Zhiqiang Liu

Mature adipocytes in bone marrow protect myeloma cells against chemotherapy through autophagy activation

A major problem in patients with multiple myeloma is chemotherapy resistance, which develops in myeloma cells upon interaction with bone marrow stromal cells. However, few studies have determined the role of bone marrow adipocytes, a major component of stromal cells in the bone marrow, in myeloma chemotherapy resistance. We reveal that mature human adipocytes activate autophagy and upregulate the expression of autophagic proteins, thereby suppressing chemotherapy-induced caspase cleavage and apoptosis in myeloma cells. We found that adipocytes secreted known and novel adipokines, such as leptin and adipsin. The addition of these adipokines enhanced the expression of autophagic proteins and reduced apoptosis in myeloma cells. In vivo studies further demonstrated the importance of bone marrow-derived adipocytes in the reduced response of myeloma cells to chemotherapy. Our findings suggest that adipocytes, adipocyte-secreted adipokines, and adipocyte-activated autophagy are novel targets for combatting chemotherapy resistance and enhancing treatment efficacy in myeloma patients.

p38 MAPK in Myeloma Cells Regulates Osteoclast and Osteoblast Activity and Induces Bone Destruction

p38 mitogen-activated protein kinase (MAPK), which is constitutively activated in human myeloma, has been implicated in bone destruction by this cancer, but the processes it recruits are obscure. In this study, we show that p38 activity in myeloma inhibits osteoblast differentiation and bone formation, but also enhances osteoclast maturation and bone resorption. p38 regulated the expression and secretion of the Wnt pathway antagonist DKK-1 and the monocyte chemoattractant MCP-1. Attenuating p38, DKK-1, or MCP-1 were each sufficient to reduce bone lesions in vivo. Although it is well known that DKK-1 inhibits osteoblast differentiation, we found that together with MCP-1, it could also promote osteoclast differentiation and bone resorption. The latter effects were mediated by enhancing expression of RANK in osteoclast progenitor cells and by upregulating secretion of its ligand RANKL from stromal cells and mature osteoblasts. In summary, our study defined the mechanisms by which p38 signaling in myeloma cells regulates osteoblastogenesis, osteoclastogenesis, and bone destruction. Our findings, which may have implications for bone invasion by other cancers where p38 is elevated, strongly suggests that targeting p38 for inhibition may offer an effective therapeutic approach to treat osteolytic bone lesions in patients with myeloma.

A critical role of autocrine sonic hedgehog signaling in human CD138+myeloma cell survival and drug resistance

Hedgehog (Hh) signaling plays an important role in the oncogenesis of B-cell malignancies such as multiple myeloma (MM). However, the source of Hh ligand sonic hedgehog (SHH) and its target cells remains controversial. Previous studies showed that stromally induced Hh signaling is essential for the tumor cells and that CD19(+)CD138(-) MM stem cells are the target cells of Hh signaling. Here we demonstrate that SHH was mainly secreted by human myeloma cells but not by stromal cells in MM bone marrow. Autocrine SHH enhanced CD138(+) myeloma cell proliferation and protected myeloma cells from spontaneous and stress-induced apoptosis. More importantly, autocrine SHH protected myeloma cells against chemotherapy-induced apoptosis in vitro and in vivo. Combinational treatment with chemotherapy and SHH-neutralizing antibody displayed synergistic antimyeloma effects. Mechanistic studies showed that SHH signaling activated the SHH/GLI1/BCL-2 axis, leading to the inhibition of myeloma cell apoptosis. Thus, this study identifies the myeloma autocrine Hh signaling pathway as a potential target for the treatment of MM. Targeting this pathway may improve the efficacy of chemotherapy in MM patients.

Immune evasion of mantle cell lymphoma: expression of B7-H1 leads to inhibited T-cell response to and killing of tumor cells.

