OBJECTIVE: To reveal the mechanisms by which miR-513b-5p inhibits metastasis of colon cancer stem cells (CCSCs) through IL-6/STAT3 in HCT116 cells.
METHODS: Sphere formation media and magnetic cell sorting were used to enrich and screen CCSCs. We used a colony formation assay, cell proliferation and viability assays, and a nude mouse transplantation tumor assay to identify CCSCs. ELISA was performed to identify IL-6 in the cell culture medium, and the growth, viability, wound healing, and transwell migration of distinct cell groups were compared to differentiate them. Dual-luciferase reporter assay, RT-PCR, and/or Western Blot analysis were conducted to determine the correlation between them.
RESULTS: CD133+CD44+ HCT116 cells were shown to have higher cloning efficiency, greater proliferation ability and viability, and stronger tumorigenicity. A dual-luciferase reporter assay revealed that miR-513b-5p negatively affected STAT3 expression. RT-PCR and/or Western Blot analysis suggested that miR-513b-5p negatively affected STAT3 and Vimentin, while positively affecting E-cadherin expression. The STAT3 overexpression vector + miR-513b-5p inhibitor cell group had the highest efficiency, greatest proliferation ability and viability, and the highest IL-6 level in the experiments.
CONCLUSIONS: Mir-513b-5p inhibited the epithelial-mesenchymal transition (EMT) of CCSCs through IL-6/STAT3. This potential mechanism may provide a new therapeutic target for colon cancer.

UNASSIGNED: To further investigate why curcumin (CUR) can attenuate psoriasis-like dermatitis of mice.
UNASSIGNED: Sixteen mice were randomized into four groups. The control group used carrier cream, and the model and the CUR group were applied with topical 5% imiquimod in the naked mice skin once a day for 6 days (62.5 mg/day/mice). Meanwhile, the control and model mice were given the same dose of saline by oral means, while mice in the CUR groups received oral drug doses of 50 and 100 mg/kg once a day for 6 days, respectively. CUR could largely improve imiquimod-induced lesions of mice. By using the ELISA and qPCR, we found that the protein and mRNA levels of epidermal TNF-α and IL-6 were inhibited by CUR. The phosphorylation levels of STAT3 and its downstream associated protein levels (eg, Cyclin D1, Bcl-2 and Pim1) in skin tissues of different groups were also inhibited by CUR. Furthermore, the results of immunohistochemistry also showed the repressed effect of CUR for the expression of TNF-α, IL-6 and p-STAT3 in psoriasis-like lesions of mice.
UNASSIGNED: CUR can effectively ameliorate the featured lesions of psoriasis mice, which may be closely associated with the involvement of IL-6/STAT3 signaling.

Background: Prostate cancer is the most common malignant tumor in men, accounting for one of the top five cancer incidences worldwide. However, there is no effective pharmacological treatment for advanced prostate cancer (APC). Herein, we aim to investigate the mechanism of Zhoushi Qiling decoction (ZQD), a traditional Chinese medicine compound, in inhibiting prostate cancer cells proliferation and tumor growth. Methods: IC50 was determined in PC3 and DU145 cells. Cell viability was determined using MTT assay after interleukin (IL) 6 stimulation. Cell proliferation ability was evaluated using colony formation assay. IL-6/signal transducer and activator of transcription 3 (STAT3) signaling pathway was analyzed using qRT-PCR and Western blot in PC3 and DU145 cells and xenograft tumor tissues. Results: It was found that ZQD significantly inhibited Il-6-induced cell viability and proliferation in PC3 and DU145 cells. Moreover, ZQD significantly reduced mRNA levels of IL-6, IL-1β, STAT3, Bcl2, and CyclinD1, stimulated by IL-6. The protein levels of p-STAT3, Bcl2 and CyclinD1 were reduced by ZQD treatment at 40 mg/mL both in PC3 and DU145 cells. Additionally, in xenograft tumor tissues, tumor volume, weight and proliferation were significantly reduced by ZQD treatment. In addition, the mRNA and protein levels of IL-6 and pSTAT3 were significantly inhibited by ZQD treatment in vivo. Conclusion: We demonstrate that ZQD can effectively reduce cell proliferation and tumor growth by inhibiting the activation of IL-6/STAT3 signaling pathway.

