[Retracted] miR­218 suppresses tumor growth and enhances the chemosensitivity of esophageal squamous cell carcinoma to cisplatin.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the western blotting data shown in Fig. 6 and the tumor images shown in Fig. 7A were strikingly similar to data appearing in different form in other articles by different authors. Owing to the fact that the contentious data in the above article had already been published elsewhere, or were already under consideration for publication, prior to its submission to Oncology Reports, the Editor has decided that this paper should be retracted from the Journal. After having been in contact with the authors, they agreed with the decision to retract the paper. The Editor apologizes to the readership for any inconvenience caused. [the original article was published in Oncology Reports 33: 981­989, 2015; DOI: 10.3892/or.2014.3657].

Dexmedetomidine upregulates microRNA-185 to suppress ovarian cancer growth via inhibiting the SOX9/Wnt/ß-catenin signaling pathway.

Dexmedetomidine (DEX) could serve as an adjuvant analgesic during cancer therapies. Abnormal expression of microRNAs (miRNAs) could lead to cancer development. This study was aimed to explore the roles of DEX in ovarian cancer (OC) development. OC cell lines SKOV3 and HO-8910 were treated with DEX, after which OC development and the miR-185, SOX9, and Wnt/ß-catenin pathway were measured. DEX-treated HO-8910 cells were transfected with miR-185 mimic, miR-185 antisense or miR-185 antisense + silenced SOX9 to further measure the OC cell growth. The target relation between miR-185 and SOX9 was identified, and SOX9 and Wnt/ß-catenin pathway were protein levels detected after miR-185 transfection. The role of miR-185 in OC in vivo was also measured. Our study found DEX had a dose-dependent inhibition on OC growth, and DEX promoted miR-185 but suppressed SOX9 expression in OC cells. miR-185 targeted SOX9. After interfering with miR-185 expression, HO-8910 cell proliferation, invasion, migration, and apoptosis were affected. SOX9 knockdown repressed OC development and Wnt/ß-catenin pathway. The volume, weight, positive rate of Ki67, CyclinD1, p53 and the degree of tumor necrosis were affected by miR-185 expression. This study demonstrated that DEX could inhibit OC development via upregulating miR-185 expression and inactivating the SOX9/Wnt/ß-catenin signaling pathway.

Effect of Dexmedetomidine-Mediated Insulin-Like Growth Factor 2 (IGF2) Signal Pathway on Immune Function and Invasion and Migration of Cancer Cells in Rats with Ovarian Cancer.

BACKGROUND The aim of this study was to explore the effect of dexmedetomidine (DEX)-mediated insulin-like growth factor 2 (IGF2) signal pathway on immune function and cancer cell invasion and migration in rats with ovarian cancer. MATERIAL AND METHODS Forty rats with ovarian cancer were divided into 4 groups: model group, and low dose (0.2 µg/kg/hour DEX), medium dose (1.0 µg/kg/hour DEX), and high dose (5.0 µg/kg/hour DEX) DEX groups. In addition, 10 Fischer344 rats were selected as a normal group. Human NUTU-19 poorly differentiated epithelial ovarian cancer cell line cells were divided into 4 groups: a blank group and low dose, medium dose, and high dose DEX NUTU-19 groups. RESULTS Compared with the normal group, in the other groups the serum interleukin (IL)-2 and interferon gamma (INF-γ) levels, CD4⁺ and CD8⁺ percentages, CD4⁺/CD8⁺ ratio, and transformation rate of splenic lymphocytes were decreased, and the serum tumor necrosis factor alpha (TNF-alpha) level, IGF2, insulin-like growth factor 1 receptor (IGF1R), insulin receptor substrate 1 (IRS1) mRNA, and protein expressions in ovarian tissue were increased (all P<0.05). Results in the DEX groups compared with model group were the opposite of those in the other groups compared with normal group (all P<0.05). Compared with the blank group, in the other groups the proliferation, invasion, and migration of ovarian cancer cells were reduced significantly (all P<0.05). Compared with the low dose DEX NUTU-19 group, in the high dose DEX NUTU-19 group the invasion and migration of ovarian cancer cells weakened significantly (both P<0.05). CONCLUSIONS A certain dose of DEX can effectively inhibit IGF2 signal pathway activation to improve the immune function of rats with ovarian cancer, inhibiting the invasion and migration of ovarian cancer cells.

