The possible regulatory effect of the PD-1/PD-L1 signaling pathway on Tregs in ovarian cancer.

Aim of this study was to investigate the possible regulatory effect of the programmed death-1 (PD-1)/programmed death ligand-1 (PD-L1) signaling pathway on Tregs in ovarian cancer. Immunohistochemistry was used to detect the expression of PD-L1 and PD-1 and the presence of FOXP3+ Tregs in ovarian cancer. Then, ovarian cancer HO8910 cells were subjected to transfection with PD-L1 siRNA in vitro. CCK-8, Transwell and wound healing assays were performed to detect the biological behaviors of ovarian cancer cells. Human T-cells isolated from human peripheral blood were cocultured with HO8910 cells, which were divided into the Control, TGF-ß, and TGF-ß+ anti-PD-L1 groups. The proportion of differentiated Tregs was detected by flow cytometry. Mouse models of ovarian cancer were established, and PD-L1 antibody therapy was administered. Tumor growth and Treg recruitment were observed. PD-L1, PD-1 and FOXP3+ Tregs were found in ovarian cancer tissue. Patients with tumors with an advanced stage and low differentiation and lymph node metastasis had significantly higher levels of PD-1, PD-L1 and FOXP3+ Tregs. After transfection with PD-L1 siRNA, HO8910 cells showed a significant reduction in PD-L1 expression, proliferation, migration and invasion. After T-cells were cocultured with ovarian cancer cells, the TGF-ß+ anti-PD-L1 group showed a substantial decline in the differentiation of T-cells into Tregs compared with the TGF-ß group. Moreover, mice in the anti-PD-L1 group had significantly reduced tumor growth rates, Treg proportions in the tumor microenvironment, and FOXP3 expression.

The role of KDR in intrauterine adhesions may involve the TGF-ß1/Smads signaling pathway.

The aim of this study was to investigate the role of kinase-insert domain-containing receptor (KDR) in intrauterine adhesions (IUA) and its mechanism. The Case group consisted of 92 patients diagnosed with IUA, and the Control group included 86 patients with uterine septum who had normal endometrium verified with an uteroscope. In addition, 50 rats were randomly assigned into Control, Sham, Model, NC-siRNA, and KDR-siRNA groups. Rats in the Model, NC-siRNA, and KDR-siRNA groups were induced by uterine curettage and lipopolysaccharide (LPS) treatment to establish the IUA model. Then, immunohistochemistry was applied for detection of VEGF and KDR expression, HE staining was used for observation of the endometrial morphology and gland counting, Masson staining for measurement of the degree of endometrial fibrosis, and qRT-PCR and western blot for the expression of KDR, VEGF, MMP-9, as well as TGF-ß1/Smads pathway-related proteins. Compared with the Control group, the mRNA and protein expressions of KDR were significantly higher in IUA endometrial tissues, and the expression of KDR was positively correlated to the severity of IUA. In addition, the injection of si-KDR increased the number of endometrial glands, reduced the area of fibrosis, inhibited mRNA and protein expression of KDR and VEGF, up-regulated the expression of MMP-9 and Smad7, and decreased the expression level of TGF-ß1, p-Smad2, p-Smad3, and Smad4 in rats with IUA. Highly-expressed KDR was related to patients' severity of IUA, and silencing KDR may prevent the occurrence and development of IUA via TGF-ß1/Smads signaling pathway and up-regulating the expression of MMP-9.

The role of KDR in intrauterine adhesions may involve the TGF-ß1/Smads signaling pathway

The aim of this study was to investigate the role of kinase-insert domain-containing receptor (KDR) in intrauterine adhesions (IUA) and its mechanism. The Case group consisted of 92 patients diagnosed with IUA, and the Control group included 86 patients with uterine septum who had normal endometrium verified with an uteroscope. In addition, 50 rats were randomly assigned into Control, Sham, Model, NC-siRNA, and KDR-siRNA groups. Rats in the Model, NC-siRNA, and KDR-siRNA groups were induced by uterine curettage and lipopolysaccharide (LPS) treatment to establish the IUA model. Then, immunohistochemistry was applied for detection of VEGF and KDR expression, HE staining was used for observation of the endometrial morphology and gland counting, Masson staining for measurement of the degree of endometrial fibrosis, and qRT-PCR and western blot for the expression of KDR, VEGF, MMP-9, as well as TGF-β1/Smads pathway-related proteins. Compared with the Control group, the mRNA and protein expressions of KDR were significantly higher in IUA endometrial tissues, and the expression of KDR was positively correlated to the severity of IUA. In addition, the injection of si-KDR increased the number of endometrial glands, reduced the area of fibrosis, inhibited mRNA and protein expression of KDR and VEGF, up-regulated the expression of MMP-9 and Smad7, and decreased the expression level of TGF-β1, p-Smad2, p-Smad3, and Smad4 in rats with IUA. Highly-expressed KDR was related to patients' severity of IUA, and silencing KDR may prevent the occurrence and development of IUA via TGF-β1/Smads signaling pathway and up-regulating the expression of MMP-9.

