Induction of breast cancer stem cells by M1 macrophages through Lin-28B-let-7-HMGA2 axis.

Proinflammatory macrophage (M1) is now being suggested as a potential therapeutic strategy for cancer because of its tumoricidal capacity. However, few studies have been focused directly on the effects of M1 macrophages on cancer cells. Here, we found that M1 induced a subpopulation of CD44high/CD24-/low or ALDH1+ cells with CSC-like phenotypes from different types of breast cancer cells (BCCs) in a paracrine manner. Stat3/NF-κB pathways in BCCs were activated by proinflammatory cytokines, igniting Lin-28B-let-7-HMGA2 axis to induce CSC through epithelial-mesenchymal transition (EMT). Previously, we reported that Stat3-coordinated Lin-28B-let-7-HMGA2 axis initiated EMT in BCCs. Here, inhibition of Stat3/NF-κB pathways or Lin-28B-let-7-HMGA2 axis suppressed EMT/CSCs program. Notably, HMGA2 knockdown directly repressed M1-induced CSC formation and expression of Klf-4 and Nanog. Meanwhile, prolonged coculture with BCCs endowed M1 with M2 properties. M1 supernatant induced CSC from non-stem cancer cells, while M2 supernatant sustained a higher proportion of ALDH1+ cells. Our data suggest that macrophages might modulate CSC formation and maintenance by transferring between M1/M2 phenotype. Given that M1 are being considered as a promising immunotherapy tool, it is important to inhibit their CSC-inducing potential by targeting key molecules and pathways.

Psychological stress promotes neutrophil infiltration in colon tissue through adrenergic signaling in DSS-induced colitis model.

Inflammatory bowel disease (IBD) is a chronic intestinal inflammatory condition. Psychological stress has been postulated to affect the clinical symptoms and recurrence of IBD. The exact molecular mechanisms are not fully understood. In the present study, we demonstrate that psychological stress promotes neutrophil infiltration into colon tissues in dextran sulfate sodium (DSS)-induced colitis model. The psychological stress resulted in abnormal expression of the proinflammatory cytokines (IL-1ß, IL-6, IL-17A, and IL-22) and neutrophil chemokines (CXCL1 and CXCL2) and overactivation of the STAT3 inflammatory signaling pathway. Under chronic unpredictable stress, the adrenergic nervous system was markedly activated, as the expression of tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis, in bone marrow and colonic epithelium was enhanced, especially in the myenteric ganglia. The ß-AR agonist isoproterenol mimicked the effects of psychological stress on neutrophilia, neutrophil infiltration, and colonic damage in DSS-induced colitis. The ß1-AR/ß2-AR inhibitor propranolol reduced the numbers of the neutrophils in the circulation, suppressed neutrophil infiltration into colonic tissues, and attenuated the colonic tissue damage promoted by chronic stress. Propranolol also abolished stress-induced upregulation of proinflammatory cytokines and neutrophil chemokines. Our data reveal a close linkage between the ß1-AR/ß2-AR activation and neutrophil trafficking and also suggest the critical roles of adrenergic nervous system in exacerbation of inflammation and damage of colonic tissues in experimental colitis. The current study provides a new insight into the mechanisms underlying the association of psychological stress with excessive inflammatory response and pathophysiological consequences in IBD. The findings also suggest a potential application of neuroprotective agents to prevent relapsing immune activation in the treatment of IBD.

A Her2-let-7-ß2-AR circuit affects prognosis in patients with Her2-positive breast cancer.