Long-acting MIC-1/GDF15 molecules to treat obesity: Evidence from mice to monkeys.

In search of metabolically regulated secreted proteins, we conducted a microarray study comparing gene expression in major metabolic tissues of fed and fasted ob/ob mice and C57BL/6 mice. The array used in this study included probes for ~4000 genes annotated as potential secreted proteins. Circulating macrophage inhibitory cytokine 1 (MIC-1)/growth differentiation factor 15 (GDF15) concentrations were increased in obese mice, rats, and humans in comparison to age-matched lean controls. Adeno-associated virus-mediated overexpression of GDF15 and recombinant GDF15 treatments reduced food intake and body weight and improved metabolic profiles in various metabolic disease models in mice, rats, and obese cynomolgus monkeys. Analysis of the GDF15 crystal structure suggested that the protein is not suitable for conventional Fc fusion at the carboxyl terminus of the protein. Thus, we used a structure-guided approach to design and successfully generate several Fc fusion molecules with extended half-life and potent efficacy. Furthermore, we discovered that GDF15 delayed gastric emptying, changed food preference, and activated area postrema neurons, confirming a role for GDF15 in the gut-brain axis responsible for the regulation of body energy intake. Our work provides evidence that GDF15 Fc fusion proteins could be potential therapeutic agents for the treatment of obesity and related comorbidities.

miR-132 targeting E2F5 suppresses cell proliferation, invasion, migration in ovarian cancer cells.

Accumulating evidence showed that microRNA-132 (miR-132) are involved in development and progression of several types of cancers, however, the function and underlying molecular mechanism of miR-132 in ovarian cancer remains unclear. In this study we investigated the biological roles and molecular mechanism of miR-132 in ovarian cancer. Here, we found that that the expression levels of miR-132 were dramatically decreased in ovarian cancer cell lines and clinical ovarian cancer tissue samples. Then, we found that introduction of miR-132 significantly suppressed the proliferation, colony formation, migration and invasion of ovarian cancer cells. Mechanism investigation revealed that miR-132 inhibited the expression of transcription factor E2F5 by specifically targeting its mRNA 3'UTR. Moreover, the expression level of E2F5 was significantly increased in ovarian cancer tissues than in the adjacent normal tissues, and its expression was inversely correlated with miR-132 expression in clinical ovarian cancer tissues. Additionally, silencing E2F5 was able to inhibit the proliferation, colony formation, migration and invasion of ovarian cancer cells, parallel to the effect of miR-132 overexpression on the ovarian cancer cells. Meanwhile, overexpression of E2F5 reversed the inhibition effect mediated by miR-132 overexpression. These results indicate that miR-132 suppresses the cell proliferation, invasion, migration in ovarian cancer cells by targeting E2F5.

Trefoil Factor 3 (TFF3) Is Regulated by Food Intake, Improves Glucose Tolerance and Induces Mucinous Metaplasia.

Trefoil factor 3 (TFF3), also called intestinal trefoil factor or Itf, is a 59 amino acid peptide found as a homodimer predominantly along the gastrointestinal tract and in serum. TFF3 expression is elevated during gastrointestinal adenoma progression and has been shown to promote mucosal wound healing. Here we show that in contrast to other trefoil factor family members, TFF1 and TFF2, TFF3 is highly expressed in mouse duodenum, jejunum and ileum and that its expression is regulated by food intake. Overexpression of TFF3 using a recombinant adeno-associated virus (AAV) vector, or daily administration of recombinant TFF3 protein in vivo improved glucose tolerance in a diet-induced obesity mouse model. Body weight, fasting insulin, triglyceride, cholesterol and leptin levels were not affected by TFF3 treatment. Induction of mucinous metaplasia was observed in mice with AAV-mediated TFF3 overexpression, however, no such adverse histological effect was seen after the administration of recombinant TFF3 protein. Altogether these results suggest that the therapeutic potential of targeting TFF3 to treat T2D may be limited.

miR-218 suppresses tumor growth and enhances the chemosensitivity of esophageal squamous cell carcinoma to cisplatin.

