Down-regulated miR-26a promotes proliferation, migration, and invasion via negative regulation of MTDH in esophageal squamous cell carcinoma.

Numerous studies have reported that the role played by miR-26a in cancer is controversial, but whether miR-26a regulates metadherin (MTDH) expression in esophageal squamous cell carcinoma (ESCC) is unclear. We performed this study to investigate the clinical relevance of miR-26a expression in ESCC. miR-26a was detected by using the in situ hybridization method. To functionally analyze the role of miR-26a in ESCC cell lines in vitro, KYSE-450 and Eca109 cells were employed, whose endogenous miR-26a was artificially down- or up-regulated, respectively, by using lentiviral-based transfection. There was significant association between miR-26a expression and clinical stage (P = 0.049), lymph node metastasis (P = 0.023), tumor volume (P = 0.003), and poor overall prognosis (P = 0.026). miR-26a was able to suppress proliferation and migration of ESCC cells in vitro Moreover, we have confirmed that miR-26a can negatively regulate MTDH in ESCC cells by using luciferase reporter assay. In addition, to investigate the role miR-26a plays in cell proliferation, we nude mice were xenografted with ESCC cells whose miR-26a was stably down- and up-regulated. Together, our results show that miR-26a is capable of suppressing the proliferation and migration of ESCC cells via negative regulation of MTDH. Moreover, miR-26a expression was clinically relevant in cancer progression and poor prognosis, which supports the idea that miR-26a acts as a tumor suppressor in ESCC.-Yang, C., Zheng, S., Liu, T., Liu, Q., Dai, F., Zhou, J., Chen, Y., Sheyhidin, I., Lu, X. Down-regulated miR-26a promotes proliferation, migration, and invasion via negative regulation of MTDH in esophageal squamous cell carcinoma.

Metabolomic profiling reveals potential biomarkers in esophageal cancer progression using liquid chromatography-mass spectrometry platform.

Esophageal cancer (EC) is one of the most common malignancies with poor prognosis. Metabolomics has been shown to be a powerful approach to discover the potential biomarkers for cancer diagnosis and prognosis. The goal of this study is to screen potential biomarkers for early diagnosis and prognosis. In this study, 40 tissue samples and the corresponding control samples from the same esophageal squamous cell carcinoma (ESCC) patients were analyzed by liquid chromatography-mass spectrometry (LC-MS)-based metabolomics. 20 potential diagnostic biomarkers were selected. Moreover, 9 metabolites were found to be closely correlated with the pathological feature such as local invasion, lymphatic metastasis and postoperative survival time. Glutamate was correlated with local invasion of tumor, and oleic acid, LysoPC(15:0), uracil, inosine and choline were closely related with the lymphatic metastasis, while glutamine, kynurenine, serine and uracil were related with postoperative survival time. The results indicated that the potential biomarkers discovered by metabolomics could reflect the metabolic characterization of ESCC, and offers a novel approach for early diagnosis, assessment and prognosis of the disease.

Partition-Defective 3 (PARD3) Regulates Proliferation, Apoptosis, Migration, and Invasion in Esophageal Squamous Cell Carcinoma Cells.

BACKGROUND Altered expression of partition-defective 3 (PARD3), a polarity-related gene associated with oncogenesis, has been identified in some cancers, but the role of PARD3 in esophageal squamous cell carcinoma (ESCC) remains unclear. MATERIAL AND METHODS PARD3 expression in Eca109 cells was silenced using siRNA and overexpressed using an expression vector. We investigated the role of PARD3 in ESCC growth and motility to evaluate its potential role in ESCC. Transwell assay was used to evaluated cell migration and invasion. PARD3 protein expression was assessed by Western blot. RESULTS PARD3 overexpression promoted apoptosis, impaired proliferation, and inhibited cell migration and invasion in Eca109 cells, while PARD3 silencing promoted proliferation and increased migration and invasion. Overexpression of PARD3 exerted its antitumor activity in vitro by impairing cell proliferation, inducing apoptosis, and inhibiting migration and invasion of Eca109 cells, suggesting that PARD3 might play a tumor suppressor role in ESCC. CONCLUSIONS Overexpression of PARD3 could be a promising new therapeutic intervention against ESCC.

