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Background: The increasing incidence and poor outcome associated with malignant pleural effusion (MPE) requires finding an effective treatment for this disease. Inhibitory B7-H4 is expressed in many different human cancers but its role in malignant pleural tissue has yet to be established.Methods: Here, patients with metastatic pleural adenocarcinoma (MPA) or with early-stage lung adenocarcinoma were clinically and statistically analyzed. Immunohistochemistry and confocal microscopy were used to determinate the expression of B7-H4 in the cancer cells. By using MPE model, we sought to a potential immunotherapy for MPE with anti-B7-H4 mAb.Results: When compared to early-stage lung adenocarcinoma, MPA possessed higher level of nuclei membranous B7-H4 and lower cytoplasmic B7-H4 expression. Also, nuclei membranous B7-H4 expression was found to be positively correlated to Ki-67 expression, and indicated a possible poor prognosis of MPA. In mouse MPE model, intra-pleurally injection of anti-B7-H4 mAb effectively suppressed MPE formation.Conclusions: Taken together, our data was in support of the significance of B7-H4 expression in MPA, which also suggest it warrants further exploration for potential immunotherapy of MPE. [ABSTRACT FROM AUTHOR]

Copper is a nutritional metal required for brain development and function. Wilson's disease (WD), or hepatolenticular degeneration, is an inherited human copper metabolism disorder caused by a mutation of the ATP7B gene. Many WD patients present with variable neurological and psychiatric symptoms, which may be related to neurodegeneration secondary to copper metabolism imbalance. The objective of this study was to explore the feasibility and use of copper-64 chloride ([C]CuCl) as a tracer for noninvasive assessment of age-dependent changes of cerebral copper metabolism in WD using an Atp7b knockout mouse model of WD and positron emission tomography/computed tomography (PET/CT) imaging. Continuing from our recent study of biodistribution and radiation dosimetry of [C]CuCl in Atp7b knockout mice, PET quantitative analysis revealed low Cu radioactivity in the brains of Atp7b knockout mice at 7th weeks of age, compared with Cu radioactivity in the brains of age- and gender-matched wild type C57BL/6 mice, at 24 h (h) post intravenous injection of [C]CuCl as a tracer. Furthermore, age-dependent increase of Cu radioactivity was detected in the brains of Atp7b knockout mice from the 13th to 21th weeks of age, based on the data derived from a longitudinal [C]CuCl-PET/CT study of Atp7b knockout mice with orally administered [Cu]CuCl as a tracer. The findings of this study support clinical use of [Cu]CuCl-PET/CT imaging as a tool for noninvasive assessment of age-dependent changes of cerebral copper metabolism in WD patients presenting with variable neurological and psychiatric symptoms. [ABSTRACT FROM AUTHOR]

Background Indoleamine 2,3-dioxygenase (IDO), the enzyme that catalyzes the first and rate-limiting step of tryptophan catabolism, suppresses T-cell responses by tryptophan depletion and accumulation of kynurenine metabolites. IDO prevents allograft rejection in various transplantations. Methods Dendritic cells (DC) highly expressing IDO (IDO + DC) were cultured through transduction of adenovirus vectors carrying the IDO sequence. IDO + DC were incubated with CD4 + CD25 − T cells to detect T cell proliferation. The effects of IDO + DC and 3-Hydroxyanthranilic acid (3-HAA) were verified in an allogeneic murine small bowel transplantation (SBT) model. Foxp3 + Treg cells of recipient mice were detected by flow cytometry and cytokines in plasma were determined by ELISA. Results IDO + DC effectively suppressed proliferation of CD4 + CD25 − T cells in vitro, and this effect could be enhanced by adding 3-HAA. In the SBT transplantation model, both 3-HAA (P < 0.05) and IDO + DC (P < 0.01) prolonged the survival time of transplanted mice. Mice treated with IDO + DC achieved longer mean survival time than 3-HAA administrated mice (11.5d vs. 18.5d). Grafts from IDO + DC, 3-HAA and combination treatment group showed reduced inflammation and minimal architectural distortion. IFN-γ production was significantly inhibited by IDO + DC and 3-HAA (P < 0.05). The expression of IL-2 was slightly lower with 3-HAA or IDO + DC treatment. However, IL-10 was higher in 3-HAA, IDO + DC and combination treatment groups, while TGF-β was elevated in all non-control groups. Conclusions IDO + DC plus 3-HAA has an immunoprotective role and represents a potential strategy to suppress acute rejection and prolong survival of grafts in SBT. [ABSTRACT FROM AUTHOR]

