Peti Thuwajit

Chronic Inflammation and Cytokines in the Tumor Microenvironment

Acute inflammation is a response to an alteration induced by a pathogen or a physical or chemical insult, which functions to eliminate the source of the damage and restore homeostasis to the affected tissue. However, chronic inflammation triggers cellular events that can promote malignant transformation of cells and carcinogenesis. Several inflammatory mediators, such as TNF-α, IL-6, TGF-β, and IL-10, have been shown to participate in both the initiation and progression of cancer. In this review, we explore the role of these cytokines in important events of carcinogenesis, such as their capacity to generate reactive oxygen and nitrogen species, their potential mutagenic effect, and their involvement in mechanisms for epithelial mesenchymal transition, angiogenesis, and metastasis. Finally, we will provide an in-depth analysis of the participation of these cytokines in two types of cancer attributable to chronic inflammatory disease: colitis-associated colorectal cancer and cholangiocarcinoma.

Periostin activates integrin α5β1 through a PI3K/AKT‑dependent pathway in invasion of cholangiocarcinoma.

Periostin (PN) is mainly produced from stromal fibroblasts in cholangiocarcinoma (CCA) and shows strong impact in cancer promotion. This work aimed to investigate the mechanism that PN uses to drive CCA invasion. It was found that ITGα5β1 and α6β4 showed high expression in non-tumorigenic biliary epithelial cells and in almost all CCA cell lines. PN had preferential binding to CCA cells via ITGα5β1 and blocking this receptor by either neutralizing antibody or siITGα5 could attenuate PN-induced invasion. After PN-ITGα5β1 binding, intracellular pAKT was upregulated whereas there was no change in pERK. Moreover, PN could not activate AKT in condition of treatment with a PI3K inhibitor. These data provide evidence that PN-activated invasion of CCA cells is through the ITGα5β1/PI3K/AKT pathway. Strategies aimed to inhibit this pathway may, thus, provide therapeutic benefits.

Cancer-associated fibroblasts induce high mobility group box 1 and contribute to resistance to doxorubicin in breast cancer cells

BackgroundCancer-associated fibroblasts and high mobility group box 1 (HMGB1) protein have been suggested to mediate cancer progression and chemotherapy resistance. The role of such fibroblasts in HMGB1 production in breast cancer is unclear. This study aimed to investigate the effects of cancer-associated fibroblasts on HMGB1 expression in breast cancer cells and its role in chemotherapeutic response.MethodsBreast cancer-associated fibroblasts (BCFs) and non-tumor-associated fibroblasts (NTFs) were isolated from human breast cancers or adjacent normal tissues and established as primary cultures in vitro. After confirmation of the activated status of these fibroblasts, conditioned-media (CM) were collected and applied to MDA-MB-231 human triple negative breast cancer cells. The levels of intracellular and extracellular HMGB1 were measured by real-time PCR and/or Western blot. The response of BCF-CM-pre-treated cancer cells to doxorubicin (Dox) was compared with those pre-treated with NTF-CM or control cultures. The effect of an HMGB1 neutralizing antibody on Dox resistance induced by extracellular HMGB1 from non-viable Dox-treated cancer cells or recombinant HMGB1 was also investigated.ResultsImmunocytochemical analysis revealed that BCFs and NTFs were alpha-smooth muscle actin (ASMA) positive and cytokeratin 19 (CK19) negative cells: a phenotype consistent with that of activated fibroblasts. We confirmed that the CM from BCFs (but not NTFs), could significantly induce breast cancer cell migration. Intracellular HMGB1 expression was induced in BCF-CM-treated breast cancer cells and also in Dox-treated cells. Extracellular HMGB1 was strongly expressed in the CM after Dox-induced MDA-MB-231 cell death and was higher in cells pre-treated with BCF-CM than NTF-CM. Pre-treatment of breast cancer cells with BCF-CM induced a degree of resistance to Dox in accordance with the increased level of secreted HMGB1. Recombinant HMGB1 was shown to increase Dox resistance and this was associated with evidence of autophagy. Anti-HMGB1 neutralizing antibody significantly reduced the effect of extracellular HMGB1 released from dying cancer cells or of recombinant HMGB1 on Dox resistance.ConclusionsThese findings highlight the potential of stromal fibroblasts to contribute to chemoresistance in breast cancer cells in part through fibroblast-induced HMGB1 production.

