responsive element and increased the expression of cellular target genes for p53 such as cell cycle regulator p21 and apoptosis inducer Puma. Consistently, induction of apoptosis was delayed by pifithrin-alpha treatment and completely blocked by increased expression of Bcl-2 in the TEL-overexpressing cells. These data collectively suggest that TEL exerts a tumor suppressive function through augmenting the p53 pathway and facilitates normal development of myelopoiesis.