[18F]-FDG uptake in brain slices prepared from an aged mouse model of Alzheimer's disease using a dynamic autoradiography technique.

OBJECTIVE: 2-[18F]fluoro-2-deoxy-D-glucose positron emission tomography ([18F]-FDG-PET) is a imaging modality that has been used to measure of glucose metabolism in the brain in Alzheimer's disease (AD). Clinically, decreased glucose uptake has been reported in the brain of AD, although the precise underlying mechanisms have not yet been elucidated. To elucidate the mechanisms of decreased [18F]-FDG uptake in the AD by PET, [18F]-FDG uptake in the brain of aged model mouse of AD was investigated using a dynamic autoradiography technique "bioradiography". A X-ray phase-contrast imaging (X-PCI) and a histopathological evaluation were also investigated to elucidate the mechanisms underlying the relationships between decreased [18F]-FDG uptake and the pathological changes in the brain of AD mouse. METHODS: In this study, AD model mouse (5XFAD, APP+/PS1+) were used. [18F]-FDG-bioradiography was conducted in fresh slices of brain tissue under the condition of resting (slices immersed in 5 mM K+ solution) and metabolically active (in 50 mM K+ solution). Amyloid ß42 (Aß42) deposition in the brain of AD mouse was confirmed by X-PCI. In addition, the positive cells of phosphated tau protein (P-tau) and deposition of Aß42 were also examined by immunohistochemical staining. RESULTS: No significant differences were observed between the two groups in the resting condition. In the activate condition of the brain, [18F]-FDG uptake was significantly decreased in AD mice compared to WT mice. In X-PCI showed Aß deposition in the AD mouse, but not in the WT. The AD mouse also showed increased P-tau, accumulation of Aß42, increase in neuronal apoptosis, and decrease in the number of neurons than that of the WT mouse. CONCLUSION: Neuronal damage, and induction of neuronal apoptosis, decreased [18F]-FDG uptake, increased Aß accumulation and P-tau induced neurofibrillary degeneration are observed in AD mouse. In clinical diagnosis, reduction of [18F]-FDG uptake by PET is one of the means of diagnosing the onset of AD. Our results suggest that decreased uptake of [18F]-FDG in the brains of AD may be associated with neuronal dysfunction and cell death in the brain.

LRRK2 negatively regulates glucose tolerance via regulation of membrane translocation of GLUT4 in adipocytes.

Epidemiological studies have shown that abnormalities of glucose metabolism are involved in leucine-rich repeat kinase 2 (LRRK2)-associated Parkinson's disease (PD). However, the physiological significance of this association is unclear. In the present study, we investigated the effect of LRRK2 on high-fat diet (HFD)-induced glucose intolerance using Lrrk2-knockout (KO) mice. We found for the first time that HFD-fed KO mice display improved glucose tolerance compared with their wild-type (WT) counterparts. In addition, high serum insulin and leptin, as well as low serum adiponectin resulting from HFD in WT mice were improved in KO mice. Using western blotting, we found that Lrrk2 is highly expressed in adipose tissues compared with other insulin-related tissues that are thought to be important in glucose tolerance, including skeletal muscle, liver, and pancreas. Lrrk2 expression and phosphorylation of its kinase substrates Rab8a and Rab10 were significantly elevated after HFD treatment in WT mice. In cell culture experiments, treatment with a LRRK2 kinase inhibitor stimulated insulin-dependent membrane translocation of glucose transporter 4 (Glut4) and glucose uptake in mouse 3T3-L1 adipocytes. We conclude that increased LRRK2 kinase activity in adipose tissue exacerbates glucose tolerance by suppressing Rab8- and Rab10-mediated GLUT4 membrane translocation.

In Vivo Imaging of Tumor Hypoxia by Maintaining Green Fluorescence of 9-Aminoanthracene Under Hypoxic Conditions.