Distal renal tubular acidosis and high urine carbon dioxide tension in a patient with southeast asian ovalocytosis

Southeast Asian ovalocytosis (SAO) is the best-documented disease in which mutation in the anion exchanger-1 (AE1) causes decreased anion (chloride [Cl-]/bicarbonate [HCO3-]) transport. Because AE1 is also found in the basolateral membrane of type A intercalated cells of the kidney, distal renal tubular acidosis (dRTA) might develop if the function of AE1 is critical for the net excretion of acid. Studies were performed in a 33-year-old woman with SAO who presented with proximal muscle weakness, hypokalemia (potassium, 2.7 mmol/L), a normal anion gap type of metabolic acidosis (venous plasma pH, 7. 32; bicarbonate, 17 mmol/L; anion gap, 11 mEq/L), and a low rate of ammonium (NH4+) excretion in the face of metabolic acidosis (26 micromol/min). However, the capacity to produce NH4+ did not appear to be low because during a furosemide-induced diuresis, NH4+ excretion increased almost threefold to a near-normal value (75 micromol/L/min). Nevertheless, her minimum urine pH (6.3) did not decrease appreciably with this diuresis. The basis of the renal acidification defect was most likely a low distal H+ secretion rate, the result of an alkalinized type A intercalated cell in the distal nephron. Unexpectedly, when her urine pH increased to 7.7 after sodium bicarbonate administration, her urine minus blood carbon dioxide tension difference (U-B Pco2) was 27 mm Hg. We speculate that the increase in U-B Pco2 might arise from a misdirection of AE1 to the apical membrane of type A intercalated cells.

Role of ERK1/2 signaling in dengue virus-induced liver injury.

The liver is considered to be an important organ of dengue virus (DENV) replication and pathogenesis. However, molecular mechanisms of hepatic injury are still poorly understood. Modulation of Mitogen Activated Protein Kinases (MAPKs) was previously shown to affect DENV-induced apoptosis of hepatocytes in vitro. However, the in vivo role of ERK1/2, a member of the MAPK family, and the question whether its activation can facilitate cell survival or cell death, has not been thoroughly investigated. Therefore, the role of ERK1/2 in a mouse model of DENV infection was examined. Our results show that DENV induces phosphorylation of ERK1/2 and increases apoptosis. Inhibition of phosphorylated ERK1/2 by the selective ERK1/2 inhibitor, FR180204, limits hepatocyte apoptosis and reduces DENV-induced liver injury. Clinical parameters, including leucopenia, thrombocytopenia, transaminases and histology, show improvements after FR180204 treatment. The expression of cell death genes was further identified using real-time PCR array and Western blot analysis. Caspase-3 was significantly decreased in FR180204 treated DENV-infected mice compared to the levels of untreated DENV-infected mice suggesting the role of ERK1/2 signaling in immune-mediated liver injury during DENV infection.

Estrogen is increased in male cholangiocarcinoma patients’ serum and stimulates invasion in cholangiocarcinoma cell lines in vitro

PurposeCholangiocarcinoma is defined as a chronic liver disease with altered estrogen metabolism and could result in estrogen retention. Estrogenic response was known as a promoting factor in progression of some cancer. In this study, we determined the significant increase of estrogen level in cholangiocarcinoma patients’ sera.MethodsThe estrogen levels in cholangiocarcinoma patients’ sera were measured and correlated with clinical presentations. Estrogen receptor-α expressions in cholangiocarcinoma tissues were detected by immunohistochemistry method. KKU-100 and KKU-M213 cholangiocarcinoma cell lines were treated with 17β-estradiol and tested the proliferative and invasive effects.ResultsThe estrogen levels showed positive correlations with serum bilirubin and alkaline phosphatase and a negative correlation with albumin. This study also showed an association with shorter survival times when patients with low and high serum estrogen levels were compared. In vitro studies demonstrated the effect of estrogen on cell proliferation and invasion in dose-dependent manners, which could be inhibited by tamoxifen, a clinical used estrogen antagonist. Invasion showed an association with the TFF1 gene expression and could be inhibited by small interfering RNA against TFF1 gene. Estrogen receptor-α was the main estrogen receptor that response to 17β-estradiol stimulation.ConclusionsTFF1 trefoil protein could be one of the effectors for estrogen-induced invasion in cholangiocarcinoma via the estrogen receptor-α. These findings could lead to an understanding of the mechanism of cholangiocarcinoma progression.

Clustering of patients with intrahepatic cholangiocarcinoma based on serum periostin may be predictive of prognosis

An effective serum biomarker may improve cholangiocarcinoma (CCA) management. Periostin (PN) has been demonstrated to be associated with aggressive CCA. The current study evaluated PN in blood serum for its diagnostic and prognostic potential in patients with CCA. Sera of 68 patients with CCA were collected prior to treatment, and PN levels were measured using an ELISA. Sera from 50 normal controls, 6 patients with benign liver diseases, 2 with hepatocellular carcinoma and 21 with breast cancer were analyzed. Immunohistochemistry of PN in CCA tissues was also investigated. The data were analyzed using the Mann-Whitney U test, Kaplan-Meier log rank tests, Cox proportional hazard regression models and Fishers exact tests. The median serum PN level in patients with CCA was significantly increased compared with that in healthy controls, patients with benign liver diseases and patients with breast cancer (all P<0.05). Using an optimal threshold value of 94 ng/ml PN, the diagnostic values for CCA compared with other conditions demonstrated a sensitivity level of 0.38 [95% confidence interval (CI), 0.27–0.51], specificity of 0.90 (95% CI, 0.81–0.96), accuracy of 0.66 (95% CI, 0.58–0.74), positive predictive value of 0.76 (95% CI, 0.59–0.89) and negative predictive value of 0.63 (95% CI, 0.53–0.72) (P<0.001). Furthermore, PN stain in stromal fibroblasts in CCA tissues was associated with serum PN levels (P=0.001), and patients with CCA were classified as low (≤94 ng/ml) or high PN (>94 ng/ml) accordingly. High serum and tissue PN levels were significantly associated with reduced survival rate (P<0.001 and P=0.033, respectively). Serum PN was an independent prognostic factor with a hazard ratio of 3.197 (P=0.001). In conclusion, serum PN may be used to divide patients with intrahepatic CCA into high and low PN groups. Elevated serum PN may be utilized as a marker of poor prognosis in patients with CCA.

