Youping Liu

miR-221/222 suppression protects against endoplasmic reticulum stress-induced apoptosis via p27(Kip1)- and MEK/ERK-mediated cell cycle regulation.

Abstract Cancer cells are relatively resistant to endoplasmic reticulum (ER) stress-induced apoptosis. However, the underlying mechanisms remain largely unclear. We observed that the microRNAs miR-221/222 are associated with apoptosis regulation under ER stress in human hepatocellular carcinoma (HCC) cells. Induction of ER stress does not trigger significant apoptosis but obviously causes downregulation of miR-221/222 in HCC cells. In these cells, ER stress-induced apoptosis is enhanced by miR-221/222 mimics and attenuated by miR-221/222 inhibitors. miR-221/222 promoted-apoptosis under ER stress is associated with p27Kip1- and MEK/ERK-mediated cell cycle regulation. Our results suggest that suppression of miR-221/222 plays a crucial role in the protection against apoptosis induced by ER stress in HCC cells.

Analysis of plasma proteome from cases of the different traditional Chinese medicine syndromes in patients with chronic hepatitis B.

A proteomic analysis method, two dimensional gel electrophoresis (2-DE) followed by matrix-assisted laser desorption/ionization time-of-flight MS (MALDI-TOF-MS), was used to explore the link between plasma proteome and the different syndromes of traditional Chinese medicine (TCM) in patients with chronic hepatitis B (CHB). In compared with the plasma proteomes from health donors, the alterations in protein expression from cases of the five TCM syndromes, including damp heat stasis in the middle-Jiao syndrome, liver Qi stagnation and spleen deficiency syndrome, spleen and kidney Yang deficiency syndrome, liver and kidney Yin deficiency syndrome, and blood stasis into collateral syndrome with CHB were identified (P<0.05). In the cases of the five TCM syndromes with CHB, immunoglobulin J-chains (IGJ) and C-reactive protein (CRP) were up-regulated, while haptoglobin (HPT), retinol binding protein (RBP) and vitronectin were down-regulated. To further confirm these results, four proteins, including CRP, IGJ, HPT and RBP, from more plasma samples were quantified by ELISA. The results showed that the changes of protein levels were consistent with those from the 2-DE experiment. Importantly, the upregulation tendency of IGJ level in plasma is related with the different TCM syndromes with CHB (P<0.05). Our results show that IGJ may serve as a novel potential biomarker for diagnosis of the different TCM syndromes in patients with CHB.

Activation of PKR/eIF2α signaling cascade is associated with dihydrotestosterone-induced cell cycle arrest and apoptosis in human liver cells.

Androgen receptor (AR) signaling plays an important role in the development and progression of several liver diseases, including hepatocellular carcinoma (HCC) and non‐alcoholic fatty liver disease (NAFLD). Dihydrotestosterone (DHT) is the active metabolite of the major circulating androgen, testosterone. In this study, we investigated the effect of DHT on human liver cells. We found that DHT not only induces cell cycle arrest but also initiates apoptosis in androgen‐sensitive liver cells, such as SMMC‐7721 and L02. Importantly, DHT/AR induces the activation of RNA‐dependent protein kinase (PKR)/eukaryotic initiation factor‐2 alpha (eIF2α) cascades in androgen‐sensitive liver cells. PKR/eIF2α activation‐induced growth arrest and DNA damage‐inducible gene 153 (GADD153) and heat shock protein 27 (Hsp27) expression contribute to cell cycle arrest in response to DHT. It is notable that DHT administration results in androgen‐sensitive liver cells apoptosis, at least in part, through PKR/eIF2α/GADD153 cascades. These results suggest that the androgen/AR pathway plays a pivotal role in liver cell growth and apoptosis regulating, whose deregulation might be involved in the pathogenesis of liver diseases. J. Cell. Biochem. 113: 1800–1808, 2012.