In this study, 9-aminoanthracene (9AA) was used as a new fluorescence reagent for the in vivo imaging of tumor hypoxia by taking advantage of the maintenance of its green fluorescence under hypoxic conditions. As 9AA is insoluble in water, polyethylene glycol (PEG)-400 was used to dissolve 9AA in saline. Each organ was successfully stained with 9AA, as observed by green fluorescence using in vivo imaging, following intragastric administration of a 9AA PEG-saline solution in mice. Therefore, the intragastric administration of 9AA can be used for in vivo imaging of normal mice. Tumor hypoxia staining using the 9AA fluorescence method was evaluated by in vivo imaging of mice subcutaneously transplanted with Ehrlich ascites carcinoma cells and compared with conventional pimonidazole (PIMO) staining under hypoxic conditions. The tumor sections were stained with green fluorescence derived from 9AA and the same sections corresponded to hypoxic areas upon immunohistochemical staining with PIMO.

Undaria pinnatifida (Wakame) Intake Ameliorates High-Fat Diet-Induced Glucose Intolerance via Promoting GLUT4 Expression and Membrane Translocation in Muscle.

Type 2 diabetes mellitus (T2DM), a lifestyle-related disease, is developed due to eating habits and decreased physical activity. Diabetes also increases the risk of cancer and major neurodegenerative diseases; controlling the onset of diabetes helps prevent various illnesses. Eating seaweed, such as Undaria pinnatifida (wakame), is a part of the Asian food culture. Therefore, we analyzed the antidiabetic effect of wakame intake using the high-fat diet-induced diabetes mouse model. Furthermore, we analyzed the effect of wakame extract on the cell membrane translocation of glucose transporter-4 (GLUT4) and activation of insulin signal molecules, such as AKT and AMPK, in insulin-sensitive tissues. Differentiated C2C12 cells were incubated with wakame components. The membrane translocation of GLUT4 and phosphorylation of AKT and AMPK were investigated with immunofluorescence staining and Western blotting, respectively. Also, male C57BL/6J mice were fed the normal diet (ND), high-fat diet (HFD), ND with 1% wakame powder (ND + W), or HFD with 1% wakame powder (HFD + W). We evaluated the effect of wakame intake on high-fat diet-induced glucose intolerance using an oral glucose tolerance test. Moreover, we analyzed insulin signaling molecules, such as GLUT4, AKT, and AMPK, in muscle using Western blotting. GLUT4 membrane translocation was promoted by wakame components. Also, GLUT4 levels and AKT and AMPK phosphorylation were significantly elevated by wakame components in C2C12 cells. In addition, the area under the curve (AUC) of the HFD + W group was significantly smaller than that of the HFD group. Furthermore, the level of GLUT4 in the muscle was increased in the wakame intake group. This study revealed that various wakame components exerted antidiabetic effects on the mice on a high-fat diet by promoting glucose uptake in the skeletal muscle, enhancing GLUT4 levels, and activating AKT and AMPK.

White matter imaging of ethanol-fixed rat brain by phase-contrast X-ray computed tomography.

BACKGROUND: Phase-contrast X-ray computed tomography imaging (PCI) based on crystal X-ray interferometry can detect minute density differences within biological soft tissues without contrast agents. Ethanol fixation yields increased tissue-background density differences due to the dehydrating and delipidifying effects of ethanol. PURPOSE: To obtain high image contrast of cerebral white matter structures in PCI, tissue fixation using ethanol and routinely used formalin have been examined. MATERIAL AND METHODS: Ethanol-fixed (EF) (n = 4) and formalin-fixed (FF) (n = 4) rat brains were imaged by crystal X-ray interferometry-based PCI. Tissue staining/microscopy was also performed for histological comparison and myelin density evaluation. Three-dimensional white matter tract images were reconstructed. RESULTS: Superior image contrast was obtained in the images of EF brains (EF images) compared to those of formalin-fixed brains (FF images), particularly for white matter structures. Significant density differences between the white matter structures and hippocampus (P < 0.01)/thalamus (P < 0.001) were observed in the EF, but not FF, images. Ethanol fixation enhanced the image contrast of white matter tracts by approximately sixfold compared to formalin fixation, and close agreement (r2 = 0.97; P < 0.05) between the density values on the CT images and the myelin density values in histological images was observed for the EF brains. Three-dimensional reconstruction of the white matter tracts was possible from the EF images, but not FF images. CONCLUSION: Ethanol fixation resulted in marked contrast enhancement of cerebral white matter structures in PCI. Thus, high-resolution PCI using ethanol for tissue fixation could be valuable for experimental neurological studies and postmortem neuropathology evaluation.

