Atherosclerosis is a lifestyle-related disease that plays a major role in cardiovascular disease. Recently, we found that gene expression of Gremlin 2, an antagonist of bone morphogenetic protein (BMP), was significantly increased in the aorta of spontaneously hypertensive and hyperlipidemic rats (SHHRs) fed a high-fat, 30% sucrose solution diet (HFDS). However, the role of Gremlin 1 (Grem1) and Gremlin 2 (Grem2) in the aortic arch of rats under hypertensive, hyperlipidemic, and hyperglycemic conditions remains unclear. Therefore, in the present study we investigated the molecular role of Gremlins in the aorta of SHHRs. Four-month-old male Sprague-Dawley rats and SHHRs were fed a normal diet or the HFDS ad libitum for 4 months. Then, gene and protein expression was analyzed using quantitative polymerase chain reaction and western blotting, respectively. Grem1 and Grem2 protein expression was increased, whereas phosphorylated Smad1/5 protein expression was low, in the aorta of SHHRs fed the HFDS. In addition, the expression of the downstream gene targets of BMP, namely inhibitor of DNA binding 1 (Id1) and atonal homolog 8 (Atoh8), was decreased in aortas of SHHRs fed the HFDS. Furthermore, mRNA expression of Snail, α-smooth muscle actin (αSMA), and Fibronectin was increased in SHHRs fed the HFDS. These findings suggest that upregulation of Gremlins attenuates the activation of BMP signaling, which contributes to fibrogenesis of the aorta.

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Endothelin-1 is a peptide with many functions including bronchoconstriction and the stimulation of fibroblasts, and myofibroblasts, and airway smooth muscle cell proliferation. These functions are related to airway remodeling and endothelin-1 is known to be upregulated in the epithelium of patients with severe asthma. We thus sought to elucidate the mechanisms underlying endothelin-1 expression in bronchial epithelial cells in vitro. The human bronchial epithelial cell line BEAS-2B was grown in culture and then treated with tumor necrosis factor-alpha (TNF-α), interleukin-4 (IL-4), interleukin-13 (IL-13), and transforming growth factor-beta (TGF-β). Expression of endothelin-1 mRNA and protein was quantified by real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. We also repressed expression of the key transcription factor in the pathogenesis of severe asthma, nuclear factor-kappa B (NF-κB), using small interfering RNA (siRNA). TNF-α and TGF-β significantly increased the release of endothelin-1 protein into the culture medium of BEAS-2B cells at 24 h after treatment compared to untreated cells; however, the Th2 cytokines, IL-4 and IL-13, had no effect. Endothelin-1 mRNA expression was also upregulated by TNF-α and TGF-β with a peak time point at 4 h after stimulation. Finally, the combination of TNF-α and TGF-β synergistically increased both endothelin-1 protein secretion and mRNA expression, and this upregulation was significantly suppressed in cells transfected with siRNA to repress NF-κB expression. TNF-α and TGF-β synergistically upregulate the expression of endothelin-1 in human bronchial epithelial cells, possibly via the activity of NF-κB. Our findings thus suggest NF-κBa as a potential therapeutic target for the regulation of airway remodeling.

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Glutathione S-transferase (GST) is a major phase II drug-metabolizing enzyme. Several isoforms of human GST as well as different GST genetic polymorphisms are known, but limited data exists concerning the relationship between GST polymorphisms and GST activity using 1-chloro-2,4-dinitrobenzene in human liver. To resolve this query, we analyzed the genetic polymorphisms of four main GST isoforms [GST mu 1 (GSTM1), GST theta 1 (GSTT1), GST alpha 1 (GSTA1), GST pi 1 (GSTP1)] and measured hepatic GST activity isolated from the same patients. We found that GSTM1 null individuals have significantly lower (P=0.0082) GST activity compared with GSTM1 positive individuals. No significant changes in GST activity were observed in individuals with GSTT1, GSTA1, and GSTP1 genotypes. Interestingly, the levels of GST activity exhibited were similar when compared with GSTA1*A/*A and GSTA1*A/*B, and GSTP1*A/*A and GSTP1*A/*B, respectively, if the genotype was GSTM1 null. Therefore, the genotypes of GSTA1*A/*B and GSTP1*A/*B individuals do not significantly affect the level of hepatic GST activity. An examination of the correlation between GST mRNA expression and GST activity subsequently revealed a significant correlation between GSTM1 mRNA levels and GST activity (r=0.626, P=0.007). These data are expected to facilitate research on the prediction of efficacy and safety of GSTM1 null-mediated drug metabolism and may establish whether genetic polymorphisms of the GST gene, specifically GSTM1, can act as a biomarker.