A growing body of evidence suggests that microRNA-218 (miR-218) acts as a tumor suppressor and is involved in tumor progression, development and metastasis and confers sensitivity to certain chemotherapeutic drugs in several types of cancer. However, our knowledge concerning the exact roles played by miR-218 in esophageal squamous cell carcinoma (ESCC) and the underlying molecular mechanisms remain relatively unclear. Thus, the aims of this study were to detect the expression of miR-218 in human ESCC tissues and explore its effects on the biological features and chemosensitivity to cisplatin (CDDP) in an ESCC cell line (Eca109), so as to provide new insights for ESCC treatment. Here, we found increased expression of miR-218 in the ESCC tissues compared with that in the matched non-tumor tissues, and its expression level was correlated with key pathological characteristics including clinical stage, tumor depth and metastasis. We also found that enforced expression of miR-218 significantly decreased cell proliferation, colony formation, migration and invasion, induced cell apoptosis and arrested the cell cycle in the G0/G1 phase, as well as suppressed tumor growth in a nude mouse model. In addition, our results showed that miR-218 mimics increased the sensitivity to the antitumor effect of CDDP in the human Eca109 cells. Importantly, this study also showed that miR-218 regulated the expression of phosphorylated PI3K, AKT and mTOR, which may contribute to suppressed tumor growth of ESCC and enhanced sensitivity of ESCC cells. These findings suggest that miR-218 is a potential therapeutic agent for the treatment of ESCC.

A ThPOK-LRF transcriptional node maintains the integrity and effector potential of post-thymic CD4+ T cells.

The transcription factor ThPOK promotes CD4(+) T cell differentiation in the thymus. Here, using a mouse strain that allows post-thymic gene deletion, we show that ThPOK maintains CD4(+) T lineage integrity and couples effector differentiation to environmental cues after antigenic stimulation. ThPOK preserved the integrity and amplitude of effector responses and was required for proper differentiation of types 1 and 2 helper T cells in vivo by restraining the expression and function of Runx3, a nuclear factor crucial for cytotoxic T cell differentiation. The transcription factor LRF acts redundantly with ThPOK to prevent the transdifferentiation of mature CD4(+) T cells into CD8(+) T cells. As such, the ThPOK-LRF transcriptional module was essential for CD4(+) T cell integrity and responses.

Thpok-independent repression of Runx3 by Gata3 during CD4+ T-cell differentiation in the thymus.

CD4(+) helper T cells are essential for immune responses and differentiate in the thymus from CD4(+) CD8(+) "double-positive" (DP) thymocytes. The transcription factor Runx3 inhibits CD4(+) T-cell differentiation by repressing Cd4 gene expression; accordingly, Runx3 is not expressed in DP thymocytes or developing CD4(+) T cells. The transcription factor Thpok is upregulated in CD4-differentiating thymocytes and required to repress Runx3. However, how Runx3 is controlled at early stages of CD4(+) T-cell differentiation, before the onset of Thpok expression, remains unknown. Here we show that Gata3, a transcription factor preferentially and transiently upregulated by CD4(+) T-cell precursors, represses Runx3 and binds the Runx3 locus in vivo. Accordingly, we show that high-level Gata3 expression and expression of Runx3 are mutually exclusive. Furthermore, whereas Runx3 represses Cd4, we show that Gata3 promotes Cd4 expression in Thpok-deficient thymocytes. Thus, in addition to its previously documented role in promoting CD4-lineage gene-expression, Gata3 represses CD8-lineage gene expression. These findings identify Gata3 as a critical pivot of CD4-CD8 lineage differentiation.

An inhibitory antibody against dipeptidyl peptidase IV improves glucose tolerance in vivo.

