Testicular teratomagenesis from primordial germ cells with overexpression of germinal center-associated nuclear protein.

Testicular teratomas are the major histologic type of testicular germ cell tumors and their incidence continues to grow. Moreover, teratomas can develop from undifferentiated cells in induced pluripotent stem (iPS) cell transplantation therapy, seriously hampering the progress of regenerative medicine. Germinal center-associated nuclear protein (GANP) is thought to be important to the biogenetic control of primordial germ cells and is among the genes susceptible to testicular germ cell tumors. Thus, we analyzed the expression of GANP in human testicular postpubertal-type teratomas and established a novel mouse model to reveal the association between GANP and teratomagenesis. We analyzed 31 cases of human testicular postpubertal-type teratomas and, in all cases, GANP was overexpressed. The aberrant expression was also detected in germ cell neoplasia in situ accompanied by the teratoma. GANP expression was particularly high in the epithelia of the epidermis, cutaneous appendages, and trachea-like ciliated epithelium. To further clarify the association between GANP and teratomagenesis, we established a novel teratomagenesis mouse model (CAG-ganpTg mice). In the GANP-teratoma mice, GANP-overexpressing teratomas were more frequent at the testes and the middle portion of the uterus than has been seen in the previously established mouse models. In conclusion, GANP is overexpressed in testicular postpubertal-type teratomas and is an essential teratomagenic factor. We also found that CAG-ganpTg mice are useful mouse models of teratomagenesis that mimics human midline teratomas and that teratomas may originate from the overexpression of GANP in primordial germ cells.

TIM-4 glycoprotein-mediated degradation of dying tumor cells by autophagy leads to reduced antigen presentation and increased immune tolerance.

Phagocytosis of apoptotic cells by myeloid cells has been implicated in the maintenance of immune homeostasis. In this study, we found that T cell immunoglobulin- and mucin domain-containing molecule-4 (TIM-4) repressed tumor-specific immunity triggered by chemotherapy-induced tumor cell death. TIM-4 was found to be highly expressed on tumor-associated myeloid cells such as macrophages (TAMs) and dendritic cells (TADCs) and danger-associated molecular patterns (DAMPs) released from chemotherapy-damaged tumor cells induced TIM-4 on tumor-associated myeloid cells recruited from bone marrow-derived precursors. TIM-4 directly interacted with AMPKα1 and activated autophagy-mediated degradation of ingested tumors, leading to reduced antigen presentation and impaired CTL responses. Consistently, blockade of the TIM-4-AMPKα1-autophagy pathway augmented the antitumor effect of chemotherapeutics by enhancing tumor-specific CTL responses. Our finding provides insight into the immune tolerance mediated by phagocytosis of dying cells, and targeting of the TIM-4-AMPKα1 interaction constitutes a unique strategy for augmenting antitumor immunity and improving cancer chemotherapy.

Cancer therapy with major histocompatibility complex-deficient and interferon ß-producing myeloid cells derived from allogeneic embryonic stem cells.

We previously established a method to generate myeloid cells with a proliferative capability from pluripotent stem cells and designated them iPS-ML. Human iPS-ML cells share features with physiological macrophages including the capability to infiltrate into cancer tissues. We observed therapeutic effects of human iPS-ML cells expressing interferon ß (iPS-ML/interferon (IFN)-ß) in xenograft cancer models. However, assessment of host immune system-mediated therapeutic and adverse effects of this therapy is impossible by xenograft models. We currently evaluated the therapeutic effects of a mouse equivalent of human iPS-ML/IFN, a mouse embryonic stem (ES) cell-derived myeloid cell line producing IFN (ES-ML/IFN). The ES-MLs producing IFN-ß (ß-ML) and IFN-γ (γ-ML) and originating from E14 ES cells derived from the 129 mouse strain (H-2b ) were generated, and the MHC (H-2Kb , Db , and I-Ab ) genes of the ES-ML/IFN were disrupted using the clustered regularly interspaced short palindromic repeats (CRISPR)/CAS9 method. We used the ES-ML/IFN to treat allogeneic BALB/c mice (H-2d ) transplanted with Colon26 cancer cells. Treatment with ß-ML but not with γ-ML cells repressed the growth of colon cancer in the peritoneal cavity and liver. The transferred ES-ML/IFN infiltrated into cancer tissues and enhanced infiltration of T cells into cancer tissues. ES-ML/IFN therapy increased the number of immune cells in the lymphoid organs. Sensitization of both cancer antigen-specific CD8+ T cells and natural killer (NK) cells were enhanced by the therapy, and CD8+ T cells were essential for the therapeutic effect, implying that donor MHC-deficient ß-ML exhibited a therapeutic effect through the activation of host immune cells derived from allogeneic recipient mice. The results suggested the usefulness of HLA-deficient human iPS-ML/IFN-ß cells for therapy of HLA-mismatched allogeneic cancer patients.

