Suppression of Lipopolysaccharide-Induced IL-1ß Gene Expression by High-Molecular-Weight Adiponectin in RAW264.7 Macrophages.

Adiponectin is an abundant adipocytokine secreted by adipocytes. It exists in the plasma in its trimeric, hexameric, high-molecular-weight (HMW), and globular (a proteolytic product) isoforms. Adiponectin's anti-inflammatory effects on macrophages remain controversial. We have previously reported a simple and effective method for purifying native HMW adiponectin from human plasma. Here, we investigated whether native HMW adiponectin from human plasma has anti-inflammatory effects on macrophages. Pretreatment with human native HMW adiponectin inhibited lipopolysaccharide (LPS)-induced interleukin-1ß (IL-1ß) gene expression, but not tumor necrosis factor (TNF)-α expression. However, simultaneous treatment with HMW adiponectin and LPS did not inhibit IL-1ß expression. Further, HMW adiponectin pretreatment decreases glycogen synthase kinase-3ß (GSK-3ß) inactivation by abrogating LPS-induced Akt (Ser473) phosphorylation, which subsequently suppresses LPS-induced CCAAT/enhancer binding protein ß (C/EBPß) protein translation and nuclear translocation. However, HMW adiponectin pretreatment did not affect LPS-induced nuclear factor-kappaB (NF-κB) activation. These results suggest that HMW adiponectin mediates potent anti-inflammatory activities in macrophages by inhibiting its Akt-C/EBPß signaling pathway, thereby suppressing IL-1ß gene expression.

MCAM+CD161- Th17 Subset Expressing CD83 Enhances Tc17 Response in Psoriasis.

Recent studies have highlighted the pathogenic roles of IL-17-producing CD8+ T cells (T-cytotoxic 17 [Tc17]) in psoriasis. However, the underlying mechanisms of Tc17 induction remain unclear. In this study, we focused on the pathogenic subsets of Th17 and their mechanism of promotion of Tc17 responses. We determined that the pathogenic Th17-enriched fraction expressed melanoma cell adhesion molecule (MCAM) and CCR6, but not CD161, because this subset produced IL-17A abundantly and the presence of these cells in the peripheral blood of patients has been correlated with the severity of psoriasis. Intriguingly, the serial analysis of gene expression revealed that CCR6+MCAM+CD161-CD4+ T cells displayed the gene profile for adaptive immune responses, including CD83, which is an activator for CD8+ T cells. Coculture assay with or without intercellular contact between CD4+ and CD8+ T cells showed that CCR6+MCAM+CD161-CD4+ T cells induced the proliferation of CD8+ T cells in a CD83-dependent manner. However, the production of IL-17A by CD8+ T cells required exogenous IL-17A, suggesting that intercellular contact via CD83 and the production of IL-17A from activated CD4+ T cells elicit Tc17 responses. Intriguingly, the CD83 expression was enhanced in the presence of IL-15, and CD83+ cells stimulated with IL-1ß, IL-23, IL-15, and IL-15Rα did not express FOXP3. Furthermore, CCR6+MCAM+CD161-CD4+ T cells expressing CD83 were increased in the peripheral blood of patients, and the CD83+ Th17-type cells accumulated in the lesional skin of psoriasis. In conclusion, pathogenic MCAM+CD161- Th17 cells may be involved in the Tc17 responses via IL-17A and CD83 in psoriasis.

A case of bronchial asthma as an immune-related adverse event of pembrolizumab treatment for bladder cancer: A case report.

