Bevacizumab induces ferroptosis and enhances CD8+ T cell immune activity in liver cancer via modulating HAT1 and increasing IL-9.

Bevacizumab is a recombinant humanized monoclonal immunoglobulin (Ig) G1 antibody of VEGF, and inhibits angiogenesis and tumor growth in hepatocellular carcinoma (HCC). Ferroptosis, a new form of regulated cell death function independently of the apoptotic machinery, has been accepted as an attractive target for pharmacological intervention; the ferroptosis pathway can enhance cell immune activity of anti-PD1 immunotherapy in HCC. In this study we investigated whether and how bevacizumab regulated ferroptosis and immune activity in liver cancer. Firstly, we performed RNA-sequencing in bevacizumab-treated human liver cancer cell line HepG2 cells, and found that bevacizumab significantly altered the expression of a number of genes including VEGF, PI3K, HAT1, SLC7A11 and IL-9 in liver cancer, bevacizumab upregulated 37 ferroptosis-related drivers, and downregulated 17 ferroptosis-related suppressors in particular. We demonstrated that bevacizumab triggered ferroptosis in liver cancer cells by driving VEGF/PI3K/HAT1/SLC7A11 axis. Clinical data confirmed that the expression levels of VEGF were positively associated with those of PI3K, HAT1 and SLC7A11 in HCC tissues. Meanwhile, we found that bevacizumab enhanced immune cell activity in tumor immune-microenvironment. We identified that HAT1 up-regulated miR-143 targeting IL-9 mRNA 3'UTR in liver cancer cells; bevacizumab treatment resulted in the increase of IL-9 levels and its secretion via VEGF/PI3K/HAT1/miR-143/IL-9 axis, which led to the inhibition of tumor growth in vivo through increasing the release of IL-2 and Granzyme B from activated CD8+ T cells. We conclude that in addition to inhibiting angiogenesis, bevacizumab induces ferroptosis and enhances CD8+ T cell immune activity in liver cancer. This study provides new insight into the mechanisms by which bevacizumab synergistically modulates ferroptosis and CD8+ T cell immune activity in liver cancer.

Aristolochic acids-hijacked p53 promotes liver cancer cell growth by inhibiting ferroptosis.

Aristolochic acids (AAs) have been identified as a significant risk factor for hepatocellular carcinoma (HCC). Ferroptosis is a type of regulated cell death involved in the tumor development. In this study, we investigated the molecular mechanisms by which AAs enhanced the growth of HCC. By conducting bioinformatics and RNA-Seq analyses, we found that AAs were closely correlated with ferroptosis. The physical interaction between p53 and AAs in HepG2 cells was validated by bioinformatics analysis and SPR assays with the binding pocket sites containing Pro92, Arg174, Asp207, Phe212, and His214 of p53. Based on the binding pocket that interacts with AAs, we designed a mutant and performed RNA-Seq profiling. Interestingly, we found that the binding pocket was responsible for ferroptosis, GADD45A, NRF2, and SLC7A11. Functionally, the interaction disturbed the binding of p53 to the promoter of GADD45A or NRF2, attenuating the role of p53 in enhancing GADD45A and suppressing NRF2; the mutant did not exhibit the same effects. Consequently, this event down-regulated GADD45A and up-regulated NRF2, ultimately inhibiting ferroptosis, suggesting that AAs hijacked p53 to down-regulate GADD45A and up-regulate NRF2 in HepG2 cells. Thus, AAs treatment resulted in the inhibition of ferroptosis via the p53/GADD45A/NRF2/SLC7A11 axis, which led to the enhancement of tumor growth. In conclusion, AAs-hijacked p53 restrains ferroptosis through the GADD45A/NRF2/SLC7A11 axis to enhance tumor growth. Our findings provide an underlying mechanism by which AAs enhance HCC and new insights into p53 in liver cancer. Therapeutically, the oncogene NRF2 is a promising target for liver cancer.

Inhibition of USP7 enhances CD8+ T cell activity in liver cancer by suppressing PRDM1-mediated FGL1 upregulation.

