Jian Sun

Blood lymphocyte-to-monocyte ratio identifies high-risk patients in diffuse large B-cell lymphoma treated with R-CHOP

Background Recent research has shown a correlation between immune microenvironment and lymphoma biology. This study aims to investigate the prognostic significance of the immunologically relevant lymphocyte-to-monocyte ratio (LMR), in diffuse large B-cell lymphoma (DLBCL) in the rituximab era. Methodology/Principal Findings We analyzed retrospective data from 438 newly diagnosed DLBCL patients treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) therapy. We randomly selected 200 patients (training set) to generate a cutoff value for LMR by receiver operating characteristic (ROC) curve analysis. LMR was then analyzed in a testing set (n = 238) and in all patients (n = 438) for validation. The LMR cutoff value for survival analysis determined by ROC curve in the training set was 2.6. Patients with low LMR tended to have more adverse clinical characteristics. Low LMR at diagnosis was associated with worse survival in DLBCL, and could also identify high-risk patients in the low-risk IPI category. Multivariate analysis identified LMR as an independent prognostic factor of survival in the testing set and in all patients. Conclusions/Significance Baseline LMR, a surrogate biomarker of the immune microenvironment, is an effective prognostic factor in DLBCL patients treated with R-CHOP therapy. Future prospective studies are required to confirm our findings.

Effects of the biological clock gene Bmal1 on tumour growth and anti-cancer drug activity

The Bmal1 gene plays a key role in controlling circadian rhythms. To better understand how the Bmal1 gene affects tumour growth and the response to anti-cancer drugs, we examined the effect of knockdown of Bmal1 by RNAi both in vitro and in vivo. Down-regulation of Bmal1 gene expression accelerated cell proliferation in vitro and promoted tumour growth in mice. Suppressing Bmal1 expression in murine colon cancer cells (C26) and fibroblast cells (L929) decreased apoptosis induced by Etoposid, reduced the distribution of cells in the G2/M phases treated by Docetaxel and decreased DNA damage induced by Cisplatin. Loss of Bmal1 reduced the expression of per1, per2, per3, wee1 and p53. The expression of p21 and c-myc was also altered in certain cell lines. However, Bmal1 deficiency increased the protein levels of cdc2, cyclin B1, cyclin D1 and cyclin E. Wee1 and cyclin A expression was minimally altered. Thus, the circadian clock gene Bmal1 plays a role in regulating tumour cell apoptosis, cell-cycle progression and DNA damage response and in homoeostasis regulation. Down-regulation of Bmal1 accelerates the development of tumours and may influence the response to anti-cancer drugs.

ApoG2, a novel inhibitor of antiapoptotic Bcl-2 family proteins, induces apoptosis and suppresses tumor growth in nasopharyngeal carcinoma xenografts

Nasopharyngeal carcinoma (NPC) is a common malignant tumor in South China. It has been reported that overexpression of antiapoptotic Bcl‐2 family proteins in NPC has caused the lack of long‐term efficacy of conventional therapies. Apogossypolone (ApoG2), a novel small‐molecule inhibitor of antiapoptotic Bcl‐2 family proteins, has been discovered as the optimized derivative of gossypol. In this study, we found that in NPC cells, ApoG2 totally blocked the antiapoptotic function of Bcl‐2 family proteins without affecting the expression levels of these proteins. ApoG2 selectively inhibited proliferation of 3 NPC cell lines (C666‐1, CNE‐1 and CNE‐2) that highly expressed the antiapoptotic Bcl‐2 proteins. This inhibitory activity was associated with release of cytochrome c, activation of caspase‐9 and caspase‐3 and apoptosis of sensitive NPC cells. However, ApoG2 had no obvious inhibitory effect on NPC cell line HONE‐1, which expressed antiapoptotic Bcl‐2 and Bcl‐xL at a low level. We further found that ApoG2 effectively suppressed tumor growth of NPC xenografts in nude mice and enhanced the antitumor effect of CDDP (cisplatin) on NPC cells in vitro and in vivo. Immunohistochemical results showed that the expression of CD31 decreased after ApoG2 treatment, which suggested inhibition of angiogenesis in NPC xenografts. Our findings strongly suggest that ApoG2 may serve as a novel inhibitor of Bcl‐2 family proteins and, by targeting these proteins, may become a promising drug for the treatment of NPC.

ApoG2 induces cell cycle arrest of nasopharyngeal carcinoma cells by suppressing the c-Myc signaling pathway.