Clinicopathological significance of p38ß, p38γ, and p38δ and its biological roles in esophageal squamous cell carcinoma.

P38ß, p38γ, and p38δ have been sporadically and scarcely reported to be involved in the carcinogenesis of cancers, compared with p38α isoform. However, little has been known regarding their clinicopathological significance and biological roles in esophageal squamous cell carcinoma (ESCC). Expression status of p38ß, p38γ, and p38δ was assayed using immunohistochemistry with ESCC tissue microarray; ensuing clinicopathological significance was statistically analyzed. To define its biological roles on proliferation, migration and invasion of ESCC cell line Eca109 in vitro, MTT, wound healing, and Transwell assays were employed, respectively. As confirmation, athymic nude mice were taken to verify the effect over proliferation in vivo. It was found that both p38ß and p38δ expression, other than p38γ, were significantly higher in ESCC tissues compared with paired normal controls. In terms of prognosis, only p38ß expression was observed to be significantly associated with overall prognosis. Clinicopathologically, there was significant association between p38γ expression and clinical stage, lymph nodes metastases, and tumor volume. No significant association was found for p38ß and p38δ between its expression and other clinicopathological parameters other than significant difference of expression between ESCC versus normal control. In Eca109, it was observed that p38ß, p38γ, and p38δ can promote the cell growth and motility. As verification, over-expression of p38δ can promote, whereas knockdown of p38γ can prevent, the tumorigenesis in nude mice model xenografted with Eca109 cells whose basal level of p38δ was stably over-expressed and p38γ was stably knocked down. Together, our results demonstrate that p38ß, p38γ, and p38δ played oncogenic roles in ESCC.

MiR-106b promotes migration and invasion through enhancing EMT via downregulation of Smad 7 in Kazakh's esophageal squamous cell carcinoma.

Accumulated evidence suggests that miR-106b played a key role in the promotion of the metastases of cancer; however, little is known about miR-106b in esophageal squamous cell carcinoma (ESCC). To investigate expression level of miR-106b in ESCC tissues, quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect miR-106b expression in 35 Kazakh's ESCC and paired normal adjacent tissues (NATs). To evaluate the role mediated by miR-106b in the proliferation, migration, and invasion, MTT, wound healing, and transwell assays were employed, respectively. Luciferase reporter assay was used to identify the downstream target through miR-106b. To understand the regulation between miR-106b and Smad 7, qRT-PCR and western blot were performed. The present study showed that miR-106b was pronouncedly upregulated in ESCC relative to paired NAT and that upregulated miR-106b was significantly associated with lymph node metastases. MiR-106b was found to be able to promote proliferation, migration, and invasion of ESCC cells in vitro. Smad 7 was confirmed as a downstream target of miR-106b in our experimental setting. Smad 7 was remarkably downregulated in ESCC compared with paired NAT. In addition, upregulation of miR-106b can promote epithelial mesenchymal transition (EMT) in ESCC cell in vitro. Our results indicated that miR-106b can promote migration and invasion of ESCC cells through enhancing EMT process via downregulation of Smad 7, suggesting that miR-106b can be a potential molecular phenotype in ESCC metastases.

M2 isoform of pyruvate kinase (PKM2) is upregulated in Kazakh's ESCC and promotes proliferation and migration of ESCC cells.

The objectives of the present study are to explore role of pyruvate kinase isoenzyme type M2 (PKM2) in progression of Kazakh's esophageal squamous cell carcinoma (ESCC) in Xinjiang, China, and to clarify mechanism of PKM2 in malignant phenotype. PKM2 expression was examined using immunohistochemistry (IHC) in 101 matched pairs of ESCC and normal adjacent tissues (NATs) and using enzyme-linked immunosorbent assay (ELISA) in 35 serum samples of Kazakh's ESCC and 8 serum samples of healthy subjects. To investigate mechanism, small interfering RNA (siRNA)-PKM2 was transfected into ESCC cells. Cell migration and invasion were evaluated by wound healing and Transwell assays. Apoptosis and cell cycle were analyzed by flow cytometry (FCM). PKM2 expression was significantly higher in ESCC tissues (77.2 %, 78/101) compared with matched NAT (P = 0.003) and also higher in serum samples of Kazakh's ESCC patients (78.84 ng/mL) compared with healthy subjects (13.55 ng/mL) (P = 0.001). Patients with overexpression of PKM2 had a poor prognosis (P = 0.032). After knockdown of PKM2, cell proliferation, migration, and invasion were significantly reduced (P = 0.001), apoptosis increased (P = 0.001), and cell cycle was arrested at G1 phase. PKM2 overexpression was significantly correlated with the worse outcome of Kazakh's ESCC. Furthermore, PKM2 was involved in progression of ESCC by promoting proliferation and suppressing apoptosis, accelerating invasion, and influencing cell cycle. PKM2 could be a potential biomarker for molecular classification of ESCC.