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Anaplastic thyroid cancer (ATC) is known to be a highly malignant cancer of the thyroid with a high mortality rate. In a previous study, we used real-time cell analysis (RTCA) to analyze cell migration and invasion of oral squamous cell carcinomas (OSCCs) of the tongue and floor of the mouth. In the present study, we investigated cell migration and invasion of ATC using RTCA, as well as their association with periostin, matrix metalloproteinases (MMPs), and integrins. Experiments were performed on TCO-1 and HTC/C3 cells, which are human ATC cell lines. OSCC cell lines were used for comparison. Using the cell analysis system, cell migration was assessed on fibronectin-coated CIM-Plates, whereas invasion was assessed on fibronectin- and matrigel-coated CIM-Plates. SCC-4 cells exhibited high cell migration and invasion activity compared with other OSCC cell lines. TCO-1 cells exhibited equivalent cell invasion but stronger migration than SCC-4 cells. Although TCO-1 cells had strong invasive activity, they did not express MMP-9, unlike SCC-4 cells. Conversely, periostin expression was high in TCO-1 cells. Therefore, periostin expression appears to be associated with the cell migration and invasion activity of ATC. The RTCA system will be useful for the analysis of the metastatic characteristics of ATC in head and neck cancer.

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Anaplastic thyroid cancer (ATC) has a poor prognosis because of immediate metastasis. Several studies in humans and animals have suggested that the ingestion of green tea or its active ingredient (–)-epigallocatechin-3-gallate (EGCG) may decrease the risk of cancer. Using a recently developed real-time cell analysis (RTCA) system, we have shown previously that EGCG inhibits cell migration and the invasion of oral cavity cancers by suppressing matrix metalloproteinases. In the present study we used RTCA to investigate the effects of EGCG on cell adhesion to fibronectin-coated plates using three cancer cell lines: one ATC cell line (TCO-1) and two poorly differentiated oral squamous cell carcinomas (OSCCs) cell lines (SAS and HO-1-u-1; originating from the tongue and floor of the mouth, respectively). EGCG (50µM) inhibited the adhesion of all three cell lines. In addition to its effects on cell adhesion, 50µM EGCG inhibited the cell proliferation of TCO-1 cells. Furthermore, EGCG decreased α_V integrin (ITGAV) mRNA levels in all three cell lines, suggesting that EGCG inhibits the cell adhesion and proliferation of OSCC and ATC cells via suppression of integrin expression. Therefore, EGCG represents a useful dietary constituent or a lead compound for counteracting metastasis of oral cavity cancers and thyroid cancers.

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Sessile serrated adenoma/polyps (SSA/Ps) are thought to be precursors of colorectal cancers. However, current endoscopic techniques for differentiating SSA/Ps from conventional hyperplastic polyps (HPs) have low diagnostic accuracy. The aim of the present study was to assess the ability of mucosal crypt patterns to distinguish SSA/Ps from HPs. We examined 140 lesions from 93 patients that had been diagnosed histologically as SSA/Ps or HPs at the Showa University Hospital between June 2010 and May 2012. Three experienced colonoscopists reviewed the endoscopic findings of magnifying colonoscopy. Type II open-shape (Type II-O) pit patterns and varicose microvascular vessels (VMVs) were identified according to previously proposed definitions. Although 140 lesions were initially identified for the study, 27 lesions were excluded from analysis because of insufficient endoscopic findings. Thus, endoscopic findings from a total of 113 lesions (68 SSA/Ps and 45 HPs) were evaluated. Of 113 serrated polyps, 51 lesions (44 SSA/Ps and 7 HPs; P<0.01) had Type II-O pit patterns. The inter- and intra-observer agreement for these patterns among three colonoscopists was κ=0.61 (range 0.57–0.65) and κ=0.68 (range 0.52–0.94), respectively. The positive predictive value (PPV), negative predictive value (NPV), sensitivity, and specificity of Type II-O pit patterns for differentiating between SSA/P and HP were 86%, 61%, 65%, and 84%, respectively. In contrast, the PPV, NPV, sensitivity, and specificity of VMVs were 68%, 43%, 37%, and 73%, respectively. The results indicate that Type II-O mucosal crypt patterns may be useful for the differential diagnosis of SSAPs and HPs.

