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Primary leptomeningeal melanomatosis is an extremely rare, diffuse, malignant melanocytic neoplasm without a localised mass. In this case, both immunocytochemistry and molecular testing were successfully completed using a CSF sample, allowing confirmation of the diagnosis and determination of the mutational status of the melanoma. [ABSTRACT FROM AUTHOR]

Background The non-essential amino acid,l-glutamine (Gln), is abundant in the human body. Gln exhibits beneficial effects on endotoxic shock through the inhibition of cytosolic phospholipase A2 (cPLA2) activity. cPLA2 has been reported to be implicated in the pathogenesis of asthma, but the effects of Gln on asthma have not yet been defined. Objective To investigate the effects of Gln on allergic bronchial inflammation and airway hyperresponsiveness (AHR), and to determine the possible action mechanisms of Gln in a murine model of asthma. Methods cPLA2 phosphorylation was assessed by immunoprecipitation and Western blotting. Smears of bronchoalveolar lavage cells were stained with Diff-Quik solution for differential cell counting. Airway levels of the proteins [T-helper type-1 (Th1) and Th2 cytokines, and mucin] were measured by ELISA. mRNA expression of cytokines was assessed by real-time RT-PCR. AHR was assessed as a change in airway resistance (RL). Histological studies were performed to assess the levels of mucin and pulmonary inflammation. Results Systemic Gln administration inhibited cPLA2 phosphorylation and its enzymatic activity in the lungs. Additionally, Gln effectively suppressed the key features of Th2-dependent asthmatic features, such as airway eosinophilia, mucus formation, and airway type 2 cytokine production, as well as late AHR. Conclusion Gln was found to be effective in the suppression of Th2-dependent phenotypes and late AHR, and this effect of Gln appeared to be at least partially attributable to its ability to suppress cLPA2 activity in the airway. Our results suggest that clinical use of Gln for patients with asthma may be beneficial. [ABSTRACT FROM AUTHOR]

This study reports the use of erythrocyte ghosts (EG) as a biocompatible nonviral delivery system for extended circulation and prolonged expression of plasmid DNA in the blood. Murine interleukin-2-expressing plasmid DNA was efficiently loaded to EG by electroporation in hypotonic condition. The presence of plasmid DNA in EG was confirmed by fluorescence-labeled plasmid DNA. At 21?min after intravenous administration into mice, the level of plasmid DNA in the blood was 92?000-fold higher following EG-mediated delivery as compared to the injection of naked form. EG-mediated gene delivery revealed higher and more prolonged mRNA expression levels of plasmid DNA in the blood until 9 days after the single intravenous injection. Moreover, plasmid DNA-loaded EG showed gene expression targeted to the blood cells. At 3 days post-dose, substantial expression levels of plasmid DNA delivered in EG were observed only in the blood and not in the other organs. Of the blood cells, the subpopulation containing granulocytes showed higher expression of plasmid DNA than mononuclear cells. These results indicate the potential of EG as a safe, prolonged and blood-targeted delivery system of therapeutic genes.Gene Therapy (2004) 11, 492-496. doi:10.1038/sj.gt.3302180 [ABSTRACT FROM AUTHOR]

