Spatially resolved transcriptomics revealed local invasion-related genes in colorectal cancer.

Objective: Local invasion is the first step of metastasis, the main cause of colorectal cancer (CRC)-related death. Recent studies have revealed extensive intertumoral and intratumoral heterogeneity. Here, we focused on revealing local invasion-related genes in CRC. Methods: We used spatial transcriptomic techniques to study the process of local invasion in four CRC tissues. First, we compared the pre-cancerous, cancer center, and invasive margin in one section (S115) and used pseudo-time analysis to reveal the differentiation trajectories from cancer center to invasive margin. Next, we performed immunohistochemical staining for RPL5, STC1, AKR1B1, CD47, and HLA-A on CRC samples. Moreover, we knocked down AKR1B1 in CRC cell lines and performed CCK-8, wound healing, and transwell assays to assess cell proliferation, migration, and invasion. Results: We demonstrated that 13 genes were overexpressed in invasive clusters, among which the expression of CSTB and TM4SF1 was correlated with poor PFS in CRC patients. The ribosome pathway was increased, while the antigen processing and presentation pathway was decreased along CRC progression. RPL5 was upregulated, while HLA-A was downregulated along cancer invasion in CRC samples. Pseudo-time analysis revealed that STC1, AKR1B1, SIRPA, C4orf3, EDNRA, CES1, PRRX1, EMP1, PPIB, PLTP, SULF2, and EGFL6 were unpregulated along the trajectories. Immunohistochemic3al staining showed the expression of STC1, AKR1B1, and CD47 was increased along cancer invasion in CRC samples. Knockdown of AKR1B1 inhibited CRC cells' proliferation, migration, and invasion. Conclusions: We revealed the spatial heterogeneity within CRC tissues and uncovered some novel genes that were associated with CRC invasion.

Transcriptional regulation of NDUFA4L2 by NFIB induces sorafenib resistance by decreasing reactive oxygen species in hepatocellular carcinoma.

Sorafenib is one a first-line therapeutic drugs for advanced hepatocellular carcinoma (HCC). However, only 30% of patients benefit from sorafenib due to drug resistance. We and other groups have revealed that nuclear factor I B (NFIB) regulates liver regeneration and carcinogenesis, but its role in drug resistance is poorly known. We found that NFIB was more upregulated in sorafenib-resistant SMMC-7721 cells compared to parental cells. NFIB knockdown not only sensitized drug-resistant cells to sorafenib but also inhibited the proliferation and invasion of these cells. Meanwhile, NFIB promoted the proliferation and invasion of HCC cells in vitro and facilitated tumor growth and metastasis in vivo. Knocking down NFIB synergetically inhibited tumor growth with sorafenib. Mechanically, gene expression profiling and subsequent verification experiments proved that NFIB could bind with the promoter region of a complex I inhibitor NDUFA4L2 and promote its transcription. Transcriptional upregulation of NDUFA4L2 by NFIB could thus inhibit the sorafenib-induced reactive oxygen species accumulation. Finally, we found that NFIB was highly expressed in HCC tissues, and high NFIB expression level was associated with macrovascular invasion, advanced tumor stage, and poor prognosis of HCC patients (n = 156). In summary, we demonstrated that NFIB could transcriptionally upregulate NDUFA4L2 to enhance both intrinsic and acquired sorafenib resistance of HCC cells by reducing reactive oxygen species induction.

Hepatocyte-Specific Knock-Out of Nfib Aggravates Hepatocellular Tumorigenesis via Enhancing Urea Cycle.