Lymphocyte activation gene 3 (LAG3), an immune checkpoint molecule expressed on activated T cells, functions as a negative regulator of immune responses. Persistent antigen exposure in the tumor microenvironment results in sustained LAG3 expression on T cells, contributing to T cell dysfunction. Fibrinogen-like protein 1 (FGL1) has been identified as a major ligand of LAG3, and FGL1/LAG3 interaction forms a novel immune checkpoint pathway that results in tumor immune evasion. In addition, ubiquitin-specific peptidase 7 (USP7) plays a crucial role in cancer development. In this study we investigated the role of USP7 in modulation of FGL1-mediated liver cancer immune evasion. We showed that knockdown of USP7 or treatment with USP7 inhibitor P5091 suppressed liver cancer growth by promoting CD8+ T cell activity in Hepa1-6 xenograft mice and in HepG2 or Huh7 cells co-cultured with T cells, whereas USP7 overexpression produced the opposite effect. We found that USP7 upregulated FGL1 in HepG2 and Huh7 cells by deubiquitination of transcriptional factor PR domain zinc finger protein 1 (PRDM1), which transcriptionally activated FGL1, and attenuated the CD8+ T cell activity, leading to the liver cancer growth. Interestingly, USP7 could be transcriptionally stimulated by PRDM1 as well in a positive feedback loop. P5091, an inhibitor of USP7, was able to downregulate FGL1 expression, thus enhancing CD8+ T cell activity. In an immunocompetent liver cancer mouse model, the dual blockade of USP7 and LAG3 resulted in a superior antitumor activity compared with anti-LAG3 therapy alone. We conclude that USP7 diminishes CD8+ T cell activity by a USP7/PRDM1 positive feedback loop on FGL1 production in liver cancer; USP7 might be a promising target for liver cancer immunotherapy.

Aspirin modulates succinylation of PGAM1K99 to restrict the glycolysis through NF-κB/HAT1/PGAM1 signaling in liver cancer.

Aspirin as a chemopreventive agent is able to restrict the tumor growth. Phosphoglycerate mutase 1 (PGAM1) is a key enzyme of glycolysis, playing an important role in the development of cancer. However, the underlying mechanism by which aspirin inhibits the proliferation of cancer cells is poorly understood. This study aims to identify the effects of aspirin on modulating PGAM1 enzymatic activities in liver cancer. Here, we found that aspirin attenuated the PGAM1 succinylation to suppress the PGAM1 enzymatic activities and glycolysis in hepatoma cells. Mechanically, aspirin remarkably reduced the global succinylation levels of hepatoma cells, including the PGAM1 succinylation, which led to the block of conversion from 3-phosphoglycerate (3-PG) to 2-phosphoglycerate (2-PG) in cells. Interestingly, RNA-seq analysis identified that aspirin could significantly decrease the levels of histone acetyltransferase 1 (HAT1), a writer of PGAM1 succinylation, in liver cancer. As a target of aspirin, NF-κB p65 could effectively up-regulate the expression of HAT1 in the system, resulting in the increase of PGAM1 enzymatic activities. Moreover, we observed that the PGAM1-K99R mutant failed to rescue the aspirin-induced inhibition of PGAM1 activities, glycolysis, and proliferation of hepatoma cells relative to PGAM1-WT. Functionally, aspirin down-regulated HAT1 and decreased the PGAM1 succinylation levels in the tumor tissues from mice treated with aspirin in vivo. Thus, we conclude that aspirin modulates PGAM1K99 succinylation to restrict the PGAM1 activities and glycolysis through NF-κB p65/HAT1/PGAM1 signaling in liver cancer. Our finding provides new insights into the mechanism by which aspirin inhibits glycolysis in hepatocellular carcinoma.

Aspirin triggers ferroptosis in hepatocellular carcinoma cells through restricting NF-κB p65-activated SLC7A11 transcription.