Backgroundapogossypolone (ApoG2) is a novel derivate of gossypol. We previously have reported that ApoG2 is a promising compound that kills nasopharyngeal carcinoma (NPC) cells by inhibiting the antiapoptotic function of Bcl-2 proteins. However, some researchers demonstrate that the antiproliferative effect of gossypol on breast cancer cells is mediated by induction of cell cycle arrest. So this study was aimed to investigate the effect of ApoG2 on cell cycle proliferation in NPC cells.ResultsWe found that ApoG2 significantly suppressed the expression of c-Myc in NPC cells and induced arrest at the DNA synthesis (S) phase in a large percentage of NPC cells. Immunoblot analysis showed that expression of c-Myc protein was significantly downregulated by ApoG2 and that the expression of c-Mycs downstream molecules cyclin D1 and cyclin E were inhibited whereas p21 was induced. To further identify the cause-effect relationship between the suppression of c-Myc signaling pathway and induction of cell cycle arrest, the expression of c-Myc was interfered by siRNA. The results of cell cycle analysis showed that the downregulation of c-Myc signaling pathway by siRNA interference could cause a significant arrest of NPC cell at S phase of the cell cycle. In CNE-2 xenografts, ApoG2 significantly downregulated the expression of c-Myc and suppressed tumor growth in vivo.ConclusionOur findings indicated that ApoG2 could potently disturb the proliferation of NPC cells by suppressing c-Myc signaling pathway. This data suggested that the inhibitory effect of ApoG2 on NPC cell cycle proliferation might contribute to its use in anticancer therapy.

Baicalein pretreatment reduces liver ischemia/reperfusion injury via induction of autophagy in rats

We previously demonstrated that baicalein could protect against liver ischemia/reperfusion (I/R) injury in mice. The exact mechanism of baicalein remains poorly understood. Autophagy plays an important role in protecting against I/R injury. This study was designed to determine whether baicalein could protect against liver I/R injury via induction of autophagy in rats. Baicalein was intraperitoneally injected 1 h before warm ischemia. Pretreatment with baicalein prior to I/R insult significantly blunted I/R-induced elevations of serum aminotransferase levels and significantly improved the histological status of livers. Electron microscopy and expression of the autophagic marker LC3B-II suggested induction of autophagy after baicalein treatment. Moreover, inhibition of the baicalein-induced autophagy using 3-methyladenine (3-MA) worsened liver injury. Furthermore, baicalein treatment increased heme oxygenase (HO)-1 expression, and pharmacological inhibition of HO-1 with tin protoporphyrin IX (SnPP) abolished the baicalein-mediated autophagy and the hepatocellular protection. In primary rat hepatocytes, baicalein-induced autophagy also protected hepatocytes from hypoxia/reoxygenation injury in vitro and the beneficial effect was abrogated by 3-MA or Atg7 siRNA, respectively. Suppression of HO-1 activity by SnPP or HO-1 siRNA prevented the baicalein-mediated autophagy and resulted in increased hepatocellular injury. Collectively, these results suggest that baicalein prevents hepatocellular injury via induction of HO-1-mediated autophagy.

Circadian Rhythm in Dihydropyrimidine Dehydrogenase Activity and Reduced Glutathione Content in Peripheral Blood of Nasopharyngeal Carcinoma Patients

Dihydropyrimidine dehydrogenase (DPD) is a rate‐limiting enzyme of 5‐fluorouracil (5‐FU) catabolism. Glutathione (GSH) is a tripeptide involved in platinum complex detoxification. This study explored the circadian rhythms of DPD activity and GSH concentration in the peripheral blood of 16 patients with histologically proven nasopharyngeal carcinoma (NPC) in order to guide the establishment of chronotherapeutic schedules for this cancer. DPD activity and GSH concentration were determined by high performance liquid chromatography (HPLC). Both variables displayed significant circadian rhythms (Cosinor analysis: p=0.009 and 0.012, respectively). Peak DPD activity occurred at about 02:30 h; whereas, peak GSH concentration occurred around 12:40 h. The differences between the peak and nadir mean values were 25.5% and 38.7%, respectively. The study showed that the circadian rhythms in DPD activity and GSH concentration in Chinese NPC are similar to those reported for western patients with colorectal cancer, despite the differences in race and kinds of cancer. These findings imply that the chronotherapeutic schedule of 5‐FU and platinum used to treat European colorectal cancer patients probably is applicable to Chinese NPC patients.

Upregulated TRIM29 promotes proliferation and metastasis of nasopharyngeal carcinoma via PTEN/AKT/mTOR signal pathway

Tripartite motif–containing 29 (TRIM29) has been reported to be dysregulated in human cancers. Up-regulation of TRIM29 was first observed in NPC cell lines by a genome-wide transcriptome analysis in our previous study. However, its expression biological function and clinical significance in nasopharyngeal carcinoma (NPC) remain unclear. In this study, TRIM29 expression was validated by qRT-PCR and immunohistochemistry in 69 NPC samples. Notably, TRIM29 protein expression was significantly and positively correlated with the tumor size, clinical stage and metastasis. TRIM29 was identified as the direct target of miR-335-5p and miR-15b-5p, both of which were down-regulated and negatively associated with TRIM29 expression in NPC cell lines and clinical samples. Ectopic TRIM29 expression promoted proliferation, epithelial-mesenchymal transition (EMT), migration and invasion in NPC cells, while its depletion inhibited cell invasion and EMT phenotype. Mechanistically, TRIM29 overexpression reduced PTEN expression and increase phosphorylated protein level of AKT, p70S6K and 4E-BP1. Correspondingly, AKT inhibitor and Rapamycin blocked the effect of TRIM29 on cell invasion. In conclusion, our results suggest that miR-335-5p and miR-15b-5p down-regulation results in TRIM29 over-expression, which induces proliferation, EMT and metastasis of NPC through the PTEN/AKT/mTOR signaling pathway.

