Increasing serum miR-223-3p indicates the onset, severe development, and adverse prognosis of bronchiectasis: a retrospective study.

BACKGROUND: miR-223-3p has been demonstrated as a Pseudomonas aeruginosa colonization-related miRNA in bronchiectasis (BE), but its clinical value in BE has not been revealed, which is of great significance for the clinical diagnosis and monitoring of BE. This study aimed to identify a reliable biomarker for screening BE and predicting patients' outcomes. METHODS: The serum expression of miR-223-3p was compared between healthy individuals (n = 101) and BE patients (n = 133) and evaluated its potential in distinguishing BE patients. The severity of BE patients was estimated by BSI and FACED score, and the correlation of miR-223-3p with inflammation and severity of BE patients was evaluated by Pearson correlation analysis. BE patients were followed up for 3 years, and the predictive value of miR-223-3p in prognosis was assessed by logistic regression analysis. RESULTS: Significant upregulation of miR-223-3p was observed in BE patients, which significantly distinguished BE patients and showed positive correlations with C-reactive protein (CRP), procalcitonin (PCT), interleukin 6 (IL-6), and neutrophil-to-lymphocyte ratio (NLR) of BE patients. Additionally, miR-223-3p was also positively correlated with BSI and FACED scores, indicating its correlation with inflammation and severity of BE. BE patients with adverse prognoses showed a higher serum miR-223-3p level, which was identified as an adverse prognostic factor and discriminated patients with different prognoses. CONCLUSION: Increasing serum miR-223-3p can be considered a biomarker for the onset, severity, and prognosis of BE.

Post-index procedural gain in body mass index is associated with recurrent ileal pouch sinus after endoscopic or surgical therapy.

BACKGROUND: Ileal pouch-anal anastomosis has become the standard surgical procedure for patients with ulcerative colitis who require colectomy. Presacral sinus is a common complication after the surgery. Pouch sinus results from chronic anastomotic or suture line leak. An excessive gain in body mass index (BMI) has been shown to be associated with poor pouch outcome. The aim of this study was to evaluate whether an increase in BMI was associated with recurrences of chronic pouch sinus after endoscopic or surgical treatment. METHODS: All consecutive ulcerative colitis patients with an ileal pouch sinus successfully treated with either endoscopic sinusotomy or redo surgery from 2006 to 2016 were identified from our IRB approved, prospectively maintained Pouch Registry. An excessive gain in BMI was defined as an increase in BMI ≥ 10% from the baseline of the initial endoscopic or surgical treatment. The primary outcome was sinus recurrence after initial complete healing. RESULTS: This study included a total of 171 patients. Sinus recurrence was seen in 48 (28.1%) patients. A higher rate of recurrence of sinus was found in patients with BMI gain (≥ 10%) (22.9% vs. 8.9%, p = 0.01). However, the recurrence-free survival in Kaplan-Meier analysis between the BMI gain and non-BMI gain groups was not statistically significant (p = 0.10). In multivariate analysis, excessive BMI gain [odds ratio (OR) 3.0, 95% confidence interval (CI) 1.0-9.0)] and Crohn's disease of the pouch (OR 2.9, 95% CI 1.0-8.1) were independently associated with sinus recurrence. The healing rate of recurrent sinus after subsequent endoscopic or surgical treatment was comparable between those who had an excessive increase in BMI and those who maintained a relatively stable weight (63.6% vs. 70.3%, p = 0.81). However, the recurrent presacral sinus-related pouch failure rate was numerically higher in patients with an excessive BMI gain (36.4% vs. 16.2%, p = 0.31). CONCLUSIONS: An excessive gain in BMI after initial successful pouch sinus treatment is associated with an increased risk for sinus recurrence. Weight control may help decrease the risk for recurrence of pouch sinus.

Combined liver-kidney perfusion enhances protective effects of normothermic perfusion on liver grafts from donation after cardiac death.