The microRNA-15a-PAI-2 axis in cholangiocarcinoma-associated fibroblasts promotes migration of cancer cells

BackgroundCholangiocarcinoma (CCA) has an abundance of tumor stroma which plays an important role in cancer progression via tumor-promoting signals. This study aims to explore the microRNA (miRNA) profile of CCA-associated fibroblasts (CCFs) and the roles of any identified miRNAs in CCA progression.MethodsmiRNA expression profiles of CCFs and normal skin fibroblasts were compared by microarray. Identified downregulated miRNAs and their target genes were confirmed by real-time PCR. Their binding was confirmed by a luciferase reporter assay. The effects of conditioned-media (CM) of miRNA mimic- and antagonist-transfected CCFs were tested in CCA migration in wound healing assays. Finally, the levels of miRNA and their target genes were examined by real-time PCR and immunohistochemistry in clinical CCA samples.ResultsmiR-15a was identified as a downregulated miRNA in CCFs. Moreover, PAI-2 was identified as a novel target gene of miR-15a. Recombinant PAI-2 promoted migration of CCA cells. Moreover, CM from miR-15a mimic-transfected CCFs suppressed migration of CCA cells. Lower expression of miR-15a and higher expression of PAI-2 were observed in human CCA samples compared with normal liver tissues. Importantly, PAI-2 expression correlated with poor prognosis in CCA patients.ConclusionsThese findings highlight the miR-15a/PAI-2 axis as a potential therapeutic target in CCA patients.

The metabolic cross-talk between epithelial cancer cells and stromal fibroblasts in ovarian cancer progression: Autophagy plays a role

Cancer and stromal cells, which include (cancer‐associated) fibroblasts, adipocytes, and immune cells, constitute a mixed cellular ecosystem that dynamically influences the behavior of each component, creating conditions that ultimately favor the emergence of malignant clones. Ovarian cancer cells release cytokines that recruit and activate stromal fibroblasts and immune cells, so perpetuating a state of inflammation in the stroma that hampers the immune response and facilitates cancer survival and propagation. Further, the stroma vasculature impacts the metabolism of the cells by providing or limiting the availability of oxygen and nutrients. Autophagy, a lysosomal catabolic process with homeostatic and prosurvival functions, influences the behavior of cancer cells, affecting a variety of processes such as the survival in metabolic harsh conditions, the invasive growth, the development of immune and chemo resistance, the maintenance of stem‐like properties, and dormancy. Further, autophagy is involved in the secretion and the signaling of promigratory cytokines. Cancer‐associated fibroblasts can influence the actual level of autophagy in ovarian cancer cells through the secretion of pro‐inflammatory cytokines and the release of autophagy‐derived metabolites and substrates. Interrupting the metabolic cross‐talk between cancer cells and cancer‐associated fibroblasts could be an effective therapeutic strategy to arrest the progression and prevent the relapse of ovarian cancer.

Characterization of liposome-containing SPIONs conjugated with anti-CD20 developed as a novel theranostic agent for central nervous system lymphoma

Despite advances in neuroscience cancer research during the past decades, the survival of cancer patients has only marginally improved and the cure remains unlikely. The blood-brain barrier (BBB) is a major obstacle protecting the entry of therapeutic agents to central nervous system, especially for primary central nervous system lymphoma (PCNSL). Thus, the use of small nanoparticle as a drug carrier may be new strategies to overcome this problem. In this study, we fabricated liposome consisting of superparamagnetic iron oxide nanoparticles (SPIONs) functionalized with anti-CD20 (Rituximab; RTX). The designed nanoparticles have a theranostic property which is not only to improve drug delivery, but also to offer diagnostic and monitoring capabilities. TEM images revealed the spherical shape of liposome with the approximately average diameters about 140-190nm with slightly negatively charge surfaces. Superparamagnetic property of SPIONs-loaded liposomes was confirmed by VSM. Liposome colloidal could be prolonged at 4°C and 25°C storages. RTX conjugated liposome induced cell internalization and apoptosis effect in B-lymphoma cells. Drug targeting and therapeutic effect was investigated in BBB model. The result confirmed that liposome nanocarrier is required as a drug carrier for effectively RTX across the BBB.