A number of studies have shown that aspirin, as commonly prescribed drug, prevents the development of hepatocellular carcinoma (HCC). Ferroptosis as a dynamic tumor suppressor plays a vital role in hepatocarcinogenesis. In this study we investigated whether aspirin affected ferroptosis in liver cancer cells. RNA-seq analysis revealed that aspirin up-regulated 4 ferroptosis-related drivers and down-regulated 5 ferroptosis-related suppressors in aspirin-treated HepG2 cells. Treatment with aspirin (4 mM) induced remarkable ferroptosis in HepG2 and Huh7 cells, which was enhanced by the ferroptosis inducer erastin (10 µM). We demonstrated that NF-κB p65 restricted ferroptosis in HepG2 and Huh7 cells through directly binding to the core region of SLC7A11 promoter and activating the transcription of ferroptosis inhibitor SLC7A11, whereas aspirin induced ferroptosis through inhibiting NF-κB p65-activated SLC7A11 transcription. Overexpression of p65 rescued HepG2 and Huh7 cells from aspirin-induced ferroptosis. HCC patients with high expression levels of SLC7A11 and p65 presented lower survival rate. Functionally, NF-κB p65 blocked the aspirin-induced ferroptosis in vitro and in vivo, which was attenuated by erastin. We conclude that aspirin triggers ferroptosis by restricting NF-κB-activated SLC7A11 transcription to suppress the growth of HCC. These results provide a new insight into the mechanism by which aspirin regulates ferroptosis in hepatocarcinogenesis. A combination of aspirin and ferroptosis inducer may provide a potential strategy for the treatment of HCC in clinic.

Heat shock protein family A member 8 serving as a co-activator of transcriptional factor ETV4 up-regulates PHLDA2 to promote the growth of liver cancer.

Heat shock protein family A member 8 (HSPA8) participates in the folding or degradation of misfolded proteins under stress and plays critical roles in cancer. In this study, we investigated the function of HSPA8 in the development of liver cancer. By analyzing the TCGA transcriptome dataset, we found that HSPA8 was upregulated in 134 clinical liver cancer tissue samples, and positively correlated with poor prognosis. IHC staining showed the nuclear and cytoplasmic localization of HSPA8 in liver cancer cells. Knockdown of HSPA8 resulted in a decrease in the proliferation of HepG2 and Huh-7 cells. ChIP-seq and RNA-seq analysis revealed that HSPA8 bound to the promoter of pleckstrin homology-like domain family A member 2 (PHLDA2) and regulated its expression. The transcription factor ETV4 in HepG2 cells activated PHLDA2 transcription. HSPA8 and ETV4 could interact with each other in the cells and colocalize in the nucleus. From a functional perspective, we demonstrated that HSPA8 upregulated PHDLA2 through the coactivating transcription factor ETV4 to enhance the growth of liver cancer in vitro and in vivo. From a therapeutic perspective, we identified both HSPA8 and PHDLA2 as novel targets in the treatment of HCC. In conclusion, this study demonstrates that HSPA8 serves as a coactivator of ETV4 and upregulates PHLDA2, leading to the growth of HCC, and is a potential therapeutic target in HCC treatment.

IFN-α inhibits HBV transcription and replication by promoting HDAC3-mediated de-2-hydroxyisobutyrylation of histone H4K8 on HBV cccDNA minichromosome in liver.

The epigenetic modification of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) plays a crucial role in cccDNA transcription and viral persistence. Interferon-α (IFN-α) is a pivotal agent against HBV cccDNA. However, the mechanism by which IFN-α modulates the epigenetic regulation of cccDNA remains poorly understood. In this study, we report that IFN-α2b enhances the histone deacetylase 3 (HDAC3)-mediated de-2-hydroxyisobutyrylation of histone H4 lysine 8 (H4K8) on HBV cccDNA minichromosome to restrict the cccDNA transcription in liver. By screening acetyltransferases and deacetylases, we identified that HDAC3 was an effective restrictor of HBV transcription and replication. Moreover, we found that HDAC3 was able to mediate the de-2-hydroxyisobutyrylation of H4K8 in HBV-expressing hepatoma cells. Then, the 2-hydroxyisobutyrylation of histone H4K8 (H4K8hib) was identified on the HBV cccDNA minichromosome, promoting the HBV transcription and replication. The H4K8hib was regulated by HDAC3 depending on its deacetylase domain in the system. The low level of HDAC3 and high level of H4K8hib were observed in the liver tissues from HBV-infected human liver-chimeric mice. The levels of H4K8hib on HBV cccDNA minichromosome were significantly elevated in the liver biopsy specimens from clinical hepatitis B patients, which was consistent with the high transcriptional activity of cccDNA. Strikingly, IFN-α2b effectively facilitated the histone H4K8 de-2-hydroxyisobutyrylation mediated by HDAC3 on the HBV cccDNA minichromosome in primary human hepatocytes and hepatoma cells, leading to the inhibition of HBV transcription and replication. Our finding provides new insights into the mechanism by which IFN-α modulates the epigenetic regulation of HBV cccDNA minichromosome.