RATIONALE: Bladder cancer is one of the most common cancers worldwide. The anti-programmed cell death protein 1 (PD-1) antibody pembrolizumab, which is an immune checkpoint inhibitor (ICI), has improved survival in bladder cancer. We report a case of bladder cancer that had a high antitumor effect with anti-programmed cell death PD-1 antibody pembrolizumab, an ICI, but asthma occurred an immune-related adverse event (irAE). PATIENT CONCERNS: A 70-year-old female patient was diagnosed as unresectable bladder cancer who was indicated for ICI treatment. DIAGNOSIS: After ICI administration as a treatment for bladder cancer, the patient had a grade 3 asthma attack. Cytotoxic T lymphocyte antigen 4 (CTLA-4) in CD4+ FOX3+ T cells was upregulated in the early phase before the development of asthma attacks. Moreover, T-cell immunoglobulin and mucin domain 3 (TIM-3) was upregulated in all memory T cells among CD4+ T cells. However, no change in the expression of TIM-3 was observed in any CD8+ T-cell subtype. In contrast, the proportion of CD161- T helper 17 cell (Th17) cells increased. INTERVENTIONS: The patient was treated with betamethasone, montelukast, salbutamol nebulization, and a combination of salmeterol (50 µg) and fluticasone (500 µg) (SFC). OUTCOMES: The patient's wheezing resolved, and her peak flow rate reached 100% of the predicted value; therefore, the patient continued treatment with SFC and montelukast and was discharged from the hospital. CONCLUSION: Increases in CTLA-4 and TIM-3 expression in CD4+ T cells (not CD8+), as well as an increase in Th17 cells, may reflect asthma-related inflammation activity. Immune-related adverse events during immune checkpoint inhibitor administration may be predictive markers of antitumor efficacy.

The Ang III/AT2R Pathway Enhances Glucose Uptake by Improving GLUT1 Expression in 3T3-L1 Adipocytes.

Angiotensin III (Ang III) is a heptapeptide derived from Ang II that has been confirmed as the preferred agonist of angiotensin II type 2 receptor (AT2R). Recent studies have revealed AT2R mainly exerts anti-inflammation effects. However, the effects of the Ang III/AT2R pathway on adipocytes remain unknown. Here, the effects of Ang III on glucose uptake were examined. The results showed that AT2R expression was upregulated during adipogenesis in 3T3-L1 preadipocytes, whereas AT1R expression was diminished. Also, Ang III (10 nM) significantly increased glucose uptake by 3T3-L1 adipocytes, which was blocked by PD123319, an AT2R blocker, but not by irbesartan, an AT1R blocker. Ang III also induced the expression of glucose transporter type 1 (GLUT1). These stimulatory effects were inhibited by pretreatment with PD123319, but not with irbesartan. Together, these results indicate that Ang III enhances glucose uptake by upregulating GLUT1 expression via AT2R.

A case report on severe nivolumab-induced adverse events similar to primary sclerosing cholangitis refractory to immunosuppressive therapy.

INTRODUCTION: Immune checkpoint inhibitors (ICIs), particularly anti-PD-1 antibody, have dramatically changed cancer treatment; however, fatal immune-related adverse events (irAEs) can develop. Here, we describe a severe case of sclerosing cholangitis-like irAE. We report the use of 3 immunosuppressive agents that resulted in the death of the patient due to treatment inefficacy. According to a postmarketing study of nivolumab, the frequency of ICI-related sclerosing cholangitis is 0.27% and that of ICI-related cholangitis is 0.20%. There have been 4 case reports of sclerosing cholangitis-like irAE, with imaging findings, including typical intrahepatic bile duct beaded constriction in primary sclerosing cholangitis. Treatment starts with prednisolone and is combined with an immunosuppressant in refractory cases. There are no reports of severe cases that ultimately led to death. PATIENTS CONCERNS: The patient is a 64-year-old male with Stage IV squamous cell lung carcinoma; he was hospitalized with abdominal pain and elevation of aspartate transaminase and alanine transaminase, approximately 4 months after ICI administration was suspended. This occurred because the patient treated with nivolumab as the second-line chemotherapy and developed type 1 diabetes mellitus after 11 courses. DIAGNOSIS: A grade 3 increase in bilirubin was observed and he was diagnosed with sclerosing cholangitis, based on magnetic resonance cholangiopancreatography imaging and pathological findings of the liver and bile duct. INTERVENTIONS: Prednisolone, mycophenolate mofetil, and tacrolimus combination therapy was administered. OUTCOMES: The treatment was difficult and failed. He died from liver failure 8 months after diagnosis. In this case, hepatitis and cholangitis, mainly alanine transaminase-dominant liver disorder, developed in the early stages of irAEs. Although he showed some improvement after prednisolone administration, bilirubin levels began rising again, and sclerosing cholangitis did not improve even with the use of 3 immunosuppressive agents recommended by the ESMO Clinical Practice Guidelines for immune-related hepatotoxicity management. Although the antitumor effect showed a complete response, liver failure led to death. CONCLUSION: This is the first case report on the ineffectiveness of triple immunosuppressant combination therapy recommended by the guidelines for immune-related hepatotoxicity. It is necessary to develop more appropriate treatment for severe sclerosing cholangitis-like irAE based on the robust evidence.