It has been shown that combined liver-kidney normothermic machine perfusion (NMP) is able to better maintain the circuit's biochemical milieu. Nevertheless, whether the combined perfusion is superior to liver perfusion alone in protecting livers from donation after circulatory death (DCD) is unclear. We aimed to test the hypothesis and explored the mechanisms. Livers from 15 DCD pig donors were subjected to either static cold storage (group A), liver-alone NMP (group B), or combined liver-kidney NMP (group C). Livers were preserved for 6 hours and reperfused ex vivo for 2 hours to simulate transplantation or were transplanted in situ. During perfusion, group C showed an improved acid-base and biochemical environment in the circuit over group B. After reperfusion, the architecture of the liver grafts was best preserved in group C, followed by group B, then group A, as shown by the histology and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining of both hepatocytes and biliary epithelium. Ki-67 staining showed substantial hepatocyte proliferation and biliary epithelial regeneration after perfusion in group B and group C. Group C produced more bile in the reperfusion phase than those in group A and group B, with more physiological bile composition and less severe biliary epithelium injury. Von Willebrand factor-positive endothelial cells and E-selectin expression decreased in both group B and group C. Combined liver-kidney NMP not only produced more adenosine triphosphate, protected the nitric oxide signaling pathway, but also diminished oxidative stress (high mobility group box-1 protein and 8-hydroxy-2-deoxy guanosine levels) and inflammatory cytokine (IL6 and IL8) release when compared with liver-alone NMP and CS. In addition, the 7-day survival rate of liver transplant recipients was higher in group C than that in groups A and B. In conclusion, combined liver-kidney NMP can better protect DCD livers from warm ischemia and reperfusion injury probably by maintaining the stability of the internal environment and by abolishing oxidative stress injury. Liver Transplantation 24 67-79 2018 AASLD.

Low expression of c-Myc protein predicts poor outcomes in patients with hepatocellular carcinoma after resection.

BACKGROUND: Embryonic Liver Fodrin (ELF) is an adaptor protein of transforming growth factor (TGF-ß) signaling cascade. Disruption of ELF results in mislocalization of Smad3 and Smad4, leading to compromised TGF-ß signaling. c-Myc is an important oncogenic transcription factor, and the disruption of TGF-ß signaling promotes c-Myc-induced hepatocellular carcinoma (HCC) carcinogenesis. However, the prognostic significance of c-Myc in HCC is less understood METHODS: The expression of c-Myc protein and mRNA were measured by immunohistochemistry (IHC) and qRT- PCR, respectively. IHC was performed to detect TGF-ß1 and ELF expression in HCC tissues. Their relationship with clinicopathological factors and overall survival (OS) and disease free survival (DFS) were examined. RESULTS: The expression of c-Myc protein and mRNA in HCC tissues were significantly higher in HCC area than those in normal liver tissues. However, the expression were low compared with those adjacent to HCC area. c-Myc protein was independently predictive of DFS and OS, and it was negatively correlated with tumor size (P = 0.031), tumor number (P = 0.038), and recurrence (P = 0.001). Low c-Myc expression was associated with short-term recurrence and poor prognosis. The predictive value of c-Myc combined with TGF-ß1 or/and ELF was higher than that of any other single marker. Low c-Myc, high TGF-ß1 or/and low ELF expression was associated with the worst DFS and OS. CONCLUSIONS: Low expression of c-Myc protein predicts poor outcomes in patients with HCC with hepatectomy. The combination of the expression of c-Myc, TGF-ß1, and ELF can be used to accurately predict outcomes of patients with HCC.

Anti-fibrogenic Potential of Mesenchymal Stromal Cells in Treating Fibrosis in Crohn's Disease.

