24(S)-Hydroxycholesterol protects the ex vivo rat retina from injury by elevated hydrostatic pressure

In the central nervous system, 24(S)-hydroxycholesterol (24(S)-HC) is an oxysterol synthesized from cholesterol by cholesterol 24-hydroxylase (CYP46A1) encoded by the cyp46a1 gene. In the present study using a rat ex vivo glaucoma model, we found that retinal 24(S)-HC synthesis is facilitated by pressure elevation. Moreover, we found that 24(S)-HC is neuroprotective against pressure mediated retinal degeneration. Quantitative real-time RT-PCR, ELISA, and immunohistochemistry revealed that elevated pressure facilitated the expression of cyp46a1 and CYP46A1. Immunohistochemically, the enhanced expression of CYP46A1 was mainly observed in retinal ganglion cells (RGC). LC-MS/MS revealed that 24(S)-HC levels increased in a pressure-dependent manner. Axonal injury and apoptotic RGC death induced by 75 mmHg high pressure was ameliorated by exogenously administered 1 μM 24(S)-HC. In contrast, voriconazole, a CYP46A1 inhibitor, was severely toxic even at normobaric pressure. Under normobaric conditions, 30 μM 24(S)-HC was required to prevent the voriconazole-mediated retinal damage. Taken together, our findings indicate that 24(S)-HC is facilitated by elevated pressure and plays a neuroprotective role under glaucomatous conditions, while voriconazole, an antifungal drug, is retinotoxic. 24(S)-HC and related compounds may serve as potential therapeutic targets for protecting glaucomatous eyes from pressure-induced injuries

24(S)-Hydroxycholesterol protects the ex vivo rat retina from injury by elevated hydrostatic pressure

In the central nervous system, 24(S)-hydroxycholesterol (24(S)-HC) is an oxysterol synthesized from cholesterol by cholesterol 24-hydroxylase (CYP46A1) encoded by the cyp46a1 gene. In the present study using a rat ex vivo glaucoma model, we found that retinal 24(S)-HC synthesis is facilitated by pressure elevation. Moreover, we found that 24(S)-HC is neuroprotective against pressure mediated retinal degeneration. Quantitative real-time RT-PCR, ELISA, and immunohistochemistry revealed that elevated pressure facilitated the expression of cyp46a1 and CYP46A1. Immunohistochemically, the enhanced expression of CYP46A1 was mainly observed in retinal ganglion cells (RGC). LC-MS/MS revealed that 24(S)-HC levels increased in a pressure-dependent manner. Axonal injury and apoptotic RGC death induced by 75 mmHg high pressure was ameliorated by exogenously administered 1 mu M 24(S)-HC. In contrast, voriconazole, a CYP46A1 inhibitor, was severely toxic even at normobaric pressure. Under normobaric conditions, 30 mu M 24(S)-HC was required to prevent the voriconazole-mediated retinal damage. Taken together, our findings indicate that 24(S)-HC is facilitated by elevated pressure and plays a neuroprotective role under glaucomatous conditions, while voriconazole, an antifungal drug, is retinotoxic. 24(S)-HC and related compounds may serve as potential therapeutic targets for protecting glaucomatous eyes from pressure-induced injuries

Additive neuroprotective effects of 24(S)-hydroxycholesterol and allopregnanolone in an ex vivo rat glaucoma model