Nuclear Factor I B (NFIB) has been reported to promote tumor growth, metastasis, and liver regeneration, but its mechanism in liver cancer is not fully elucidated. The present study aims to reveal the role of NFIB in hepatocellular carcinogenesis. In our study, we constructed hepatocyte-specific NFIB gene knockout mice with CRISPR/Cas9 technology (Nfib-/-; Alb-cre), and induced liver cancer mouse model by intraperitoneal injection of DEN/CCl4. First, we found that Nfib-/- mice developed more tumor nodules and had heavier livers than wild-type mice. H&E staining indicated that the liver histological severity of Nfib-/- group was more serious than that of WT group. Then we found that the differentially expressed genes in the tumor tissue between Nfib-/- mice and wild type mice were enriched in urea cycle. Furthermore, ASS1 and CPS1, the core enzymes of the urea cycle, were significantly upregulated in Nfib-/- tumors. Subsequently, we validated that the expression of ASS1 and CPS1 increased after knockdown of NFIB by lentivirus in normal hepatocytes and also promoted cell proliferation in vitro. In addition, ChIP assay confirmed that NFIB can bind with promoter region of both ASS1 and CPS1 gene. Our study reveals for the first time that hepatocyte-specific knock-out of Nfib aggravates hepatocellular tumor development by enhancing the urea cycle.

Idiopathic acute superior mesenteric venous thrombosis after renal transplantation: A case report.

BACKGROUND: Acute superior mesenteric venous thrombosis (MVT) is a rare condition associated with a high mortality rate. The treatment strategy for MVT is clinically challenging due to its insidious onset and rapid development, especially when accompanied by kidney transplantation. CASE SUMMARY: Here we present a rare case of acute MVT developed 3 years after renal transplantation. A 49-year-old patient was admitted with acute abdominal pain and diagnosed as MVT with intestinal necrosis. An emergency exploratory laparotomy was performed to remove the infarcted segment of the bowel. Immediate systemic anticoagulation was also initiated. During the treatment, the patient experienced bleeding, anastomotic leakage, and sepsis. However, after aggressive treatment was administered, all thrombi were completely resolved, and the patient recovered with his renal graft function unimpaired. CONCLUSION: The present case suggests that accurate diagnosis and timely surgical treatment are important to improve the survival rate of MVT patients. Bleeding with anastomotic fistula needs to be treated with caution because of grafts. Also, previously published cases of mesenteric thrombosis after renal transplantation were reviewed.

Development Trend of Infectious Disease Hospitals in China 2002-2018: A Longitudinal Analysis of National Survey Data.

BACKGROUND: Infectious disease hospitals (IDHs) play very important roles in the battle against the infectious disease. The present study aims to systematically analyze the development trends and possible problems of IDHs in China. METHODS: Most of the data came from the China Health Statistics Yearbook 2003-2019. Joinpoint Regression Model was used to analyze the development trends of IDHs between 2002 and 2018. RESULTS: From 2002 to 2018, the number of IDHs in China increased from 126 to 167, with an average annual percent change (AAPC) of 1.82%. The ratio of nurses to beds increased from 0.38 to 0.46 with the AAPC of 0.88%, and average business housing area per bed increased with an AAPC of 1.97%. The percentage of liabilities to total assets increased year by year and the percentage of medical business costs to total expenditure decreased. The segmented trend of daily visits per physician from 2014 to 2018 was stable, and the segmented trend of daily inpatients per physician from 2012 to 2018 decreased significantly. In 2017, the rates of surgical inpatients leaving the hospital without the doctor's advice and surgical inpatients mortality were higher than 2016. CONCLUSION: Although the development of IDHs was generally good in China, the scale of IDHs was generally small, the ability to respond to major emergencies was weak, the problem of irrational resource allocation was still prominent, and the operation of IDHs was facing a dilemma.

LncRNA-LALR1 upregulates small nucleolar RNA SNORD72 to promote growth and invasion of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most prevalent cancers and currently the second leading cause of cancer-related mortality worldwide. One recent study reported that lncRNA-LALR1 promotes liver regeneration, the role and underlying mechanisms of lncRNA-LALR1 in HCC remain largely unknown. In this study, we demonstrated that lncRNA-LALR1 was significantly upregulated in HCC tissues compared with adjacent tissues and high expression of lncRNA-LALR1 was associated with advanced TNM stage, poor differentiation, and distant metastasis. RNA Fluorescence in situ hybridization analysis showed lncRNA-LALR1 was expressed not only in cytoplasm but also in nucleolus. Knockdown of lncRNA-LALR1 obviously inhibited HCC cells growth and invasion in vivo and in vitro. Besides, transcriptomic analysis and subsequent confirmation revealed that lncRNA-LALR1 upregulated small nucleolar RNA SNORD72 via binding with SNORD72 and stabilized ID2 mRNA. SNORD72 was overexpressed in HCC tissues and enhanced HCC cells proliferation, colony formation and invasion. Overexpression of SNORD72 could also stabilize ID2 mRNA and rescue the inhibitory effect of silencing lncRNA-LALR1. In conclusion, lncRNA-LALR1 is highly expressed in HCC and promotes tumor growth and invasion by upregulating SNORD72 to stabilize ID2 mRNA, implying that lncRNA-LALR1 might be a novel target for intervention of HCC.