Rare Nivolumab-associated Super Hyper Progressive Disease in Patients With Advanced Gastric Cancer.

BACKGROUND/AIM: Rapid tumor growth after administration of immune checkpoint inhibitors is designated hyper progressive disease (HPD). In this study, besides the conventional HPD category, we proposed the "super HPD" category where the disease is naturally rapidly growing. PATIENTS AND METHODS: Patients treated for advanced gastric cancer with irinotecan or nivolumab as a third-line treatment were retrospectively compared. RESULTS: Eighteen and 26 patients were treated with irinotecan or nivolumab, respectively. There were 3 HPD cases (16.7%) in the irinotecan group, 6 cases (23.1%) in the nivolumab group, and the frequency of HPD was not significantly different. Two cases satisfied the super HPD definition only in the nivolumab group. When one of them was analyzed immunologically, the number of regulatory T cells was found to be increased, resulting in a low neutrophil-to-lymphocyte ratio. CONCLUSION: Our proposed super HPD was likely to represent a true HPD, with a frequency of 7.7%.

Transfer and Enzyme-Mediated Metabolism of Oxidized Phosphatidylcholine and Lysophosphatidylcholine between Low- and High-Density Lipoproteins.

Oxidized low-density lipoprotein (oxLDL) and oxidized high-density lipoprotein (oxHDL), known as risk factors for cardiovascular disease, have been observed in plasma and atheromatous plaques. In a previous study, the content of oxidized phosphatidylcholine (oxPC) and lysophosphatidylcholine (lysoPC) species stayed constant in isolated in vivo oxLDL but increased in copper-induced oxLDL in vitro. In this study, we prepared synthetic deuterium-labeled 1-palmitoyl lysoPC and palmitoyl-glutaroyl PC (PGPC), a short chain-oxPC to elucidate the metabolic fate of oxPC and lysoPC in oxLDL in the presence of HDL. When LDL preloaded with d13-lysoPC was mixed with HDL, d13-lysoPC was recovered in both the LDL and HDL fractions equally. d13-LysoPC decreased by 50% after 4 h of incubation, while d13-PC increased in both fractions. Diacyl-PC production was abolished by an inhibitor of lecithin-cholesterol acyltransferase (LCAT). When d13-PGPC-preloaded LDL was incubated with HDL, d13-PGPC was transferred to HDL in a dose-dependent manner when both LCAT and lipoprotein-associated phospholipase A2 (Lp-PLA2) were inhibited. Lp-PLA2 in both HDL and LDL was responsible for the hydrolysis of d13-PGPC. These results suggest that short chain-oxPC and lysoPC can transfer between lipoproteins quickly and can be enzymatically converted from oxPC to lysoPC and from lysoPC to diacyl-PC in the presence of HDL.

Circulating oxidized LDL, increased in patients with acute myocardial infarction, is accompanied by heavily modified HDL.