The association between oral microbiota and cancer development has been a topic of intense research in recent years, with compelling evidence suggesting that the oral microbiome may play a significant role in cancer initiation and progression. However, the causal connections between the two remain a subject of debate, and the underlying mechanisms are not fully understood. In this case-control study, we aimed to identify common oral microbiota associated with several cancer types and investigate the potential mechanisms that may trigger immune responses and initiate cancer upon cytokine secretion. Saliva and blood samples were collected from 309 adult cancer patients and 745 healthy controls to analyze the oral microbiome and the mechanisms involved in cancer initiation. Machine learning techniques revealed that six bacterial genera were associated with cancer. The abundance of Leuconostoc, Streptococcus, Abiotrophia, and Prevotella was reduced in the cancer group, while abundance of Haemophilus and Neisseria enhanced. G protein-coupled receptor kinase, H+-transporting ATPase, and futalosine hydrolase were found significantly enriched in the cancer group. Total short-chain fatty acid (SCFAs) concentrations and free fatty acid receptor 2 (FFAR2) expression levels were greater in the control group when compared with the cancer group, while serum tumor necrosis factor alpha induced protein 8 (TNFAIP8), interleukin-6 (IL6), and signal transducer and activator of transcription 3 (STAT3) levels were higher in the cancer group when compared with the control group. These results suggested that the alterations in the composition of oral microbiota can contribute to a reduction in SCFAs and FFAR2 expression that may initiate an inflammatory response through the upregulation of TNFAIP8 and the IL-6/STAT3 pathway, which could ultimately increase the risk of cancer onset.

The tumor microenvironment is a complex and dynamic ecosystem composed of various physical cues and biochemical signals that facilitate cancer progression, and tumor-associated macrophages are especially of interest as a treatable target due to their diverse pro-tumorigenic functions. Engineered three-dimensional models of tumors more effectively mimic the tumor microenvironment than monolayer cultures and can serve as a platform for investigating specific aspects of tumor biology within a controlled setting. To study the combinatorial effects of tumor-associated macrophages and microenvironment mechanical properties on osteosarcoma, we co-cultured human osteosarcoma cells with macrophages within biomaterials-based bone tumor niches with tunable stiffness. In the first 24 h of direct interaction between the two cell types, macrophages induced an inflammatory environment consisting of high concentrations of tumor necrosis factor alpha (TNFα) and interleukin (IL)-6 within moderately stiff scaffolds. Expression of Yes-associated protein (YAP), but not its homolog, transcriptional activator with PDZ-binding motif (TAZ), in osteosarcoma cells was significantly higher than in macrophages, and co-culture of the two cells slightly upregulated YAP in both cells, although not to a significant degree. Resistance to doxorubicin treatment in osteosarcoma cells was correlated with inflammation in the microenvironment, and signal transducer and activator of transcription 3 (STAT3) inhibition diminished the inflammation-related differences in drug resistance but ultimately did not improve the efficacy of doxorubicin. This work highlights that the biochemical cues conferred by tumor-associated macrophages in osteosarcoma are highly variable, and signals derived from the immune system should be considered in the development and testing of novel drugs for cancer.

