Akihide Tanimoto

Shortened microsatellite d(CA)21 sequence down-regulates promoter activity of matrix metalloproteinase 9 gene.

One characteristic elements in the promoter of the matrix metalloproteinase 9 (MMP‐9) gene is the d(CA) repeat. To investigate whether this element regulates the transcription of the MMP‐9 gene and its enzymatic activities, we sequenced the promoter region isolated from esophageal carcinoma cell lines. TE9 cells with low MMP‐9 enzymatic activity had the number of d(CA) repeats shortened from 21 to 14 or 18. TE8, TE10 and TE11 cells with high MMP‐9 activities had 21 or 23 d(CA) repeats. Luciferase assays using MMP‐9 promoter containing 18, 14 or 0 d(CA) repeats showed transcriptional activities which were 50, 50 or 5%, respectively, of the level achieved with promoter containing 21 d(CA) repeats. Sequence analysis of the promoter of 223 Japanese subjects revealed that most had two alleles with 20, 21 or 22 d(CA) repeats, whereas six had one or two alleles with 14, 18 or 19 d(CA) repeats. We postulate that length alteration of the d(CA) repeat causes phenotypic differences among carcinoma cells and that microsatellite instability may contribute to the polymorphism of d(CA) repeat length.

Programmed Cell Death Protein 4 Down-regulates Y-Box Binding Protein-1 Expression via a Direct Interaction with Twist1 to Suppress Cancer Cell Growth

Programmed cell death protein 4 (PDCD4) has recently been shown to be involved in both transcription and translation, and to regulate cell growth. However, the mechanisms underlying PDCD4 function are not well understood. In this study, we show that PDCD4 interacts directly with the transcription factor Twist1 and leads to reduced cell growth through the down-regulation of the Twist1 target gene Y-box binding protein-1 (YB-1). PDCD4 interacts with the DNA binding domain of Twist1, inhibiting its DNA binding ability and YB-1 expression. Immunohistochemical analysis showed that an inverse correlation between nuclear PDCD4 and YB-1 expression levels was observed in 37 clinical prostate cancer specimens. Growth suppression by PDCD4 expression was completely recovered by either Twist1 or YB-1 expression. Moreover, PDCD4-overexpressing cells are sensitive to cisplatin and paclitaxel but not to etoposide or 5-fluorouracil. In summary, PDCD4 negatively regulates YB-1 expression via its interaction with Twist1 and is involved in cancer cell growth and chemoresistance.

Vasculoprotective roles of neuronal nitric oxide synthase

Nitric oxide (NO) has multiple important actions that contribute to the maintenance of vascular homeostasis. NO is synthesized by three different isoforms of NO synthase (NOS), all of which have been reported to be expressed in human atherosclerotic vascular lesions. Although the regulatory roles of endothelial NOS (eNOS) and inducible NOS (iNOS) on the development of atherosclerosis have been described, little is known about the role of neuronal NOS (nNOS). Here, we show that nNOS also exerts important vasculoprotective effects in vivo. In a carotid artery ligation model, nNOS gene‐deficient (nNOS‐KO) mice exhibited accelerated neointimal formation and constrictive vascular remodeling caused by blood flow disruption. In a rat balloon injury model, the selective inhibition of nNOS activity potently enhanced vasoconstrictor responses to a variety of calcium‐mobilizing stimuli, suppressed tissue cGMP concentrations, a marker of vascular NO production, and exacerbated neointimal formation. In both models, nNOS was absent before injury and was up‐regulated only after the injury, and was predominantly expressed in the neointima and medial smooth muscle cells. These results provide the first direct evidence that nNOS plays important roles in suppressing arteriosclerotic vascular lesion formation in vivo.