KCNK levels are prognostic and diagnostic markers for hepatocellular carcinoma.

Two-pore-domain (KCNK, K2P) K+ channels are transmembrane protein complexes that control the flow of ions across biofilms, which underlie many essential cellular functions. Because KCNK family members are known to contribute to tumorigenesis in various types of cancer, we hypothesized that they might be differentially expressed in hepatocellular carcinoma (HCC) cells as compared to healthy tissue and serve as diagnostic or prognostic biomarkers. We tested this hypothesis through bioinformatic analyses of publicly available data for the expression of various KCNK subunits in HCC. We observed reduced expression of KCNK2, KCNK15, and KCNK17 in liver cancer, as well as overexpression of KCNK9, all of which correlated with a better prognosis for HCC patients per survival analyses. Moreover, ROC curves indicated that KCNK2, KCNK9, KCNK15, and KCNK17 levels could be used as a diagnostic biomarker for HCC. Finally, our western blot and qRT-PCR results were consistent with those obtained from bioinformatic analyses. Taken together, these results suggest that KCNK2, KCNK9, KCNK15, and KCNK17 could serve as potential diagnostic and prognostic biomarkers of HCC.

NAMPT and NAPRT, Key Enzymes in NAD Salvage Synthesis Pathway, Are of Negative Prognostic Value in Colorectal Cancer.

Nicotinamide adenine dinucleotide (NAD) is a profoundly important cofactor in redox reactions. Nicotinamide phosphoribosyltransferase (NAMPT) and nicotinate phosphoribosyltransferase (NAPRT) are key enzymes for NAD salvage biosynthesis pathway, which reciprocally synthesize NAD to supply the main source of NAD biosythesis. However, the prognostic value of NAMPT and NAPRT in colorectal cancer (CRC) remains largely unknown. Our present study detected NAMPT and NAPRT protein expression in cancer and adjacent tissues from 261 CRC using immunohistochemical staining. We found that high expression of NAMPT or NAPRT was associated with vascular invasion, invasion depth and advanced TNM stage in CRC. High expression of NAMPT or NAPRT predicts short overall survival and disease-free survival time in CRC patients, which were further confirmed by public datasets. Furthermore, positive correlation between expression of NAMPT and NAPRT was revealed in CRC tissues and cell lines. NAPRThigh/NAMPThigh patients tended to have the shortest survival time. Using the TCGA RNA-sequencing data, we showed that gene amplification, mutation, and methylation of NAPRT are more common than NAMPT. On the other hand, NAMPT gene might be targeted by more miRNAs. Finally, genes that are correlated with NAPRT or NAMPT are enriched in different pathways. In conclusion, we found that high expression of NAMPT or NAPRT predicts poor prognosis of CRC patients, but the regulatory mechanism might be distinct from each other.

Chromatin accessibility changes are associated with enhanced growth and liver metastasis capacity of acid-adapted colorectal cancer cells.

The acidic extracellular microenvironment, namely acidosis, is a biochemical hallmark of solid tumors. However, the tumorigenicity, metastatic potential, gene expression profile and chromatin accessibility of acidosis-adapted colorectal cancer cells remain unknown. The colorectal cancer cell SW620 was cultured in acidic medium (pH 6.5) for more than 3 months to be acidosis-adapted (SW620-AA). In comparison to parental cells, SW620-AA cells exhibit enhanced tumorigenicity and liver metastatic potential in vivo. Following mRNA and lncRNA expression profiling, we validated that OLMF1, NFIB, SMAD9, DGKB are upregulated, while SESN2, MAP1B, UTRN, PCDH19, IL18, LMO2, CNKSR3, GXYLT2 are downregulated in SW620-AA cells. The differentially expressed mRNAs were significantly enriched in DNA remodeling-associated pathways including HDACs deacetylate histones, SIRT1 pathway, DNA methylation, DNA bending complex, and RNA polymerase 1 chain elongation. Finally, chromatin accessibility evaluation by ATAC-sequencing revealed that the differentially opened peaks were enriched in pathways such as small cell lung cancer, pathways in cancer, ErbB signaling, endometrial cancer, and chronic myeloid leukemia, which were mainly distributed in intergenic regions and introns. These results suggest that the chromatin accessibility changes are correlated with enhanced growth and liver metastasis capacity of acid-adapted colorectal cancer cells.