Visualization Ability of Phase-Contrast Synchrotron-Based X-Ray Imaging Using an X-Ray Interferometer in Soft Tissue Tumors.

Phase-contrast synchrotron-based X-ray imaging using an X-ray interferometer provides high sensitivity and high spatial resolution, and it has the ability to depict the fine morphological structures of biological soft tissues, including tumors. In this study, we quantitatively compared phase-contrast synchrotron-based X-ray computed tomography images and images of histopathological hematoxylin-eosin-stained sections of spontaneously occurring rat testicular tumors that contained different types of cells. The absolute densities measured on the phase-contrast synchrotron-based X-ray computed tomography images correlated well with the densities of the nuclear chromatin in the histological images, thereby demonstrating the ability of phase-contrast synchrotron-based X-ray imaging using an X-ray interferometer to reliably identify the characteristics of cancer cells within solid soft tissue tumors. In addition, 3-dimensional synchrotron-based phase-contrast X-ray computed tomography enables screening for different structures within tumors, such as solid, cystic, and fibrous tissues, and blood clots, from any direction and with a spatial resolution down to 26 µm. Thus, phase-contrast synchrotron-based X-ray imaging using an X-ray interferometer shows potential for being useful in preclinical cancer research by providing the ability to depict the characteristics of tumor cells and by offering 3-dimensional information capabilities.

LRRK2 Inhibition Ameliorates Dexamethasone-Induced Glucose Intolerance via Prevents Impairment in GLUT4 Membrane Translocation in Adipocytes.

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are associated with Parkinson's disease. LRRK2 is a large protein with multiple functional domains, including a guanosine 5'-triphosphate (GTP)-binding domain and a protein kinase domain. Recent studies indicated that the members of the Rab GTPase family, Rab8a and Rab10, which are involved in the membrane transport of the glucose transporter type 4 (GLUT4) during insulin-dependent glucose uptake, are phosphorylated by LRRK2. However, the physiological role of LRRK2 in the regulation of glucose metabolism is largely unknown. In the present study, we investigated the role of LRRK2 using dexamethasone (DEX)-induced glucose intolerance in mice. LRRK2 knockout (KO) mice exhibited suppressed glucose intolerance, even after treatment with DEX. The phosphorylation of LRRK2, Rab8a and Rab10 was increased in the adipose tissues of DEX-treated wild-type mice. In addition, inhibition of the LRRK2 kinase activity prevented the DEX-induced inhibition of GLUT4 membrane translocation and glucose uptake in cultured 3T3-L1 adipocytes. These results suggest that LRRK2 plays an important role in glucose metabolism in adipose tissues.

Evaluation of cell viability and metabolic activity of a 3D cultured human epidermal model using a dynamic autoradiographic technique with a PET radiopharmaceutical.