Cell death and regeneration of renal proximal tubular cells in rats with subchronic cadmium intoxication

Male Sprague-Dawley rats were injected subcutaneously with 0.6 mg cadmium (Cd)/kg/day for 8 wk. The subsequent changes in renal proximal tubules were studied histologically, histochemically, and ultrastructurally. The urinary and tissue Cd concentrations were determined by atomic absorption spectrophotometry. After 4 wk of exposure, apoptosis was observed predominantly in segment S3 along with epithelial regeneration in the affected tubules, and these changes gradually became more pronounced as the experimental period was prolonged. The apoptotic cells were shed into the lumen and were found to contain a large quantity of Cd. Apoptotic cells were counted in paraffin sections after various periods of exposure to Cd. Nuclear bromodeoxyuridine uptake, mitotic count, and nuclear density were used as indicators of tubular regeneration. A correlation was found between the numerical increase of apoptotic cells and the rate of urinary Cd excretion, and the rate of increase in the tissue Cd concentration had a tendency to reduce after 4 wk as the rate of urinary Cd increased. These observations suggest that apoptosis might be helpful for the efficient excretion of Cd into urine. Progressive increases in the preceding indicators of regeneration were observed. From our results, it appears that Cd-induced tubular damage, i.e., cell deletion due to apoptosis, is reversible as a result of marked epithelial regeneration. On the basis of these histological changes, the critical concentration of Cd required to produce renal tubular damage was estimated to be 600 μg/g dry tissue.

Statin Treatment Upregulates Vascular Neuronal Nitric Oxide Synthase Through Akt/NF-κB Pathway

Objective—Three-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) are known to enhance vascular expression of endothelial (eNOS) and inducible nitric oxide synthase (iNOS). In this study, we examined whether statins also upregulate vascular expression of neuronal NOS (nNOS). Methods and Results—In cultured rat aortic smooth muscle cells, treatment with atorvastatin significantly increased nNOS expression, associated with activation of Akt and NF-&kgr;B. Inhibition of Akt by dominant-negative Akt suppressed atorvastatin-induced nNOS expression as well as Akt and NF-&kgr;B activation. Inhibition of NF-&kgr;B by dominant-negative I&kgr;B also attenuated atorvastatin-induced nNOS expression and NF-&kgr;B activation, but not Akt activation. We further examined whether atorvastatin also enhances nNOS expression in isolated mouse aorta, and if so, how much nNOS-derived NO accounts for atorvastatin-induced NOx production. In isolated aortas of wild-type mice, atorvastatin significantly increased all three NOS isoform expression and NOx production. In isolated aortas of doubly i/eNOS−/−, n/eNOS−/−, and n/iNOS−/− mice, which express only nNOS, iNOS, and eNOS, respectively, atorvastatin-induced NOx production was approximately 25%, 25%, and 50% to that of wild-type mice, respectively, suggesting that nNOS accounts for 25% of the atorvastatin-mediated NOx production. Conclusions—These results indicate that atorvastatin upregulates vascular nNOS through Akt/NF-&kgr;B pathway, demonstrating a novel nNOS-mediated vascular effect of the statin.

Switch of histamine receptor expression from H2 to H1 during differentiation of monocytes into macrophages

It is known that histamine suppresses gene expression and synthesis of tumor necrosis factor alpha (TNF‐α) induced by lipopolysaccharide (LPS) in human peripheral blood mononuclear monocytes (HPM) or alveolar macrophages via histamine H2 receptors. We investigated the effect of histamine and differentiation in macrophages on the expression and secretion of TNF‐α, TNF‐α‐converting enzyme (TACE), and histamine H1 and H2 receptors by use of a leukemia cell line, U937, and HPM. Differentiation of U937 and HPM cells with 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) enhanced the H1 receptor expression and rather suppressed the H2 receptor, resulting in up‐regulation of the histamine‐induced expression and secretion of TNF‐α, modulated via TACE. Therefore, histamine failed to inhibit up‐regulated expression of TNF‐α induced by LPS in macrophages. The switch from H2 to H1 receptors during differentiation in the monocyte/macrophage lineage could participate in the pathogenic processes of atherosclerosis and inflammatory reactions in the arterial wall.