Oxidized LDL (oxLDL) is a known risk factor for atherogenesis. This study aimed to reveal structural features of oxLDL present in human circulation related to atherosclerosis. When LDL was fractionated on an anion-exchange column, in vivo-oxLDL, detected by the anti-oxidized PC (oxPC) mAb, was recovered in flow-through and electronegative LDL [LDL(-)] fractions. The amount of the electronegative in vivo-oxLDL, namely oxLDL in the LDL(-) fraction, present in patients with acute MI was 3-fold higher than that observed in healthy subjects. Surprisingly, the LDL(-) fraction contained apoA1 in addition to apoB, and HDL-sized particles were observed with transmission electron microscopy. In LDL(-) fractions, acrolein adducts were identified at all lysine residues in apoA1, with only a small number of acrolein-modified residues identified in apoB. The amount of oxPC adducts of apoB was higher in the LDL(-) than in the L1 fraction, as determined using Western blotting. The electronegative in vivo-oxLDL was immunologically purified from the LDL(-) fraction with an anti-oxPC mAb. The majority of PC species were not oxidized, whereas oxPC and lysoPC did not accumulate. Here, we propose that there are two types of in vivo-oxLDL in human circulating plasma and the electronegative in vivo-oxLDL accompanies oxidized HDL.

Cooperative Action of Oxidized Low-Density Lipoproteins and Neutrophils on Endothelial Inflammatory Responses Through Neutrophil Extracellular Trap Formation.

The function of oxidatively modified low-density lipoprotein (oxLDL) in the progression of cardiovascular diseases has been extensively investigated and well-characterized with regards to the activation of multiple cellular responses in macrophages and endothelial cells. Although accumulated evidence has revealed the presence of neutrophils in vascular lesions, the effect of oxLDL on neutrophil function has not been properly investigated. In the present decade, neutrophil extracellular traps (NETs) gained immense attention not only as a primary response against pathogenic bacteria but also due to their pathological roles in tissue damage in various diseases, such as atherosclerosis and thrombosis. In this study, we investigated if oxLDL affects NET formation and if it is a risk factor for inflammatory reactions in endothelial cells. HL-60-derived neutrophils were stimulated with phorbol 12-myristate 13-acetate (PMA) for 30 min to induce NET formation, followed by incubation with 20 µg/mL native or oxidized LDL for additional 2 h. Culture media of the stimulated cells containing released NETs components were collected to evaluate NET formation by fluorometric quantitation of released DNA and detection of myeloperoxidase (MPO) by western blot analysis. NET formation of HL-60-derived neutrophils induced by PMA was significantly enhanced by additional incubation with oxLDL but not with native LDL. Treatment of HL-60-derived neutrophils with oxLDL alone in the absence of PMA did not induce NET formation. Furthermore, the culture media of HL-60-derived neutrophils after NET formation were then transferred to human aortic endothelial cell (HAECs) culture. Treatment of HAECs with the culture media containing NETs formed by HL-60-derived neutrophils increased the expression of metalloproteinase-1 protein in HAECs when HL-60-derived neutrophils were incubated with native LDL, and the expression was accelerated in the case of oxLDL. In addition, the culture media from NETs formed by HL-60-derived neutrophils caused the elongation of HAECs, which was immensely enhanced by coincubation with native LDL or oxLDL. These data suggest that oxLDL may act synergistically with neutrophils to form NETs and promote vascular endothelial inflammation.

[A Questionnaire Survey on Cooperation between Community Pharmacies and Hospitals in Outpatient Chemotherapy-Comparison of Roles of Pharmacists in Community Pharmacy and Hospitals].

