FOXP4-AS1 promotes CD8+ T cell exhaustion and esophageal cancer immune escape through USP10-stabilized PD-L1.

Esophageal cancer (EC) is the 9th most frequently diagnosed malignancy globally with unfavorable prognosis. Immune escape is one of the principal factors leading to poor survival, however, the mechanism underlying immune escape remains largely uninvestigated. The xenograft mouse model and EC cell-CD8+ cytotoxic T lymphocytes (CTLs) co-culture system were established. Immunohistochemistry, qRT-PCR or western blot were employed to detect the levels of long non-coding RNA (lncRNA) FOXP4-AS1, PD-L1, USP10 and other molecules. The abundance of T cells, cytokine production and cell apoptosis were monitored by flow cytometry. The viability of CTLs was assessed by Trypan blue staining. The binding between FOXP4-AS1 and USP10 was validated by RNA pull-down assay, and the interaction between USP10 and PD-L1, as well as the ubiquitination of PD-L1, were detected by co-immunoprecipitation. The elevation of FOXP4-AS1 in EC was associated with decreased CTL abundance, and upregulated PD-L1 facilitated CTL apoptosis in EC. FOXP4-AS1 accelerated EC tumor growth by decreasing the abundance of tumor infiltrating CTLs in vivo. FOXP4-AS1 inhibited the viability of CTLs and facilitated the cytotoxicity and exhaustion of CTLs. In Kyse 450 cell-CTL co-culture system, FOXP4-AS1 suppressed the viability and abundance of CTLs, and inhibited EC cell apoptosis via PD-L1. Mechanistically, FOXP4-AS1 regulated the ubiquitination of PD-L1 through deubiquitinating enzyme USP10. FOXP4-AS1 promoted CTL exhaustion and EC immune escape through USP10-stabilized PD-L1. HIGHLIGHTS: PD-L1 facilitated CD8+ T cell apoptosis in EC. Upregulated FOXP4-AS1 promoted EC tumor growth by inhibiting the viability and facilitating the cytotoxicity and exhaustion of tumor infiltrating CD8+ T cells. FOXP4-AS1 suppressed the viability and abundance of CD8+ T cells through USP10-mediated deubiquitination of PD-L1.

Development and Validation of a Prognostic Model for Postoperative Anastomotic Recurrence in Siewert II or III Adenocarcinomas Without Neoadjuvant Therapy in an East Asian Population.

PURPOSE: Anastomotic recurrence leads to poor prognosis in patients with Siewert II or III adenocarcinoma who undergo radical gastrectomy and do not receive neoadjuvant therapy. We aimed to establish a prognostic model to evaluate the risk of postoperative anastomotic recurrence in patients with Siewert II or III adenocarcinoma who did not receive neoadjuvant therapy. METHODS: We included 366 patients with Siewert II or III adenocarcinoma who were treated with radical gastrectomy without neoadjuvant therapy at Fujian Provincial Hospital (FPH) between 2012 and 2018 as the development cohort. Cox regression was used to verify prognostic factors for anastomotic recurrence, and a nomogram was established. The nomogram was externally validated using a combined cohort of two external centers. Patients were classified into high- or low-risk groups according to the diagnostic threshold and nomogram scores, and recurrence-related survival analysis was analyzed. RESULTS: The average age was 64.6 years, and 285 patients were male. All surgeries were successfully performed (185 open vs 181 laparoscopic). The 3-year anastomotic recurrence rate was significantly lower in the low-risk group (3.5% vs 18.8%, P < 0.001). The predictive performance was verified in the external validation cohort. This model better stratified patient survival than the American Joint Committee on Cancer (AJCC) TNM staging system. CONCLUSIONS: This novel nomogram with surgical margin, postoperative tumor node metastasis (pTNM) stage, and neural invasion as prognostic factors has a significant predictive performance for the risk of anastomotic recurrence after radical gastrectomy in patients with Siewert II or III adenocarcinoma.

High SNHG expression may predict a poor lung cancer prognosis based on a meta-analysis.

