Recently, it has been demonstrated that ultrafine particles (UFPs) are able to translocate from the lung into the systemic circulation. Precise mechanisms of the anatomical translocation (crossing the air-blood barrier) of inhaled UFPs at the alveolar wall are not fully understood. In this study, we examined the translocation pathway of the intratracheally instilled ultrafine carbon black (UFCB) from the lung into the blood circulation in mouse. Electron microscopy demonstrated accumulation of intratracheally instilled UFCB in the large-sized gaps developing between the cytoplasmic processes of the alveolar epithelial cells, possibly as a result of shrinkage of cytoplasm, by receiving stimulus/signals generated and released following UFCB attachment on the alveolar epithelial cells. Occasional penetration of the accumulated UFCB into the alveolar basement membrane, exposing to the air space, was observed at the gap. These results suggest that inhaled UFPs may, in part, pass the air-blood barrier through the large-sized gap formed between the alveolar epithelial cells.

Cancer-induced immunosuppression is a major problem as it reduces the anti-tumor effects of immunotherapies. In cancer tissues, cancer cells, immune cells, and other stromal cells interact and create an immunosuppressive microenvironment through a variety of immunosuppressive factors. Some cancer subpopulations such as cancer cells undergoing epithelial-mesenchymal transition and cancer stem-like cells have immunosuppressive and immunoresistant properties. The production of immunosuppressive factors by cancer cells is mechanistically attributed to oncogenic signals frequently activated in cancer cells, including the STAT3, MAPK, NF-κB, and Wnt/β-catenin signals, which are upstream events leading to immunosuppressive cascades. Moreover, some of these signals are also activated in immunosuppressive immune cells stimulated by cancer-derived factors and contribute to their immunosuppressive activities. Therefore, targeting these signals both in cancer cells and immunosuppressive immune cells may result in the restoration of immunocompetence in cancer patients and improve current immunotherapy.

To compare the pulmonary toxicity between ultrafine colloidal silica particles (UFCSs) and fine colloidal silica particles (FCSs), mice were intratracheally instilled with 3 mg of 14 nm UFCSs and 230 nm FCSs and pathologically examined from 30 minutes to 24 hour postexposure. Histopathologically, lungs exposed to both sizes of particles showed bronchiolar degeneration and necrosis, neutrophilic inflammation in alveoli with alveolar type II cell swelling and particle-laden alveolar macrophage accumulation. UFCSs, however, induced extensive alveolar hemorrhage compared to FCSs from 30 minutes onwards. UFCSs also caused more severe bronchiolar epithelial cell necrosis and neutrophil influx in alveoli than FCSs at 12 and 24 hours postexposure. Laminin positive immunolabellings in basement membranes of bronchioles and alveoli of UFCSs treated animals was weaker than those of FCSs-treated animals in all observation times. Electron microscopy demonstrated UFCSs and FCSs on bronchiolar and alveolar wall surface as well as in the cytoplasm of alveolar epithelial cells, alveolar macrophages and neutrophils. Type I alveolar epithelial cell erosion with basement membrane damage in UFCSs treated animals was more severe than those in FCSs-treated animals. At 12 and 24 hours postexposure, bronchiolar epithelial cells in UFCSs-treated animals showed more intense vacuolation and necrosis compared to FCSs-treated animals. These findings suggest that UFCSs have greater ability to induce lung inflammation and tissue damages than FCSs.

STAT3 signaling constitutes an important negative feedback mechanism for the maintenance of immune homeostasis, a suppressive signal for the Th1 immune response in murine macrophages, and a cancer immune evasion signal in various immune cells. The strategy for STAT3 signal inhibition should be considered, because these features could impede effective cancer immunotherapy. We have evaluated the effects of STAT3 inactivation in dendritic cells (DCs) on immune responses in mice and humans. DCs derived from LysMcre/STAT3flox/flox mice displayed higher cytokine production in response to TLR stimulation, activated T cells more efficiently, and were more resistant to the suppression of cytokine production by cancer-derived immunosuppressive factors compared with DCs from control littermates. Antitumor activities of STAT3-depleted and control DCs were compared by intratumoral administration of gp70 Ag peptide-pulsed DCs in the therapeutic MC38 tumor model. Intratumoral administration of STAT3-depleted DCs significantly inhibited MC38 tumor growth of both injected and nontreated remote tumors. The inhibition was accompanied by an increase in gp70-specific T cell response as well as in systemic Th1 immune response. STAT3-depleted human DCs with adenoviral STAT3 short hairpin RNA were also capable of producing more cytokines with TLR stimulation and more resistant to cancer-derived factors, and they induced tumor Ag-specific T cells more efficiently than control DCs. The identified role of DC STAT3 signaling in both in vivo therapeutic tumor models in mice and in vitro-specific T cell induction in humans indicates that STAT3-inactivated DCs may be a promising approach for cancer immunotherapy.