Metabonomic signature analysis in plasma samples of glioma patients based on (1)H-nuclear magnetic resonance spectroscopy.

OBJECTIVE: The presence of a glioma is associated with increasing mortality. In this study, nuclear magnetic resonance (NMR) based metabonomics has been applied to investigate the metabolic signatures of a glioma in plasma. The purpose of this study was to assess the diagnostic potential of this approach and gain novel insights into the metabolism of glioma and its systemic effects. METHODS: Plasma samples were collected prospectively by centrifugation of blood samples from patients with a glioma (n = 70) or a control group (n = 70). NMR spectra of these plasma samples were analyzed using orthogonal partial least square discriminant analysis (OPLS-DA) to identify the potential biomarkers. RESULTS: The OPLS-DA model showed a good differentiation between the glioma and the control groups. A total of 20 metabolites were identified, which are closely correlating with the presence of a glioma. Compared to the control group, patients with a glioma were associated with lower concentrations of isoleucine, leucine, valine, lactate, alanine, glycoprotein, glutamate, citrate, creatine, myo-inositol, choline, tyrosine, phenylalanine, 1-methylhistidine, α-glucose, ß-glucose, and higher concentrations of very low density lipoprotein, low density lipoprotein (LDL), unsaturated lipid, and pyruvate. These 20 metabolites, which are involved in energy, fatty acid, and amino acid metabolism, may be associated with a human glioma. CONCLUSION: Our study is the first one to identify the plasma metabolites that have the potential to distinguish between patients with a glioma and healthy subjects. NMR-based metabonomics provides a good sensitivity and selectivity in differentiating the healthy control group from patients suffering form the disease. Plasma metabolic profiling may have a potential in diagnosing a glioma in the early phase and may help in enhancing our understanding of its underlying mechanisms.

Under-expression of annexin A2 is associated with Kazakh's esophageal squamous cell carcinoma.

The aim of the study was to identify candidate biomarkers for esophageal squamous cell carcinoma (ESCC) in Kazakh ethnic in Xinjiang as well as to reveal the potential role of Annexin A2 in ESCC carcinogenesis and progression. Five paired of Kazakh's ESCC tissues (T) and matched adjacent morphologically normal tissues (N) were separated by two-dimensional electrophoresis (2-DE) and differential proteins were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Annexin A2 was identified as a down-regulated protein in Kazakh's ESCC and further validated by immunohistochemistry (IHC) in 77 Kazakh's ESCC formalin-fixed paraffin-embeded (FFPE) samples. The expression level of Annexin A2 protein significantly correlated with the degree of ESCC differentiation and depth of invasion. For clarification of the role of Annexin A2 in regulating cell phenotype, in vitro eukaryotic expression vectors harboring full length Annexin A2 (pCMV-XL5-Annexin A2) was tranfected into Eca109 cells, and transfection effects were evaluated by RT-PCR and Western blotting analysis, respectively. Functionally, there was a significant decrease in cell proliferation, migration, and invasion capability in Eca109 with transfected pCMV-XL5-Annexin A2 compared to the controls. Furthermore, up-regulating Annexin A2 can significantly cause cell cycle arrest at the G2 phase, but no apoptosis was induced. Together, our findings suggested that Annexin A2 was involved in malignant phenotype and was a potential biomark for molecular classification in ESCC.

Roles of phosphorylated JNK in esophageal squamous cell carcinomas of Kazakh ethnic.