Background: Hepatic ischemia-reperfusion injury (IRI) is an important determinant of the outcome of hepatic surgery, including re-section and transplantation. Previous studies have shown that nitric oxide (NO) has a protective effect against IRI. Therefore, many studies have examined methods for supplying NO. In this study, we investigated the effect of NO-releasing nanofibers on hepatic IRI in a rat model., Methods: Male Sprague-Dawley rats were divided into 4 groups: control, IRI only (n = 3); group 1, hepatic IRI and liver-wrapping with nanofiber lacking NO (n = 4); group 2, hepatic IRI and liver-wrapping with NO rapid-releasing nanofiber (n = 4); and group 3, hepatic IRI and liver-wrapping with NO slow-releasing nanofiber (n = 5)., Results: The levels of aspartate aminotransferase and alanine aminotransferase were not significantly different between groups. On the basis of Western blots, Bax/β-actin levels were significantly lower in group 2 than in group 3 (P < .01). Cleaved Caspase-3/β-actin levels were significantly lower in group 2 than in the control, group 1, and group 3 (P < .05, .01, and .01, respectively). However, there were no significant differences in Bcl-2/β-actin between groups., Conclusions: The liver-wrapping NO rapid-releasing nanofiber downregulated cleaved Caspase-3 and Bax expression. It has a protective effect by reducing apoptosis in hepatic IRI in rats., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Osteoprotegerin (OPG) is secreted by stromal and osteoblastic lineage cells and inhibits osteoclastogenesis by preventing the interaction of receptor activator of nuclear factor-κB ligand (RANKL) with receptor activator of nuclear factor-κB (RANK). In this study, the expression of OPG in osteoclasts themselves and its biological functions during osteoclastogenesis were investigated for the first time. OPG expression in vivo in the developing rat maxilla was examined by immunofluorescence analysis. OPG expression in osteoclasts during in vitro osteoclastogenesis was determined by reverse-transcription polymerase chain-reaction (RT-PCR), Western blot, and immunofluorescence staining. We determined the function of OPG produced by osteoclasts during osteoclastogenesis by silencing the OPG gene. The effects of OPG on bone-resorbing activity and apoptosis of mature osteoclasts were examined by the assay of resorptive pit formation on calcium-phosphate-coated plate and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining, respectively. In the immunofluorescence findings, strong immunoreactivities were unexpectedly seen in multinucleated tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts around the growing and erupting tooth germs in the rat alveolar bone. In vitro, OPG expression was significantly increased during the differentiation of osteoclasts from mouse bone-marrow-derived cells treated with a combination of macrophage colony-stimulating factor (M-CSF) and RANKL. Interestingly, it was found that OPG small interfering (si)RNA treatment during osteoclastogenesis enhanced the sizes of osteoclasts, but attenuated their bone-resorbing activity. Also, the increased chromosomal DNA fragmentation and caspase-3 activity in the late phase of osteoclastogenesis were found to be decreased by treatment with OPG siRNA. Furthermore, effects of OPG siRNA treatment on osteoclastogenesis and bone-resorbing activity were recovered by the treatment of exogenous OPG. These results suggest that OPG, expressed by the osteoclasts themselves, may play an auto-regulatory role in the late phase of osteoclastogenesis through the induction of apoptosis., (© International & American Associations for Dental Research.)

This study was performed to determine effects of alendronate on the tibial proximal epiphyseal cartilage undergoing endochondral ossification and the expression of vascular endothelial growth factor (VEGF) from the cartilage. Alendronate was injected subcutaneously every other day in postnatal Day 1 Sprague Dawley rats. The rats were sacrificed 3, 5, 7, and 10 days after the first injection. The effect of alendronate treatment for 10 days was demonstrated from the morphological change that the area of the secondary ossification center in the epiphysis was significantly smaller in the alendronate group than that in the control group (P < 0.05). Strong immunoreactivity to VEGF was observed in the hypertrophied chondrocytes and some proliferating chondrocytes in the epiphyseal cartilage at postnatal Day 5 and was decreased after the alendronate treatment for 5 days. Immunoreactivity was observed in not only hypertrophied cells but also the peripheral cartilaginous matrix adjacent to the vascular canals invading into the central portion of the cartilage at postnatal Day 7. This reactivity was also reduced considerably by the alendronate treatment for 7 days. The level of VEGF expression was reduced by the alendronate treatment at both the transcription and translation levels. However, the transcriptional level of the flt-1 and flk-1 receptors was relatively unaltered by the treatment. These results suggest that VEGF expression is required for vascular invasion into the developing cartilage and alendronate can affect its resorption by downregulating VEGF expression.

Objective: The chorda-lingual (CL) nerve carries parasympathetic fibers to the hilum of the sublingual and submandibular glands (SMGs) and evokes the secretion of saliva. The effect of cutting the CL nerve on the biological processes in SMGs was investigated by examining the gene-expression profiles in the SMGs after a surgical parasympathectomy., Methods: At day 3 after the CL nerve cut, the changes in the SMGs at both the experimental cut and contralateral control sides were analysed by microarray and light microscopy. The expression levels of 6 selected genes were confirmed by real-time PCR, Western blot and immunofluorescence staining., Results: The wet weight of the parasympathectomised SMGs decreased significantly compared to that of the contralateral side (p<0.05). Histological analyses after the parasympathectomy showed a widened interacinar space as well as some atropic changes to the acini of the SMGs in the cut side. Microarray analysis revealed that twofold differential expression in mRNA expression in the parasympathectomized SMGs were detected in 88 genes (0.004%): 41 genes were overexpressed, 11 were underexpressed and 36 were unknown. Changes of the expression of 6 selected genes detected by Western blot and/or real-time PCR were consistent with the microarray data., Conclusion: The important genes involved in biological processes for salivation were identified through a large-scale gene expression analysis., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Objective: This study examined the expression pattern of the Deleted-in-colorectal-carcinoma (DCC) gene in developing rat tooth germs., Methods: Rat pups at 4, 7 and 10 d postpartum were used in this study. Reverse transcription-polymerase chain reaction (RT-PCR) and immunofluorescent localization were used to determine the level of DCC expression during tooth development., Results: There was more than 2-fold higher level of DCC mRNA in the rat 2nd maxillary molar tooth germs on 10 d postpartum, which was the root stage, than in the rat 3rd maxillary molar tooth germ, which was at the cap/early bell development stage. In addition, the levels of DCC mRNA in the 2nd maxillary molar germs at 4, 7 and 10 d postpartum increased gradually according to tooth development. Interestingly, immunoreactivity against DCC was specifically detected in the differentiating ameloblasts. DCC was observed in the lateral and apical sides of the newly differentiating and secretory stage ameloblasts. Afterwards, DCC was localized only in the apical side of the maturation stage ameloblasts, not in the lateral side., Conclusion: DCC is expressed in the differentiating ameloblasts, which suggests that this molecule plays a crucial role in amelogenesis.