BACKGROUND: Intestinal fibrosis is a major complication of CD and may result in stricture formation leading to intestinal obstruction. MSCs play multiple roles in active CD and fibrosis-associated diseases. AIMS: This study was designed to investigate the role of MSCs in CD-associated intestinal fibrosis. METHODS: Intestinal fibrosis was induced over 7 weeks of enema with increasing doses of TNBS and assessed by Masson's trichrome staining. Transcriptome sequencing and gene set enrichment analysis were conducted to reveal the transcriptome changes among groups at the mRNA level. Immunofluorescence assays were used to validate the role of EMT in intestinal fibrosis. Quantitative real-time PCR and immunohistochemistry analyses were performed to clarify the association between the anti-fibrogenic properties of MSCs and the immune microenvironment. Western blotting was used to verify the potential signaling pathways. RESULTS: Fibrotic tissue accumulation and inflammatory cell infiltration were detected in the colon tissue after TNBS induction treatment. Prophylactic MSCs treatment inhibited colon shortening, while therapeutic treatment decreased colon weight. Prophylactic treatment with MSCs inhibited the accumulation of fibrotic tissue, the expression of fibrotic proteins and EMT. Therapeutic MSCs treatment reversed the established intestinal fibrosis and reduced EMT. The secretion of the fibrogenic factors IL-1beta, IL-6 and IL-13 was down-regulated after both MSCs treatment approaches, while IL-10, an anti-fibrogenic factor, was up-regulated. Both MSCs therapies inhibited the expression of TGF-beta and the phosphorylation of Smad2 and Smad3 after TNBS induction. CONCLUSION: MSCs exert anti-fibrogenic activity against CD-associated fibrosis by regulating the inflammatory environment, inhibiting the TGF-beta/Smad signaling pathway and ameliorating EMT.

miR-17-5p suppresses cell proliferation and invasion by targeting ETV1 in triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is the malignancy with the worst outcome among all breast cancer subtypes. We reported that ETV1 is a significant oncogene in TNBC tumourigenesis. Consequently, investigating the critical regulatory microRNAs (miRNAs) of ETV1 may be beneficial for TNBC targeted therapy. METHODS: We performed in situ hybridization (ISH) and immunohistochemistry (IHC) to detect the location of miR-17-5p and ETV1 in TNBC patient samples, respectively. miR-17-5p expression in TNBC tissues and cell lines was assessed by quantitative real-time PCR (qRT-PCR). ETV1 expression was evaluated by qRT-PCR, western blotting and IHC. Cell Counting Kit-8 (CCK-8), colony formation, Transwell and wound closure assays were utilized to determine the TNBC cell proliferation and migration capabilities. In vivo tumour metastatic assays were performed in a zebra fish model. RESULTS: The abundance of miR-17-5p was significantly decreased in TNBC cell lines and clinical TNBC tissues. The miR-17-5p expression levels were closely correlated with tumour size (P < 0.05) and TNM stage (P < 0.05). By contrast, the expression of ETV1 was significantly up-regulated in TNBC cell lines and tissues. There is an inverse correlation between the expression status of miR-17-5p and ETV1 (r = -0.28, P = 3.88 × 10-3). Luciferase reporter assay confirmed that ETV1 was a direct target of miR-17-5p. Forced expression of miR-17-5p in MDA-MB-231 or BT549 cells significantly decreased ETV1 expression and suppressed cell proliferation, migration in vitro and tumour metastasis in vivo. However, rescuing the expression of ETV1 in the presence of miR-17-5p significantly recovered the cell phenotype. High miR-17-5p expression was associated with a significantly favourable prognosis, in either the ETV1-positive or ETV1-negative groups (log-rank test, P < 0.001; P < 0.001). Both univariate and multivariate analyses showed that miR-17-5p and ETV1 were independent risk factors in the prognosis of TNBC patient. CONCLUSIONS: Our data indicate that miR-17-5p acts as a tumour suppressor in TNBC by targeting ETV1, and a low-abundance of miR-17-5p may be involved in the pathogenesis of TNBC. These findings indicate that miR-17-5p may be a therapeutic target for TNBC.

MicroRNA-129-5p inhibits hepatocellular carcinoma cell metastasis and invasion via targeting ETS1.

MiR-129-5p is deregulated in various human cancers and has been associated with hepatocellular carcinoma (HCC) progression. However, the underlying mechanisms of miR-129-5p involvement in the development and progression of HCC and the effects of miR-129-5p deregulation on the clinical characteristics observed in HCC patients remain poorly understood. We therefore investigated the correlation between low miR-129-5p expression and vascular invasion, intrahepatic metastasis, and poor patient survival. Ectopic restoration of miR-129-5p expression in HCC cells suppressed cellular migration and invasion and the expression of v-ets erythroblastosis virus E26 oncogene homolog 1 (ETS1), while inhibition of endogenous miR-129-5p caused an increase in these parameters. We identified the ETS1 gene as a novel direct target of miR-129-5p. SiRNA-mediated ETS1 knockdown rescued the effects of anti-miR-129-5p inhibitor in HCC cell lines, while the effects of miR-129-5p overexpression were partially phenocopied in the knockdown model. In addition, miR-129-5p levels inversely correlated with those of ETS1 in HCC cells and tissues. Taken together, our findings indicate an important role for miR-129-5p in the molecular etiology of invasive HCC and suggest that miR-129-5p could have potential therapeutic applications in HCC.