Sorafenib-induced acute-on-chronic liver failure in a patient with hepatocellular carcinoma after transarterial chemoembolization and radiofrequency ablation: A case report.

Comprehensive treatments together with sorafenib provide a survival benefit for patients with advanced hepatocellular carcinoma (HCC). Acute-on-chronic liver failure (ACLF) is an increasingly recognized distinct entity; however, only few cases induced by sorafenib have been reported to date. We herein present a rare case of ACLF in a patient under treatment with sorafenib. A 63-year-old woman who suffered from hepatitis B for 10 years was admitted to our hospital. A computed tomography (CT) scan led to a diagnosis of HCC in the V/VIII hepatic segment, with invasion of the middle hepatic and the right portal veins. The patient received multidisciplinary treatment, including transarterial chemoembolization, radiofrequency ablation and sorafenib. Two months after sorafenib treatment, the patient was readmitted due to progressive jaundice and ACLF was diagnosed by liver biopsy shortly thereafter. Although aggressive treatment was administered, the patient succumbed to the disease following rapid deterioration of her clinical condition. The majority of patients with HCC have underlying liver disease and combination therapy with sorafenib should be administered with caution, as it may increase the risk of severe hepatotoxicity. The present case suggests that patients treated with sorafenib may require a dose reduction following interventional treatment.

The significance of Delphian lymph node in papillary thyroid cancer.

AIM: To investigate the clinical significance of the DLN metastasis in papillary thyroid cancer (PTC). METHODS: A total of 231 PTC patients who underwent first surgical treatment in the Department of Hand and Neck Surgery of Zhejiang Cancer Hospital from January 2013 to June 2014 were enrolled. The relationship between Delphian lymph node (DLN) metastasis and patient age, gender, tumor size, tumor number, unilateral or bilateral, capsular invasion, pretracheal and paratracheal node metastasis, and lateral node metastasis was analyzed. RESULTS: Within 231 cases, 69 showed DLN, but only 19 (8.23%) were found with metastasis. In the univariate analysis, DLN metastasis was significantly associated with tumor size (P = 0.023), capsular invasion (P = 0.001), pretracheal or paratracheal node metastasis (P = 0.003) and lateral node metastasis (P = 0.001), while there were no significant correlation between DLN metastasis and gender (P = 0.976), age (P = 0.976), tumor number (P = 0.234) and unilateral or bilateral (P = 0.724). In the multivariate analysis, capsular invasion was an independent risk factor of DLN metastasis (P < 0.05, odds ratio = 10.15). CONCLUSION: Capsular invasion is an independent risk factor of DLN metastasis and DLN metastasis could be used as a predictor of lateral node metastasis. The dissection of DLN in PTC patients is recommended and lateral lymph node should be evaluated for patients with DLN positive.

Vascular invasion is an independent prognostic factor for distant recurrence-free survival in papillary thyroid carcinoma: a matched-case comparative study.