In a rat ex vivo acute glaucoma model, high pressure (75 mmHg) causes swelling of ganglion cell axons and elevates levels of the endogenous steroids 24(S)-hydroxycholesterol (24SH) and allopregnanolone (AlloP). Furthermore, 24SH (0.1 mu M) alone elevates AlloP levels via NMDA receptors. With this model, we now investigate possible interactions between 24SH and AlloP. We found that inhibition of AlloP synthesis with dutasteride under high pressure results in severe excitotoxicity in addition to axonal swelling. The excitotoxicity is prevented by exogenous AlloP but not 24SH, indicating that endogenous AlloP is crucial for protection. However, inhibition of 24SH synthesis with voriconazole induces severe excitotoxicity under normal pressure. Paradoxically, the excitotoxicity by voriconazole is better prevented by AlloP than 24SH. These findings suggest that inhibition of 24SH synthesis becomes excitotoxic in the absence of AlloP. We also observed that co-administration of sub-micromolar 24SH (0.1 mu M) and AlloP (0.1 mu M), concentrations that are only partially effective when administered alone, prevents axonal swelling under high pressure. This apparent enhanced protection indicates strong interaction between the two neurosteroids to preserve neuronal integrity, with 24SH contributing to AlloP synthesis via NMDA receptors and with AlloP playing an essential role in neuroprotection via GABA(A) receptors

The enantiomer of allopregnanolone prevents pressure-mediated retinal degeneration via autophagy

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Effect of polyglycolic acid sheets with fibrin glue (MCFP technique) on the healing of wounds after partial resection of the border of the tongue in rabbits: a preliminary study

The aim of this study was to examine the effectiveness of covering wounds to the tongue with a polyglycolic acid (PGA) sheet and fibrin glue. Eighteen mature male Japanese white rabbits had a unilateral glossectomy involving an area 10 mm × 10 mm × 2 mm. After glossectomy the tongues were covered with PGA sheets 8 mm × 8 mm in size and fibrin glue (mucosal defect covered with fibrin glue and polyglycolic acid sheet = MCFP) 1 week after the operation (n = 3), after 2 weeks (n = 3), and after 4 weeks (n = 3). In control groups, after 1, 2, and 4 weeks (n = 3 in each group), the partially resected tongues were closed with absorbable sutures (polyglactin 910). One week (experimental and control groups 1), 2 weeks (experimental and control groups 2) and 4 weeks (experimental and control groups 3) after operation the tongues were harvested and stained for microscopic examination. Histological examination showed that the covered wound surface had not epithelialised and the basal layer had yet to form in experimental group 1, but had formed in experimental group 2. However, in control group 1, epithelialisation of the sutured wound had begun. Immunohistochemical examination showed that, in experimental group 1, the non-uniform epithelial layer of the covered wound surface expressed cytokeratin AE1/AE3, and the epithelial and connective tissue layers stained strongly for FGF-2. Similar results were obtained in experimental group 2, whereas in experimental group 3, FGF-2 was expressed only in the connective tissue layer, and epithelialisation was complete. However, in control group 1, AE1/AE3 was expressed in the epithelial layer, and FGF was expressed in the connective tissue layer beneath the basal layer. In control groups 2 and 3, AE1/AE3 and FGF-2 were expressed in patterns similar to those in experimental groups 2 and 3. We suggest that this method is useful and the operation is simple. However, further testing of the method is needed and it should be widely used clinically before it is recommended

Comprehensive Prospective Analysis of the Factors Contributing to Aspiration Pneumonia Following Endoscopic Submucosal Dissection in Patients with Early Gastric Neoplasms

Endoscopic submucosal dissection (ESD) has become the first-line treatment for early gastric neoplasms; however, a subset of patients treated by this method develop aspiration pneumonia. We conducted a comprehensive prospective analysis of the factors contributing to post-ESD aspiration pneumonia in early gastric neoplasms in this study, with special focus on whether pre-treatment oral care can prevent aspiration pneumonia. Sixty-one patients who underwent ESD for gastric neoplasms were randomly assigned to the oral care or control groups. ESD was performed under deep sedation. Of 60 patients whose data were available for analysis, 5 (8.3%) experienced pneumonia confirmed either by chest radiography or computed tomography. Although no difference in the rate of pneumonia was found between the control and oral care groups, the post-oral care bacteria count was significantly higher in the saliva of patients who developed pneumonia compared to those without pneumonia. In addition, the presence of vascular brain diseases and the dose of meperidine were also significantly associated with the occurrence of pneumonia. These results suggest that the number of oral bacteria as well as pre-existing vascular brain diseases and high-dose narcotics can affect the incidence of post-ESD pneumonia