The prognostic value of combined TGF-ß1 and ELF in hepatocellular carcinoma.

BACKGROUND: Tumor suppression of Transforming Growth Factor (TGF-ß) signaling pathway requires an adaptor protein, Embryonic Liver Fodrin (ELF). Disruption of ELF expression resulted in miscolocalization of Smad3 and Smad4, then disruption of TGF-ß signaling. However, the prognostic significance of ELF for hepatocellular carcinoma (HCC) hasn't been clarified. This study aimed to investigate whether measuring both TGF-ß1 and ELF provides a more powerful predictor for HCC prognosis than either marker alone. METHODS: TGF-ß1 and ELF protein were detected by immunohistochemistry. The relationship between TGF-ß1/ELF expression and patients' clinicopathologic factors was analyzed. The association between TGF-ß1/ELF expression and disease-free survival and overall survival was analyzed by Kaplan-Meier curves, the log-rank test, and Multivariate Cox regression analyses. RESULTS: The expression of TGF-ß1 in HCC tissues was significantly higher than that in normal liver tissues. Conversely, the expression of ELF in HCC tissues declined markedly. ELF protein was correlated with HBsAg, tumor size, tumor number, TNM and recurrence. Data also indicated a significant negative correlation between ELF and TGF-ß1. Patients with high TGF-ß1 expression or/and low ELF expression appeared to have a poor postoperative disease-free survival and overall survival compared with those with low TGF-ß1 expression or/and high ELF expression. Furthermore, the predictive range of ELF combined with TGF-ß1 was more sensitive than that of either one alone. CONCLUSIONS: TGF-ß1 and ELF protein are potential and reliable biomarkers for predicting prognosis in HCC patients after hepatic resection. Our current study has demonstrated that the prognostic accuracy of testing can be enhanced by their combination.

Transducin ß-like 1 X-linked receptor 1 suppresses cisplatin sensitivity in nasopharyngeal carcinoma via activation of NF-κB pathway.

BACKGROUND: Transducin ß-like 1 X-linked receptor 1 (TBL1XR1) is an important transcriptional cofactor involved in the regulation of many signaling pathways, and is associated with carcinogenesis and tumor progression. However, the precise role of TBL1XR1 in these processes is not well understood. METHODS: We detected the expression of TBL1XR1 protein and mRNA in nasopharyngeal carcinoma (NPC) cell lines and biopsies by western blotting, real-time PCR and immunohistochemical staining (IHC). Overexpression of TBL1XR1 in NPC enhanced chemoresistance to cisplatin using two NPC cell lines in vitro and in vivo. RESULTS: TBL1XR1 was upregulated in NPC cell lines and clinical samples. The expression of TBL1XR1 was correlated with several clinicopathological factors including clinical stage, T classification, N classification and patient survival. Univariate and multivariate analysis revealed that TBL1XR1 was an independent prognostic factor for patient survival. In vitro and in vivo studies demonstrated that TBL1XR1 high expression induced resistance to cisplatin-induced apoptosis in NPC cells. Furthermore, we found that TBL1XR1 activated the NF-κB pathway and promoted transcription of genes downstream of NF-κB, especially anti-apoptotic genes. CONCLUSIONS: Upregulation of TBL1XR1 induces NPC cells resistance to cisplatin by activating the NF-κB pathway, and correlates with poor overall survival of NPC patients. TBL1XR1 has a pivotal role in NPC and could be a valuable prognostic factor as well as a novel biomarker for tailoring appropriate therapeutic regimes.

Interleukin-32 contributes to invasion and metastasis of primary lung adenocarcinoma via NF-kappaB induced matrix metalloproteinases 2 and 9 expression.