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Bony atrophy around the components following total knee arthroplasty (TKA) is often observed by radiography. To investigate the underlying cause of this incidence, we sought to quantitatively evaluate postoperative bone changes around the total knee components using dual-energy X-ray absorptiometry (DXA) over time. A retrospective study was conducted to compare bone mineral density (BMD) around the components in 22 patients pre-operatively and at 1, 3, 6, 12, and 24 months after cementless TKA for the treatment of osteoarthritis. In the coronal view, the medial tibia showed that a significant decrease in BMD at 12 months compared with 1 month after surgery (P<0.05), while the sagittal view showed that a significant decrease in BMD only at the anterior femoral condyle at 24 months compared with 1 month postoperatively (P<0.05). TKA improved the leg alignment and load-bearing axis in all patients, but we found that BMD tended to decrease on the medial side of the tibia and the anterior femoral condyle area over time following surgery. Our results suggest that recreating proper valgus alignment of the knee joint provides physiological balancing and condition. A larger prospective study is warranted.

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Compared with the conventional open surgery, thoracoscopic esophagectomy results in decreased thorax destruction, fewer postoperative complications, and shorter hospitalisation. However, preoperative fasting causes hyperglycemia, prompting attempts to improve postoperative hyperglycemia by preoperatively administering carbohydrate orally or intravenously. Herein, we examined the effect of preoperative carbohydrate and amino acid infusion on counter-regulatory hormone levels in patients undergoing elective thoracoscopic esophagectomy. The glucose and amino acid (GA) group (n=12) were infused with a low concentration of sugar accelerant and amino acid, and the control (GAF) group (n=12) was infused with a sugar-free extracellular fluid, until entering the operating room. We evaluated plasma catecholamine 3 fractions, cortisol, and glucose, as well as 3-methylhistidine in the urine. Adrenaline levels were significantly higher in the GAF group (263.0±201.8µIU/ml) than in the GA group (114.7±127.0µIU/ml) at the end of the surgery (P=0.042), and at postoperative day (POD) 1 (200.8±137.4 vs. 80.5±64.3µIU/ml; P=0.013). The noradrenalin level was also significantly higher in the GAF group (517.9±523.6µIU/ml) than in the GA group (254.3±205.4µIU/ml) at POD1 (P=0.028), as was the cortisol level (20.0±10.6µIU/ml vs. 10.2±8.0µIU/ml; P=0.015). No significant differences were observed between the two groups in levels of blood glucose or 3-methylhistidine in the urine. Preoperative glucose-amino acid administration improved catabolism suppression in this study.

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Psoriasis is a chronic immune-mediated disease in which the interleukin (IL)-23/IL-17 axis plays a key role in the inflammatory cascade. We recently reported that co-expression of CCR6 and melanoma cell adhesion molecule (MCAM) in effector memory CD4 T cells (T_EM) of peripheral blood might be a useful marker for T helper (Th) 17 cells. In this study, we monitored changes in T_EM expressing CCR6 and MCAM using the Psoriasis Area and Severity Index (PASI) score during anti-tumor necrosis factor (TNF) therapy. We also studied CCR6⁺ MCAM⁺ Th17 cells histologically in skin biopsy samples from psoriasis patients. In psoriasis patients treated with anti-TNF therapy, the PASI score and the percentage of CCR6⁺ MCAM⁺ T_EM cells in the blood changed almost in parallel. In immunohistochemical analyses, the proportions of IL-17⁺ T cells and MCAM⁺ T cells in the lesional skin of severely psoriatic patients were significantly higher than in mildly psoriatic patients (P<0.05), and the number of IL17⁺ T cells correlated with the PASI score (r=0.400, P<0.05). Taken together, these results indicate that CCR6⁺ MCAM⁺ Th17 cells in peripheral blood and lesional skin might play an important role in the pathology of psoriasis.

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