Quality control of tissues and organs for transplant is important to confirm their safety and effectiveness for regenerative medicine. However, quality evaluation is only carried out using a limited range of inspection criteria, because many of the available evaluation tests are invasive. In order to explore the potential of 2-[18F]fluoro-2-deoxy-D-glucose ([18F]FDG)-bioradiography as a non-invasive test for estimation of the safety, soundness, and effectiveness of tissues for transplantation, [18F]FDG uptake and cell viability or metabolism were investigated using a reconstructed human epidermal model (RHEM). We developed an imaging system, and suitable bioradiographic image acquisition conditions and its effectiveness were investigated. [18F]FDG uptake increased in agreement with DNA content as a marker of cell numbers and for histological assessment during cell proliferation and keratinization. [18F]FDG uptake was significantly decreased in good agreement with the viability of tissues used with various hazardous chemical treatments. [18F]FDG uptake by the tissues was decreased by hypothermia treatment and increased by hypoxia treatment while maintaining cell viability in the tissue. Therefore, [18F]FDG-bioradiography can be useful to estimate cell viability or metabolism in this RHEM. This method might be utilized as a non-invasive test for quality evaluation of tissues for transplantation.

Testicular seminoma in the aged rat visualized by phase-contrast X-ray computed tomography.

Spontaneously growing testicular seminoma in the aged rat was imaged by one of the most sensitive imaging modalities, namely, phase-contrast X-ray computed tomography (CT) with crystal X-ray interferometry. Phase-contrast X-ray CT clearly depicted the detailed inner structures of the tumor and provided 20× magnified images compared to light-microscopic images. Phase-contrast X-ray CT images are generated based on density variations in the object, whereas pathological images are based on differentiation of cellular structures, such as the cellular nuclei and cytoplasm. The mechanism of image generation differs between the two techniques: phase-contrast X-ray CT detects even minute differences in the density among pathological structures, depending, for example, on the number and sizes of the nuclei, variations of the cytoplasmic components, and presence/absence of fibrous septa, cystic changes, and hemorrhage. Thus, phase-contrast X-ray CT with a spatial resolution of 26 µm might allow prediction of the morphological characteristics of a tumor even before histopathological processing.

Biochemical Characterization of Ferulic Acid and Caffeic Acid Which Effectively Inhibit Melanin Synthesis via Different Mechanisms in B16 Melanoma Cells.

In this study, we examined the inhibitory effects of ferulic acid and caffeic acid on melanin production using a murine B16 melanoma cell line. The mechanisms by which the two acids inhibit melanin production were investigated by evaluating their effects on the activity of tyrosinase, which is involved is the first step of melanin biosynthesis. Ferulic acid showed no toxicity against the melanoma cells at any dose, whereas caffeic acid exerted cellular toxicity at concentrations higher than 0.35 mM. Both ferulic and caffeic acids effectively inhibited melanin production in the B16 melanoma cells. Ferulic acid reduced tyrosinase activity by directly binding to the enzyme, whereas no binding was observed between caffeic acid and tyrosinase. Both ferulic acid and caffeic acid inhibited casein kinase 2 (CK2)-induced phosphorylation of tyrosinase in a dose-dependent manner in vitro. Ferulic acid was found to be a more effective inhibitor of melanin production than caffeic acid; this difference in the inhibitory efficacy between the two substances could be attributable to the difference in their tyrosine-binding activity. Our analysis revealed that both substances also inhibited the CK2-mediated phosphorylation of tyrosinase.

Maintaining of the Green Fluorescence Emission of 9-Aminoanthracene for Bioimaging Applications.

The green fluorescence emission of 9-aminoanthracence (9AA) was maintained by controlling the oxidation of 9AA with oxygen in the solid state and in solution. The solid-state fluorescence of 9AA was maintained for a longer time when lauric acid was used because the equilibrium between 9AA and 9-anthrylammonium salt (9AAH + ) inclines toward the right-hand side in the presence of an acid. A solution of 9AA in CDCl3, to which nitrogen had been bubbled through for 5 min, continued to emit green fluorescence for more than 3 days, whereas the fluorescence emission disappeared within 3 days for the solution that had been bubbled with oxygen for 5 min. 9AA is oxidized by oxygen in MeOH under dark conditions to give almost nongreen fluorescent anthraquinone monoimine (AQNH), whereas dimerization of 9AA occurs under UV irradiation at 365 nm, much faster than the generation of AQNH. These results suggest that 9AA is oxidized by the triplet rather than the singlet oxygen in MeOH. Some of the organic molecules, proteins, and biological tissues were successfully stained with 9AA on microscope slides within 10 min because the green fluorescence emission of 9AA was successfully maintained in the presence of an acid and under hypoxic conditions of the used materials.