Pelvic organ cross-sensitization, also termed as viscero-visceral referred hyperalgesia, is a major contributor to painful endometriosis. Its underlying mechanism is poorly understood. Clinical and basic studies have shown that gabapentin, a drug that binds to the αδ−1 subunit of voltage-dependent calcium channels (Caαδ−1), is effective in treating chronic visceral pain. Accordingly, we hypothesized that pelvic organ cross-sensitization in painful endometriosis is mediated by an upregulation of Caαδ−1 in the spinal cord. We examined if the dysregulation of spinal Caαδ−1 subunit may play an important role in the development of ectopic growths-to-colon cross-sensitization in a rat model of experimentally-induced endometriosis. Our findings suggest that there was an increased Caαδ−1 expression in the dorsal horn and an ectopic growths-to-colon cross-sensitization in female rats with established endometriosis. Intrathecal administration of gabapentin (300 μg) remarkably reduced the ectopic growths-to-colon cross-sensitization in rats with established endometriosis. Furthermore, intrathecal injection of Caαδ−1 antisense oligodeoxynucleotides reversed the ectopic growths-to-colon cross-sensitization and also normalized the upregulation of spinal Caαδ−1 expression in endometriosis rats. The current study suggests that the upregulation of Caαδ−1 in the spinal cord may contribute to pelvic organ cross-sensitization in painful endometriosis. Our study may provide a biological basis for selectively targeting this pathway to relieve viscero-visceral referred hyperalgesia in patients with painful endometriosis. [ABSTRACT FROM AUTHOR]

ASPP2 is a pro-apoptotic member of the p53 binding protein family. ASPP2 has been shown to inhibit autophagy, which maintains energy balance in nutritional deprivation. We attempted to identify the role of ASPP2 in the pathogenesis of non-alcoholic fatty liver disease ( NAFLD). In a NAFLD cell model, control treated and untreated HepG2 cells were pre-incubated with GFP-adenovirus ( GFP-ad) for 12 hrs and then treated with oleic acid ( OA) for 24 hrs. In the experimental groups, the HepG2 cells were pre-treated with ASPP2-adenovirus ( ASPP2-ad) or ASPP2-si RNA for 12 hrs and then treated with OA for 24 hrs. BALB/c mice fed a methionine- and choline-deficient ( MCD) diet were used to generate a mouse model of NAFLD. The mice with fatty livers in the control group were pre-treated with injections of GFP-ad for 10 days. In the experimental group, the mice that had been pre-treated with ASPP2-ad were fed an MCD diet for 10 days. ASPP2-ad or GFP-ad was administered once every 5 days. Liver tissue from fatty liver patients and healthy controls were used to analyse the role of ASPP2. Autophagy, apoptosis markers and lipid metabolism mediators, were assessed with confocal fluorescence microscopy, immunohistochemistry, western blot and biochemical assays. ASPP2 overexpression decreased the triglyceride content and inhibited autophagy and apoptosis in the HepG2 cells. ASPP2-ad administration suppressed the MCD diet-induced autophagy, steatosis and apoptosis and decreased the previously elevated alanine aminotransferase levels. In conclusion, ASPP2 may participate in the lipid metabolism of non-alcoholic steatohepatitis and attenuate liver failure. [ABSTRACT FROM AUTHOR]

The tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), which is abundant in tobacco smoke, is a potent lung procarcinogen. The present study was aimed to prove that transgenic expression of human cytochrome P450 2A13 (CYP2A13), known to be selectively expressed in the respiratory tract and be the most efficient enzyme for NNK bioactivation in vitro, will enhance NNK bioactivation and NNK-induced tumorigenesis in the mouse lung. Kinetic parameters of NNK bioactivation in vitro and incidence of NNK-induced lung tumors in vivo were determined for wild-type, Cyp2a5-null and CYP2A13-humanized (CYP2A13-transgenic/Cyp2a5-null) mice. As expected, in both liver and lung microsomes, the loss of CYP2A5 resulted in significant increases in Michaelis constant (Km) values for the formation of 4-oxo-4-(3-pyridyl)-butanal, representing the reactive intermediate that can lead to the formation of O6-methylguanine (O6-mG) DNA adducts; however, the gain of CYP2A13 at a fraction of the level of mouse lung CYP2A5 led to recovery of the activity in the lung, but not in the liver. The levels of O6-mG, the DNA adduct highly correlated with lung tumorigenesis, were significantly higher in the lungs of CYP2A13-humanized mice than in Cyp2a5-null mice. Moreover, incidences of lung tumorigenesis were significantly greater in CYP2A13-humanized mice than in Cyp2a5-null mice, and the magnitude of the differences in incidence was greater at low (30mg/kg) than at high (200mg/kg) NNK doses. These results indicate that CYP2A13 is a low Km enzyme in catalyzing NNK bioactivation in vivo and support the notion that genetic polymorphisms of CYP2A13 can influence the risks of tobacco-induced lung tumorigenesis in humans. [ABSTRACT FROM AUTHOR]