The c-Jun NH2 -terminal kinase (JNK) signal pathway has been implicated in the growth, cellular proliferation, and apoptosis in many kinds of carcinomas. However, the role of JNK in the development of esophageal squamous cell carcinomas (ESCCs) is unknown. To investigate the role of JNK in ESCC, in vitro, esophageal cancer cell line Eca109 was pretreated using SP600125, JNK specific inhibitor, then was subjected to MTT assay to examine cellular proliferation, flow cytometric analysis to detect apoptosis and cell cycle, and wound healing assay to evaluate cell migration. Meanwhile, the mRNA and protein expression of JNK in Eca109 cells pretreated with SP600125 were examined by real-time quantitative reverse transcription PCR (qRT-PCR) and Western blotting, respectively. In vivo, 12 paired of fresh ESCC and normal adjacent tissues (NAT) from Kazakh patients were used to validate the expression of JNK by qRT-PCR and Western blotting. Furthermore, to reconfirm the expression trend of activation JNK (p-JNK), enlarged 72 paired of Kazakh's ESCC and NAT were subjected to immunohistochemistry. Our results showed that the suppression of p-JNK could lead to apoptosis and reduce proliferation in Eca109 cells. However, there was an elevated expression of p-JNK protein in NAT compared with ESCC tissues, and there was significant difference between p-JNK expression and pathological differentiation (P < 0.05) in Kazakh populations. Together, all the data we obtained in the present study indicated that the p-JNK MAPK pathway was involved in pathogenesis of Kazakh's ESCC, and played a different roles in carcinogenesis and development of Kazakh's ESCC.

HMGA2 is down-regulated by microRNA let-7 and associated with epithelial-mesenchymal transition in oesophageal squamous cell carcinomas of Kazakhs.

AIMS: To investigate the expression of let-7 and its regulation of high-mobility group A2 protein (HMGA2), and to verify the relationship between let-7, HMGA2 and the process of epithelial-mesenchymal transition (EMT), in oesophageal squamous cell carcinomas (OSCC) of Kazakh patients. METHODS AND RESULTS: Expression of let-7 was significantly lower in Eca109 cells than in normal oesophageal squamous epithelium (P = 2.4 × 10(-7) ). Increased accumulation of let-7 after transfection of Eca109 cells with synthetic let-7 inhibited cell proliferation. Let-7 could repress expression of HMGA2 after co-transfection with let-7 and HMGA2 (P = 0.002). Moreover, let-7 expression was observed less frequently (P = 2.0 × 10(-8) ), and HMGA2 expression more frequently (P = 1.0 × 10(-10) ), in OSCC than in normal adjacent tissues; and let-7 expression was observed less frequently in OSCC from Kazakh patients than in those from Han and Uygur patients (P = 0.041). There was a reverse correlation between expression of let-7 and HMGA2 (P = 0.018). Expression of Snail was statistically higher in Kazakhs' OSCC (P = 0.029), and was correlated with depth of invasion (P = 0.021) and HMGA2 expression (P = 0.026). CONCLUSIONS: Expression of let-7 can suppress cell proliferation by acting directly on regulation of HMGA2 in OSCC. High expression of Snail and its correlation with HMGA2 expression and tumour invasion suggest that HMGA2 may be involved in EMT in the OSCC of Kazakh patients.

MicroRNA-21 promotes the proliferation and inhibits apoptosis in Eca109 via activating ERK1/2/MAPK pathway.