PRMT5 confers lipid metabolism reprogramming, tumour growth and metastasis depending on the SIRT7-mediated desuccinylation of PRMT5 K387 in tumours.

The protein arginine methyltransferase 5 (PRMT5), which is highly expressed in tumour tissues, plays a crucial role in cancer development. However, the mechanism by which PRMT5 promotes cancer growth is poorly understood. Here, we report that PRMT5 contributes to lipid metabolism reprogramming, tumour growth and metastasis depending on the SIRT7-mediated desuccinylation of PRMT5 K387 in tumours. Mass spectrometric analysis identified PRMT5 lysine 387 as its succinylation site. Moreover, the desuccinylation of PRMT5 K387 enhances the methyltransferase activity of PRMT5. SIRT7 catalyses the desuccinylation of PRMT5 in cells. The SIRT7-mediated dessuccinylation of PRMT5 lysine 387 fails to bind to STUB1, decreasing PRMT5 ubiquitination and increasing the interaction between PRMT5 and Mep50, which promotes the formation of the PRMT5-Mep50 octamer. The PRMT5-Mep50 octamer increases PRMT5 methyltransferase activity, leading to arginine methylation of SREBP1a. The symmetric dimethylation of SREBP1a increases the levels of cholesterol, fatty acid, and triglyceride biogenesis in the cells, escaping degradation through the ubiquitin-proteasome pathway. Functionally, the desuccinylation of PRMT5 K387 promotes lipid metabolism reprogramming, tumour growth and metastasis in vitro and in vivo in tumours.

Aspirin modulates 2-hydroxyisobutyrylation of ENO1K281 to attenuate the glycolysis and proliferation of hepatoma cells.

Aspirin can efficiently inhibit the glycolysis and proliferation of cancer cells, however, the underlying mechanism is poorly understood. Here, we report that aspirin attenuates the glycolysis and proliferation of hepatoma cells through modulating the levels of lysine 2-hydroxyisobutyrylation (Khib) of enolase 1 (ENO1). We found that aspirin decreased the levels of glucose consumption and lactate production in hepatoma cells. Moreover, 4 mM aspirin reduced the activities of ENO1, a key enzyme of glycolysis, and decreased the levels of ENO1 Khib in the cells. Interestingly, we identified that 4 mM aspirin could decrease the levels of Khib on many proteins by using pan Khib antibody in the cells. Interestingly, the activities of ENO1 could be rescued by the transient overexpression of ENO1, but not by ENO1 mutant (K281R). Moreover, we identified that the C646, an inhibitor of p300 which is a writer of Khib, could reduce the levels of ENO1 Khib, resulting in the decrease of ENO1 activities. The treatment with PDTC, an inhibitor of NF-κB which is a target of aspirin, could work well as C646 in the cells. Both of aspirin and C646 (or PDTC) displayed a stronger effect than the single treatment in the system. Functionally, ENO1, but not ENO1 mutant (K281R), could rescue the aspirin-induced inhibition of proliferation of liver cancer cells in vitro, suggesting that ENO1K281 is involved in the aspirin-mediated inhibition of liver cancer. Our finding provides new insights into the mechanism by which aspirin attenuates the glycolysis and proliferation of hepatoma cells.

Plasma miR-6089 as potential diagnostic biomarker for retinoblastoma.