Dipeptidyl peptidase IV (DPP-IV) degrades the incretin hormone glucagon-like peptide 1 (GLP-1). Small molecule DPP-IV inhibitors have been used as treatments for type 2 diabetes to improve glucose tolerance. However, each of the marketed small molecule drugs has its own limitation in terms of efficacy and side effects. To search for an alternative strategy of inhibiting DPP-IV activity, we generated a panel of tight binding inhibitory mouse monoclonal antibodies (mAbs) against rat DPP-IV. When tested in vitro, these mAbs partially inhibited the GLP-1 cleavage activity of purified enzyme and rat plasma. To understand the partial inhibition, we solved the co-crystal structure of one of the mAb Fabs (Ab1) in complex with rat DPP-IV. Although Ab1 does not bind at the active site, it partially blocks the side opening, which prevents the large substrates such as GLP-1 from accessing the active site, but not small molecules such as sitagliptin. When Ab1 was tested in vivo, it reduced plasma glucose and increased plasma GLP-1 concentration during an oral glucose tolerance test in rats. Together, we demonstrated the feasibility of using mAbs to inhibit DPP-IV activity and to improve glucose tolerance in a diabetic rat model.

Control of the development of CD8αα+ intestinal intraepithelial lymphocytes by TGF-ß.

The molecular mechanisms that direct the development of TCRαß+CD8αα+ intestinal intraepithelial lymphocytes (IELs) are not thoroughly understood. Here we show that transforming growth factor-ß (TGF-ß) controls the development of TCRαß+CD8αα+ IELs. Mice with either a null mutation in the gene encoding TGF-ß1 or T cell-specific deletion of TGF-ß receptor I lacked TCRαß+CD8αα+ IELs, whereas mice with transgenic overexpression of TGF-ß1 had a larger population of TCRαß+CD8αα+ IELs. We observed defective development of the TCRαß+CD8αα+ IEL thymic precursors (CD4⁻CD8⁻TCRαß+CD5+) in the absence of TGF-ß. In addition, we found that TGF-ß signaling induced CD8α expression in TCRαß+CD8αα+ IEL thymic precursors and induced and maintained CD8α expression in peripheral populations of T cells. Our data demonstrate a previously unrecognized role for TGF-ß in the development of TCRαß+CD8αα+ IELs and the expression of CD8α in T cells.

Generation of novel long-acting globular adiponectin molecules.

Adiponectin is an adipocyte-derived hormone that has been shown to play important roles in the regulation of glucose and energy homeostasis. It exists as homotrimers or complexes containing multiple homotrimer units in plasma. The recombinant adiponectin proteins have been difficult to produce, making it challenging for both research as well as potential therapeutic development. Here, we show a novel approach for the generation of globular adiponectin that involves linking three monomer sequences together in tandem to generate one continuous polypeptide, which we have termed single-chain globular adiponectin (sc-gAd). To improve the pharmacokinetic properties of sc-gAd further, we fused it to an Fc fragment. The combined effects of single-chain and Fc fusion improved the plasma half-life from less than 2 h to close to 2 weeks. Using adeno-associated virus as a delivery method, we demonstrate that Fc-sc-gAd improved both fasting glucose levels and the tolerance to a glucose challenge in ob/ob mice without changes in body weight. Therefore, our results demonstrated the feasibility of generating globular adiponectin trimers from a single polypeptide and a long-acting globular adiponectin that could serve as a starting point for adiponectin-based therapeutics. This novel approach could also be applied to other complement factor C1q family members; in particular, this opens the possibility to study the biological functions of precisely defined heterotrimers of various family members that had not been previously possible.

The transcription factor Ets1 is important for CD4 repression and Runx3 up-regulation during CD8 T cell differentiation in the thymus.

The transcription factor Ets1 contributes to the differentiation of CD8 lineage cells in the thymus, but how it does so is not understood. In this study, we demonstrate that Ets1 is required for the proper termination of CD4 expression during the differentiation of major histocompatability class 1 (MHC I)-restricted thymocytes, but not for other events associated with their positive selection, including the initiation of cytotoxic gene expression, corticomedullary migration, or thymus exit. We further show that Ets1 promotes expression of Runx3, a transcription factor important for CD8 T cell differentiation and the cessation of Cd4 gene expression. Enforced Runx3 expression in Ets1-deficient MHC I-restricted thymocytes largely rescued their impaired Cd4 silencing, indicating that Ets1 is not required for Runx3 function. Finally, we document that Ets1 binds at least two evolutionarily conserved regions within the Runx3 gene in vivo, supporting the possibility that Ets1 directly contributes to Runx3 transcription. These findings identify Ets1 as a key player during CD8 lineage differentiation and indicate that it acts, at least in part, by promoting Runx3 expression.