Deficiency in either COX-1 or COX-2 genes does not affect amyloid beta protein burden in amyloid precursor protein transgenic mice.

Epidemiologic studies indicate that chronic use of non-steroidal anti-inflammatory drugs (NSAIDs) is associated with a lower risk for developing Alzheimer's disease (AD). Because the primary mode of action of NSAIDs is to inhibit cyclooxygenase (COX) activity, it has been proposed that perturbed activity of COX-1 or COX-2 contributes to AD pathogenesis. To test the role of COX-1 or COX-2 in amyloid deposition and amyloid-associated inflammatory changes, we examined amyloid precursor protein (APP) transgenic mice in the context of either COX-1 or COX-2 deficiency. Our studies showed that loss of either COX-1 or COX-2 gene did not alter amyloid burden in brains of the APP transgenic mice. However, one marker of microglial activation (CD45) was decreased in brains of COX-1 deficient/APP animals and showed a strong trend in reduction in COX-2 deficient/APP animals. These results suggest that COX activity and amyloid deposition in brain are likely independent processes. Further, if NSAIDs do causally reduce the risks of AD, then our findings indicate that the mechanisms are likely not due primarily to their inhibition on COX or γ-secretase modulation activity, the latter reported recently after acute dosing of ibuprofen in humans and nonhuman primates.

Spontaneous brain tumor imaging of aged rat by crystal X-ray interferometer-based phase-contrast X-ray CT.

BACKGROUND: Crystal X-ray interferometer-based phase-contrast X-ray computed tomography (C-PCCT) enables the depiction of internal structures of biological tissue without contrast agents. PURPOSE: To determine the advantage of this technique in visualizing detailed morphological structures of a rare spontaneous brain tumor in an aged rat. MATERIAL AND METHODS: An aged rat's spontaneous brain tumor was imaged by C-PCCT without contrast agent. Three-dimensional (3D) images of the tumor microvasculature were reconstructed and compared with pathological pictures. RESULTS: C-PCCT depicted the tumor's various pathological features clearly, e.g. its cell density and vasculature, and blood clots caused by hemorrhaging and/or hematomas. The obtained images resembled pathological pictures with a magnification of ×20 and were used to reconstruct 3D images of the tumor vascularity up to approximately 26 µm in diameter. CONCLUSION: Since C-PCCT is able to depict various pathological conditions, it might be useful for cancer research.

γ-Secretase Modulators and APH1 Isoforms Modulate γ-Secretase Cleavage but Not Position of ε-Cleavage of the Amyloid Precursor Protein (APP).

The relative increase in Aß42 peptides from familial Alzheimer disease (FAD) linked APP and PSEN mutations can be related to changes in both ε-cleavage site utilization and subsequent step-wise cleavage. Cleavage at the ε-site releases the amyloid precursor protein (APP) intracellular domain (AICD), and perturbations in the position of ε-cleavage are closely associated with changes in the profile of amyloid ß-protein (Aß) species that are produced and secreted. The mechanisms by which γ-secretase modulators (GSMs) or FAD mutations affect the various γ-secretase cleavages to alter the generation of Aß peptides have not been fully elucidated. Recent studies suggested that GSMs do not modulate ε-cleavage of APP, but the data were derived principally from recombinant truncated epitope tagged APP substrate. Here, using full length APP from transfected cells, we investigated whether GSMs modify the ε-cleavage of APP under more native conditions. Our results confirmed the previous findings that ε-cleavage is insensitive to GSMs. In addition, fenofibrate, an inverse GSM (iGSM), did not alter the position or kinetics of ε-cleavage position in vitro. APH1A and APH1B, a subunit of the γ-secretase complex, also modulated Aß42/Aß40 ratio without any alterations in ε-cleavage, a result in contrast to what has been observed with PS1 and APP FAD mutations. Consequently, GSMs and APH1 appear to modulate γ-secretase activity and Aß42 generation by altering processivity but not ε-cleavage site utilization.