OBJECTIVE: It is still unclear whether the clinicopathological and outcome characteristics of vascular invasion (VI) (+) papillary thyroid carcinoma (PTC) differ from VI (-) PTC. This study aims to establish distinguishing features of patients with VI (+) and VI (-) PTC and to investigate the biological and clinical aggressiveness of the disease in these patient groups. DESIGN: A matched-case comparative study. METHODS: 412 patients (VI (+) PTC study group n=103, and VI (-) PTC control group n=309). These two patient groups were matched 1:3 for variables of age, gender, histological variants, tumour/node/metastasis (TNM) staging and approximate duration of follow-up. Clinicopathological factors and prognosis were compared across the two patient groups. RESULTS: The median age at the time of diagnosis was 42.0 years, and 68.9% were females. Across the patient groups, the incidence of tumour multifocality in patients with VI (+) PTC was slightly higher than in those with VI (-) PTC (p=0.049). In addition, when undergoing more aggressive therapy regimens patients with VI (+) PTC showed decreased distant recurrence-free survival (DRFS), but not regional recurrence-free survival (RRFS) and disease-specific survival (DSS) compared with patients who were VI (-). VI was found to be an independent predictor of DRFS, combined with tumour size >3 cm and total thyroidectomy. CONCLUSIONS: VI was an independent risk factor for DRFS, necessitating the need for appropriate postoperative treatment and careful follow-up.

Inhibition of mTOR promotes hyperthermia sensitivity in SMMC-7721 human hepatocellular carcinoma cell line.

The mammalian target of rapamycin (mTOR) is a critical mediator of the phosphoinositide 3-kinase/protein kinase B/mTOR signaling pathway, and mTOR activity is induced following heat shock. Thermotherapy is used to treat hepatocellular carcinoma (HCC). However, the role of mTOR in modulating thermosensitivity in HCC has yet to be elucidated. In the present study, the antisense plasmid pEGFP-C1-mTOR was transfected into SMMC-7721 cells, and the expression levels of mTOR were analyzed by reverse transcription-polymerase chain reaction and western blot analysis. The thermal responses of the transfected cells were also examined. The results revealed that SMMC-7721 cells were sensitive to heat treatment, and cell viability was significantly inhibited following hyperthermia treatment (P<0.01). The mRNA and protein expression levels of mTOR decreased post-transfection. Cell proliferation, colony-forming ability and motility were all significantly decreased following hyperthermia treatment in the transfected cells. Flow cytometry analysis demonstrated that apoptosis was significantly increased following treatment (P<0.01). The number of cells in S phase was increased, and the cell cycle was arrested in S phase. In conclusion, inhibition of mTOR increased the thermosensitivity of SMMC-7721 cells by increasing cellular apoptosis and inducing S phase arrest.

Clinicopathological features and prognosis of familial papillary thyroid carcinoma--a large-scale, matched, case-control study.

OBJECTIVE: It remains controversial whether or not the aggressiveness of familial nonmedullary thyroid cancer (FNMTC) differs from sporadic carcinoma. The aim of this study was to determine the clinicopathological features and prognosis of FNMTC. DESIGN: A matched-case comparative study. METHODS: Three hundred and seventy-two patients with familial papillary thyroid carcinoma (FPTC) were enrolled as the study group, and another 372 patients with sporadic PTC were enrolled as controls and matched for gender, age, tumour/node/metastasis (TNM) staging and approximate duration of follow-up. We compared the differences in the clinicopathological features and prognosis between the subgroups. RESULTS: Compared with sporadic PTC, patients with FPTC were more likely to present tumour multicentricity, bilateral growth and a concomitant nodular goitre (P < 0·05). In papillary thyroid microcarcinoma (PTMC), a higher recurrence rate was noted in patients with a family history of PTC, and this remained independently predictive on multivariate analysis. The patients with FPTC in the second generation showed an earlier age of onset, more frequent Hashimoto's thyroiditis and a higher recurrence rate than the first generation, while the first-generation offspring of patients had a higher incidence of nodular goitre than the second generation. CONCLUSIONS: The presence of familial history in PTC indicates an increase in biological aggressiveness, and patients in the second generation may exhibit the 'genetic anticipation' phenomenon. At present, the available data are not sufficient to support a more aggressive approach for FPTC. However, a family history of PTC is an independent risk factor for recurrence in patients with PTMC.

Elevated expression of ASCL2 is an independent prognostic indicator in lung squamous cell carcinoma.