Germ cells divide and differentiate in a unique local microenvironment under the control of somatic cells. Signals released in this niche instruct oocyte reentry into the meiotic cell cycle. Once initiated, the progression through meiosis and the associated programme of maternal messenger RNA translation are thought to be cell autonomous. Here we show that translation of a subset of maternal mRNAs critical for embryo development is under the control of somatic cell inputs. Translation of specific maternal transcripts increases in oocytes cultured in association with somatic cells and is sensitive to EGF-like growth factors that act only on the somatic compartment. In mice deficient in amphiregulin, decreased fecundity and oocyte developmental competence is associated with defective translation of a subset of maternal mRNAs. These somatic cell signals that affect translation require activation of the PI(3)K-AKT-mTOR pathway. Thus, mRNA translation depends on somatic cell cues that are essential to reprogramme the oocyte for embryo development. [ABSTRACT FROM AUTHOR]

Abstract: Previous studies have shown that high-intensity focused ultrasound (HIFU) ablation can enhance host antitumor immune response, though the mechanism is still unknown. In the present study, we investigated whether HIFU ablation could activate tumor-specific T lymphocytes and then induce antitumor cellular immunity. We studied 70 C57BL/6J mice bearing the H22 tumor; they were randomly divided into a HIFU group and a sham-HIFU group. Of the mice, 35 in the HIFU group underwent HIFU ablation of the H22 hepatic tumor, and the remaining 35 received a sham-HIFU procedure. In addition, 35 female, naïve syngeneic C57BL/6J mice were used as controls. All mice were sacrificed 14 days after HIFU, and the spleens were harvested. The function of T lymphocytes was determined. As a valuable tool for detecting and characterizing peptide-specific cells, the frequency of MHC class I tetramer/CD8-positive cells was quantified, which could help to determine the response and number of T lymphocytes. The therapeutic effect of the HIFU-activated lymphocytes on tumor-bearing mice was investigated after adoptive transfer of the lymphocytes. The results showed that compared to sham-HIFU and control groups, HIFU ablation significantly increased the cytotoxicity of cytotoxic T lymphocytes (p < 0.05), with a significant increase of IFN-γ and TNF-α secretion (p < 0.001). The frequency of the MHC class I tetramer/CD8-positive cells was significantly higher in the HIFU group (p < 0.05). A stronger inhibition of tumor progression and higher survival rates were observed to be significant after adoptive immunotherapy in the HIFU group as compared to the sham-HIFU and control groups (p < 0.01). It is concluded that HIFU ablation could activate tumor-specific T lymphocytes, thus inducing antitumor cellular immune responses in tumor-bearing mice. [Copyright &y& Elsevier]

Aim:To investigate whether fluvastatin is able to ameliorate the impaired cardiac function or baroreflex sensitivity (BRS) in rats with type 1 diabetes.Methods:Type 1 diabetic rats were induced by intraperitoneal injection of streptozotocin (STZ) and then administered fluvastatin (1.5, 3.0, and 6.0 mg·kg−1·d−1) for 30 d. Food and drink intake was recorded every day. Fasting blood glucose (FBG) level, blood lipid level, cardiac function and BRS were measured in diabetic rats after fluvastatin treatment for 30 d.Results:The polydipsia, polyphagia and abnormal biochemical indexes of blood were significantly ameliorated by the the 3.0- and 6.0-mg doses of fluvastatin in STZ-induced diabetic rats. FBG was decreased in diabetic rats after fluvastatin treatment for 30 d. The left ventricular systolic pressure (LVSP) and the maximum rate of change of left ventricular pressure in the isovolumic contraction and relaxation period (±dp/dtmax) were elevated, and left ventricular diastolic pressure (LVEDP) was decreased by fluvastatin. The attenuated heart rate responses to arterial blood pressure (ABP) increase induced by phenylephrine (PE) and ABP decrease induced by sodium nitroprusside (SNP) were reversed by the 3.0-mg dose of fluvastatin.Conclusion:Fluvastatin regulates blood lipid levels and decreases the FBG level in diabetic rats. These responses can protect the diabetic heart from complications by improving cardiac function and BRS. [ABSTRACT FROM AUTHOR]