Lymph node metastasis is a poor prognostic factor for patients with head and neck squamous cell carcinoma (HNSCC). However, its molecular mechanism has not yet been fully understood. In our study, we investigated the expression of CCR4 and its ligand CCL22 in the HNSCC tumor microenvironment and found that the CCR4/CCL22 axis was involved in lymph node metastasis of HNSCC. CCR4 was expressed in 20 of 31 (64.5%) human tongue cancer tissues, and its expression was significantly correlated with lymph node metastasis (p < 0.01) and lymphatic invasion (p < 0.05 HNSCC is promoted by CCL22 in an autocrine or M2-like macrophage-dependent paracrine manner. Therefore, the CCR4/CCL22 axis may be an attractive target for the development of diagnostic and therapeutic strategies for patients with HNSCC.Head and neck squamous cell carcinoma (HNSCC) has a global incidence of 630,000 cases and causes 350,000 deaths annually. 1 Recent advances in multimodal combined therapy have improved loco-regional control of HNSCC.2 However, over the last decade, the overall 5-year survival rate of HNSCC has been only moderately improved, in part due to metastasis.2-4 Lymph nodes are the primary sites of metastasis of HNSCC before distant metastasis occurs.3-5 The presence of lymph node metastasis is strongly associated with poor clinical outcome in patients with HNSCC. 4,[6][7][8] Therefore, it is essential to elucidate the mechanisms underlying lymph node metastasis of HNSCC for improvement of diagnostic and therapeutic strategies for patients with HNSCC.Chemokines have recently been implicated in organspecific metastasis of cancer cells. The role of ectopic Key words: head and neck cancer, lymph node metastasis, CCR4, CCL22, macrophage Abbreviations: a/iLN: axilla and inguinal lymph nodes; ADCC: antibody dependent cellular cytotoxicity; cLN: cervical lymph nodes; FBS: fetal bovine serum; GFP: green fluorescent protein; HNSCC: head and neck squamous cell carcinoma; mAb: monoclonal antibody; pAb: polyclonal antibody; RT-PCR: reverse transcriptase-polymerase chain reaction; SD: standard deviations; siRNA: short-interfering RNA Additional Supporting Information may be found in the online version of this article.

Background:Although T-cell immunity is thought to be involved in the prognosis of epithelial ovarian cancer (EOC) patients, immunosuppressive conditions hamper antitumour immune responses. Thus, their mechanisms and overcoming strategies need to be investigated.Methods:The role of NF-κB in human EOC cells and macrophages was evaluated by in vitro production of immunosuppressive IL-6 and IL-8 by EOC cells and in vivo analysis of immune responses in nude mice implanted with human EOC cells using an NF-κB inhibitor DHMEQ.Results:In EOC patients, increased plasma IL-6, IL-8, and arginase were observed. The NF-κB inhibitor DHMEQ inhibited the production of IL-6 and IL-8 by EOC cell lines. Immunosuppression of human DCs and macrophages by culture supernatant of EOC cells was reversed with the pretreatment of DHMEQ. Administration of DHMEQ to nude mice implanted with human EOC resulted in the restoration of T-cell stimulatory activity of murine DCs along with the reduction of tumour accumulation and arginase expression of MDSCs. Nuclear factor-κB inhibition in tumour-bearing mice also enhanced antitumour effects of transferred murine naive T cells.Conclusions:NF-κB is involved in the immunosuppression induced by human EOC, and its inhibitor may restore antitumour immune responses, indicating that NF-κB is an attractive target for EOC treatment.

To improve current cancer immunotherapies, strategies to modulate various immunosuppressive cells including myeloid derived suppressor cells (MDSC) which were shown to be negative factors in immune‐checkpoint blockade therapy, need to be developed. In the present study, we evaluated the role of the local renin‐angiotensin system (RAS) in the tumor immune‐microenvironment using murine models bearing tumor cell lines in which RAS was not involved in their proliferation and angiogenetic ability. Giving angiotensin II receptor blockers (ARB) to C57BL/6 mice bearing murine colon cancer cell line MC38 resulted in significant enhancement of tumor antigen gp70 specific T cells. ARB administration did not change the numbers of CD11b+ myeloid cells in tumors, but significantly reduced their T‐cell inhibitory ability along with decreased production of various immunosuppressive factors including interleukin (IL)‐6, IL‐10, vascular endothelial growth factor (VEGF), and arginase by CD11b+ cells in tumors. ARB also decreased expression of immunosuppressive factors such as chemokine ligand 12 and nitric oxide synthase 2 in cancer‐associated fibroblasts (CAF). Last, combination of ARB and anti‐programmed death‐ligand 1 (PD‐L1) antibodies resulted in significant augmentation of anti‐tumor effects in a CD8+ T cell‐dependent way. These results showed that RAS is involved in the generation of an immunosuppressive tumor microenvironment caused by myeloid cells and fibroblasts, other than the previously shown proliferative and angiogenetic properties of cancer cells and macrophages, and that ARB can transform the immunosuppressive properties of MDSC and CAF and could be used in combination with PD‐1/PD‐L1 immune‐checkpoint blockade therapy.

Lymph node metastasis is the major clinicopathologic feature associated with poor prognosis in patients with head and neck squamous cell carcinoma (HNSCC). Here, web-based bioinformatics meta-analysis was performed to elucidate the molecular mechanism of lymph node metastasis of human HNSCC. Preferential upregulation of Myosin 1b (MYO1B) transcript in HNSCC datasets was identified. Myo1b mRNA was highly expressed in human HNSCC cells and patient tissue specimens compared with their normal counterparts as shown by quantitative PCR (qPCR) analyses. Immunohistochemistry (IHC)-detected Myo1b expression was significantly correlated with lymph node metastases in patients with oral cancer of the tongue. HNSCC with high expression of Myo1b and chemokine receptor 4 (CCR4), another metastasis-associated molecule, was strongly associated with lymph node metastasis. RNA interference (RNAi) of Myo1b in HNSCC cells, SAS and HSC4, significantly inhibited migratory and invasive abilities through decreased large protrusion formation of cell membranes. Finally, Myo1b knockdown in SAS cells significantly inhibited in vivo cervical lymph node metastases in a cervical lymph node metastatic mouse model system. Implications: Myo1b is functionally involved in lymph node metastasis of human HNSCC through enhanced cancer cell motility and is an attractive target for new diagnostic and therapeutic strategies for patients with HNSCC.