c‐Met function requires n‐linked glycosylation modification of pro‐Met

c‐Met, the receptor for hepatocyte growth factor (HGF), is cell surface tyrosine kinase that controls cancer cell growth, survival, invasion, and metastasis. Post‐translational modification, such as glycosylation, plays an essential role in regulating the function of cell surface molecules. Whether glycosylation modification regulates the enzymatic properties of c‐Met is unknown. In this study, we investigated the effect of glycosylation on the function of c‐Met. We found that c‐Met is an N‐linked glycosylated protein. Both pro‐Met and p145Met (the β subunit of mature c‐Met) have N‐linked glycosylation. Glycosylation inhibitor studies revealed that the N‐glycosylation modification of p145Met is from pro‐Met, but not due to the further modification of pro‐Met. Importantly, blocking the N‐glycosylation targets pro‐Met to cytoplasm and initiates its phosphorylation independent of HGF engagement. Nonglycosylated pro‐Met activates c‐Met downstream pathways to a certain extent to compensate for the degradation of p145Met induced by glycosylation blocking‐mediated endoplasmic reticulum (ER) stress. J. Cell. Biochem. 114: 816–822, 2013.

JNK Contributes to the Tumorigenic Potential of Human Cholangiocarcinoma Cells through the mTOR Pathway Regulated GRP78 Induction

Less is known about the roles of c-Jun N-terminal kinase (JNK) in cholangiocarcinoma (CCA). Here, we report that JNK exerts its oncogenic action in human CCA cells, partially due to the mammalian target of rapamycin (mTOR) pathway regulated glucose-regulated protein 78 (GRP78) induction. In human CCA cells, the phosphorylation of eukaryotic initiation factor alpha (eIF2α) results in the accumulation of activating transcription factor 4 (ATF4) and GRP78 independent of unfolded protein response (UPR). Suppression of GRP78 expression decreases the proliferation and invasion of human CCA cells. Its notable that mTOR is required for eIF2α phosphorylation-induced ATF4 and GRP78 expression. Importantly, JNK promotes eIF2α/ATF4-mediated GRP78 induction through regulating the activity of mTOR. Thus, our study implicates JNK/mTOR signaling plays an important role in cholangiocarcinogenesis, partially through promoting the eIF2α/ATF4/GRP78 pathway.

Synergistic antitumor activity of the combination of salubrinal and rapamycin against human cholangiocarcinoma cells

Less is known about the roles of eukaryotic initiation factor alpha (eIF2α) in cholangiocarcinoma (CCA). Here, we report that eIF2α inhibitor salubrinal inhibits the proliferation of human CCA cells. Clinical application of mammalian target of rapamycin (mTOR) inhibitors only has moderate antitumor efficacy. Therefore, combination approaches may be required for effective clinical use of mTOR inhibitors. Here, we investigated the efficacy of the combination of salubrinal and rapamycin in the treatment of CCA. Our data demonstrate a synergistic antitumor effect of the combination of salubrinal and rapamycin against CCA cells. Rapamycin significantly inhibits the proliferation of CCA cells. However, rapamycin initiates a negative feedback activation of Akt. Inhibition of Akt by salubrinal potentiates the efficacy of rapamycin both in vitro and in vivo. Additionally, rapamycin treatment results in the up-regulation of Bcl-xL in a xenograft mouse model. It is notable that salubrinal inhibits rapamycin-induced Bcl-xL up-regulation in vivo. Taken together, our data suggest that salubrinal and rapamycin combination might be a new and effective strategy for the treatment of CCA.

The suppressive role of p38 MAPK in cellular vacuole formation

Vacuolization of the cytoplasm is one of the dramatic and frequently observed phenomena in various cell types. Cellular vacuoles occur spontaneously or via a wide range of inductive stimuli, but the molecular mechanism involved in this process remains largely unknown. In this study, we investigated the role of the p38 and JNK pathways in the formation of cytoplasmic vacuoles. We found that p38 and JNK agonist anisomycin abolishes spontaneous cytoplasmic vacuolization of HepG2 cells through p38 activation, but not through JNK activation. Importantly, blocking the activity of p38 or suppression the expression of p38 elicits cytoplasmic vacuoles formation in various cancer cells. Furthermore, cytoplasmic vacuoles induced by p38 blocking are derived from the perinuclear region. These observations provide direct evidence for a role of p38 signaling in regulating the formation of cytoplasmic vacuoles. J. Cell. Biochem. 114: 1789–1799, 2013.