Atractylodes macrocephala Koidz. is one of the most frequently used traditional Chinese medicines for the treatment of ulcerative colitis (UC). The beneficial effect of polysaccharide from Atractylodes macrocephala Koidz. (PAMK) on UC has been reported, while the underlying mechanism and target remain unclear. In this study, we systematically investigated the therapeutic effect and the underlying mechanism of PAMK in UC based on a mouse model of dextran sodium sulfate (DSS)-induced colitis. PAMK treatment (100 mg/kg, 200 mg/kg and 400 mg/kg) significantly ameliorated DSS-induced colitis, manifested as a reduction in weight loss, disease activity index (DAI), colon shortening, spleen index and histological score. Moreover, PAMK treatment inhibited inflammation and improved the integrity of the intestinal barrier in colitis mice. Mechanistically, microarray analysis determined the critical role of the immunoregulatory effect of PAMK in alleviating UC. Flow cytometry analysis further demonstrated that PAMK treatment regulated the balance between T helper (Th) 17 and regulatory T (Treg) cells in the mesenteric lymph nodes (MLN) and spleen in mice with colitis. In addition, PAMK treatment downregulated the expression of IL-6 and suppressed the phosphorylation of STAT3. Together, these data revealed that PAMK treatment alleviated DSS-induced colitis by regulating the Th17/Treg cell balance, which may be dependent on the inhibition of the IL-6/STAT3 signaling pathway. Our study is the first to elucidate that the underlying mechanism by which PAMK treatment alleviates DSS-induced colitis is associated with an improved the Th17/Treg cell balance. Collectively, the study provides evidence for the potential of PAMK to treat UC.

Background & Aims: Peroxisome proliferator-activated receptor α (PPARα) is a ligand-activated transcription factor abundantly expressed in liver. PPARα activator has been previously reported to protect against acetaminophen-induced hepatotoxicity, but fenofibrate, a lipid-lowering drug that activates PPARα, has a common side-effect causing liver injury. Thus, the exact effect of liver PPARα on drug-induced liver injury remains obscure. Methods: Hepatocyte-specific Ppara knockout mice and littermate wild-type control mice were intraperitoneally injected with acetaminophen (400 mg/kg body weight). Blood and liver samples were collected at different time points. We measured phase I and II cytochrome P450 enzymes, glutathione, reactive oxygen species, cytokines including Il6, and pSTAT3 by reverse transcriptase quantitative PCR, colorimetric, immunohistochemistry analyses and Western blotting. Results: Hepatic expression of PPARα was significantly decreased in DILI patients. Disruption of the Ppara gene in hepatocytes significantly reduced acetaminophen-induced liver injury in mice. ROS production rather than the expression levels of phase I and II cytochrome P450 enzymes was reduced in hepatocyte-specific Ppara knockout mice compared to control mice after acetaminophen administration. Mechanistically, hepatocyte-specific Ppara knockout mice had upregulated activation of the hepatoprotective pathway IL-6/STAT3 compared to wild-type mice, as evidenced by hepatic Il6 mRNA levels, hepatic protein levels of STAT3 and phosphorylated STAT3 were much higher in hepatocyte-specific Ppara knockout mice than in wild-type mice post acetaminophen injection. Conclusions: Hepatocyte-specific disruption of the Ppara gene protects against acetaminophen-induced liver injury by reducing oxidative stress and upregulating the hepatoprotective IL-6/STAT3 signaling pathway.

Numerous studies have found that chronic stress could promote tumor progression and this may be related to inhibtion of immune system. Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of cells with immunosuppressive activity. MDSCs may represent a key link between chronic stress and tumor progression. However, the role of stress-induced MDSCs in breast cancer progression is unclear. The present study showed that pre-exposure of chronic stress could lead to MDSCs elevation and facilitated breast cancer metastasis in tumor-bearing mice. Adoptive transfer of MDSCs could significantly increase lung metastatic foci. In contrast, lung metastasis could be alleviated by depleting endogenous MDSCs with Gr-1 antibody. The concentration of norepinephrine in serum and the expression of tyrosine hydroxylase in bone marrow could be significantly elevated by chronic stress. Moreover, propranolol, an inhibitor of β-adrenergic signaling, could inhibit breast carcinoma metastasis and prevent the expansion of chronic stress-induced MDSCs. Further study revealed that the expressions of IL-6 and JAK/STAT3 signaling pathways were upregulated by chronic stress in mice, and this upregulation could be inhibited by propranolol. Blocking the IL-6 signal or inhibiting the activation of the JAK/STAT3 signaling pathway could reduce tumor growth and metastasis by attenuating the accumulation of MDSCs in vivo. Besides, propranolol inhibited the expression of IL-6 in supernatant of 4T1 cells induced by isoproterenol and reduced the proportion of inducible MDSCs in vitro. Taken together, these data indicated that chronic stress may accumulate MDSCs via activation of β-adrenergic signaling and IL-6/STAT3 pathway, thereby promoting breast carcinoma metastasis.