Suppressive Effect of Dietary Fucoidan on Proinflammatory Immune Response and MMP-1 Expression in UVB-Irradiated Mouse Skin.

It is well known that ultraviolet B irradiation leads to dermal inflammation. In this study, we found that Mekabu fucoidan suppressed edema, decreased the thickness of the prickle cell layer, and decreased matrix metalloproteinase 1 in the skin of mice irradiated with ultraviolet B. Moreover, we found that the mean level of interferon gamma of Mekabu fucoidan-treated, ultraviolet B-irradiated mice (approximately 2.2 ng/mL) was not significantly different from that in normal mice (approximately 2.5 ng/mL). In contrast, a significant decrease in the mean level of interferon gamma (approximately 1.3 ng/mL) in ultraviolet B-irradiated control mice was observed compared with that in Mekabu fucoidan-treated, ultraviolet B-irradiated mice. The mean thickness of the prickle cell layer in the skin of Mekabu fucoidan-treated, ultraviolet B-irradiated mice was less than that in the ultraviolet B-irradiated control mice. Metalloproteinase 1 activity was significantly higher in the skin of ultraviolet B-irradiated mice than in the skin of untreated, nonirradiated normal mice. Metalloproteinase 1 in the skin of ultraviolet B-irradiated, Mekabu fucoidan- or L(+)-ascorbic acid (vitamin C)-treated mice was significantly lower than that in the ultraviolet B-irradiated control mice. Mitigation of the morphological changes in Mekabu fucoidan-treated mice was correlated with a decrease in metalloproteinase 1 levels. These data indicate that Mekabu fucoidan is an effective suppressor of inflammation in an ultraviolet B-irradiated mouse model.

Enhanced renal image contrast by ethanol fixation in phase-contrast X-ray computed tomography.

Phase-contrast X-ray imaging using a crystal X-ray interferometer can depict the fine structures of biological objects without the use of a contrast agent. To obtain higher image contrast, fixation techniques have been examined with 100% ethanol and the commonly used 10% formalin, since ethanol causes increased density differences against background due to its physical properties and greater dehydration of soft tissue. Histological comparison was also performed. A phase-contrast X-ray system was used, fitted with a two-crystal X-ray interferometer at 35 keV X-ray energy. Fine structures, including cortex, tubules in the medulla, and the vessels of ethanol-fixed kidney could be visualized more clearly than that of formalin-fixed tissues. In the optical microscopic images, shrinkage of soft tissue and decreased luminal space were observed in ethanol-fixed kidney; and this change was significantly shown in the cortex and outer stripe of the outer medulla. The ethanol fixation technique enhances image contrast by approximately 2.7-3.2 times in the cortex and the outer stripe of the outer medulla; the effect of shrinkage and the physical effect of ethanol cause an increment of approximately 78% and 22%, respectively. Thus, the ethanol-fixation technique enables the image contrast to be enhanced in phase-contrast X-ray imaging.

Leucine-rich repeat kinase 2 regulates tau phosphorylation through direct activation of glycogen synthase kinase-3ß.