Abstract: The herbicide 2,6-dichlorobenzonitril (DCBN) is a potent and tissue-specific toxicant to the olfactory mucosa (OM). The toxicity of DCBN is mediated by cytochrome P450 (P450)-catalyzed bioactivation; however, it is not known whether target-tissue metabolic activation is essential for toxicity. CYP2A5, expressed abundantly in both liver and OM, was previously found to be one of the P450 enzymes active in DCBN bioactivation in vitro. The aims of this study were to determine the role of CYP2A5 in DCBN toxicity in vivo, by comparing the extents of DCBN toxicity between Cyp2a5-null and wild-type (WT) mice, and to determine whether hepatic microsomal P450 enzymes (including CYP2A5) are essential for the DCBN toxicity, by comparing the extents of DCBN toxicity between liver-Cpr-null (LCN) mice, which have little P450 activity in hepatocytes, and WT mice. We show that the loss of CYP2A5 expression did not alter systemic clearance of DCBN (at 25mg/kg); but it did inhibit DCBN-induced non-protein thiol depletion and cytotoxicity in the OM. Thus, CYP2A5 plays an essential role in mediating DCBN toxicity in the OM. In contrast to the results seen in the Cyp2a5-null mice, the rates of systemic DCBN clearance were substantially reduced, while the extents of DCBN-induced nasal toxicity were increased, rather than decreased, in the LCN mice, compared to WT mice. Therefore, hepatic P450 enzymes, although essential for DCBN clearance, are not necessary for DCBN-induced OM toxicity. Our findings form the basis for a mechanism-based approach to assessing the potential risks of DCBN nasal toxicity in humans. [ABSTRACT FROM AUTHOR]

Abstract: A novel method was established for simultaneous quantitation of testosterone (T) and dihydrotestosterone (DHT) in murine tissue and serum samples. Endogenous T and DHT, together with the internal standards 17α-methyl-T and 17α-methyl-DHT, were extracted from tissues and then derivatized by reaction with 2-hydrazino-4-(trifluoromethyl)-pyrimidine (HTP). Analysis by liquid chromatography–electrospray ionization tandem mass spectrometry (LC–ESI–MS/MS) resulted in product ion spectra of HTP derivatives of both T and DHT that showed analyte-specific fragmentations; the latter fragmentations were characterized by the use of high-resolution Orbitrap MS/MS. These specific fragmentations enabled quantitation of T and DHT in the multiple-reaction monitoring (MRM) mode. The method was validated with charcoal-stripped serum as the matrix. The lower limit of quantitation (LLOQ) was 0.10ng/ml for T and 0.50ng/ml for DHT. The method was then used for determination of serum and tissue levels of T and DHT in transgenic mice carrying a hypomorphic NADPH–cytochrome P450 reductase gene (Cpr-low mice). Remarkably, ovarian T levels in Cpr-low mice were found to be 25-fold higher than those in wild-type mice, a finding that at least partly explains the female infertility seen in the Cpr-low mice. In conclusion, our method provides excellent sensitivity and selectivity for determination of endogenous levels of T and DHT in mouse tissues. [Copyright &y& Elsevier]