OBJECTIVE: The purpose of this study was to screen target miRNA related to RB and explore the expression levels of target miRNA in RB and its potential value of diagnosis. METHODS: The Affymetrix GeneChip miRNA 4.0 Array was used to screen the differential miRNAs in the plasma of 5 RB patients before and after intravenous chemotherapy, and the most significant down-regulated miRNA was selected for target miRNA. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) is used to verify the expression levels of plasma target miRNA in 30 RB patients. Then, qRT-PCR was performed to further verify the expression of target miRNA in plasma of RB patients and RB tumor tissues. Finally, receiver-operating-characteristic (ROC) curve and the area under the ROC curve (AUC) were used to evaluate the diagnostic power of plasma target miRNA. RESULTS: The miRNA Array obtain 8 core miRNAs, 1 up-regulated and 7 down-regulated, of which miR-6089 was the most significantly down-regulated. Plasma miR-6089 levels were significantly up-regulated in RB patients. Besides, in RB tumor tissues, miR-6089 levels were also obviously up-regulated. After intravenous chemotherapy, the expression of plasma miR-6089 was significantly decreased. Furthermore, ROC curve analysis showed that miR-6089 in the plasma had a good sensitivity and specificity for distinguishing RB from the healthy control group. CONCLUSIONS: MiR-6089 may be considered as a novel potential diagnostic biomarker for RB. TRIAL REGISTRATION NUMBER: ChiCTR2000040154; date of registration: 2020/11/22; retrospectively registered.

Aspirin inhibits the proliferation of hepatoma cells through controlling GLUT1-mediated glucose metabolism.

Aspirin can efficiently inhibit liver cancer growth, but the mechanism is poorly understood. In this study, we report that aspirin modulates glucose uptake through downregulating glucose transporter 1 (GLUT1), leading to the inhibition of hepatoma cell proliferation. Our data showed that aspirin significantly decreased the levels of reactive oxygen species (ROS) and glucose consumption in hepatoma cells. Interestingly, we identified that GLUT1 and HIF1α could be decreased by aspirin. Mechanically, we demonstrated that the -1008/-780 region was the regulatory element of transcriptional factor NF-κB in GLUT1 promoter by luciferase report gene assays. PDTC, an inhibitor of NF-κB, could suppress the expression of GLUT1 in HepG2 and H7402 cells, followed by affecting the levels of ROS and glucose consumption. CoCl2-activated HIF1α expression could slightly rescue the GLUT1 expression inhibited by aspirin or PDTC, suggesting that aspirin depressed GLUT1 through targeting NF-κB or NF-κB/HIF1α signaling. Moreover, we found that GLUT1 was highly expressed in clinical HCC tissues relating to their paired adjacent normal tissues. Importantly, we observed that high level of GLUT1 was significantly correlated with the poor relapse-free survival of HCC patients by analysis of public data. Functionally, overexpression of GLUT1 blocked the PDTC-induced or aspirin-induced inhibition of glucose metabolism in HepG2 cells. Conversely, aspirin failed to work when GLUT1 was stably knocked down in the cells. Administration of aspirin could depress the growth of hepatoma cells through controlling GLUT1 in vitro and in vivo. Thus, our finding provides new insights into the mechanism by which aspirin depresses liver cancer.

Clinical diagnosis and treatment of intraorbital wooden foreign bodies.