BACKGROUND: Our previous studies show that ß2-adrenergic receptor (ß2-AR) is highly expressed in most Her2-overexpressing breast cancers. However, the mechanisms underlying upregulation of the ß2-AR expression in Her2-overexpressing breast cancer cells are not fully understood. The clinical significance of the ß2-AR overexpression in breast cancer is unclear. METHODS: Human breast cancer cells MCF-7 and MCF-7/Her2 were transfected with the let-7 mimics or inhibitors. The expression of ß2-AR was analyzed by Western blot. The ß2-AR status in primary and metastatic sites of breast cancer and the human breast cancer tissue microarrays containing 49 primary tumors and 50 metastatic lymph node tissues was analyzed by immunohistochemistry. The correlation of lymph node metastasis with the ß2-AR level was determined in 59 primary tumor tissues from the patients with Her2-positive breast cancer. The clinical prognostic significance of the ß2-AR overexpression in the patients with Her2-positive breast cancers was evaluated by a retrospective study. RESULTS: The let-7f level in Her2-overexpressing breast cancer cells SKBR3 and BT474 was significantly lower than that in MCF-7 cells, which express low level of Her2. Ectopic expression of Her2 in MCF-7 cells (MCF-7/Her2) represses the expression of microRNA let-7f, which is previously identified to regulate baseline ß2-AR expression. The treatment with MEK1/2 inhibitors PD98059 or PD184352 effectively restored the let-7f level, suggesting that Her2-overexpression-mediated ERK constitutive activation inhibited let-7f, leading to the upregulation of the ß2-AR expression. The transfection with the let-7f mimics markedly downregulated the ß2-AR level, whereas the let-7 inhibitor significantly upregulated the ß2-AR expression in both parental MCF-7 and MCF-7/Her2 cells. In addition, treatment of MCF-7/Her2 cells with isoproterenol resulted in a concentration-dependent reduction of the let-7f expression, demonstrating that the inhibitory effect of Her2 overexpression on let-7f can be reinforced by agonist-triggered ß2-AR activation. We further demonstrate that high level of ß2-AR associates with lymph node metastasis and poor outcome in the patients with Her2-positive breast cancer. CONCLUSIONS: The mutual and reciprocal interaction between Her2, ß2-AR, and let-7f may maintain a high level of ß2-AR in breast cancer cells. Our data suggest that ß2-AR may be a new useful biomarker for predicting prognosis in Her2-positive breast cancer and may also be a promising selective therapeutic target for the aggressive subtype of breast cancer.

Acquisition of resistance to trastuzumab in gastric cancer cells is associated with activation of IL-6/STAT3/Jagged-1/Notch positive feedback loop.

In the present study, we demonstrate that prolonged treatment by trastuzumab induced resistance of NCI-N87 gastric cancer cells to trastuzumab. The resistant cells possessed typical characteristics of epithelial to mesenchymal transition (EMT)/cancer stem cells and acquired more invasive and metastatic potentials both in vitro and in vivo. Long term treatment with trastuzumab dramatically inhibited the phosphorylation of Akt, but triggered the activation of STAT3. The level of IL-6 was remarkably increased, implicating that the release of IL-6 that drives the STAT3 activation initiates the survival signaling transition. Furthermore, the Notch activities were significantly enhanced in the resistant cells, companied by upregulation of the Notch ligand Jagged-1 and the Notch responsive genes Hey1 and Hey2. Inhibiting the endogenous Notch pathway reduced the IL-6 expression and restored the sensitivities of the resistant cells to trastuzumab. Blocking of the STAT3 signaling abrogated IL-6-induced Jagged-1 expression, effectively inhibited the growth of the trastuzumab resistant cells, and enhanced the anti-tumor activities of trastuzumab in the resistant cells. These findings implicate that the IL-6/STAT3/Jagged-1/Notch axis may be a useful target and that combination of the Notch or STAT3 inhibitors with trastuzumab may prevent or delay clinical resistance and improve the efficacy of trastuzumab in gastric cancer.

Adrenergic signaling promotes angiogenesis through endothelial cell-tumor cell crosstalk.

Angiogenesis is an important factor in invasive tumor growth, progression, and metastasis. Multiple proangiogenic mechanisms are involved in tumor angiogenesis. In this study, we showed that the neurotransmitter norepinephrine upregulated VEGF (VEGFA) expression in breast cancer cells and that the culture supernatant from norepinephrine-treated breast cancer cells promoted the formation of the capillary-like network of endothelial cells. However, the effects of norepinephrine were further enhanced when the endothelial cells were cocultured with breast cancer cells, indicating a critical role of tumor cell-endothelial cell contacts in norepinephrine-induced tumor angiogenesis. Interestingly, norepinephrine dramatically induced the activation of the Notch pathway, which is a cell-contact-mediated intercellular signaling pathway and tightly linked to tumor cell-stromal cell interaction and angiogenesis, in the endothelial cells that had been cocultured with breast cancer cells. Furthermore, the expression of the Notch ligand Jagged 1 was significantly upregulated by norepinephrine at both mRNA and protein levels in breast cancer cells. Inhibitors of ß2-adrenergic receptor (ß2-AR), protein kinase A (PKA), and mTOR could reverse norepinephrine-induced Jagged 1 upregulation, indicating that the ß2-AR-PKA-mTOR pathway participates in this process. Knockdown of Jagged 1 expression in breast cancer cells not only repressed norepinephrine-induced activation of the Notch pathway in cocultured endothelial cells but also evidently impaired the effects of norepinephrine on capillary-like sprout formation. These data demonstrate that tumor angiogenesis mediated by the Jagged 1/Notch intercellular signaling is governed by the norepinephrine-activated ß2-AR-PKA-mTOR pathway.