Inhibition of Local Macrophage Growth Ameliorates Focal Inflammation and Suppresses Atherosclerosis.

OBJECTIVE: Macrophages play a central role in various stages of atherosclerotic plaque formation and progression. The local macrophages reportedly proliferate during atherosclerosis, but the pathophysiological significance of macrophage proliferation in this context remains unclear. Here, we investigated the involvement of local macrophage proliferation during atherosclerosis formation and progression using transgenic mice, in which macrophage proliferation was specifically suppressed. APPROACH AND RESULTS: Inhibition of macrophage proliferation was achieved by inducing the expression of cyclin-dependent kinase inhibitor 1B, also known as p27kip, under the regulation of a scavenger receptor promoter/enhancer. The macrophage-specific human p27kip Tg mice were subsequently crossed with apolipoprotein E-deficient mice for the atherosclerotic plaque study. Results showed that a reduced number of local macrophages resulted in marked suppression of atherosclerotic plaque formation and inflammatory response in the plaque. Moreover, fewer local macrophages in macrophage-specific human p27kip Tg mice helped stabilize the plaque, as evidenced by a reduced necrotic core area, increased collagenous extracellular matrix, and thickened fibrous cap. CONCLUSIONS: These results provide direct evidence of the involvement of local macrophage proliferation in formation and progression of atherosclerotic plaques and plaque stability. Thus, control of macrophage proliferation might represent a therapeutic target for treating atherosclerotic diseases.

Evaluation of neurobehavioral impairment in methylmercury-treated KK-Ay mice by dynamic weight-bearing test.

Methylmercury (MeHg) is known to cause neurobehavioral impairment in human and experimental animals. We previously reported that MeHg (5 mg Hg/kg) induced severe neurobehavioral dysfunction in 4-week-old KK-Ay mice, although it is difficult to evaluate quantitatively the neurobehavioral impairment in MeHg-treated KK-Ay mice because of their obesity. The aim of this study was to evaluate MeHg-induced neurobehavioral dysfunction in KK-Ay mice using the dynamic weight-bearing test, which analyzes the animal's weight distribution between the four limbs. Male 12-week-old KK-Ay mice were treated with MeHg (5 mg Hg/kg) three times per week for 5 weeks. Body weight loss began after approximately 2 weeks of MeHg treatment, and decreased significantly at 4 weeks. Seven of the nine MeHg-treated mice exhibited overt neurological symptoms such as ataxia and gait disturbance. The weight-bearing load was lower for the forelimb than for the hindlimb at baseline and until 1 week after MeHg treatment was initiated. In weeks 2-4, the dynamic weight-bearing loads on the forelimb and hindlimb were similar. The load on the forelimb exceeded the load on the hindlimb after 5 weeks of treatment. This finding indicates that the dynamic weight-bearing test is useful for semi-quantitative evaluation of neurobehavioral impairment in MeHg-treated rodents, and is less stressful for the animals. Infiltration of CD204-positive macrophages was observed in the sciatic nerve of MeHg-treated mice, suggesting that CD204 can serve as a useful marker of tissue injury in peripheral nerves and a possible target in regenerating peripheral nerves and controlling neuropathies.