　Previous reports suggested that sharing outpatient information during chemotherapy is very important for managing pharmaceutical usage between community pharmacies and hospitals. We herein examined using a questionnaire survey whether pharmaceutical management for outpatient chemotherapy is desired by community and hospital pharmacists. The response rates were 44.3% (133/300) for pharmacists in community pharmacies and 53.7% (161/300) for pharmacists in hospitals. Prescriptions for outpatients during chemotherapy were issued at 88.2% of the hospitals. Currently, 28.9% of hospital pharmacists rarely provide pharmaceutical care, such as patient guidance and adverse effect monitoring, for outpatients receiving oral chemotherapy. Furthermore, whereas 93.7% of hospital pharmacists conducted prescription audits based on the chemotherapy regimen, audits were only performed by 14.8% of community pharmacists. Thus, outpatients, particularly those on oral regimens, were unable to receive safe pharmaceutical care during chemotherapy. Community pharmacists suggested that hospital pharmacists should use "medication notebooks" and disclose prescription information when providing clinical information to community pharmacists. They also suggested sending clinical information to hospital pharmacists by fax. On the other hand, hospital pharmacists suggested the use of "medication notebooks" and electronic medical records when providing clinical information to community pharmacists. In addition, they suggested for community pharmacists to use electronic medical records when providing clinical information to hospital pharmacists. As there may be differences in opinion between community and hospital pharmacists, mutual preliminary communication is important for successful outpatient chemotherapy.

Killer cell immunoglobulin-like receptor 2DL4 is expressed in and suppresses the cell growth of Langerhans cell histiocytosis.

Killer cell immunoglobulin-like receptor (KIR) 2DL4 (CD158d) is a receptor for human leukocyte antigen-G. The function of KIR2DL4 has been reported in human natural killer cell lymphoma and mastocytosis, but not in Langerhans cell histiocytosis (LCH). Herein, we examined the expression and function of KIR2DL4 in LCHs. In pathological specimens, 27 of 36 LCH cases (75.0%) were immunohistochemically positive for KIR2DL4. Its expression was independent of age, gender, location, multi- or single-system, and the status of BRAFV600E immunostaining. We also confirmed the expression of KIR2DL4 mRNA and protein in the human LCH-like cell lines ELD-1 and PRU-1. KIR2DL4 protein was distributed in the membrane and cytoplasm of ELD-1 cells, but only in the cytoplasm of PRU-1 cells. An agonistic antibody against KIR2DL4 reduced phosphorylation of extracellular signal-regulated kinases (ERKs) and suppressed the cell growth of ELD-1 cells in a Src homology region 2 domain-containing phosphatase-2 dependent manner, but it had no effect in PRU-1 cells. These results suggest that KIR2DL4-mediated ERK suppression is a possible therapeutic target for LCH cells.

Efforts by Pharmacists in Community Pharmacy in Peacetime for Headache Medical Care in Emergencies, Such as Great Earthquake and Disaster.

We herein analyzed the issues that pharmacists in a community pharmacy in peacetime need to prepare for regarding headache medical care in emergencies (the state that supply of medical supplies is difficult) using a questionnaire intended for doctors and pharmacists in a community pharmacy. Recovery rates were 48.0% (96/200) for doctors and 37.3% (112/300) for pharmacists. In order to distinguish between patients for whom pharmacists need to "recommend OTC drugs" and those who need to be encouraged "to consult a hospital or clinic", doctors indicated that pharmacists need to use an "assistance tool to diagnosis headaches, such as a migraine screener" and "guidelines for chronic headaches". However, few pharmacists used these tools. Approximately 66.7% of doctors indicated that it is "meaningful" for pharmacists to distinguish patients with headaches. Moreover, doctors indicated the need for guidance by pharmacists in peacetime regarding headache medical care in emergencies. Although 73.2% of pharmacists instructed the patients with headaches of the importance of medication notebooks in emergencies, guidance ("understanding the triggers of headaches", "understanding the importance of removing the cause of the headache", "standing OTC drugs" and "standing prescription drugs") by pharmacists to prepare for an emergency was insufficient. These results provide useful information to improve the efforts by pharmacists in community pharmacies in peacetime for headache medical care in emergencies.

Ex vivo expansion of natural killer cells from human peripheral blood mononuclear cells co-stimulated with anti-CD3 and anti-CD52 monoclonal antibodies.