BACKGROUND: O-linked β-N-acetylglucosamine transferase (O-GlcNAc transferase, OGT) is a key enzyme that regulates O-GlcNAc modification, which is significantly up-regulated and participates in the regulation of tumorigenesis. Although previous research indicated that OGT promotes epithelial-mesenchymal transition (EMT) of lung cancer, the underlying molecular mechanisms, especially within the tumor inflammatory microenvironment, require further elucidation.
METHODS: The role of the inflammatory signaling Interleukin 6/Signal Transducer and activator of transcription 3 (IL-6/STAT3) in Non-small cell lung cancer (NSCLC) cells A549 were confirmed by Transwell assay, Scratch wound healing assay, Western blot, Immunofluorescence staining, and Nuclear and cytoplasmic extraction experiment. Western blot detected OGT expression and whole protein O-GlcNacylation after IL-6 stimulation in NSCLCs cells. The biological effects and related mechanism of OGT in NSCLC cells were investigated by Western blot, Transwell assay, Immunofluorescence staining and Immunoprecipitation. The up-stream mechanism of OGT expression was explored by employing the specific chemical inhibitors, and the expression and distribution of OGT and phosphorylated STAT3 in NSCLC samples were confirmed by immunohistochemical analysis.
RESULTS: IL-6/STAT3 promoted the migration and invasion of NSCLC cells. IL-6 stimulation elevated OGT expression and the total protein O-GlcNacylation in A549 cells. Silencing OGT by shRNA significantly inhibited the IL-6 induced EMT marker (N-cadherin and Slug) expression, migration and invasion in A549 cells. OGT interacted with and mediated O-GlcNacylation of STAT3, which promoted STAT3 Y705 phosphorylation in IL-6 treated NSCLC cells. OGT expression was positively regulated by NF-κB p65 signaling pathway after IL-6 stimulation, instead of STAT3 signaling. OGT and phosphorylated STAT3 had an obviously higher expression in human NSCLC tissues, and phosphorylated STAT3 was mainly expressed in the nucleus.
CONCLUSIONS: The above results showed that OGT regulated O-GlcNacylation promoted migration and invasion by activating IL-6/STAT3 signaling in lung cancer.

Currently available therapies for hepatocellular carcinoma (HCC), with a high morbidity and high mortality, are only marginally effective and with sharp adverse side effects, which makes it compulsory to explore novel and more effective anticancer molecules. Chinese medicinal herbs exhibited prominent anticancer effects and were applied to supplement clinical cancer treatment. Here, we reported a compound, trilobolide-6-O-isobutyrate (TBB), isolated from the flowers of Wedelia trilobata with a markedly cytotoxic effect on HCC cells. We found that TBB time- and dose-dependently inhibited HCC cells\' growth and colony formation in vitro. Moreover, TBB induced cell cycle arrest at the G2/M phase, mitochondrial caspase-dependent apoptosis, and suppressed migration and invasion, as well as the glycolysis of HCC cells. Mechanistically, our data indicated that TBB inhibited the STAT3 pathway activation by directly interacting with the TYR 640/657 sites of the STAT3 protein and decreasing the level of p-STAT3. TBB also regulated the expression of PCNA, Ki67, Cyclin B1, Cyclin E, Bax, Bcl2, MMP2/9, and PGK1 through the inhibition of the IL-6/STAT3 signaling pathway. Lastly, we confirmed that TBB effectively eliminated tumor growth without causing overt toxicity to healthy tissues in the xenograft tumor model. The exploration of anticancer activity and the underlying mechanism of TBB suggested its usage as a promising chemotherapeutic agent for HCC.