The zinc finger transcription factor Zbtb7b represses CD8-lineage gene expression in peripheral CD4+ T cells.

How CD4-CD8 differentiation is maintained in mature T cells is largely unknown. The present study has examined the role in this process of the zinc finger protein Zbtb7b, a critical factor for the commitment of MHC II-restricted thymocytes to the CD4+ lineage. We showed that Zbtb7b acted in peripheral CD4+ T cells to suppress CD8-lineage gene expression, including that of CD8 and cytotoxic effector genes perforin and Granzyme B, and was important for the proper repression of interferon-gamma (IFN-gamma) during effector differentiation. The inappropriate expression of IFN-gamma by Zbtb7b-deficient CD4+ T cells required the activities of Eomesodermin and Runx transcription factors. Runx activity was needed for Granzyme B expression, indicating that Runx proteins control expression of the cytotoxic program. We conclude that a key function of Zbtb7b in the mature CD4+ T cell compartment is to repress CD8-lineage gene expression.

Dual function for a unique site within the beta2I domain of integrin alphaMbeta2.

Integrin activation has been postulated to occur in part via conformational changes in the I domain of the beta subunit (the betaI domain), especially near the F-alpha(7) loop, in response to "inside-out" signaling. However, direct evidence for a role of the F-alpha(7) loop in ligand binding and activity modulation is still lacking. Here, we report our finding that the F-alpha(7) loop (residues 344-358) within the beta(2)I domain has dual functions in ligand binding by alpha(M)beta(2). On the one hand, it supports intercellular adhesion molecule 1 (ICAM-1) binding to alpha(M)beta(2) directly as part of a recognition interface formed by five noncontiguous segments (Pro(192)-Glu(197), Asn(213)-Glu(220), Leu(225)-Leu(230), Ser(324)-Thr(329), and Glu(344)-Asp(348)) on the apex of the beta(2)I domain. On the other hand, it controls the open and closed conformation of the alpha(M)beta(2) receptor, thereby indirectly affecting alpha(M)beta(2) binding to other ligands. Switching the five constituent sequences of the ICAM-1-binding site within the beta(2)I domain to their beta(1) counterparts destroyed ICAM-1 binding but had no effect on the gross conformations of the receptor. Of the five ICAM-1 binding-defective mutants, four had normal or even stronger interaction with Fg and C3bi, as reported in our previous study. Synthetic peptides derived from the identified site inhibited alpha(M)beta(2)-ICAM-1 interaction and supported direct binding to ICAM-1. Most importantly, perturbation of the F-alpha(7) loop caused conformational changes within the beta(2)I domain, which was further propagated to other regions of alpha(M)beta(2). Altogether, our data demonstrate that inside-out signaling could modulate ligand binding directly by changing the ligand-binding pocket per se and/or indirectly by inducing multiple conformational changes within the receptor.

Short-chain fatty acids stimulate leptin production in adipocytes through the G protein-coupled receptor GPR41.

Leptin is an adipose-derived hormone that regulates a wide variety of physiological processes, including feeding behavior, metabolic rate, sympathetic nerve activity, reproduction, and immune response. Circulating leptin levels are tightly regulated according to energy homeostasis in vivo. Although mechanisms for the regulation of leptin production in adipocytes are not well understood, G protein-coupled receptors may play an important role in this adipocyte function. Here we report that C2-C6 short-chain fatty acids, ligands of an orphan G protein-coupled receptor GPR41, stimulate leptin expression in both a mouse adipocyte cell line and mouse adipose tissue in primary culture. Acute oral administration of propionate increases circulating leptin levels in mice. The concentrations of short-chain fatty acids required to stimulate leptin production are within physiological ranges, suggesting the relevance of this pathway in vivo.