BACKGROUND AIMS: This study developed a new method to expand CD3(-)CD56(+) natural killer (NK) cells from human peripheral blood mononuclear cells (PBMCs) without feeder cells for clinical trials. METHODS: PBMCs from healthy subjects were co-stimulated with anti-CD3 and anti-CD52 monoclonal antibodies and cultured for 14 days in newly developed NKGM-1 medium containing autologous plasma and interleukin-2. Expanded NK cells were examined for cell number, phenotype, in vitro and in vivo cytotoxicity and interferon (IFN)-γ secretion. We also evaluated the proliferative ability of NK cells after cryopreservation. A patient with advanced pancreatic cancer was treated with autologous-expanded NK cells through the use of this method in combination with chemotherapy. RESULTS: Expanded NK cells expressed higher levels of activating molecules compared with resting NK cells and exhibited potent cytotoxicity against K562 cells and IFN-γ secretion by cytokine stimulation. Significant anti-tumor activity was observed in immunodeficient mice injected with the human pancreatic cancer cell line BxPC-3. Large-scale cultures generated a median 5.7 × 10(9) NK cells from 20 mL of peripheral blood (n = 38) after 14 days of culture and 8.4 × 10(9) NK cells after 18 days of culture through the use of a cryopreservation procedure. The number of NK cells and cytotoxic activity in the peripheral blood of the patient with pancreatic cancer greatly increased, and successful clinical responses were observed after multiple infusions of expanded NK cells. CONCLUSIONS: These data demonstrate that this simple and safe methodology for the ex vivo expansion of NK cells can be used for cancer immunotherapy.

Time course-changes in phosphatidylcholine profile during oxidative modification of low-density lipoprotein.

BACKGROUND: Oxidized phosphatidylcholines (oxPC) and lysophosphatidylcholine (lysoPC) generated during the formation of oxidized low-density lipoprotein (oxLDL) are involved in atherosclerotic lesion development. We investigated the time course-changes in phosphatidylcholine (PC) molecular species during oxidation of LDL to determine how those atherogenic PCs are produced. METHODS: Human and rabbit LDLs were pretreated with or without a selective platelet-activating factor acetylhydrolase (PAF-AH) inhibitor. LDL was oxidized by incubation with copper sulfate, and PC profiles were analyzed by liquid chromatography-tandem mass spectrometry. RESULTS: When human LDL was oxidized, the peak areas for polyunsaturated fatty acid (PUFA)-containing PC species dramatically decreased after a short lag period, concomitantly lysoPC species increased sharply. Although a variety of oxPC species containing oxidized fatty acyl groups or cleaved acyl chains are formed during LDL oxidation, only a few oxPC products accumulated in oxLDL: 1-palmitoyl-2-(9-oxo-nonanoyl) PC and long-chain oxPC with two double bonds. Pretreatment of LDL with the PAF-AH inhibitor greatly reduced lysoPC production while it had no effect on lipid peroxidation reactions and oxPC profiles. Rabbit LDL, which has a different composition of PC molecular species and needs a longer time to reach achieve full oxidation than human LDL, also accumulated lysoPC during oxidation. The increase in lysoPC in rabbit oxLDL was suppressed by pretreatment with the PAF-AH inhibitor. The major oxPC species formed in rabbit oxLDL were almost the same as human oxLDL. CONCLUSIONS: These results suggest that lysoPC species are the major products and PAF-AH activity is crucial for lysoPC generation during oxidation of LDL. The oxPC species accumulated are limited when LDL is oxidized with copper ion in vitro.

A case of pulmonary papillary adenoma: possible relationship between tumor histogenesis/tumorigenesis and fibroblast growth factor receptor 2 IIIb.