PURPOSE: The intraorbital wooden foreign body is often misdiagnosed or missed on computed tomog- raphy (CT) scan, due to the invisible or unclear images. The residual foreign bodies often occur during surgical removal. The clinical manifestations, imaging features and treatment of intraorbital wooden foreign bodies were discussed in this study. METHOD: We retrospectively analyzed 14 cases of intraorbital wooden foreign bodies managed at our hospital between January 2007 and May 2015. All patients underwent orbital CT examination before surgery, and surgery was performed under general anesthesia with orbital wound debridement and suture, as well as exploration and removal of wooden foreign bodies. RESULTS: At first, 11 cases underwent removal of foreign bodies, including 1 case with incomplete removal and then receiving a secondary surgery. Foreign bodies were not found in three cases with preoperative misdiagnosis and orbital MRI found residual foreign bodies in the orbit. Operations were performed via primary wound approach in eight cases, conjunctival approach in two cases, and anterior orbitotomy in four cases. Postoperatively, one case was complicated with eye injuries, three cases with ocular muscle injuries, eight cases with visual loss, and eight cases with orbital abscess. The length of foreign bodies ranged from 1.8 cm to 11.0 cm. The maximum of four foreign bodies were removed at the same time. CONCLUSION: Because the imaging of orbital wooden foreign bodies is complex and varied, MRI should be combined when they are invisible on CT scan. At the same time injuries trajectory and clinical mani- festations of patients should be taken into account. Surgical exploration should be extensive and thor- ough, and foreign bodies and orbital abscess must be cleared.

Hepatocellular carcinoma-associated mesenchymal stem cells promote hepatocarcinoma progression: role of the S100A4-miR155-SOCS1-MMP9 axis.

UNLABELLED: Cancer-associated mesenchymal stem cells (MSCs) play a pivotal role in modulating tumor progression. However, the interactions between liver cancer-associated MSCs (LC-MSCs) and hepatocellular carcinoma (HCC) remain unreported. Here, we identified the presence of MSCs in HCC tissues. We also showed that LC-MSCs significantly enhanced tumor growth in vivo and promoted tumor sphere formation in vitro. LC-MSCs also promoted HCC metastasis in an orthotopic liver transplantation model. Complementary DNA (cDNA) microarray analysis showed that S100A4 expression was significantly higher in LC-MSCs compared with liver normal MSCs (LN-MSCs) from adjacent cancer-free tissues. Importantly, the inhibition of S100A4 led to a reduction of proliferation and invasion of HCC cells, while exogenous S100A4 expression in HCC cells resulted in heavier tumors and more metastasis sites. Our results indicate that S100A4 secreted from LC-MSCs can promote HCC cell proliferation and invasion. We then found the expression of oncogenic microRNA (miR)-155 in HCC cells was significantly up-regulated by coculture with LC-MSCs and by S100A4 ectopic overexpression. The invasion-promoting effects of S100A4 were significantly attenuated by a miR-155 inhibitor. These results suggest that S100A4 exerts its effects through the regulation of miR-155 expression in HCC cells. We demonstrate that S100A4 secreted from LC-MSCs promotes the expression of miR-155, which mediates the down-regulation of suppressor of cytokine signaling 1, leading to the subsequent activation of STAT3 signaling. This promotes the expression of matrix metalloproteinases 9, which results in increased tumor invasiveness. CONCLUSION: S100A4 secreted from LC-MSCs is involved in the modulation of HCC progression, and may be a potential therapeutic target. (HEPATOLOGY 2013).

Mesenchymal stem cells from primary breast cancer tissue promote cancer proliferation and enhance mammosphere formation partially via EGF/EGFR/Akt pathway.

Mesenchymal stem cells (MSCs) play a critical role in promoting cancer progression. However, it is not clear whether MSCs are located in breast cancer tissues and correlated with tumor proliferation. The aim of this study was to investigate the presence of MSCs in breast cancer tissues and evaluate their interactions with cancer cells. We successfully isolated and identified MSCs from primary breast cancer tissues. Breast cancer-associated MSCs (BC-MSCs) showed homogenous immunophenotype, and possessed tri-lineage differentiation potential (osteoblast, adipocyte, and chondrocyte). When co-transplanted with cancer cells in a xenograft model in vivo, BC-MSCs significantly increased the volume and weight of tumors. We observed that BC-MSCs stimulated mammosphere formation in the transwell co-culture system in vitro. This effect was significantly suppressed by the EGF receptor inhibitor. We verified that BC-MSCs could secrete EGF and activate cancer cell's EGF receptors. Furthermore, our data showed that EGF derived from BC-MSCs could promote mammosphere formation via the PI3K/Akt signaling pathway. Our results confirmed the presence of MSC in primary breast cancer tissues, and they could provide a favorable microenvironment for tumor cell growth in vivo, partially enhance mammosphere formation via the EGF/EGFR/Akt pathway.