The aim of this study was to investigate how miR-21 promotes proliferation and inhibits apoptosis in esophageal squamous cell carcinoma (ESCC). MTT, wound healing assay and cell cycle showed that proliferation and migration of ESCC cell line Eca109 cells were increased in miR-21 mimics group, and decreased in anti-miR-21 Oligonucleotide (AMO) group after transfection into Eca109 cells with miR-21 mimics, AMO and scramble sequence, respectively. Cell apoptosis assay indicated that cell apoptosis can be obviously inhibited by overexpression of miR-21 and promoted by downregulation of miR-21. Meanwhile, western-blot results showed that p-ERK1/2 expression was elevated in miR-21 mimics group, whereas decreased in AMO group. Furthermore, the ERK1/2, a key component of MAPK signaling pathway, was knocked down, and overexpressed successfully using shRNA-ERK1/2 and overexpressing plasmids containing full length cDNA of ERK1/2, respectively. It was observed that shRNA-ERK1/2 can significantly decreased the level of miR-21 expression, while overexpression of ERK1/2 can up-regulate expression of miR-21. As further confirmation, Eca109 cells were treated with gradient concentration of U0126, a kind of MEK inhibitor, and expression of miR-21 was subsequently examined. It was found that U0126 can significantly decreased endogenous expression of miR-21. In parallel, U0126 decreased cell proliferation, migration and increased the apoptosis in Eca109 cells, with the expression of miR-21 being reduced significantly in U0126 group as compared with control groups. Our findings indicated that miR-21 promoted the proliferation, migration and inhibited apoptosis of Eca109 cells through activating ERK1/2/MAPK pathway, and that targeting miR-21 could be a promising therapeutic strategy in ESCC.

Association of the plasma riboflavin levels and riboflavin transporter (C20orf54) gene statuses in Kazak esophageal squamous cell carcinoma patients.

To evaluate the association of the plasma riboflavin level in Kazak esophageal cancer patients and their riboflavin transporter (C20orf54) gene statuses. Plasma riboflavin levels were detected by high performance liquid chromatography in Kazak patients with esophageal squamous cell carcinoma (ESCC) and healthy controls. C20orf54 mRNA and protein expression were analyzed by real-time fluorogenic quantitative polymerase chain reaction and immunohistochemistry in samples from 61 ESCC patients consisting of both tumor and normal tissue, respectively. C20orf54 mRNA expression was decreased in ESCC (0.279 ± 0.102) than in normal counterpart tissue (0.479 ± 0.287; P = 0.049) significantly. Tumors exhibited low C20orf54 protein expression (42.6, 26.2, 18.0 and 13.1% for no C20orf54 staining, weak staining, medium staining and strong staining, respectively), which was significantly lower than that in the normal mucous membrane (13.1, 26.2, 41.0 and 19.7% for no C20orf54 staining, weak staining, medium staining and strong staining, respectively). Defective expression of C20orf54 in tumor cells was significantly associated with poor differentiation. However, other parameters such as depth of invasion and lymph node metastasis had no significant relationship with C20orf54 expression. The average blood concentration of riboflavin was 2.6468 ± 1.3474 ng/ml in ESCC patients lower than control group (4.2960 ± 3.2293 ng/ml, P = 0.015). A positive correlation of plasma riboflavin levels with defective expression of C20orf54 protein was found in ESCC patients (F = 8.626; P = 0.038). Defective expression of C20orf54 is associated with the development of Kazak esophageal squamous cell carcinoma and this may represent a mechanism underlying the decreased plasma riboflavin levels in ESCC.

LMP gene promoter hypermethylation is a mechanism for its down regulation in Kazak's esophageal squamous cell carcinomas.

Abnormal hypermethylation of CpG islands not only associated with tumor suppressor genes can lead to repression of gene expression, but also contribute to escape of the tumor from immune surveillance and contribute significantly to tumorigenesis. In the present study, we studied the hypermethylation of low molecular-weight protein (LMP) gene and its regulation on protein expression in biopsies from resected tissues from Kazak's esophageal squamous cell carcinoma (ESCC) patients and their neighboring normal tissues. LMP2 and LMP7 genes promoter region methylation sequences were maped in esophageal cancer cell line Eca109 by bisulfite-sequencing PCR and quantitative detection of methylated DNA from 30 pairs of Kazak's ESCC and adjacent normal tissues by MassARRAY (Sequenom, San Diego, CA, USA) and LMP2 and LMP7 protein expression were analyzed with immunohistochemistry. In Eca109, we identified 6 CG sites methylated from all of 22 CpG sites of LMP7 gene. However, no methylation was found for LMP2. The analysis of the data resulted from the quantitative analysis of single CpG site methylation by Sequenom MassARRAY platform, has shown that the methylation level between two groups CpG sites (CpG_5, CpG_9, CpG_20, CpG_21 and CpG_20) from CpG_1, CpG_2, CpG_3, CpG_4, CpG_5, CpG_6, CpG_7, CpG_8, CpG_9, CpG_10.11, CpG_12.13.14, CpG_15.16.17.18, CpG_19, CpG_20, CpG_21 and CpG_22 significant differences between ESCC and neighboring normal tissues. The analysis of methylation level of whole target CpG fragment indicated that the methylation level of LMP7 was significant higher in ESCC (0.0517 ± 0.0357) than in neighboring normal tissues (0.0380 ± 0.0214, P < 0.05). there was a tendency of decreasing the LMP7 proteins expression as the increasing the methylation level of LMP7 gene promoter regions (F = 7.69, P = 0.041). The LMP7 gene promoter methylation and protein downregulation were correlated at high extent in Kazakh's ESCC patients, and may explain the epigenetic regulation on gene expression.