Interleukin (IL)-32 is a novel proinflammatory cytokine, which has been shown to play an important role in tumor growth and metastasis. Here, we discovered that IL-32 was aberrantly over-expressed in lung adenocarcinoma tissues and cell lines. Positive expression of IL-32 significantly correlated with the clinical staging, and lymph node and distant metastases. High expression of IL-32 was an independent indicator of poor prognosis in lung adenocarcinoma patients. Moreover, IL-32-facilitated cell migration and invasion in vitro was mediated through transactivation of the nuclear transcription factor (NF)-κB signaling pathway and subsequent upregulation of matrix metalloproteinase (MMP)-2 and MMP9 expression. These studies demonstrate that IL-32 plays a role in the tumor-associated inflammatory microenvironment and that overexpression of IL-32 contributes to invasion and metastasis in primary lung adenocarcinoma, suggesting that it may have clinical utility as a prognostic biomarker and potential target for immunotherapy in lung adenocarcinoma.

URG4 overexpression is correlated with cervical cancer progression and poor prognosis in patients with early-stage cervical cancer.

BACKGROUND: Upregulator of cell proliferation 4 (URG4) has been implicated in the oncogenesis of certain cancers. However, the correlation between URG4 expression and clinicopathological significance in human cancer remains unclear. Therefore, this study investigated its expression and clinicopathological significance in cervical cancer patients. METHODS: URG4 expression was examined using quantitative PCR (qPCR) and western blotting in normal cervical epithelial cells, cervical cancer cells, and eight matched pairs of cervical cancer tissues and adjacent noncancerous tissues from the same patient. In addition, immunohistochemistry (IHC) was used to examine URG4 expression in paraffin-embedded tissues from 167 cervical cancer patients (FIGO stages Ib1-IIa2). Statistical analyses were performed to evaluate associations between URG4 expression and prognostic and diagnostic factors. RESULTS: URG4 was significantly upregulated in the cervical cancer cell lines and tissues compared with the normal cells and adjacent noncancerous cervical tissues. IHC revealed high URG4 expression in 59 out of the 167 (35.13%) cervical cancer specimens. Its expression was significantly correlated with clinical stage (P < 0.0001), tumour size (P = 0.012), T classification (P = 0.023), lymph node metastasis (P = 0.001) and vaginal involvement (P = 0.002). Patients with high URG4 expression, particularly those who received concurrent chemotherapy and radiotherapy (P < 0.0001), showed a shorter overall survival (OS) and disease-free survival (DFS) compared to those with the low expression of this protein. Multivariate analysis revealed that URG4 expression is an independent prognostic factor for cervical cancer patients. CONCLUSIONS: Our results demonstrated that elevated URG4 protein expression is associated with a poor outcome in patients with early-stage cervical cancer. URG4 may be a novel prognostic marker and therapeutic target for the treatment of cervical cancer.

Overexpression of prostate tumor overexpressed 1 correlates with tumor progression and predicts poor prognosis in breast cancer.

BACKGROUND: Prostate tumor overexpressed 1 (PTOV1) was demonstrated to play an important role in cancer progression and was correlated with unfavorable clinical outcome. However, the clinical role of PTOV1 in cancer remains largely unknown. This study aimed to investigate the expression and clinicopathological significance of PTOV1 in breast cancer. METHODS: The mRNA and protein expression levels of PTOV1 were analyzed in 12 breast cancer cell lines and eight paired breast cancer tumors by semi-quantitative real time-PCR and western blotting, respectively. Immunohistochemistry was performed to assess PTOV1 protein expression in 169 paraffin-embedded, archived breast cancer samples. Survival analysis and Cox regression analysis were performed to investigate the clinicopathological significance of PTOV1 expression. RESULTS: Our data revealed that PTOV1 was frequently overexpressed in breast cancer cell lines compared to normal human breast epithelial cells and in primary breast cancer samples compared to adjacent noncancerous breast tissues, at both the mRNA and protein levels. Moreover, high expression of PTOV1 in breast cancer is strongly associated with clinicopathological characteristics and estrogen receptor expression status (P = 0.003). Breast cancer patients with higher PTOV1 expression had substantially shorter survival times than patients with lower PTOV1 expression (P < 0.001). Univariate and multivariate analysis revealed that PTOV1 might be an independent prognostic factor for breast cancer patients (P = 0.005). CONCLUSIONS: Our study showed that PTOV1 is upregulated in breast cancer cell lines and clinical samples, and its expression was positively associated with progression and aggressiveness of breast cancer, suggesting that PTOV1 could serve as an independent prognostic marker.