Pulmonary papillary adenoma is a rare tumor. We analyzed a tumor which appeared in a 16-year-old Japanese woman. The tumor histologically showed papillary proliferation of one-layered tumor cells coating inflammatory fibrovascular cores. At the periphery of the tumor, the tumor cells grew in a lepidic fashion. The tumor cells were confirmed as type-II pneumocytes with electron-microscope. In this study, using immunohistochemistry, in situ hybridization and real-time reverse transcription polymerase chain reaction, we examined the expressions and quantities of fibroblast growth factor 10 (FGF10), keratinocyte growth factor (KGF) and fibroblast growth factor receptor 2 (FGFR2) IIIb, based on the extent of their abilities of proliferation and differentiation of type II pneumocytes. The tumor cells expressed FGFR 2 and produced 350 times more FGFR2IIIb messenger RNA (mRNA) than did the nontumorous lung. The quantity of KGF mRNA in the tumor tissue was twice that of the nontumorous lung. Moreover, there was dysregulation of FGFR2IIIb transcription in the tumor. According to these findings, we expect overexpression of FGFR2IIIb to play an important role in causing tumor. Because FGFR is suspected to be connected with lung carcinoma, we also treat similar tumorigenesis via FGFR as carcinoma; complete resection of adenoma might be indicated.

Coexpression of CCR6 and CD146 (MCAM) is a marker of effector memory T-helper 17 cells.

Effector memory T (T(EM)) cells are a subpopulation of memory T cells that express receptors mediating migration to inflamed tissues and produce various cytokines. Effector memory T-helper (Th)17 (Th17(EM)) cells are thought to be essential for inflammation in Th17-mediated diseases, but have not been studied in detail. To identify superior surface markers to isolate a homogeneous population of Th17(EM) cells from peripheral blood, CD4(+) T cells were isolated from the peripheral blood of healthy donors based on the expression of CCR7, CCR6 and CD146 using six-color flow cytometry. After 4days of culture in the presence of anti-CD3/28 beads, intracellular cytokines were determined by flow cytometric analysis. To investigate the relevance of Th17(EM) cells in Th17-mediated disease, the frequencies of T(EM) -cell subsets in psoriasis were quantified using six-color flow cytometry. An enzyme-linked immunosorbent assay was performed to confirm the interleukin (IL)-17-producing capacity of T(EM) -cell subsets from the peripheral blood of a patient with psoriasis. CCR6(+) CD146(+) T(EM) (CD4(+) CD45RA(-) CCR7(-)) cells had a greater capacity to produce IL-17 than CCR6(+) CD146(-) or CCR6(-) CD146(+) T(EM) cells. Although the percentage of CCR6(+) CD146(+) cells in T(EM) cells was not significantly different between patients with psoriasis and controls, three of eight patients had a higher percentage of CCR6(+) CD146(+) T(EM) cells than the mean +5 standard deviations of the controls. Coexpression of CCR6 and CD146 is a useful marker for Th17(EM) cells. Increasing the number of CCR6(+) CD146(+) Th17(EM) cells in peripheral blood may facilitate estimation of systemic Th17-cell activity in Th17-mediated diseases.

Oxidized low-density lipoprotein-induced periodontal inflammation is associated with the up-regulation of cyclooxygenase-2 and microsomal prostaglandin synthase 1 in human gingival epithelial cells.

Periodontitis is characterized by chronic gingival tissue inflammation, and inflammatory mediators such as IL-8 and prostaglandin E(2) (PGE(2)) are associated with disease progression. Previously we showed that oxidatively modified low-density lipoprotein (oxLDL) was present in gingival crevicular fluid. In this study, the role of oxLDL in the gingival epithelial cell inflammatory response was further investigated using Ca9-22 cells and primary human oral keratinocytes (HOK). Treatment of Ca9-22 cells and HOK with oxLDL induced an up-regulation of IL-8 and the PGE(2)-producing enzymes, cyclooxygenase-2 and microsomal PGE(2) synthase-1. These responses induced by oxLDL were significantly suppressed by a nuclear factor-kappa B (NF-κB) inhibitor. However, unlike the result in macrophages, oxLDL did not lead to an increase in CD36 expression in these two cells. These results suggest that oxLDL elicits gingival epithelial cell inflammatory responses through an activation of the NF-κB pathway. These data suggest a mechanistic link between periodontal disease and lipid metabolism-related disorders, including atherosclerosis.

A case of granulocyte colony-stimulating factor and interleukin 6 receptor-producing mediastinal mature cystic teratoma with somatic-type malignancy.