In this study, we have investigated the mechanisms underlying organ susceptibility to candida infection. Infection of BALB/c mice with Candida albicans led to both an early (1-8 h) and late (24-48 h) activation of NF-kappaB in the organs resistant to C. albicans, including the lung and spleen. In susceptible organs such as the kidneys, early activation of NF-kappaB was not observed. The kinetics of TNF-alpha mRNA expression paralleled those of NF-kappaB activation in all organs examined. Blocking the effects of endogenous platelet-activating factor (PAF) by pretreatment with the PAF antagonist BN50739 or antioxidants significantly reduced the early activity of NF-kappaB and TNF-alpha mRNA expression, and increased the recovery of C. albicans in the lung and spleen. Importantly, administration of PAF 5 min prior to the infection resulted in the appearance of early activities of NF-kappaB and TNF-alpha mRNA expression, followed by a nearly complete clearance of the organisms in the kidneys. Pretreatment with anti-TNF-alpha Ab resulted in an enhanced susceptibility to C. albicans, and the PAF-mediated resistance was abrogated by anti-TNF-alpha in all organs examined. These data indicated that endogenously produced PAF in response to C. albicans is a key molecule involved in the early activation of NF-kappaB, which, in turn, renders the organ resistant to the fungus by promoting the production of anti-candidal proinflammatory cytokines such as TNF-alpha. Susceptible organs, including the kidneys, lack the capacity to generate a sufficient PAF-induced early NF-kappaB response.

We report a case of Sertoli cell adenoma in complete androgen insensitivity syndrome (CAIS) in a 22-year-old woman. Polymerase chain reaction-single strand conformation polymorphism and DNA sequencing revealed a single nucleotide substitution on exon 7 of the human androgen receptor (hAR) gene, resulting in a change of CGA (arginine) to CAA (glutamine) in codon 831.

We report three autopsy cases of congenital cytomegalovirus (CMV) infection in fetuses with a review of literature. The clinical manifestations in these cases of congenital CMV infection include intrauterine fetal death, hydrops fetalis, and CMV pneumonia associated with cardiovascular defect. The pathological characteristics were as follows: 1) the kidney was the most frequently involved organ, followed by lung and liver, 2) CMV inclusions were found predominantly in epithelial cells and to a lesser degree in endothelial cells, 3) intrahepatic bile duct epithelial cells were frequently involved, and 4) inflammatory reaction around CMV inclusions was not prominent in the early stage of pregnancy. Diagnostic confirmation was obtained by in situ hybridization (ISH) using a biotinylated CMV-DNA probe, which demonstrated intranuclear inclusions and sometimes recognized cells that did not show intranuclear inclusion.

Lipopolysaccharide (LPS) is a known inducer of numerous pro-inflammatory events including the production of platelet-activating factor (PAF). PAF released in response to LPS is a major contributor to the pathological events associated with endotoxemia. The present study demonstrates that dexmethasone (DEX) inhibited the LPS-induced early plasma PAF raise in a dose- and time-dependent manner. In addition, DEX prevented the subsequent PAF-mediated pathological phenomena such as anaphylactic shock-like symptoms, symptoms of disseminated intravascular coagulation and hemorrhage in renal medullae. DEX or the PAF antagonist BN 50739 significantly inhibited LPS-induced NF-kappaB activation. The inhibition of NF-kappaB activation by DEX was overcome by the injection of exogenous PAF. Administration of PAF or LPS resulted in a rapid loss of IkappaBalpha protein. The LPS-induced degradation of IkappaBalpha was prevented by pretreatment with BN 50739, suggesting that PAF is a critical intermediate in the LPS-triggered degradation of IkappaBalpha protein. DEX prevented the LPS-induced IkappaBalpha degradation, which was also reversed by exogenous PAF. Administration of DEX or BN 50739 caused an increase in cytoplasmic IkappaBalpha level. Our results indicate that DEX inhibits IkappaBalpha degradation and subsequent NF-kappaB activation through blocking the initial release of PAF.