AIMS: ASCL2, a basic helix-loop-helix (bHLH) transcription factor, is putatively involved in tumour progression. This study aimed to evaluate ASCL2 expression level in non-small-cell carcinoma (NSCLC) and assess its prognostic value for patients. METHODS: ASCL2 protein expression was detected by immunohistochemistry (IHC cohort) in 79 cases of squamous cell carcinoma (SCC) and 67 cases of adenocarcinoma (AC). Kaplan-Meier analysis and Cox regression analysis were performed to evaluate the prognostic significance of ASCL2. The same analyses were conducted in a cohort (n=790) from The Cancer Genome Atlas database (TCGA) to validate the expression pattern and prognostic value of ASCL2. RESULTS: ASCL2 expression levels were significantly increased in SCC compared with normal lung tissue (p<0.001) and AC (p=0.008). High ASCL2 expression was associated with advanced tumour-node-metastasis (TNM) stage (p=0.023) and worse differentiation status (p=0.001) in SCC, but a positive correlation between ASCL2 expression level and advanced TNM stage (p=0.016) was observed in AC. Kaplan-Meier analysis showed that ASCL2 was prognostic in SCC (p=0.004) but not in AC (p=0.183). Multivariable Cox regression analysis indicated that elevated expression of ASCL2 was an independent prognostic factor (HR 2.764; p=0.030) in SCC patients. The expression pattern and prognostic significance of ASCL2 in SCC and AC were validated using the TCGA cohort. CONCLUSIONS: Elevated expression of ASCL2 may identify an aggressive subgroup in SCC and serve as an independent prognostic indicator in these patients.

Parathyroid carcinoma: A report of six cases with a brief review of the literature.

Parathyroid carcinoma (PC) is an uncommon endocrine malignancy and constitutes a rare cause of hyperparathyroidism. The current study presents the clinical features, laboratory findings, sensitivity of imaging modalities, surgical treatment and the long-term outcome of six patients, who were diagnosed with PC and treated in the Department of Head and Neck Surgery, Zhejiang Province Cancer Hospital (Hangzhou, China) over 13 years (February 1999-January 2012). Pre-operative recognition and intraoperative identification of this rare endocrine malignancy is extremely important, but require a high index of clinical suspicion. The primary treatment is surgical en bloc resection of the tumor and any involved surrounding structures, and it is of great importance, as the prognosis depends on the initial surgery. Radiation therapy and chemotherapy showed no evidence of effectiveness on PC, although certain data show a decreased risk of localized disease recurrence with the addition of radiation therapy. The prognosis of PC is variable and post-operative parathyroid hormone levels that do not decrease often indicate a poor prognosis or presence of other metastases.

Synchronous double primary cancer - intrahepatic cholangiocarcinoma with bone metastases and thyroid carcinoma: A case report.

There is a low incidence of multiple primary cancer, particularly when the cancer is synchronous. The present report presents a case of synchronous double primary malignancies. A 58-year-old woman was admitted to Ling Nan Hospital (Guangzhou, China) complaining of pain in the left hip. X-ray revealed an osteolytic lesion and further examination indicated the presence of double primary cancer, consisting of hepatic cholangiocarcinoma and thyroid carcinoma. Biopsy of the osteolytic lesion showed a metastatic adenocarcinoma of unknown origin. Subsequently, final diagnosis was confirmed by I-131 scan and liver lesion biopsy. The patient received positive multidisciplinary treatments and survived for 9 months following diagnosis. The results of the present case suggest that multiplicity of primary malignancy is not necessarily an indicator of poor prognosis, as long as effective diagnosis and adequate disease management are achieved.

Clinical Characteristics Related to Central Lymph Node Metastasis in cN0 Papillary Thyroid Carcinoma: A Retrospective Study of 916 Patients.

Background. Papillary thyroid carcinoma (PTC) is a form of thyroid cancer with high risk of cervical lymph node metastasis. Aim. The aim of this study was to investigate the incidence and the predictive factors for occult ipsilateral central lymph node (CLN) metastasis in the patients with papillary thyroid carcinoma. Methods. A total of 916 PTC patients (1017 lesions) undergoing central lymph node dissection in our hospital from 2005 to 2011 were enrolled. The relationship between CLN metastasis and clinical factors such as gender, age, tumor size, tumor number, capsule invasion, and tumor location was analyzed. Results. Occult CLN metastasis was observed in 52.41% (533/1017) of PTC lesions, respectively. Multivariate analysis showed that age ≤ 35 years, tumor size > 1.5 cm, present capsule invasion/extracapsular invasion, and tumor located in upper/middle pole/whole lobe were risk factors of CLN metastasis. Conclusions. Tumor located in upper/middle pole/whole lobe, less than 35 years old, tumor size > 1.5 cm, and present capsule invasion/extracapsular invasion were risk factors of CLN metastasis. We recommend performing ipsilateral prophylactic CLN dissection in cN0 PTC patients.