2-aminopurine suppresses the TGF-ß1-induced epithelial-mesenchymal transition and attenuates bleomycin-induced pulmonary fibrosis.

The epithelial-mesenchymal transition (EMT) is a multifunctional cell process involved in the pathogenesis of numerous conditions, including fibrosis and cancer. Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal disease characterized by fibroblast accumulation and collagen deposition in the lungs. The fibroblasts involved in this process partially originate from lung epithelial cells via the EMT. Evidence suggests that the EMT contributes to progression, invasion, and metastasis of various types of cancer. We screened a series of 80 compounds for the ability to interfere with the EMT and potentially be applied as a therapeutic for IPF and/or lung cancer. We identified 2-aminopurine (2-AP), a fluorescent analog of guanosine and adenosine, as a candidate in this screen. Herein, we demonstrate that 2-AP can restore E-cadherin expression and inhibit fibronectin and vimentin expression in TGF-ß1-treated A549 lung cancer cells. Moreover, 2-AP can inhibit TGF-ß1-induced metastasis of A549 cells. This compound significantly attenuated bleomycin (BLM)-induced pulmonary inflammation, the EMT, and fibrosis. In addition, 2-AP treatment significantly decreased mortality in a mouse model of pulmonary fibrosis. Collectively, we determined that 2-AP could inhibit metastasis in vitro by suppressing the TGF-ß1-induced EMT and could attenuate BLM-induced pulmonary fibrosis in vivo. Results of this study suggest that 2-AP may have utility as a treatment for lung cancer and pulmonary fibrosis.

Urinary metabonomics of rats with ketamine-induced cystitis using GC-MS spectroscopy.

Ketamine abuse has dramatically increased in recently years. With the widely application of ketamine, its side effects, especially cystitis induced by long-term use, have attracted more and more attention from the public. In the present study, we aimed to explore the potential generative mechanism of ketamine-induced cystitis by determining the endogenous metabolites at different time points after ketamine treatment. Body weight, bladder/body coefficient, urinary frequency, urinary potassium, serum IL-6, and TNF-α were determined at different time points after ketamine treatment. H&E staining was used to observe the changes of histopathology. Metabonomics was performed to determine the changes of endogenous metabolites. After 12 weeks of treatment, obvious inflammatory reaction was noticed in the KET group; the body weight and urinary potassium of the KET group were significantly lower than the NS group (P < 0.05) and other factors, such as urinary frequency, bladder/body coefficient, serum TNF-α and IL-6 were higher than the NS group (P < 0.05). A total of 30, 28, and 32 significantly changed metabolites were identified at the 1st week, 4th week and 12th week, respectively. Metabolic pathway analysis showed that different metabolic pathways were affected during the treatment process. Linoleic acid metabolism, beta-alanine metabolism, glyoxylate and dicarboxylate metabolism were only affected following long-term administration of ketamine. Those metabolic pathways may have a close relationship with cystitis induced by ketamine.

Predictive value of microRNA-143 in evaluating the prognosis of patients with hepatocellular carcinoma.

OBJECTIVE: The study aims to evaluate the predictive value of microRNA-143 (miR-143) for the prognosis of patients with hepatocellular carcinoma (HCC). METHODS: Between October 2010 and October 2012, 131 HCC patients were selected as a case group; meanwhile, 122 healthy controls were enrolled as a control group. The miR-143 expression in serum was detected by quantitative real-time polymerase chain reaction (qRT-PCR). These HCC patients were divided into the high miR-143 expression group and the low miR-143 expression group based on the threshold of receiver operating characteristic (ROC) curve. Kaplan-Meier method was applied to analyze the prognosis of HCC patients. RESULTS: MiR-143 exhibited decreased expression in the case group significantly compared to the control group. The areas under the ROC curve (AUC), sensitivity value and specificity value of the miR-143 expression for the diagnosis of HCC were 0.831, 80.30% and 82.40%, respectively. The miR-143 expression was negatively correlated with vascular invasion, TNM staging, tumor recurrence, metastasis and survival of HCC patients. CONCLUSIONS: Our study provides evidence that miR-143 may be negatively correlated with the prognosis of HCC and provides a promising strategy for HCC treatment and prognosis improvement.

Porphyromonas gingivalis infected macrophages upregulate CD36 expression via ERK/NF-κB pathway.

CD36, a scavenger receptor, plays an important role in the progression of atherosclerosis through its interaction with oxidized low-density lipoprotein (ox-LDL). Porphyromonas gingivalis (P. gingivalis, Pg) has been shown to promote macrophage-derived foam cell formation by affecting the expression of CD36. However, the regulatory role of CD36 in macrophages infected with Pg remains largely unknown. Therefore, the aim of the present study was to explore the molecular mechanism of Pg induced CD36 expression in macrophages. Our results showed that Pg promoted ox-LDL uptake by macrophages and the formation of foam cells. Pg infection increased CD36 mRNA and protein levels in ox-LDL-untreated macrophages. Moreover, small interferon RNA (siRNA) targeting CD36 significantly reduced foam cell formation induced by Pg. Additionally, Pg stimulated nuclear translocation of p65, which directly bound to the promoters of CD36 to facilitate its transcription. Inhibition of p65, NF-κB or ERK1/2 blocked Pg-induced CD36 production; whereas, overexpression of NF-κB subunits p65 and p50 upregulated CD36. Furthermore, Ras inhibitors significantly attenuated ERK1/2 activation and CD36 expression. Taken together, the data indicated that stimulation of the ERK/NF-κB pathway by Pg led to transactivation of the CD36 promoters, thereby upregulating CD36 expression in the infected macrophages. These findings may help design new treatment strategies in atherosclerosis.