Mediastinal germ cell tumor with somatic-type malignancy is a rare neoplasm. We describe one such case in a 49-year-old Japanese man who had shown an elevated serum concentration of granulocyte colony-stimulating factor (GCSF) and leukocytosis without a shift to the left. Histologically, the tumor formed a teratomatous cyst whose wall contained benign epithelial components, well-differentiated tubular and mucinous adenocarcinoma, and poorly-differentiated pleomorphic carcinoma. Immunohistochemically, both the well differentiated adenocarcinoma and poorly differentiated pleomorphic carcinoma expressed GCSF. Immunohistochemistry and molecular analysis revealed that both components also produced interleukin 6 receptor (IL6R). We diagnosed this tumor as a GCSF- and IL6R-producing mediastinal mature cystic teratoma with somatic-type malignancy. The tumor showed immunohistochemical expression of activated signal transducer and activator of transcription 3. The patient died 6 months after developing systemic symptoms. For a GCSF-producing tumor, complete resection appears to offer the best outcome at present. For any patient presenting with leukocytosis without a shift to the left, a thorough analysis should be conducted, and the tumor diagnosed as early as possible.

Enhancement of IL-2-induced cytotoxicity by interferon-alpha in renal cell carcinoma.

Metastatic renal cell carcinoma (mRCC) treatment consists of molecular targeted agents and cytokines that have fundamentally different mechanisms of action. Clinical responses also differ; complete response is rare with molecular targeted agents but is sometimes achieved with cytokine therapies. Because of the relatively high efficacy of combination therapy with low-dose interleukin-2 (IL-2) and interferon-alpha (IFN-alpha) against mRCC, it is important to reevaluate cytokine therapies in vitro. Here, we show that when IL-2 is administered in combination with IFN-alpha, a stronger cytotoxic effect of PBMCs on RCC cell lines is observed than when IL-2 is administered alone. The upregulation of TNF-related apoptosis-inducing ligand on NK cell by IL-2 and suppression of regulatory T cells (Tregs) by IFN-alpha were recognized at the same time when cytotoxicity of peripheral blood mononuclear cells (PBMCs) was enhanced. IL-2 is known to activate natural killer cell cytotoxicity; however, IL-2 also stimulates Treg expansion, which enhances immunosuppression. On the other hand, IFN-alpha negatively regulates Treg cells, thereby increasing the function of immune effector cells. Our in vitro results may explain, at least in part, the clinical efficacy of combination low-dose IL-2 and IFN-alpha therapy against mRCC.

Zoledronate stimulates gamma delta T cells in prostate cancer patients.

Androgen deprivation therapy is the mainstay of treatment for prostate cancer. Given its frequent failure, new therapy that reduces prostate cancer progression would be a breakthrough in treating this disease. Bisphosphonates are well-established agents for treating skeletal-related events (SREs) in prostate cancer patients with bone metastases. Exposure to bisphosphonates may not only reduce the incidence of SREs, but also have anticancer effects by modulating a patient's immunity. The purpose of this study was to examine the effect of zoledronate (ZOL) on gamma delta T cells, serum prostate-specific antigen (PSA) levels, and velocities. The effect of ZOL, with and without IL-2, on gamma delta T cell activation was examined in vitro. Furthermore, the activated state and the number of gamma delta T cells and changes in serum PSA levels were examined for patients who received ZOL infusion for the prevention of SREs. We found that ZOL activated gamma delta T cells, and the number of gamma delta T cell was increased when IL-2 was administered with ZOL in vitro. Comparisons before and after the first ZOL infusion revealed that gamma delta T cells in peripheral blood were activated by ZOL. Moreover, after the first ZOL treatment, reduction in serum PSA was observed in 3 of 11 patients, and reduction in PSA velocity was observed in 5 of 10 patients. Our findings indicate that ZOL stimulates gamma delta T cells in vivo and in vitro. This study provides further insight into the ability of gamma delta T cells to induce an antitumor immune response.