Mtu1-Mediated Thiouridine Formation of Mitochondrial tRNAs Is Required for Mitochondrial Translation and Is Involved in Reversible Infantile Liver Injury.

Reversible infantile liver failure (RILF) is a unique heritable liver disease characterized by acute liver failure followed by spontaneous recovery at an early stage of life. Genetic mutations in MTU1 have been identified in RILF patients. MTU1 is a mitochondrial enzyme that catalyzes the 2-thiolation of 5-taurinomethyl-2-thiouridine (τm5s2U) found in the anticodon of a subset of mitochondrial tRNAs (mt-tRNAs). Although the genetic basis of RILF is clear, the molecular mechanism that drives the pathogenesis remains elusive. We here generated liver-specific knockout of Mtu1 (Mtu1LKO) mice, which exhibited symptoms of liver injury characterized by hepatic inflammation and elevated levels of plasma lactate and AST. Mechanistically, Mtu1 deficiency resulted in a loss of 2-thiolation in mt-tRNAs, which led to a marked impairment of mitochondrial translation. Consequently, Mtu1LKO mice exhibited severe disruption of mitochondrial membrane integrity and a broad decrease in respiratory complex activities in the hepatocytes. Interestingly, mitochondrial dysfunction induced signaling pathways related to mitochondrial proliferation and the suppression of oxidative stress. The present study demonstrates that Mtu1-dependent 2-thiolation of mt-tRNA is indispensable for mitochondrial translation and that Mtu1 deficiency is a primary cause of RILF. In addition, Mtu1 deficiency is associated with multiple cytoprotective pathways that might prevent catastrophic liver failure and assist in the recovery from liver injury.

Positive correlation between the density of macrophages and T-cells in undifferentiated sarcoma.

Undifferentiated sarcoma (US) is a frequent soft tissue sarcoma. Although the 10-year survival rate is around 60%, advanced US is highly resistant to chemo/radiotherapy. The tumor microenvironment (TME) is closely associated with tumor progression. However, few studies of infiltrated immune cells in US have been published. In this study, we evaluated tumor-associated macrophages (TAMs) and CD8-positive cytotoxic T lymphocytes (CTLs) in 28 cases of US. Iba1, CD163, and CD204 were used as markers for TAMs. The density of CTLs was positively correlated with the density of TAMs. However, a negative correlation was seen between the density of CTLs and the percentage of CD204-positive TAMs. We found no significant association between the density of Iba1-/CD204-/CD8-positive cells and clinicopathological factors. No significant correlation between immune cell infiltration and clinical outcome was observed. Although we found no significant association between immune cells and clinicopathological factors, these findings may provide new insight into the characterization of immune cells in the TME of US.

MUC1/KL-6 expression confers an aggressive phenotype upon myeloma cells.

The sialic glycoprotein, MUC1, is known to be involved in the pathogenesis of various types of cancers. KL-6 is one of the surface antigens of MUC1 and also a marker of interstitial pneumonitis. A fraction of patients with myeloma (3.9%) have elevated serum KL-6 levels without any evidence of interstitial pneumonitis and their myeloma cells have high MUC1 expression. We established a myeloma cell line designated EMM1 from a patient with multiple myeloma accompanied with elevated serum KL-6. EMM1 cells expressed high levels of MUC1 compared with other myeloma cell lines. Knockdown of MUC1 in EMM1 cells induced cell cycle arrest during S phase and apoptosis, suggesting that the MUC1 expression is involved in accelerated growth of EMM1 cells. RNA-seq analysis suggests that MUC1 expression activates k-ras and TNFα-induced NFκB pathways in EMM1 cells. We injected EMM1 cells subcutaneously into Rag2-/-Jak3-/- Balb/c mice to establish a mouse xenograft model. These mice had aggressive tumor growth that was accompanied by high serum KL-6 levels. In addition, MUC1 knockdown in EMM1 cells led to inhibited tumor growth. These findings demonstrate that MUC1 serves as a potential target for developing drugs for treatment of patients with KL-6+ myeloma, and EMM1 cells and EMM1-engrafted mice are useful tools for the development of such novel agents.