Down-regulation of Gli-1 inhibits hepatocellular carcinoma cell migration and invasion.

Glioma-associated oncogene homolog-1 (Gli-1) is considered a marker of Hedgehog pathway activation and is associated with the progression of several cancers. We have previously reported that Gli-1 was correlated with invasion and metastasis in hepatocellular carcinoma (HCC). However, the exact roles and mechanisms of Gli-1 in HCC invasion are unclear. In this study, we found that small interfering RNA mediated down-regulation of Gli-1 expression significantly suppressed adhesion, motility, migration, and invasion of both SMMC-7721 and SK-Hep1 cells. Furthermore, down-regulation of Gli-1 significantly reduced expressions and activities of both matrix metalloproteinase (MMP)-2 and MMP-9. In addition, we found that down-regulation of Gli-1 resulted in up-regulation of E-cadherin and concomitant down-regulation of Snail and Vimentin, consistent with inhibition of epithelial-mesenchymal transition (EMT). Taken together, our results suggest that down-regulation of Gli-1 suppresses HCC cell migration and invasion likely through inhibiting expressions and activations of MMP-2, 9 and blocking EMT.

miR-145 suppresses cell invasion in hepatocellular carcinoma cells: miR-145 targets ADAM17.

AIM: miR-145 is a candidate tumor suppressor miRNA. However, it is unknown whether miR-145 is involved in the invasion of hepatocellular carcinoma (HCC). Therefore, we aimed to explore the effect and mechanism of miR-145 in the control of HCC cell invasion. METHODS: HCC cell invasion was evaluated by transwell assays after transfection with pre-miR-145 or anti-miR-145. A luciferase reporter assay was used to determine whether a disintegrin and metalloprotease 17 (ADAM17) were a target of miR-145. The levels of miR-145 and ADAM17 mRNA were detected by a real-time polymerase chain reaction assay, and the level of ADAM17 protein was measured by western blot analysis. Pearson's correlation test was used to assess the correlation between ADAM17 mRNA expression and miR-145 expression in 20 HCC tissue samples. RESULTS: miR-145 was significantly downregulated in HCC tissues and cell lines. The loss of miR-145 expression was associated with the tumor-node-metastasis stage, vascular invasion and intrahepatic metastasis. The overexpression of miR-145 was able to suppress tumor MHCC-97H cell invasion, whereas the knockdown of miR-145 expression induced SMMC-7721 cell invasion. We demonstrated that miR-145 bound directly to the 3'-untranslated region of ADAM17 and inhibited the expression of ADAM17. The knockdown of ADAM17 in SMMC-7721 cells could partially reverse the effects of anti-miR-145. miR-145 expression was inversely associated with ADAM17 expression in 20 HCC tissue specimens. CONCLUSION: Our findings indicate that miR-145 could inhibit HCC cell invasion by regulating the expression of ADAM17.

Pathological characteristics of liver allografts from donation after brain death followed by cardiac death in pigs.