Optimal cut-off values for CYFRA 21-1 expression in NSCLC patients depend on the presence of benign pulmonary diseases.

BACKGROUND: Serum cytokeratin fragment 21-1 (CYFRA 21-1) expression levels are reported to be useful in the diagnosis of lung cancer, especially non-small cell lung cancer (NSCLC). However, the clinical value of CYFRA 21-1 as a tumor marker remains unclear, and no optimal cut-off value has been determined thus far. The purpose of this study was to establish a potential clinical cut-off value for serum CYFRA 21-1 as a diagnostic marker in patients with NSCLC. METHODS: A total of 90 patients with NSCLC, 237 patients with benign pulmonary disease (BPD), and 1296 healthy controls were enrolled in this study. Among BPD there are 84 with chronic obstructive pulmonary disease (COPD), 81 with pneumonia, 38 with tuberculosis and 34 with chronic bronchitis. CYFRA 21-1 was measured in sera with an electrochemiluminescence (ECL) E170 analyzer. Comparisons were conducted using the chi-squared test and the Mann-Whitney test (two-sided). A receiver operating characteristic (ROC) curve was constructed to investigate the diagnostic power of CYFRA 21-1 expression, and the recommended cut-off value was chosen to calculate its sensitivity and specificity. RESULTS: The cut-off values of CYFRA 21-1 in NSCLC by the ROC curve were 4.70 ng/mL when compared with COPD, which was obviously greater than that found with pneumonia (2.79 ng/mL) (P<0.05), tuberculosis (2.66 ng/mL) (P<0.05), and chronic bronchitis (3.94 ng/mL) (P<0.05) patients. Therefore, a cut-off value of 4.24 ng/mL in NSCLC was suggested. CONCLUSIONS: The presence of various BPDs may be one of the main reasons that no optimal cut-off value for CYFRA 21-1 expression in NSCLC has been determined previously.

Association of supervillin with KIR2DL1 regulates the inhibitory signaling of natural killer cells.

Inhibitory signaling is crucial in the regulation of the cytotoxicity of natural killer (NK) cells. Here, we show that KIR2DL1, an inhibitory receptor of NK cells, associates with supervillin, an F-actin binding protein. Interaction of supervillin with KIR2DL1 is dependent on the KIR2DL1 receptor stimulation and requires the phosphorylation of tyrosines in both ITIM motifs. "Knockdown" of expression of supervillin by RNA interference (RNAi) restores the KIR2DL1-suppressed cytotoxicity of NK cells. Inhibition of supervillin by RNAi also enhances the polarization of cytolytic granules (both granzyme B and perforin) to the synapse formed between YTS-GFP-KIR2DL1 NK cells and 721.221-HLA-Cw4 target cells. Further study reveals that supervillin is required for KIR2DL1-mediated inhibition of Vav1 and ERK phoshorylation. Moreover, we have found that binding of supervillin with KIR2DL1 facilitates the recruitment of SHPs especially SHP-2 to KIR2DL1 receptor. Thus, our findings demonstrate that supervillin is a novel molecule that associates with KIR2DL1 receptor and regulates the inhibitory signaling in NK cells.

MicroRNA-101 targets MAPK phosphatase-1 to regulate the activation of MAPKs in macrophages.

MAPK phosphatase-1 (MKP-1) is an archetypical member of the dual-specificity phosphatase family that deactivates MAPKs. Induction of MKP-1 has been implicated in attenuating the LPS- or peptidoglycan-induced biosynthesis of proinflammatory cytokines, but the role of noncoding RNA in the expression of the MKP-1 is still poorly understood. In this study, we show that MKP-1 is a direct target of microRNA-101 (miR-101). Transfection of miR-101 attenuates induction of MKP-1 by LPS as well as prolonged activation of p38 and JNK/stress-activated protein kinase, whereas inhibition of miR-101 enhances the expression of MKP-1 and shortens p38 and JNK activation. We also found that expression of miR-101 is induced by multiple TLR ligands, including LPS, peptidoglycan, or polyinosinic-polycytidylic acid, and that inhibition of PI3K/Akt by LY294002 or Akt RNA interference blocks the induction of miR-101 by LPS in RAW264.7 macrophage cells. Moreover, treatment of cells with dexamethasone, a widely used anti-inflammatory agent, markedly inhibits miR-101 expression and enhances the expression of MKP-1 in LPS-stimulated macrophages. Together, these results indicate that miR-101 regulates the innate immune responses of macrophages to LPS through targeting MKP-1.