Epimorphin promotes human hepatocellular carcinoma invasion and metastasis through activation of focal adhesion kinase/extracellular signal-regulated kinase/matrix metalloproteinase-9 axis.

UNLABELLED: The high incidence rate of hepatocellular carcinoma (HCC) is mainly the result of frequent metastasis and tumor recurrence. Unfortunately, the underlying molecular mechanisms driving HCC metastasis are still not fully understood. It has been demonstrated that tumor stroma cells contribute to primary tumor growth and metastasis. Within the HCC environment, activated hepatic stellate cells (HSCs) can release a number of molecules and enhance cancer cell proliferation and invasiveness in a paracrine manner. Here, for the first time, we demonstrate that epimorphin (EPM; also called syntaxin-2), an extracellular protein, is strongly elevated in activated HSCs within tumor stroma. We show that knockdown of EPM expression in HSCs substantially abolishes their effects on cancer cell invasion and metastasis. Ectopic expression of EPM in HCC cancer cells enhances their invasiveness; we demonstrate that the cells expressing EPM have markedly increased metastasis potential. Furthermore, EPM-mediated invasion and metastasis of cancer cells is found to require up-regulation of matrix metalloproteinase-9 (MMP-9) through the activation of focal adhesion kinase (FAK)/extracellular signal-regulated kinase (ERK) axis. CONCLUSION: Our results show that EPM, secreted by activated HSCs within HCC stroma, promotes invasion and metastasis of cancer cells by activating MMP-9 expression through the FAK-ERK pathway.

Epidemiology and risk factors of retinoblastoma in Chongqing area.

AIM: To investigate the risk factors of retinoblastoma in Chongqing and offer the bases for scientific measures. METHODS: One hundred and thirty-three retinoblastoma patients were taken as case group, and 133 children were taken as control group in 1:1 ratio. The relevant factors were collected from a questionnaire survey which was made by our research group. First, Chi-square and t-test were used to discuss the various factors, and then the logistic regression analysis was made by statistics software SPSS17.0 based on the result of 266 people. RESULTS: Single factor analysis results showed the differences between the two groups were statistically significant (P<0.05) in 17 factors which were education level of their parents, occupation of their parents, exposure to harmful chemicals of their parents 6mo before pregnancy, the history of pelvic inflammatory disease of mother before pregnancy, childbearing history of their parents, pregnant age of their parents, the medication history of father 6 mo before pregnancy, living place of their parents, the pollution living place of mother, hobbies of their parents. Multivariate analysis showed that the living place of parents, mother who feed pets before pregnancy, and exposure to harmful chemicals of father before pregnancy were the independent risk factors of retinoblastoma (P<0.05). CONCLUSION: The living place of parents, mother who feed pets before pregnancy, exposure to harmful chemicals of father before pregnancy were the risk factors of retinoblastoma which affects the occurrence of retinoblastoma in a certain extent.

[Migration and differentiation of human bone marrow mesenchymal stem cells in the rat brain].

Bone marrow mesenchymal stem cells (MSCs) are multipotent tissue stem cells that can be induced in vitro to differentiate into a variety of cells such as osteoblasts, chondrocytes and adipocytes. MSCs are useful vehicles for both cell and gene therapy for a variety of diseases. Here, we injected human MSCs with enhanced green fluorescent protein (EGFP) into the striatum of Parkinson disease (PD) rat and examined their survival, migration, differentiation, and the behavior changes in PD rats, which will provide a theoretical foundation and technical method for clinic PD therapy by stem cells. The results showed that human bone marrow MSCs can survive in rat brain for a long time (exceeding 70 d). MSCs were found in multiple areas of the rat brain including the striatum, the corpus callosum, contralateral cortex and even the brain vascular wall. Immunocytochemical staining suggested that implanted cells expressed human neurofilament (NF), neuron-specific enolase (NSE) and glial fibrillary acid protein (GFAP). At the same time, remission in abnormal behavior of the PD rats appeared. Rotation scores decreased gradually from 8.86+/-2.09 r/min pre-transplantation to 4.87+/-2.06 r/min 90 d post-transplantation (statistic result showed P<0.05).