Donation after brain death followed by circulatory death (DBCD) is a unique practice in China. The aim of this study was to define the pathologic characteristics of DBCD liver allografts in a porcine model. Fifteen male pigs (25-30 kg) were allocated randomly into donation after brain death (DBD), donation after circulatory death (DCD) and DBCD groups. Brain death was induced by augmenting intracranial pressure. Circulatory death was induced by withdrawal of life support in DBCD group and by venous injection of 40 mL 10% potassium chloride in DCD group. The donor livers were perfused in situ and kept in cold storage for 4 h. Liver tissue and common bile duct samples were collected for hematoxylin and eosin staining, TUNEL testing and electron microscopic examination. Spot necrosis was found in hepatic parenchyma of DBD and DBCD groups, while a large area of necrosis was shown in DCD group. The apoptosis rate of hepatocytes in DBD [(0.56±0.30)%] and DBCD [(0.50 ± 0.11)%] groups was much lower than that in DCD group [(3.78±0.33)%] (P<0.05). And there was no significant difference between DBD group and DBCD group (P>0.05)). The structures of bile duct were intact in both DBD and DBCD groups, while the biliary epithelium was totally damaged in DCD group. Under electron microscope, the DBD hepatocytes were characterized by intact cell membrane, well-organized endoplasmic reticulum, mild mitochondria edema and abundant glycogens. Broken cell membrane, mild inflammatory cell infiltration and sinusoidal epithelium edema, as well as reduced glycogen volume, were found in the DBCD hepatocytes. The DCD hepatocytes had more profound cell organelle injury and much less glycogen storage. In conclusion, the preservation injury of DBCD liver allografts is much less severe than that of un-controlled DCD, but more severe than that of DBD liver allografts under electron microscope, which might reflect post-transplant liver function to some extent.

Sonic hedgehog signaling pathway induces cell migration and invasion through focal adhesion kinase/AKT signaling-mediated activation of matrix metalloproteinase (MMP)-2 and MMP-9 in liver cancer.

The aberrant activation of sonic hedgehog (SHH) pathway contributes to initiation and progression of various malignancies. However, the roles and underlying mechanisms of SHH signaling pathway in invasion and metastasis of liver cancer have not been well understood. In this study, we found that SHH signaling was activated and correlated with invasion and metastasis in hepatocellular carcinoma (HCC). Enhanced SHH signaling by recombinant human SHH N-terminal peptide (rSHH-N) promoted hepatoma cell adhesion, migration and invasion, whereas blockade of SHH signaling with SHH neutralizing antibody or cyclopamine suppressed hepatoma cell adhesion, migration and invasion. Furthermore, matrix metalloproteinase (MMP)-2 and MMP-9 expressions and activities were upregulated and downregulated by rSHH-N and SHH signaling inhibitor, respectively. The rSHH-N-mediated hepatoma cell migration and invasion was blocked by MMP-specific inhibitors or neutralizing antibodies to MMP-2 and MMP-9. In addition, phosphorylations of AKT and focal adhesion kinase (FAK) were increased and decreased by rSHH-N and SHH signaling inhibitor, respectively. Further investigations showed that activation of AKT and FAK were required for rSHH-N-mediated upregulation of MMP-2 and MMP-9, cell migration and invasion. Finally, we found that SHH protein expression was positively correlated with phosphorylatd FAK Tyr397, phosphorylatd AKT Ser473, MMP-2 and MMP-9 protein expressions in HCC samples. Taken together, our findings suggest that SHH pathway induces cell migration and invasion through FAK/AKT signaling-mediated MMP-2 and MMP-9 production and activation in liver cancer.

Phosphoglycerate dehydrogenase induces glioma cells proliferation and invasion by stabilizing forkhead box M1.

Phosphoglycerate dehydrogenase (PHGDH) is the first enzyme branching from glycolysis in the three-step serine biosynthetic pathway. Recent evidence has shown that PHGDH is amplified in human breast cancer and melanoma and plays a key role in cancer metabolism. However, PHGDH expression in glioma and a potential non-metabolic role in tumorigenesis have not been reported. We analyzed PHGDH levels in specimens from glioma patients and found that PHGDH, although negative in normal brain tissues, was highly expressed in astrocytic tumors and increasingly expressed in more aggressive cancer types. Inhibition of PHGDH expression in glioma cells downregulated the expression of VEGF, MMP-2, CHK2 and cyclin D1 and reduced glioma cell proliferation, invasion and tumorigenicity in vitro and in vivo. Interestingly, we found that the oncogenic transcription factor FOXM1 was also downregulated in PHDGH-silenced glioma cells. Using LC/LC MS analysis, we identified PHGDH as a novel binding partner of FOXM1. PHGDH interacted with and stabilized FOXM1 at the protein level, promoting the proliferation, invasion and tumorigenicity of glioma cells. Our data identified PHGDH as a potential prognostic marker of glial brain tumors and identified a non-metabolic role for PHGDH in glioma tumorigenesis, providing a novel angle of targeting the PHGDH-FOXM1 axis in future brain tumor therapy.