BACKGROUND: An increasing number of small nucleolar RNA host genes (SNHGs) have been revealed to be dysregulated in lung cancer tissues, and abnormal expression of SNHGs is significantly correlated with the prognosis of lung cancer. The purpose of this study was to conduct a meta-analysis to explore the correlation between the expression level of SNHGs and the prognosis of lung cancer. METHODS: A comprehensive search of six related databases was conducted to obtain relevant literature. Relevant information, such as overall survival (OS), progression-free survival (PFS), TNM stage, lymph node metastasis (LNM), and tumor size, was extracted. Hazard ratios (HRs) and 95% confidence intervals (CIs) were pooled to evaluate the relationship between SNHG expression and the survival outcome of lung cancers. Sensitivity and publication bias analyses were performed to explore the stability and reliability of the overall results. RESULTS: Forty publications involving 2205 lung cancer patients were included in this meta-analysis. The pooled HR and 95% CI values indicated a significant positive association between high SNHG expression and poor OS (HR: 1.890, 95% CI: 1.595-2.185), disease-free survival (DFS) (HR: 2.31, 95% CI: 1.57-3.39) and progression-free survival (PFS) (HR: 2.01, 95% CI: 0.66-6.07). The pooled odds ratio (OR) and 95% CI values indicated that increased SNHG expression may be correlated with advanced TNM stage (OR: 1.509, 95% CI: 1.267-1.799), increase risk of distant lymph node metastasis (OR: 1.540, 95% CI: 1.298-1.828), and large tumor size (OR: 1.509, 95% CI: 1.245-1.829). Sensitivity analysis and publication bias results showed that each result had strong reliability and robustness, and there was no significant publication bias or other bias. CONCLUSION: Most SNHGs are upregulated in lung cancer tissues, and high expression of SNHGs predicts poor survival outcomes in lung cancer. SNHGs may be potential prognostic markers and promising therapeutic targets.

Epigallocatechin-3-gallate inhibits proliferation and induces apoptosis in odontogenic keratocyst keratinocytes.

OBJECTIVE: The aim of this study was to investigate the effects of epigallocatechin-3-gallate on the proliferation and apoptosis of odontogenic keratocyst (OKC) keratinocytes in vitro. MATERIALS AND METHODS: Keratinocytes isolated from the epithelial lining of the OKC were cultured in keratinocyte serum-free medium and identified by CK10, CK14, pan-cytokeratin and vimentin immunofluorescence staining. The cells were exposed to EGCG at different concentrations, and proliferation inhibition was measured by cell counting kit 8 assay. Cell cycle and apoptosis were assessed by flow cytometry, and expression of the WNT signalling pathway-related proteins FZD3 and JNK3 was detected by quantitative real-time PCR and Western blotting. Human oral keratinocytes (HOKs) were used as the control. RESULTS: The OKC keratinocytes were successfully cultured. The primary cells were tile-like and expressed the epithelial biomarkers CK10, CK14 and pan-cytokeratin. Epigallocatechin-3-gallate inhibited cell proliferation in a dose- and time-dependent manner, arrested cell cycle in the G1 phase and induced apoptosis of OKC keratinocytes. FZD3 and JNK3 were overexpressed in OKC keratinocytes compared with HOKs and were downregulated by epigallocatechin-3-gallate treatment. CONCLUSION: Epigallocatechin-3-gallate inhibited proliferation and induced apoptosis in OKC keratinocytes, possibly by suppressing the WNT/JNK signalling pathway. It may thus be potentially used for OKC treatment.

From pure C36 fullerene to cagelike nanocluster: a density functional study.

The geometrical structures, energetics properties, and aromaticity of C(36-n) Si(n) (n ≤ 18) fullerene-based clusters were studied using density functional theory calculations. The geometries of C(36-n) Si(n) clusters undergo strong structural deformation with the increase of Si substitution. For the most energy favorable structures of C(36-n) Si(n) , the silicon and carbon atoms form two distinct homogeneous segregations. Subsequently, the binding energy, HOMO-LUMO energy gap, vertical ionization potential, vertical electron affinity, and chemical hardness for the energetic favorable C(36-n) Si(n) geometries were computed and analyzed. In addition, the aromatic property of C(36-n) Si(n) cagelike clusters was investigated, and the result demonstrate that these C(36-n) Si(n) cagelike structures possess strong aromaticity.

Synergistic analgesic effects between neuronostatin and morphine at the supraspinal level.

Neuronostatin, a 13-amino acid peptide, is encoded in the somatostatin pro-hormone. I.c.v. administration of neuronostatin produces a significant antinociceptive effect in the mouse tail-flick test, which is mediated by endogenous opioid receptor. However, the direct functional interaction between morphine and neuronostatin has not been characterized. In the present study, effect of neuronostatin on morphine analgesia was investigated in the tail-flick test. Our findings showed that i.c.v. administration of neuronostatin (0.3nmol/mouse i.c.v.) significantly enhanced the antinociceptive effect of morphine (2.5, 5 or 10µg/kg) at the supraspinal level. Results of antagonism experiments suggested that the synergistic analgesia induced by morphine and neuronostatin was mediated by µ- and к-opioid receptors not δ-opioid receptor. In conclusion, there may be a cascade amplification phenomenon when morphine and neuronostatin were co-administered in acute pain model. The above results provide evidence for the potential use of neuronostatin in combination with morphine to control pain and addiction.