CAFs and TAMs: maestros of the tumour microenvironment.

Many non-tumour host cells such as inflammatory cells, fibroblasts, and endothelial cells are present in the tumour microenvironment and affect the malignant potential and chemo-resistance of the tumour cells. Macrophages and fibroblasts are the main components of infiltrating stromal cells and are referred to as tumour-associated macrophages (TAMs) and cancer-associated fibroblasts (CAFs), respectively. TAMs and CAFs are reported to be involved in tumour progression, although their functions change to those of an anti-tumour phenotype under specific conditions. Notably, recent work published in The Journal of Pathology by Hashimoto and colleagues provided critical evidence indicating the significance of collaboration between TAMs and CAFs for tumour progression. They showed that cell-cell interaction between these two cell types induced recruitment and activation of each other, and that the combined activities of these cells were involved in neuroblastoma progression. Although many research groups are now interested in the significance of stromal cells such as CAFs and TAMs for tumour progression, only a few studies have been published describing the cell-cell interactions of these cells. Cell-cell interactions of stromal cells potentially play important roles in tumour progression and should be a focus for further oncology research. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

High CD169 expression in lymph node macrophages predicts a favorable clinical course in patients with esophageal cancer.

Recent findings indicate CD169-positive lymph node sinus macrophages (LySMs) in the regional lymph nodes (RLNs) play an important role in anti-cancer immunity. In the present study, we investigated the correlation between CD169 expression in RLNs and clinicopathologic factors. Higher CD169 expression in LySMs was significantly associated with longer cancer-specific survival (CSS). The density of tumor-infiltrating lymphocytes (TILs) in the cancer nest and CD169 expression on LySMs were positively associated in patients who underwent pretreatment. As CD169 expression is thought to reflect a high interferon signature in RLNs, we tried to identify immunity-related genes that are up-regulated by interferon in macrophages as well as CD169. Indoleamine 2,3-dioxygenase (IDO1) was found to be elevated by interferon, and expression of IDO1 was tested using immunohistochemistry. IDO1 expression on LySMs was positively correlated with CD169 expression; however, there was no significant correlation between IDO1 and clinicopathologic factors. These results suggest that high expression of CD169 in LySMs reflects a high potential for anti-cancer immune responses in esophageal cancer patients and that monitoring CD169 expression would be useful for evaluating the potential of anti-cancer immune reactions.

CD163-positive cancer cells are potentially associated with high malignant potential in clear cell renal cell carcinoma.

CD163 is preferentially expressed by monocyte/macrophages; however, recent studies using immunohistochemistry (IHC) have reported that some cancer cells also express CD163. In the present IHC study, we investigated CD163 staining of cancer cells and macrophages in clear cell renal cell carcinoma (ccRCC) tissues and determined the relationship between cancer cell CD163 expression and clinical prognosis in patients with ccRCC. IHC for CD163 was performed in ccRCC tissues from 103 patients. CD163-positive cancer cells were detected in 35% of the patients (36/103); however, the positive signals on cancer cells were significantly lower than those on macrophages. CD163-positive cancer cells were preferentially detected in patients with high T classification, and females, and were significantly associated with shortened progression-free survival and a lower overall survival ratio. Notably, a high intensity of CD163-positive macrophage infiltration was detected in the CD163-positive cancer cell-high tumor areas. Although CD163 mRNA was detected in cultured macrophages, no CD163 mRNA was detected in two cultured RCC cell lines. The detailed mechanism by which a positive signal is detected on cancer cells has not been clarified. Detection of the CD163 antigen on cancer cells might be a useful marker for evaluating the clinical course of patients with ccRCC.