Clinical trials of immunotherapy in mantle cell lymphoma have not yet delivered desirable results, partly because of the inhibitory machinery of the tumor and its microenvironment. Here we investigated the role of B7-H1, a member of the B7 family of co-stimulatory/co-inhibitory ligands, in mantle cell lymphoma-mediated immunosuppression. Allogeneic CD3+, CD4+ and CD8+ T cells were purified and co-cultured with irradiated mantle cell lymphoma cells. Mantle cell lymphoma-reactive T-cell lines from HLA-A*0201+ healthy blood donors were generated after in vitro restimulation, and were subjected to functional tests. We found that B7-H1 expressed on mantle cell lymphoma cells was able to inhibit T-cell proliferation induced by the tumor cells, impair the generation of antigen-specific T-cell responses, and render mantle cell lymphoma cells resistant to T-cell-mediated cytolysis. Blocking or knocking down B7-H1 on mantle cell lymphoma cells enhanced T-cell responses and restored tumor-cell sensitivity to T-cell-mediated killing in vitro and in vivo. Knocking down B7-H1 on mantle cell lymphoma cells primed more CD4+ or CD8+ memory effector T cells. Our study demonstrates for the first time that lymphoma cell-expressed B7-H1 may lead to the suppression of host anti-tumor immune responses in mantle cell lymphoma and targeting tumor cell B7-H1 may represent a novel approach to improve the efficacy of immunotherapy in patients with mantle cell lymphoma.

Autocrine Sonic Hedgehog Attenuates Inflammation in Cerulein-Induced Acute Pancreatitis in Mice via Upregulation of IL-10

Hedgehog signaling plays critical roles in pancreatic oncogenesis and chronic pancreatitis, but its roles in acute pancreatitis (AP) are largely ambiguous. In this study, we provide evidence that Sonic hedgehog (Shh), but neither Desert hedgehog (Dhh) nor Indian hedgehog (Ihh), is the main protein whose expression is activated during the development of cerulein-induced acute pancreatitis in mice, and the Shh serves as an anti-inflammation factor in an autocrine manner. Blocking autocrine Shh signaling with anti-Shh neutralizing antibody aggravates the progression of acute pancreatitis. Mechanistic insight into Shh signaling activation in acute pancreatitis indicates that inflammatory stimulation activates Shh expression and secretion, and subsequently upregulates the expression and secretion of interleukin-10 (IL-10). Moreover, inhibition of Shh signaling with neutralizing antibody abolishes IL-10 production in vivo and in vitro. Molecular biological studies show that autocrine Shh signaling activates the key transcriptional factor Gli1 so that the target gene IL-10 is upregulated, leading to the protective and anti-inflammatory functions in the mouse model of acute pancreatitis. Thus, this study suggests autocrine Shh signaling functions as a protective signaling in the progression of acute pancreatitis.

Gankyrin is frequently overexpressed in cervical high grade disease and is associated with cervical carcinogenesis and metastasis.

Our previous studies have showed that Gankyrin expression is correlated with a malignant phenotype in endometrial carcinoma. Here, we investigated the possible role of Gankyrin in cervical disease. The increasing protein level of Gankyrin was observed in high-grade cervical intraepithelial neoplasia and carcinoma compared with benign cervical tissues and low-grade cervical intraepithelial neoplasia. In para-carcinoma tissues, it was found interestingly that there was no lymph node metastasis when nuclei Gankyrin was positively expressed, but lymph node metastasis rate was 30% (6/20) when nuclei Gankyrin was negatively expressed. In vitro, the transfection of Gankyrin resulted in markedly up-regulating of Vimentin, β-catenin and Twist2, as well as down-regulating of E-cadherin in cervical carcinoma cells. Our results suggested that Gankyrin may be functional in cervical carcinogenesis and metastasis.

Resistin induces multidrug resistance in myeloma by inhibiting cell death and upregulating ABC transporter expression

Despite advances in therapy, multiple myeloma remains incurable, with a high frequency of relapse. This suggests the need to identify additional factors that contribute to drug resistance. Our previous studies revealed that bone marrow adipocytes promote resistance to chemotherapy in myeloma through adipocyte-secreted adipokines, but the mechanism underlying this effect and the specific adipokines involved are not well understood. We proposed to determine the role of resistin, an adipokine that is secreted by adipocytes, in chemotherapy resistance in myeloma. We found that resistin abrogated chemotherapy-induced apoptosis in established myeloma cell lines and primary myeloma samples. Resistin inhibited chemotherapy-induced caspase cleavage through the NF-κB and PI3K/Akt pathways. Resistin also increased the expression and drug efflux function of ATP-binding cassette (ABC) transporters in myeloma cells through decreasing the expression of both DNA methyltransferases DNMT1 and DNMT3a and the methylation levels of ABC gene promoters. In vivo studies further demonstrated the protective effect of resistin in chemotherapy-induced apoptosis. Our study thus reveals a new biological function of resistin in the pathogenesis of myeloma, with the implication that targeting resistin could be a potential strategy to prevent or overcome multidrug resistance in myeloma.