[Effects of Bmi1 gene on endothelial cells promoting glioma stem cell-like phenotype].

OBJECTIVE: To explore the effects of B-cell specific Maloney leukemia virus integration site 1 (Bmi1) gene on endothelial cells promoting glioma stem cell (GSC)-like phenotype. METHODS: Glioblastoma cell line GL261 and brain micro-vessel endothelial cell line b.END3 were used. Transwell co-culture system, limit dilution assay, xenograft, real-time polymerase chain reaction (PCR), Western blot, fluorescence activating cell sorter (FACS) and gene knock-down assay were used to determine the GSC-like phenotype and Bmi1 gene expression in glioma cells. RESULTS: Compared with the control of GL261 cell alone, (1) more and larger tumor spheres formed after co-culturing with endothelial cells (62.5% ± 1.5% vs 25.0% ± 4.6% at 40 cells/well, P = 0.000). Xenografts generated by GL261 cells with b.END3 cells appeared earlier and were larger than that by GL261 cells alone ((0.798 ± 0.297) cm(3) vs (0.362 ± 0.123) cm(3), P = 0.000); (2) CD133 positive glioma cells increased after co-culturing with endothelial cells (8.48% ± 0.78% vs 4.81% ± 0.37%, P = 0.000); (3) the expression of Bmi1 in co-cultured glioma cells was up-regulated at mRNA level (2.72 ± 0.18 vs 1.00 ± 0.15, P = 0.000) and at protein level; (4) the above phenomenon was attenuated when Bmi1 gene expression was inhibited by siRNA in glioma cells, CD133 positive portion of Bmi1-knockdown GL261 cells co-culturing with b.END3 cells decreased than that of wildtype GL261 cells (0.34% ± 0.21% vs 1.70% ± 0.69%, P = 0.025). CONCLUSION: Endothelial cells promote GSC-like phenotype by up-regulating the expression of Bmi1 in glioma cells.

ß-Catenin/POU5F1/SOX2 transcription factor complex mediates IGF-I receptor signaling and predicts poor prognosis in lung adenocarcinoma.

Cancer stem-like cells (CSLC) are crucial in tumor initiation and progression; however, the underlying mechanism for the self-renewal of cancer cells remains undefined. In the study, immunohistochemical analysis of specimens freshly excised from patients with lung adenocarcinoma showed that high expression of insulin-like growth factor I receptor (IGF-IR) in lung adenocarcinoma cells was positively correlated with the expressions of cancer stem cell markers CD133 and aldehyde dehydrogenase 1 family member A1 (ALDH1A1). IGF-IR activation enhanced POU class 5 homeobox 1 (POU5F1) expression on human lung adenocarcinoma stem-like cells (LACSLC) through PI3K/AKT/GSK3ß/ß-catenin cascade. POU5F1 could form a novel complex with ß-catenin and SOX2 to bind Nanog promoter for transcription to maintain self-renewal of LACSLCs, which was dependent on the functional IGF-IR. Genetic and pharmacologic inhibition of IGF-IR abrogated LACSLC capabilities for self-renewal and tumorigenicity in vitro. In an in vivo xenograft tumor model, knockdown of either IGF-IR or POU5F1 impeded tumorigenic potentials of LACSLCs. By analyzing pathologic specimens excised from 200 patients with lung adenocarcinoma, we found that colocalization of highly expressed IGF-IR with ß-catenin and POU5F1 predicted poor prognosis. Taken together, we show that IGF-IR-mediated POU5F1 expression to form a complex with ß-catenin and SOX2 is crucial for the self-renewal and oncogenic potentials of LACSLCs, and the integrative clinical detection of the expressions of IGF-IR, ß-catenin, and POU5F1 is indicatory for predicting prognosis in the patients of lung adenocarcinoma.