Tumor necrosis factor (TNF)-alpha and platelet-activating factor (PAF) are important mediators of inflammatory reactions, and their release is controlled by a positive feedback network. However, the regulatory mechanisms underlying the interaction of these two molecules are unknown. Within 10 min of the injection of lipopolysaccharide (LPS) into C57BL/6 mice, effects inducible by PAF such as anaphylactic shock-like symptoms, disseminated intravascular coagulation, and hemorrhage in renal medullae were observed, and all these pathological changes were prevented by the PAF antagonist, BN 50739. The plasma level of PAF after LPS injection reached a peak at 5 min. TNF-alpha gene expression was evident 20 min after LPS injection and was maximal at 40 min, and the level of serum TNF-alpha reached a peak at 1 h. Pretreatment with BN 50739 inhibited LPS-induced TNF-alpha gene expression and protein synthesis in a dose-dependent manner. Injection of PAF or treatment of the macrophage cell line, J774A.1, with PAF activated the transcription factor, nuclear factor (NF)-kappa B, which is essential for inducible TNF-alpha transcription. The activation of NF-kappa B by PAF preceded the LPS-mediated TNF-alpha gene expression. Pretreatment with BN 50739 inhibited LPS-induced mobilization of NF-kappa B in a dose-dependent manner in vivo as well as in vitro. These data suggest that PAF, which is released immediately or shortly after LPS injection, induces the expression of TNF-alpha through the activation of NF-kappa B.

The effect of platelet-activating factor (PAF) on experimental pulmonary metastasis by the B16F10 murine melanoma and the possible involvement of PAF in the activities of tumor necrosis factor alpha (TNF-alpha) and interleukin 1alpha (IL-1alpha) in tumor metastasis were investigated. i.p. injection of PAF enhanced the lung colonization in a dose- and time-dependent manner. PAF enhanced lung colonization when it was administered after, but not before, B16F10 inoculation. Multiple injections of PAF were more effective than a single injection. Neutralization of endogenous PAF with PAF antagonist BN50739 decreased lung colonization, suggesting that endogenous PAF plays an important role in pulmonary metastases. A single i.p. injection of TNF-alpha or IL-1alpha caused a marked enhancement in lung colonization. TNF-alpha- and IL-1alpha-mediated enhancement in lung colonies was significantly inhibited by BN50739. These results demonstrate that PAF has a metastasis-enhancing effect and is a mediator of the metastatic activities of TNF-alpha and IL-1alpha.

To improve drug permeation through skin, extracts of crude drugs were evaluated using in vitro and in vivo penetration techniques with rabbit skin as a model membrane. The acetone extract of Amomum cardamomum (ae-AC) and Elettaria cardamomum (ae-EC) had the best effect in enhancing the penetration of Indomethacin (IDM). The flux of IDM for the formulation containing ae-AC from pH 7.4 buffer-alcohol donor solution was 3.6 times higher than that from the 50% alcohol donor solution. In order to confirm enhancer/skin interaction, a microscopic study was carried out in order to determine the presence of physical evidence in the skin caused by enhancers. A pharmacokinetic model of percutaneous absorption of IDM after topical administration was employed on the assumption of a zero-order penetration rate through the stratum corneum followed by absorption into the blood by a first-order process. Good correlation was observed between the experimental data point and simulated plasma concentration. From the results of the study, the in vitro permeability coefficient correlates with the in vivo penetration rate through the stratum corneum.

Two commercial slow-release potassium chloride tablets, Slow-K and Addi-K have the characteristics of slow-release in the different dissolution conditions. The two tablets had similar dissolution profiles in simulated intestinal fluid (pH 6.8), simulated gastric fluid (pH 1.2) and water; which indicates that, the dissolution rates of the two commercial potassium chloride tablets are independent of the pH values of the dissolution medium. The release patterns of potassium chloride fitted the First order model and Higuchi matrix model. These were distinguished by differential rate treatment, which showed that the release followed the First order pattern. It was found that the dissolution rate of Addi-K were more rapid than Slow-K in different dissolution conditions.