Leucine-rich repeat kinase 2 (LRRK2) has been identified as the causal molecule for autosomal-dominant Parkinson's disease (PD). Experimental evidence indicates that LRRK2 may play an important role in the pathology induced by abnormal phosphorylation of tau. In the present study, we demonstrated that LRRK2 directly associates with GSK-3ß, and that this interaction enhances the kinase activity of GSK-3ß. Furthermore, we found that LRRK2-mediated activation of GSK-3ß induces high phosphorylation of tau at Ser396 in SH-SY5Y cells. From our present findings, we conclude that LRRK2 may function as a novel enhancer for GSK-3ß and as a physiological regulator of neurite outgrowth and axonal transport through regulation of the GSK-3ß-mediated phosphorylation of tau at the cellular level. Since LRRK2 is detected in tau-positive inclusions in brain tissue affected by various neurodegenerative disorders, including PD, LRRK2-stimulated phosphorylation of tau by GSK-3ß may be involved in development of pathological features in the initial stage of PD.

Modulation of chemokine expression on intestinal epithelial cells by Kampo (traditional Japanese herbal) medicine, Hochuekkito, and its active ingredients.

The intestinal epithelial cells sit at the interface between a lumen and a lamina propria or lymph nodes such as Peyer's patches, where they play important roles in maintaining intestinal homeostasis through chemokine secretion. This study investigated the effect of Hochuekkito (TJ-41)-a traditional Japanese herbal (Kampo) formula used as a tonic for weakness-on chemokine expression in intestinal epithelial cells in order to explore the mechanism of its modulating effect against mucosal immunity. When cells from the rat normal small intestinal epithelial cell-line IEC-6 were stimulated with TJ-41, mRNA expression of CC chemokine ligand (CCL) 11 (eotaxin), CCL20 (MIP-3α) and CCL25 (TECK) was enhanced. Oral administration of TJ-41 to methotrexate-treated mice enhanced mRNA expression of CCL25 and keratinocyte growth factor in the jejunum with, decreasing mRNA expression of the inflammatory marker tumor necrosis factor (TNF)-α. Although oral administration of TJ-41 did not affect CCL20 mRNA expression in villus epithelium of methotrexate-treated mice, enhancement of CCL20 mRNA expression was observed in Peyer's patches. Immunohistochemical analysis detected dense staining with anti-CCL20 antibody in the follicle-associated epithelium region of Peyer's patches in mice administered TJ-41. Analysis of active ingredients indicates that polysaccharide-containing macromolecules in TJ-41 contribute to the enhancement of CCL20 mRNA expression through an intracellular signal cascade via nuclear factor kappa B (NF-κB) activation.

Tumor apoptosis in prostate cancer by PGD(2) and its metabolite 15d-PGJ(2) in murine model.

Fifteen-deoxy-Δ(12,14)-PGJ(2) (15d-PGJ(2)) is one of non-enzymatically converted metabolite from prostaglandin D(2) (PGD(2)). Anti-tumor effects of 15d-PGJ(2) in various tumors are partially known, but the detail of in vivo mechanisms of action is still unclear. In this study, we investigated the effects of 15d-PGJ(2) and PGD(2) on murine prostate cancer in vitro and in vivo. Murine prostate cancer cells RM9 were transfected with murine prostaglandin D(2) synthase (mPGDS) gene by using defective retrovirus vector, designated as RM9-mPGDS. In addition, RM9 was also transfected with only defective retrovirus vector, designated as RM9-EV and used as control in this study. The expression and production of the gene were confirmed by RT-PCR and ELISA, respectively. For in vivo study, RM9-mPGDS was injected into the back of C57BL/6 mice, then resulted tumor was used for pathological analysis 14days after the inoculation. Tumor cell apoptosis in the tissue was detected by TUNEL staining. Retrovirally transfected mPGDS in RM9 significantly induced apoptosis in vivo but not in vitro, by TUNEL staining and cell death ELISA, respectively. Our results strongly suggested that the apoptosis induced in RM9-mPGDS in vivo was probably achieved in tumor environment such as hypoxic condition. The introduction of PGDS gene into cancer cells might be a novel therapy against cancer.