Viral load of HPV 16/18 in esophageal squamous cell carcinoma in three ethnic groups living in Xinjiang Autonomous Region, China.

To investigate the viral load of human papillomavirus (HPV) in esophageal squamous cell carcinoma (ESCC) patients from three ethnic groups in Xinjiang. Using Gp5+/Gp6+ consensus primers, the prevalence of HPV DNA was examined in 253 paraffin-embedded ESCC samples. The presence and viral load of HPV 16 and HPV 18 were detected in Kazakhs, Uygurs and Hans using type-specific primers by quantitative real-time PCR (qRT-PCR). Among the 253 ESCC samples, 52 cases were positive for HPV DNA, all the 52 positive cases displayed HPV 16 infection, and six of the 52 cases were co-infected by HPV 16 and 18. HPV 16-positive rate and viral load were higher in lesions, and was inversely correlated with differentiation grades. However, there was no statistic significance among different differentiation grades. Also, there were no significant difference between detection rates of HPV types, viral load and age, gender, ethnic group, and lymph node metastasis. HPV 16 and HPV 18 genotypes could simultaneously be detected in ESCC specimens in three main ethnic groups in Xinjiang. The viral load of HPV 16 is higher in the ESCC lesions, and is inversely correlated with the differentiation grades. These observations reinforce the suggestion that HPV infection may involved in ESCC carcinogenesis; however, high prevalence or viral load of HPV infection does not seem to be related with high incidence of ESCC in Kazakhs, which may be the one element among the multiple risk factors contributing to ESCC.

Clinicopathological pattern and Annexin A2 and Cdc42 status in patients presenting with differentiation and lymphnode metastasis of esophageal squamous cell carcinomas.

Annexin A2 and Cdc42 were identified by 2-dimensional electrophoresis (2-DE) and MALDI-TOF-MS between esophageal squamous cell carcinomas (ESCC) and corresponding normal esophagus mucosa in our previous study. To assess clinico-pathological pattern and Annexin A2 and Cdc42 status with respect to cell differentiation and lymphnode metastasis in patients with ESCC. The expression of Annexin A2 and Cdc42 in 22 pairs of fresh ESCC and matched tissues were detected by qRT-PCR and western blot, respectively. And it was further confirmed by immunohistochemistry with 175 pairs of formalin-fixed, paraffin-embedded ESCC. Results showed that Annexin A2 expression was significantly down-regulated, and Cdc42 was up-regulated in ESCC compared to matched control on both mRNA and protein level (P < 0.05), which was in accordance with our previous results on proteomics data. Additionally, Annexin A2 and Cdc42 expression was significantly correlated with lymphoid node metastasis (P < 0.05) and pathological differentiation (P < 0.05). Taken together, we proposed that the aberrant expression of Annexin A2 and Cdc42 played a role in carcinogenesis, differentiation and metastasis of ESCC, which implied its potential target for clinical biomarkers in differentiation and lymph node metastasis.

Revealing the metabonomic variation of EC using ¹H-NMR spectroscopy and its association with the clinicopathological characteristics.