Overexpression of peroxiredoxin 4 protects against high-dose streptozotocin-induced diabetes by suppressing oxidative stress and cytokines in transgenic mice.

Peroxiredoxin 4 (PRDX4) is one of a newly discovered family of antioxidative proteins. We generated human PRDX4 (hPRDX4) transgenic (Tg) mice, displaying a high level of hPRDX4 expression in the pancreatic islets, and then focused on the functions of PRDX4 in a type 1 diabetes mellitus (T1DM) model using a single high dose of streptozotocin (SHDS). After SHDS-injection, Tg mice showed significantly less hyperglycemia and hypoinsulinemia and a much faster response on glucose tolerance test than wild-type (WT) mice. Morphologic and immunohistochemical observation revealed that the pancreatic islet areas of Tg mice were larger along with less CD3-positive lymphocyte infiltration compared with WT mice. Upon comparison between these two mouse models, β-cell apoptosis was also repressed, and reversely, β-cell proliferation was enhanced in Tg mice. Real-time RT-PCR demonstrated that the expression of many inflammatory-related molecules and their receptors and transcription factors were significantly downregulated in Tg mice. These data indicate that PRDX4 can protect pancreatic islet β-cells against injury caused by SHDS-induced insulitis, which strongly suggests that oxidative stress plays an essential role in SHDS-induced diabetes. This study, for the first time, implicates that PRDX4 has a pivotal protective function against diabetes progression in this T1DM model.

Aldehyde dehydrogenase 1 expression predicts poor prognosis in triple-negative breast cancer

Ohi Y, Umekita Y, Yoshioka T, Souda M, Rai Y, Sagara Y, Sagara Y, Sagara Y & Tanimoto A
(2011) Histopathology59, 776–780

Role of Histamine Produced by Bone Marrow–Derived Vascular Cells in Pathogenesis of Atherosclerosis

To clarify the role of histamine-producing cells and its origin in atherosclerosis, we investigated histidine decarboxylase (HDC; histamine-producing enzyme) expression in murine arteries with vascular injuries after the animal had received transplanted bone marrow (BM) from green fluorescent protein (GFP)–transgenic mice. The neointima in the ligated carotid arteries contained BM-derived HDC+ cells that expressed macrophage (Mac-3) or smooth muscle cell antigen (&agr;-SMA). In contrast, the HDC+ BM-derived cells, which were positive for Mac-3, were mainly located in the adventitia in the cuff replacement model. In apolipoprotein E-knockout mice on a high cholesterol diet, BM-derived cells expressing Mac-3 in the atheromatous plaques were also positive for HDC. In comparison with wild-type mice, HDC−/− mice showed reduced neointimal thickening and a decreased intima-to-media ratio after ligation and cuff replacement. These results indicate that histamine produced from BM-derived progenitor cells, which could transdifferentiate into SMC- or macrophage-like cells, are important for the formation of neointima and atheromatous plaques.

Effects of histamine and interleukin-4 synthesized in arterial intima on phagocytosis by monocytes/macrophages in relation to atherosclerosis

We investigated the localization of histidine decarboxylase (HDC), which is the rate‐limiting enzyme that generates histamine from histidine, in human aorta/coronary artery. RT‐PCR and immunohistochemical staining revealed that the HDC gene was expressed in monocytes/macrophages and T cells in the arterial intima but not in smooth muscle cells in either the arterial intima or the media. A luciferase promoter assay with U937 and Jurkat cells demonstrated that interleukin‐4 (IL‐4) inhibited the expression of the HDC gene. In contrast, among a scavenger receptor family, IL‐4 as well as histamine up‐regulated U937 cells to express the LOX‐1 gene but not the SR‐A gene, which genes encode receptors that scavenge oxidized lipids. These findings suggest that histamine synthesized in the arterial wall participates in the initiation and progression of atherosclerosis and that IL‐4 can act as an important inhibitory and/or stimulatory factor in the function of monocytes/macrophages modulated by histamine in relation to the process of atherosclerosis.