Phenotypical change of tumor-associated macrophages in metastatic lesions of clear cell renal cell carcinoma.

Macrophages are the main immune cells of the tumor microenvironment in clear cell renal cell carcinoma (ccRCC). A high density of CD163+ or CD204+ tumor-associated macrophages (TAMs), rather than the density of total TAMs, is known to be linked to poor clinical outcome. In the present study, we investigated the phenotypical differences between the paired primary and metastatic lesions in ccRCC cases. Using immunostaining, the densities of CD163+ and CD204+ TAMs in metastatic lesions were found to be significantly lower compared to primary lesions, although the total number of TAMs was increased in metastatic lesions. Since CD163 and CD204 are considered to be the markers of an M2/protumor phenotype in macrophages, TAMs in metastatic lesions are suggested to have a greater M1/inflammatory function compared with those from primary lesions. These findings give new insights in regard to the immunological status of metastatic lesions of ccRCC.

Possible functions of CD169-positive sinus macrophages in lymph nodes in anti-tumor immune responses.

The lymph node (LN) is an important immune system in which a number of antigen-presenting cells are present that induce rapid immune responses to foreign antigens. While a great number of macrophages exist in lymph nodes, recent studies using animal models have shown that lymph node sinus macrophages are associated with the induction of anti-tumor immunity, playing a significant role in host immune responses against tumor cells. In colorectal tumor, malignant melanoma, and endometrial tumor, it was shown that a high density of CD169-positive macrophages in the LN sinus was a predictive factor for better clinical prognosis. The observations that the density of CD169-positive macrophages in the LN sinus was positively associated with the density of infiltrating T or NK cells in tumor tissues, indicates the significance of CD169-positive macrophages in anti-tumor immune reactions of tumor patients. Moreover, antigen delivery targeting LN macrophages is also considered to be promising approach for vaccination. In this article, we have summarized the significance of CD169-positive LN macrophages in anti-tumor immunity.

High density of CD204-positive macrophages predicts worse clinical prognosis in patients with breast cancer.

Recent studies have indicated the clinical significance of tumor-associated macrophages (TAM) in several malignant tumors including breast cancer. Although recent studies have focused on CD68-positive or CD163-positive TAM in breast cancer, no study has investigated the significance of CD204-positive TAM in breast cancer. We found that CD204 expression on macrophages was evaluated following stimulation with the conditioned medium (CM) of breast cancer cell lines. Paraffin sections of 149 breast cancer samples which were diagnosed as invasive ductal carcinoma were immunohistochemically analyzed for CD68, CD163 and CD204 expression. The results of analyses indicated that a high number of CD204-positive TAM was associated with worse clinical prognoses, including relapse-free survival, distant relapse-free survival and breast cancer-specific survival; however, neither the numbers of CD68-positive or CD163-positive TAM were associated with clinical courses. Of the clinicopathological factors investigated, estrogen receptor, Ki-67 index, hormone subtype, and histological grade were significantly related to the increased number of CD163-positive and CD204-positive TAM. These data indicate the clinical significance of CD204-positive TAM in breast cancer progression and CD204 is a marker for predicting clinical prognosis in breast cancer.

A xenograft model reveals that PU.1 functions as a tumor suppressor for multiple myeloma in vivo.

We previously demonstrated that PU.1 expression is down-regulated in the majority of myeloma cell lines and primary myeloma cells from patients. We introduced the tet-off system into the human myeloma cell lines U266 and KMS12PE that conditionally express PU.1 and demonstrated that PU.1 induces cell cycle arrest and apoptosis in myeloma cells in vitro. Here, we established a mouse xenograft model of myeloma using these cell lines to analyze the effects of PU.1 on the phenotype of myeloma cells in vivo. When doxycycline was added to the drinking water of mice engrafted with these myeloma cells, all mice had continuous growth of subcutaneous tumors and could not survived more than 65 days. In contrast, mice that were not exposed to doxycycline did not develop subcutaneous tumors and survived for at least 100 days. We next generated mice engrafted with subcutaneous tumors 5-10 mm in diameter that were induced by exposure to doxycycline. Half of the mice stopped taking doxycycline-containing water, whereas the other half kept taking the water. Although the tumors in the mice taking doxycycline continued to grow, tumor growth in the mice not taking doxycycline was significantly suppressed. The myeloma cells in the tumors of the mice not taking doxycycline expressed PU.1 and TRAIL and many of such cells were apoptotic. Moreover, the expression of a cell proliferation marker Ki67 was significantly decreased in tumors from the mice not taking doxycycline, compared with that of tumors from the mice continuously taking doxycycline. The present data strongly suggest that PU.1 functions as a tumor suppressor of myeloma cells in vivo.