In this study, (1)H NMR-based metabonomics has been applied to investigate esophageal cancer metabolic signatures in plasma and urine, purpose of assessing the diagnostic potential of this approach and gaining novel insights into esophageal cancer metabolism and systemic effects. Plasma and urine samples from esophageal cancer patients (n = 108) and a control healthy group (n = 40) were analyzed by Nuclear Magnetic Resonance (NMR) spectroscopy (600 MHz), and their spectral profiles subjected to Orthogonal Projections to Latent Structures (OPLS-DA) for multivariate statistics. Potential metabolic biomarkers were identified using data base comparisons used for examining the significance of metabolites. Compared to healthy controls, esophageal cancer plasma had higher levels of dimethylamine, α-glucose, ß-glucose, citric acid, together with lower levels of Leucine, alanine, isoleucine, valine, glycoprotein, lactate, acetone, acetate, choline, isobutyrate, unsaturated lipid, VLDL, LDL, 1-methylhistidine; Compared to healthy controls, esophageal cancer urine had higher levels of Mannitol, glutamate, γ-propalanine, phenylalanine, acetate, allantoin, pyruvate, tyrosine, ß-glucose and guinolinate, together with lower levels of N-acetylcysteine, valine, dihydrothymine, hippurate, methylguanidine, 1-methylnicotin- amide and Citric acid; Very good discrimination between cancer and control groups was achieved by multivariate modeling of plasma and urinary profiles. (1)H NMR-based metabolite profiling analysis was shown to be an effective approach to differentiating between patients with EC and healthy subjects. Good sensitivity and selectivity were shown by using the metabolite markers discovered to predict the classification of samples from the healthy control group and the patients with the disease. Plasma and urine metabolic profiling may have potential for early diagnosis of EC and may enhance our understanding of its mechanisms.

The status of phosphorylated p38 in esophageal squamous cell carcinoma.

The p38 mitogen-activated protein kinase (MAPK) is a member of the MAPK family, which is initially found to be activated by stress stimuli, proinflammatory cytokines, and growth factors. However, its role in the pathogenesis of esophageal squamous cell carcinoma (ESCC) is largely unkown, so we investigate the role of phosphorylated p38 (p-p38) MAPK in ESCC. First of all, in vitro cell line ECa109, SB203580 as selective inhibitor of p38, can suppress the growth of esophageal cancer cell in a dose- and time-dependent way, suggesting that ECa109 cell growth and proliferation was closely associated with p-p38. Using western-blot analysis of fresh 16 paired surgically resected ESCC and matched non-tumor adjacent tissues (NAT), we showed that p-p38 was significantly expressed higher in NAT compared to ESCC. Moreover, expressions of p-p38 were further confirmed by 162 paired of formalin-fixed paraffin-embedded (FFPE) ESCC and NAT by immunohistochemistry, the same trend result was obtained through statistical analysis that there was increased expression of p-p38 in NAT as compared with ESCC (P < 0.01), and expression of p-p38 was not significantly associated with lymph nodes metastasis (P > 0.05) and ESCC differentiation degree (P > 0.05). Taken together, all the results we obtained demonstrated that p-p38 plays a key role in the malignant transformation of ESCC.

Alpha Protein Kinase 2 Promotes Esophageal Cancer via Integrin Alpha 11.

Background: As a common disease around the world, esophageal cancer (EC) primarily includes two subclasses: esophageal adenocarcinoma and esophageal squamous cell carcinoma. Mortality has been rising over the years; hence, exploring the mechanism of EC development has become critical. Among the alpha protein kinases, alpha protein kinase 2 (ALPK2) presumably has a connection with EC, but it has never been revealed before. Methods: In this study, IHC analysis was used for ALPK2 expression quantification in ES tissues. TE-1 and Eca-109, which are both human EC cell lines, were used for in vitro analysis of cell proliferation, migration, apoptosis, and colony formation. Results: ALPK2 was found to have an abundant expression within EC tissues (P < 0.001), as well as in the two selected human EC cell lines (P < 0.05). The data showed that ALPK2 depletion suppressed EC cell proliferation, migration, and colony formation, meanwhile stimulating apoptosis (P < 0.001). The in vivo experiments also displayed inhibitory effects caused by ALPK2 depletion on EC tumorigenesis (P < 0.001). It was further validated that ALPK2 depletion made the phosphorylation of Akt and mTOR, as well as CDK6 and PIK3CA levels downregulated (P < 0.001). Mechanistically, we identified integrin alpha 11 (ITGA11) as a downstream gene of ALPK2 regulating EC. More importantly, we found that ITGA11 elevation promoted cell proliferation and migration and rescued the suppression effects caused by ALPK2 depletion (P < 0.001). Conclusions: ALPK2 promotes esophageal cancer via integrin its downstream gene alpha 11; ALPK2 can potentially act as a target for the treatment of EC.