Granulocyte Colony Stimulating Factor Induces Lipopolysaccharide (LPS) Sensitization via Upregulation of LPS Binding Protein in Rat

Liver is the main organ for lipopolysaccharide (LPS) clearance. Sensitization to LPS is associated with the upregulation of LPS-binding protein (LBP) in animal models. Therefore, we hypothesized that LBP could induce LPS sensitization through enhancing hepatic uptake of LPS. In this study, we examined the role of LBP in pathogenesis of LPS induced systemic inflammatory response syndrome (SIRS). LBP expression was upregulated after granulocyte colony stimulating (G-CSF) pretreatment. The effect of LBP was further confirmed by blockade of LBP using LBP blocking peptide – LBPK95A. After G-CSF pretreatment, upregulation of LBP was observed in bone marrow cells and liver. The G-CSF induced LBP upregulation caused LPS hypersensitization in rats as indicated by higher mortality and severer liver damage. Of note, LBP blockade increased the survival rate and attenuated the liver injury. The LBP induced LPS hypersensitization was associated with increased hepatic uptake of LPS and augmented hepatic expression of LPS receptors, such as toll-like receptor (TLR)-4. Furthermore, LBP mediated early neutrophil infiltration, which led to increased monocyte recruitment in liver after LPS administration. In conclusion, G-CSF induced LBP expression could serve as a new model for investigation of LPS sensitization. We demonstrated the crucial role of LBP upregulation in pathogenesis of LPS induced SIRS.

ApoG2 inhibits antiapoptotic Bcl-2 family proteins and induces mitochondria-dependent apoptosis in human lymphoma U937 cells

Lymphoma is one of the most common types of hematological malignancies and proteins from the Bcl-2 family are highly expressed in human lymphomas. Apogossypolone (ApoG2), the most potent gossypol derivative, has been classified as a novel small-molecule inhibitor of antiapoptotic Bcl-2 family proteins. Here, we assessed the in-vitro cytotoxicity of ApoG2 on human U937 lymphoma cells, and explored the underlying intracellular molecular mechanisms of ApoG2. Using the WST-8 assay, we found that ApoG2 inhibited growth of U937 cells in a dose-dependent and time-dependent manner, and the IC50 values were 30.08, 14.81, and 9.26 μmol/l for 24, 48, and 72 h treatments, respectively. ApoG2 also induced apoptosis in U937 cells, as noted through changes in morphological characteristics, including cellular internucleosomal DNA fragmentation and the appearance of a sub-G1 apoptotic peak. Treatment with ApoG2 downregulated Bcl-xL and Mcl-1 protein expression and blocked the binding of Bcl-2 with Bax protein. Furthermore, ApoG2 led to an abundant release of cytochrome c from mitochondria and a five-fold increase in the activity of caspase-3 and caspase-9. Taken together, our results suggest that ApoG2 could effectively suppress the growth of human lymphoma cell line U937 through the inhibition of the antiapoptotic Bcl-2 family proteins and the induction of mitochondria-dependent apoptotic cell death.

Ischemic preconditioning attenuates ischemia/reperfusion injury in rat steatotic liver: role of heme oxygenase-1-mediated autophagy

Steatotic livers are more susceptible to ischemia/reperfusion (I/R) injury, which is ameliorated by ischemic preconditioning (IPC). Autophagy possesses protective action on liver I/R injury and declines in steatotic livers. The aim of this study was to test the hypothesis that the increased susceptibility of steatotic livers to I/R injury was associated with defective hepatic autophagy, which could be restored by IPC via heme oxygenase-1 (HO-1) signaling. Obesity and hepatic steatosis was induced using a high fat diet. Obesity impaired hepatic autophagy activity and decreased hepatic HO-1 expression. Induction of HO-1 restored autophagy activity and inhibited calpain 2 activity. Additionally, suppression of calpain 2 activity also restored autophagy activity. Mitochondrial dysfunction and hepatocellular injury were significantly increased in steatotic livers compared to lean livers in response to I/R injury. This increase in sensitivity to I/R injury was associated with defective hepatic autophagy activity in steatotic livers. IPC increased autophagy and reduced mitochondrial dysfunction and hepatocellular damage in steatotic livers following I/R injury. Furthermore, IPC increased HO-1 expression. Inhibition of HO-1 decreased the IPC-induced autophagy, increased calpain 2 activity and diminished the protective effect of IPC against I/R injury. Inhibition of calpain 2 restored autophagic defect and attenuated mitochondrial dysfunction in steatotic livers after I/R. Collectively, IPC might ameliorate steatotic liver damage and restore mitochondrial function via HO-1-mediated autophagy.