PD-L1 expression in papillary renal cell carcinoma.

BACKGROUND: The immune escape or tolerance of cancer cells is considered to be closely involved in cancer progression. Programmed death-1 (PD-1) is an inhibitory receptor expressed on activating T cells, and several types of cancer cells were found to express PD-1 ligand 1 (PD-L1) and ligand 2 (PD-L2). METHODS: In the present study, we investigated PD-L1/2 expression in papillary renal cell carcinoma (pRCC). RESULT: We found PD-L1 expression in 29 of 102 cases, but no PD-L2 expression was seen. PD-L1 expression was not significantly correlated with any clinicopathological factor, including progression-free survival and overall survival. The frequency of PD-L1-positive cases was higher in type 2 (36%) than in type 1 (22%) pRCC; however, there was no significant difference in the percentages of score 0 cases (p value = 0.084 in Chi-square test). The frequency of high PD-L1 expression cases was higher in type 2 (23%) than in type 1 (11%), and the frequency of high PD-L1 expression cases was higher in grade 3/4 (21%) than in grade 1/2 (13%). However, no significant association was found between PD-L1 expression and all clinicopathological factors in pRCC. CONCLUSION: High expression of PD-L1 in cancer cells was potentially associated to highly histological grade of malignancy in pRCC. The evaluation of the PD-L1 protein might still be useful for predicting the efficacy of anti-cancer immunotherapy using immuno-checkpoint inhibitors, however, not be useful for predicting the clinical prognosis.

Neutrophil-to-lymphocyte ratio predicts metachronous liver metastasis of pancreatic neuroendocrine tumors.

BACKGROUND: Pancreatic neuroendocrine tumors (PNETs) are clinically malignant, having metastatic potential. Histological tumor grade is an accepted indicator of malignant potential, but noninvasive prognostic markers have not yet been identified. This study assessed whether the preoperative neutrophil-to-lymphocyte ratio (NLR) could predict clinical outcomes of PNET patients. METHODS: Fifty-eight patients who underwent curative resection for PNETs between 2001 and 2015 were retrospectively evaluated. The correlations between the preoperative NLR and clinicopathological parameters, including patient baseline clinical characteristics, tumor progression, and postoperative oncological outcome were evaluated. RESULTS: A high preoperative NLR was significantly associated with large tumor size (P = 0.0015) and high tumor grade (P < 0.0001). Overall survival and relapse-free survival of patients with a high NLR (≥2.4) were significantly shorter than those of patients with a low NLR (<2.4, P = 0.0481 and P < 0.0001, respectively). Multivariate analysis revealed that NLR ≥2.4 and tumor size ≥2 cm were independent predictors of postoperative recurrence (hazard ratio 6.012, P = 0.0035 and 6.760, P = 0.0049, respectively). Interestingly, a high NLR independently predicted postoperative liver, but not lymph node, metastasis. CONCLUSIONS: In this patient series, a high NLR (≥2.4) was a noninvasive marker that independently predicted postoperative liver metastasis in patients with PNETs, and thereby could be clinically useful.

The significance of TIMD4 expression in clear cell renal cell carcinoma.