A dual AP-1 and SMAD decoy ODN suppresses tissue fibrosis and scarring in mice.

The transforming growth factor-ß (TGF-ß) signaling pathway promotes tissue fibrosis and scarring through SMAD (small mothers against decapentaplegic)-dependent and SMAD-independent mechanisms. However, inhibition of SMAD-mediated signal transduction alone induces an excessive inflammatory response that impairs the antifibrotic effects of TGF-ß inhibitors. In this study, we designed and characterized a dual-functional transcription activator protein 1 (AP-1) and SMAD decoy oligodeoxynucleotide, antifibrosis oligodeoxynucleotide 4 (AFODN4) in vitro and in vivo. AFODN4 binds directly to recombinant AP-1 and SMAD with high affinity. AFODN4 significantly inhibited the DNA-binding and transcriptional activities of both AP-1 and SMAD, as well as the production of fibrotic mediators stimulated by TGF-ß1 or TGF-ß2 in L929 murine fibroblasts. Local administration of AFODN4 significantly inhibited fibrosis associated with acute dermal wounds in mice. Intriguingly, AFODN4 inhibited AP-1-mediated production of proinflammatory mediators, which can be caused by blockage of SMAD alone in vitro and in vivo. Collectively, these findings suggest that dual inhibition of SMAD and AP-1 signaling by AFODN4 is a useful strategy for the development of new antifibrotic agents.

SPINDLIN1 promotes cancer cell proliferation through activation of WNT/TCF-4 signaling.

SPINDLIN1, a new member of the SPIN/SSTY gene family, was first identified as a gene highly expressed in ovarian cancer cells. We have previously shown that it is involved in the process of spindle organization and chromosomal stability and plays a role in the development of cancer. Nevertheless, the mechanisms underlying its oncogenic role are still largely unknown. Here, we first showed that expression of SPINDLIN1 is upregulated in clinical tumors. Ectopic expression of SPINDLIN1 promoted cancer cell proliferation and activated WNT/T-cell factor (TCF)-4 signaling. The Ser84 and Ser99 amino acids within SPINDLIN1 were further identified as the key functional sites in WNT/TCF-4 signaling activation. Mutation of these two sites of SPINDLIN1 abolished its effects on promoting WNT/TCF-4 signaling and cancer cell proliferation. We further found that Aurora-A could interact with and phosphorylate SPINDLIN1 at its key functional sites, Ser84 and Ser99, suggesting that phosphorylation of SPINDLIN1 is involved in its oncogenic function. Collectively, these results suggest that SPINDLIN1, which may be a novel substrate of the Aurora-A kinase, promotes cancer cell growth through WNT/TCF-4 signaling activation.

SIRT1 is required for long-term growth of human mesenchymal stem cells.

Human mesenchymal stem cells (MSCs) have therapeutic potential because of their ability to self-renew and differentiate into multiple tissues. However, senescence often occurs in MSCs when they are cultured in vitro and the molecular mechanisms underlying this effect remain unclear. In this study, we found that NAD-dependent protein deacetylase SIRT1 is differentially expressed in both human bone marrow-derived MSCs (B-MSCs) and adipose tissue-derived MSCs after increasing passages of cell culture. Using lentiviral shRNA we demonstrated that selective knockdown of SIRT1 in human MSCs at early passage slows down cell growth and accelerates cellular senescence. Conversely, overexpression of SIRT1 delays senescence in B-MSCs that have undergone prolonged in vitro culturing and the cells do not lose adipogenic and osteogenic potential. In addition, we found that the delayed accumulation of the protein p16 is involved in the effect of SIRT1. However, resveratrol, which has been used as an activator of SIRT1 deacetylase activity, only transiently promotes proliferation of B-MSCs. Our findings will help us understand the role of SIRT1 in the aging of normal diploid cells and may contribute to the prevention of human MSCs senescence thus benefiting MSCs-based tissue engineering and therapies.