Reactive astrocytes play an important role in the development of Alzheimer's disease and primary tauopathies. Here, we aimed to investigate the relationships between reactive astrocytes. Microgliosis and glucose metabolism with Tau and amyloid beta pathology by using multi-tracer imaging in widely used tauopathy and familial Alzheimer's disease mouse models. Positron emission tomography imaging using [18F]PM-PBB3 (tau), [18F]florbetapir (amyloid-beta), [18F]SMBT-1 (monoamine oxidase-B), [18F]DPA-714 (translocator protein) and [18F]fluorodeoxyglucose was carried out in 3- and 7-month-old rTg4510 tau mice, 5 × FAD familial Alzheimer's disease mice and wild-type mice. Immunofluorescence staining was performed to validate the pathological distribution in the mouse brain after in vivo imaging. We found increased regional levels of [18F]PM-PBB3, [18F]SMBT-1, and [18F]DPA-714 and hypoglucose metabolism in the brains of 7-month-old rTg4510 mice compared to age-matched wild-type mice. Increased [18F]SMBT-1 uptake was observed in the brains of 3, 7-month-old 5 × FAD mice, with elevated regional [18F]florbetapir and [18F]DPA-714 uptakes in the brains of 7-month-old 5 × FAD mice, compared to age-matched wild-type mice. Positive correlations were shown between [18F]SMBT-1 and [18F]PM-PBB3, [18F]DPA-714 and [18F]PM-PBB3 in rTg4510 mice, and between [18F]florbetapir and [18F]DPA-714 SUVRs in 5 × FAD mice. In summary, these findings provide in vivo evidence that reactive astrocytes, microglial activation, and cerebral hypoglucose metabolism are associated with tau and amyloid pathology development in animal models of tauopathy and familial Alzheimer's disease. • Increased levels of [18F]SMBT-1 in the brains of 7-month-old rTg4510 mice compared to age-matched wild-type mice. • Higher [18F]SMBT-1 uptake in the brains of 3-, 7-month-old 5 × FAD mice compared to age-matched wild-type mice. • Positive correlations between the regional levels of [18F]SMBT-1 and [18F]DPA-714 in the brain of 5 × FAD mice. • Positive correlations between uptakes of [18F]SMBT-1 and [18F]PM-PBB3, and [18F]DPA-714 in the brain of rTg4510 mice. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: Folate receptor (FR) is a potential molecular target for radionuclide imaging since it is overexpressed in many human epithelial tumor cells. In this study, a novel folate conjugate was synthesized and labeled with 99mTc using different coligands. In vitro and in vivo evaluations of these complexes have been done to explore the effect of coligands on the stable, affinity and pharmacokinetic properties. Methods: A novel folate conjugate, HYNIC-NHHN-FA, was synthesized and characterized. This conjugate was radiolabeled with 99mTc using tricine, tricine /diphenylphosphinobenzene-3-sulfonic acid sodium (TPPMS) and tricine /trisodium triphenylphosphine-3,3′,3″-trisulfonate (TPPTS) as coligands, respectively. The complexes were purified by high-pressure liquid chromatography (HPLC). In vitro and in vivo evaluations were performed with FR-positive KB cells, normal Kunming mice and athymic nude mice bearing KB tumors. Results: Labeling with 99mTc using different coligands resulted in three complexes, 99mTc (HYNIC-NHHN-FA)(tricine), 5, 99mTc (HYNIC-NHHN-FA)(tricine/TPPMS), 6 and 99mTc (HYNIC-NHHN-FA)(tricine/TPPTS), 7. Complex 5 showed at least two isomers and was unstable after being purified by HPLC. Complexes 6 and 7 displayed high stability and similar affinity to FR in vitro. Biodistribution results in athymic nude mice bearing KB tumor showed that complex 7 had a high uptake in FR-positive tumor (9.79±1.66%ID/g at 4 h postinjection), and the results of blockade studies confirmed the specific accumulation of the radiotracer in vivo. However, complex 6 showed a low tumor uptake due to its fast excretion via the gastrointestinal tract. Conclusion: The modification of the coligands can significantly alter the pharmacokinetic properties of the corresponding 99mTc-HYNIC complexes. 99mTc (HYNIC-NHHN-FA)(tricine/TPPTS), 7 could be a promising radiotracer for FR imaging. [Copyright &y& Elsevier]

The laboratory mouse has emerged as a primary model organism for studying axon regeneration after experimental spinal cord injury, owing to its genetic amenability. Mutant mouse models are contributing significantly to our understanding of the molecular mechanisms of axon regeneration failure in the adult mammalian central nervous system (CNS), in particular regarding the role of axon-growth inhibitors. Here, we discuss recent advances in understanding axon regeneration failure that have been made using genetically modified mice, focusing on the inhibitory influences in the CNS, and we illustrate the advantages of using the mouse as a surrogate organism to study axon regeneration and spinal cord repair. [Copyright &y& Elsevier]