T-cell immunoglobulin and mucin domain (TIMD) family genes are related to innate immune responses. TIMD4 is a receptor for phosphatidylserine and is involved in the phagocytosis of apoptotic cells by macrophages. In the present study, we found that TIMD4 is expressed on the cancer cells of patients with clear cell renal cell carcinoma (ccRCC). TIMD4 was immunostained in the resected samples of 89 patients diagnosed as ccRCC. High expression of TIMD4 in cancer cells was closely related to short progression free survival time; however, it was not correlated with other clinicopathological factors. Intracellular expression of TIMD4 was observed in the RCC cell line, 786-O. In vitro studies using 786-O cells and shRNA targeting TIMD4 indicated that TIMD4 expression was associated with resistance to sorafenib but not with cell proliferation. TIMD4 might be useful as a prognostic factor and may also be a new target for therapy of ccRCC.

Prognostic significance of CD169-positive lymph node sinus macrophages in patients with endometrial carcinoma.

Lymph node (LN) macrophages play critical roles in anti-tumor immunity, which develops via the activation of cytotoxic T cells (CTL) and NK cells. The present study aims to determine the prognostic significance of CD169(+) LN macrophages in patients with endometrial carcinoma (EC). The number of CD169(+) cells or the CD169(+) -to-CD68(+) macrophage ratio in regional LN (RLN), and the number of CD8(+) CTL or CD57(+) NK cells in tumor tissues were investigated by immunohistochemistry in paraffin-embedded tissue samples from 79 patients with EC. A high density of CD169(+) cells in the RLN of patients with EC was correlated with an early clinical stage or no LN metastasis. A high number of CD169(+) cells and a high CD169(+) -to-CD68(+) macrophage ratio were significantly associated with longer overall survival in EC. We also found that the density of CD169(+) macrophages was positively correlated with the number of CD8(+) CTL and CD57(+) NK cells that infiltrated into tumor tissues. A high density of CD57(+) cells in EC tissues was associated with a better prognosis, while a high density of CD8(+) cells was not linked to an altered prognosis. The present study showed that the density of CD169(+) macrophages in RLN was associated with an improved prognosis in EC patients. CD169(+) macrophages in RLN might represent a useful marker for assessing clinical prognoses and monitoring anti-tumor immunity in patients with EC.

An IL-27/Stat3 axis induces expression of programmed cell death 1 ligands (PD-L1/2) on infiltrating macrophages in lymphoma.

Immune escape and tolerance in the tumor microenvironment are closely involved in tumor progression, and are caused by T-cell exhaustion and mediated by the inhibitory signaling of immune checkpoint molecules including programmed death-1 (PD-1), cytotoxic T-lymphocyte associated protein 4, and T-cell immunoglobulin and mucin domaincontaining molecule-3. In the present study, we investigated the expression of the PD-1 ligand 1 (PD-L1) in a lymphoma microenvironment using paraffin-embedded tissue samples, and subsequently studied the detailed mechanism of upregulation of PD-L1 on macrophages using cultured human macrophages and lymphoma cell lines. We found that macrophages in lymphoma tissues of almost all cases of adult T-cell leukemia/lymphoma (ATLL), follicular lymphoma and diffuse large B-cell lymphoma expressed PD-L1. Cell culture studies showed that the conditioned medium of ATL-T and SLVL cell lines induced increased expression of PD-L1/2 on macrophages, and that this PD-L1/2 overexpression was dependent on activation of signal transducer and activator of transcription 3 (Stat3). In vitro studies including cytokine array analysis showed that IL-27 (heterodimer of p28 and EBI3) induced overexpression of PD-L1/2 on macrophages via Stat3 activation. Because lymphoma cell lines produced IL-27B (EBI3) but not IL-27p28, it was proposed that the IL-27p28 derived from macrophages and the IL-27B (EBI3) derived from lymphoma cells formed an IL-27 (heterodimer) that induced PD-L1/2 overexpression. Although the significance of PD-L1/2 expressions on macrophages in lymphoma progression has never been clarified, an IL-27-Stat3 axis might be a target for immunotherapy for lymphoma patients.