Intracerebroventricular administration of neuronostatin delays gastric emptying and gastrointestinal transit in mice.

Neuronostatin is a 13-amino acid amidated peptide widely distributed in various organs including gastrointestinal tract. However, the effect of neuronostatin on gastrointestinal motility has not been well characterized. In the present work, effects of central administration of neuronostatin on gastric emptying and gastrointestinal transit were investigated. The results indicated that intracerebroventricular (i.c.v.) administration of neuronostatin (1, 5, 10 or 20nmol/mouse) delayed gastric emptying and gastrointestinal transit in a dose-related manner in mice. The effects were significantly reversed by melanocortin 3/4 receptor antagonist SHU9119 or classical opioid receptor antagonist naloxone, suggesting that the central melanocortin system and opioid system may be involved in the gastrointestinal effects elicited by i.c.v. administration of neuronostatin. In addition, we found that C-terminal amidation modification of neuronostatin is essential to exert its gastrointestinal effects. These results indicated that neuronostatin may play an important role in regulating gastrointestinal function.

Neuronostatin induces hyperalgesia in formalin test in mice.

Neuronostatin, a newly identified peptide encoded by the somatostatin (SST) gene, was proved to produce significant antinociceptive effect in mouse tail immersion test. However, the effect of neuronostatin on tonic pain was still not clear. The aim of this study was to investigate the effect of neuronostatin in the formalin test and its possible mechanism. We found that intracerebroventricular (i.c.v.) administration of neuronostatin (1, 3, 6, 12nmol/mouse) increased licking in a dose-related manner during the late phase, but did not affect the early phase of formalin test in mice. In addition, the hyperalgesic effect during the late phase was completely reversed by melanocortin 3/4 receptor antagonist SHU9119 (50pmol/mouse) or opioid receptor antagonist naloxone (5nmol/mouse), but not GABAA receptor antagonist bicuculline (1086pmol/mouse). These data suggested that the hyperalgesic response induced by neuronostatin was dependent upon the central melanocortin system and endogenous opioid system. In conclusion, these results indicated that neuronostatin may be a new neuropeptide with important role in the modulation of acute and tonic pain.

Intracerebroventricular administration of neuronostatin induces depression-like effect in forced swim test of mice.

Neuronostatin is a recently discovered endogenous bioactive peptide that is encoded by pro-mRNA of somatostatin. In the present study, we investigated the effect of neuronostatin on mood regulation in the forced swim test of mice. Our results showed intracerebroventricular (i.c.v.) administration of neuronostatin produced an increase in the immobility time, suggesting that neuronostatin induced depression-like effect. In order to rule out the possibility that neuronostatin had increased immobility time by a non-specific reduction in general activity, the effect of neuronostatin on locomotor activity was examined. Neuronostatin had no influence on locomotor activity in mice. In addition, the depression-like effect of neuronostatin was completely reversed by melanocortin 3/4 receptor antagonist SHU9119 or GABAA receptor antagonist bicuculline, but not by opioid receptor antagonist naloxone. These data suggested that the depression-like effect induced by i.c.v. administered neuronostatin was dependent upon the central melanocortin system and GABAA receptor. In conclusion, the results of this study report that neuronostatin induces depression-like effect. These findings reveal that neuronostatin is a new neuropeptide with an important role in regulating depressive behavior.

Central administration of neuronostatin induces antinociception in mice.

Neuronostatin, a recently discovered endogenous bioactive peptide, was encoded by pro-mRNA of somatostatin that contributes to modulation of nociception. However, nociceptive effect of neuronostatin is still not fully known. The aim of this study was to evaluate effect of neuronostatin on nociception and elucidate its possible mechanism of action. Intracerebroventricular (i.c.v.) administration of neuronostatin (0.3, 3, 6, 12nmol/mouse) produced a dose- and time-related antinociceptive effect in the tail immersion assay in mice, an acute pain model. The antinociceptive effect of neuronostatin was significantly antagonized by naloxone, and was strongly inhibited by co-injection with ß-funaltrexamine or nor-binaltorphimine, but not by naltrindole. Also, melanocortin 3/4 receptor antagonist, SHU9119, completely blocked the effect of neuronostatin. These data indicated the involvement of both µ- and κ-opioid receptors and central melanocortin system in the analgesic response induced by neuronostatin. In addition, neuronostatin (6nmol, i.c.v.) increased c-Fos protein expression in the periaqueductal gray (PAG) and the nucleus raphe magnus (NRM) that have a pivotal role in regulating descending pain pathways. Taken together, this study is the first to reveal that neuronostatin produces antinociceptive effect via opioid and central melanocortin systems, which is associated with an increase in neuronal activity the PAG and NRM.