Analysis of the genome-wide DNA methylation profile of side population cells in hepatocellular carcinoma.

BACKGROUND: DNA methylation plays an important role in maintaining pluripotency and regulating the differentiation of stem cells, but the DNA methylation profile of stem cells in hepatocellular carcinoma (HCC) remains unclear. AIMS: To investigate the genome-wide DNA methylation profile of side population (SP) cells of HCC, a special subpopulation of cells enriched with cancer stem cells, by DNA methylation microarray analysis and to analyze the functions and signal pathways of the aberrantly methylated genes in SP cells. METHODS: Side population cells were isolated from HCC cell lines Huh7 and PLC/PRF/5 using flow cytometry, and the tumorigenicity of these SP cells was assessed in NOD/SCID mice. The genome-wide DNA methylation status of SP cells and non-SP (NSP) cells was detected and compared by DNA methylation microarray analysis. Genes with differential methylation between SP and NSP cells were further analyzed for their functions and roles in related signaling pathways. RESULTS: Subcutaneous inoculation of 1 × 10(3) SP cells yielded tumors in 60 % NOD/SCID mice, whereas no tumor was developed after the inoculation of 1 × 10(6) NSP cells. Genome-wide DNA methylation microarray analysis showed that 72 and 181 genes were hypermethylated and hypomethylated, respectively, in both Huh7 and PLC/PRF/5 SP cells as compared with their corresponding NSP cells. Analyses of signaling pathways revealed that hypermethylated and hypomethylated genes were related to four and eight pathways, respectively. CONCLUSIONS: Hepatocellular carcinoma SP cells possessed a differential DNA methylation status compared with NSP cells, and the differentially methylated genes in SP cells were involved in 12 signaling pathways. Our results provide valuable clues for further investigations in elucidating the importance of epigenetic regulation in sustaining HCC SP cells and tumorigenesis.

Expression and prognostic relevance of PRAME in primary osteosarcoma.

The preferentially expressed antigen of melanoma (PRAME), a cancer-testis antigen with unknown function, is expressed in many human malignancies and is considered an attractive potential target for tumor immunotherapy. However, studies of its expression and function in osteosarcoma have rarely been reported. In this study, we found that PRAME is expressed in five osteosarcoma cell lines and in more than 70% of osteosarcoma patient specimens. In addition, an immunohistochemical analysis showed that high PRAME expression was associated with poor prognosis and lung metastasis. Furthermore, PRAME siRNA knockdown significantly suppressed the proliferation, colony formation, and G1 cell cycle arrest in U-2OS cells. Our results suggest that PRAME plays an important role in cell proliferation and disease progression in osteosarcoma. However, the detail mechanisms of PRAME function in osteosarcoma require further investigation.

Celastrol inhibits vasculogenesis by suppressing the VEGF-induced functional activity of bone marrow-derived endothelial progenitor cells.

Bone marrow (BM)-derived endothelial progenitor cells (EPCs) play a critical role in tumor vasculogenesis because they provide both instructive (release of pro-angiogenic cytokines, such as VEGF) and structural (vessel incorporation and stabilization) functions. Celastrol, derived from Trypterygium wilfordii Hook F., a traditional Chinese medicine plant, has been studied for its antitumorigenic properties, but its mechanism of action is not well understood. The aims of this study are to investigate the effects of Celastrol on VEGF-induced functional activity of BM-EPCs and to identify any mechanisms associated with this process. Here, we show that Celastrol attenuates VEGF secretion in BM-EPCs in vitro. This attenuation, in turn, inhibits the in vitro VEGF-induced cell viability, cell-cell adhesion, cell-ECM adhesion, migration response and vascular tube formation of BM-EPCs. Additionally, Celastrol inhibits the phosphorylation of VEGFR2, endothelial nitric oxide synthase (eNOS), and Akt to attenuate cell functions. Taken together, the present study demonstrates that Celastrol decreases Akt/eNOS signaling in BM-EPCs in vitro. These findings identify novel mechanisms that regulate EPC function and may provide new insights for the medicinal use of Celastrol.