Socs1 and Socs3 degrades Traf6 via polyubiquitination in LPS-induced acute necrotizing pancreatitis

Mechanisms involved in inflammatory development during acute pancreatitis (AP) are largely vague, especially in the transformation of acute edematous pancreatitis (AEP) into acute necrotizing pancreatitis (ANP). This current study aims to investigate the functions of Traf6 in different AP models in vitro and in vivo, and to identify the possible regulatory mechanism in the progression of inflammation from mild to severe. Our data revealed that the level of Traf6 expression was significantly increased in the mild AP induced by caerulein, and the upregulation of Traf6 played a protective role in acinar cells against caerulein-induced apoptosis. In contrast, only Traf6 protein but not mRNA was downregulated in the severe ANP induced by combination treatment of caerulein and LPS. Mechanistic studies showed that LPS upregulated the levels of Socs1 and Socs3 expressions in acinar cells, Socs1 and Socs3 interacted Traf6 directly and degraded Traf6 protein via polyubiquitination, thereby counteracted the protective function of Traf6. In vivo study further showed that combination treatment of caerulein and LPS failed to induce an ANP model in the TLR4 knockout mice, and the level of Traf6 expression in the pancreatic tissues remained the same as that from the acute edematous pancreatitis (AEP) mouse. Taken together, our study reveals that Traf6 functioned as a protective factor in the progression of AP, and LPS-induced Socs1 and Socs3 exacerbate mild AP to severe AP, which provides evidence for developing a new therapeutic target to combat AP.

The indicative function of Twist2 and E-cadherin in HPV oncogene-induced epithelial-mesenchymal transition of cervical cancer cells.

High-risk human papillomavirus (HR-HPV) infections are among the most important factors for cervical carcinogenesis. However, whether patients infected with HR-HPV eventually develop a malignant tumor, largely depends on epithelial-mesenchymal transition (EMT), which plays an extraordinary role in the process of carcinogenesis and metastasis. Therefore, we evaluated the protein levels of EMT-related genes in normal cervical squamous epithelium, cervical intraepithelial neoplasia (CIN), and cervical squamous cell carcinoma (SCC) by tissue microarray and immunohistochemical staining. By comparing the expression of EMT-related proteins in 31 cases of cervical tumors and tumor adjacent tissues and exploring the relationship between HPV16 oncogenes and EMT in vitro, we found that Twist2 protein levels were significantly higher in CIN and cervical cancer than in normal cervical squamous epithelial samples (p<0.01 and p<0.001, respectively). This finding corresponded with the decreased expression of E-cadherin in cervical cancer. The difference in the expression of Twist2 and E-cadherin between 31 cases of cervical tumors and tumor adjacent tissues was statistically significant (p<0.01). HPV16 oncogenes were able to induce morphological alterations in the SiHa cell line, upregulate the expression of Twist2 and vimentin, downregulate E-cadherin in vitro, and exert an effect on invasion. Thus, joint detection of Twist2 and E-cadherin expression can help evaluate and provide greater insight into cervical carcinogenesis and progression.

The protein levels of mcm7 and p63 in evaluating lesion severity of cervical disease

Objectives The objective of this study was to analyze the relationship among the protein levels of MCM7, p63, and human papillomavirus (HPV) in different cervical lesion tissues and appraise their predictive value in evaluating severity of cervical disease. Methods Twelve normal cervix or chronic cervicitis, 42 squamous intraepithelial lesions, and 53 cervical carcinoma tissues were enrolled, and the protein levels of MCM7, p63, and HPV were detected by immunohistochemistry. Results The positive examination rates of all the MCM7, p63, and HPV proteins increased gradually and significantly from normal cervix and chronic cervicitis tissues, low-grade squamous intraepithelial lesions, high-grade squamous intraepithelial lesions to cervical carcinomas, respectively. As to predict high-grade squamous intraepithelial lesions and carcinogenesis is concerned, the MCM7 protein had a sensitivity of 94.0%, a specificity of 56.5%, a positive predictive value of 88.8%, and a negative predictive value of 72.2%. The p63 protein had a sensitivity of 78.6%, a specificity of 81.8%, a positive predictive value of 94.3%, and a negative predictive value of 50.0%. Protein level of MCM7 was positively correlated with that of p63 in cervical tissues (r = 0.806, P < 0.01), and the p63 was also positively correlated with histopathologic type (P < 0.05). Conclusions Protein levels of MCM7 and p63 were associated significantly with high-grade cervical lesion, and aberrant p63 protein level may distinguish different histopathologic types of cervical carcinoma. They may act as co–predictive index in both HPV-dependent and HPV-independent high-grade cervical lesion with high sensitivity and specificity.