The diagnostic significance of blood-derived circRNAs in NSCLC: Systematic review and meta-analysis.

Background: Using blood-derived circular RNAs (circRNAs) may be an efficient tool for noninvasive fluid biopsy in diagnosing non-small cell lung cancer (NSCLC). However, no relevant systemic meta-analysis has been conducted so far to support the diagnostic value of using blood-derived circRNAs in NSCLC clinically. The aim of this study is to clarify the issue through a meta-analysis. Methods: A systematic search strategy was used to search relevant literature in the databases of PubMed, Web of Science, and Cochrane Library from 2017 to 2022. The relationship between the diagnostic accuracy of circRNAs and NSCLC was analyzed. For the purpose of evaluating the quality of the literature, Quality Assessment of Diagnostic Accuracy Studies-2 (QUADAS-2) was used. Statistical analyses were assessed using Stata software (version 17.0) and META-DISC (version 1.4). Results: The meta-analysis included 1,093 patients with NSCLC and 959 controls. Results are as follows: pooled sensitivity, 0.78 (95% CI = 0.71-0.83, I2 = 71.86); pooled specificity, 0.76 (95% CI = 0.70-0.82, I2 = 70.12); pooled positive likelihood ratio (PLR), 3.3 (95% CI = 2.6-4.2, I2 = 37.56); pooled negative likelihood ratio (NLR), 0.29 (95% CI = 0.23-0.37, I2 = 64.67); diagnostic odds ratio (DOR), 11.42 (95% CI = 7.88-16.56, I2 = 99.05); area under the receiver operating characteristic curve (AUC), 0.84 (95% CI = 0.80-0.87). Based on the subgroup analysis, it appears that the heterogeneity is primarily caused by the NSCLC subgroup. Conclusion: circRNAs are highly useful diagnostic biomarkers for NSCLC in China. Further prospective studies on the diagnostic value of circRNAs should be conducted in multiple countries. Systematic review registration: https://www.crd.york.ac.uk/prospero/, identifier CRD42022323804.

Highly Expressing SCARA5 Promotes Proliferation and Migration of Esophageal Squamous Cell Carcinoma.

Background: Thrombospondin type 1 domain-containing 7A (THSD7A) was reported to play a procancer role in esophageal squamous cell carcinoma (ESCC). The aim of the study was to screen the downstream functional genes of THSD7A and explore their functions in ESCC, based on the reported research into THSD7A function and on gene microarrays. Methods: We adopted quantitative reverse-transcription polymerase chain reaction (qRT-PCR) and Celigo high-content screening (HCS) technology to screen the downstream genes of THSD7A. The expression level of target genes was examined by PCR, western blot, and immunohistochemistry (IHC). The effects of these target genes on ESCC malignant biological behavior were performed in vivo and in vitro. The Kaplan-Meier (K-M) survival analysis and Cox regression were used to analyze the prognostic significance of target genes in ESCC patients. Experiments in the literature on liver cancer (LC) were repeated to verify the functions of these genes in different tumors. We further explored the cancer-promoting mechanism of target genes in ESCC by sequencing of the genes' exons. Results: Scavenger receptor class A member 5 (SCARA5) was proved to be the downstream driving gene of THSD7A. SCARA5 promoted cell proliferation and migration but inhibited apoptosis in ESCC. IHC results confirmed that SCARA5 expression in ESCC exceeded that in normal tissues. The K-M survival analysis indicated that SCARA5 expression quantity was not related to prognosis, but tumor volume and T classification were both the independent prognostic factors. Repetition of experiments in LC in the literature confirmed that SCARA5 had exactly opposite functions in EC and LC. Conclusion: SCARA5 was related to the development and occurrence of ESCC. Our findings suggested that it was a potentially diagnostic individualized therapeutic target for ESCC in the future and that its application could possibly be combined with that of upstream THSD7A gene.