Cross-Linguistic Influence on L2 before and after extreme reduction in input:The case of Japanese returnee children

This study investigates the choice of genitive forms (the woman’s book vs. the book of the woman) in the English of Japanese-English bilingual returnees (i.e., children who returned from a second language dominant environment to their first language environment). The specific aim was to examine whether change in language dominance/exposure influences choice of genitive form in the bilingual children; the more general question was the extent to which observed behavior can be explained by cross linguistic influence (CLI). First, we compared the choice of genitive form between monolingual English speakers and bilinguals who had recently returned to Japan from an English speaking environment. Second, we tracked changes in genitive preference within bilingual children, comparing their performances upon return to Japan to those of one year later. Results show that CLI alone is insufficient to explain the difference in genitive evaluation between bilinguals and monolinguals, as well as the intra-group bilingual variation over time. We suggest that both CLI and general processing considerations couple together to influence the changes in genitive preference

Silencing of the p53R2 gene by RNA interference inhibits growth and enhances 5-fluorouracil sensitivity of oral cancer cells

The p53R2 gene encodes the ribonucleotide reductase (RR) small subunit 2 homologue, and is induced by several stress signals activating p53, such as DNA-damaging agents. The p53R2 gene product causes an increase in the deoxynucleotide triphosphate (dNTP) pool in the nucleus, which facilitates DNA repair and synthesis. We hypothesized that p53R2 would be a good molecular target for cancer gene therapy. In this study, three human oral cancer cell lines (SAS, HSC-4 and Ca9-22), a human breast cancer cell line MCF-7, and a normal human fibroblast cell line NHDF were tested. We silenced the expression of p53R2 with the highly specific post-transcriptional suppression of RNA interference (RNAi). We investigated p53R2 expression with the reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. The sensitivity to anticancer agents was evaluated by a 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The expression of p53R2 showed no association with the mutational status of p53. The cancer cell lines with higher p53R2 expression were more resistant to 5-FU. RNAi-mediated p53R2 reduction selectivity inhibited growth and enhanced chemosensitivity in cancer cell lines but not in normal fibroblasts. These results suggest that basal transcription of p53R2 could be associated with the sensitivity to anticancer agents. Moreover, we assessed the possibility that p53R2 would be a good molecular target, and report that RNAi targeting of p53R2 could be useful for oral cancer gene therapy

Isolation and characterization of cancer stem-like side population cells in human oral cancer cells.

Recent studies suggest that cancer stem cells may be responsible for tumorigenesis and contribute to some individuals\u27 resistance to cancer therapy. Some studies demonstrate that side population (SP) cells isolated from diverse cancer cell lines harbor stem cell-like properties; however, there are few reports examining the role of SP cells in human oral cancer. To determine whether human oral cancer cell lines contain a SP cell fraction, we first isolated SP cells by fluorescence activated cell sorting, followed by culturing in serum-free medium (SFM) using the SCC25 tongue cancer cell line, so that SP cells were able to be propagated to maintain the CSC property. Differential expression profile of stem cell markers (ABCG2, Oct-4 and EpCAM) was examined by RT-PCR in either SP cells or non-SP cells. Growth inhibition by 5-FU was determined by the MTT assay. Clonogenic ability was evaluated by colony formation assay. SCC25 cells contained 0.23% SP cells. The fraction of SP cells was available to grow in SFM cultures. SP cells showed higher mRNA expression of stem cell markers (ABCG2, Oct-4 and EpCAM) as compared with non-SP cells. Moreover, SP cells demonstrated more drug resistance to 5-FU, as compared with non-SP cells. The clone formation efficiency of SP cells was significantly higher than non-SP cells at an equal cell number (P<0.01). We isolated cancer stem-like SP cells from an oral cancer cell line. SP cells possessed the characteristics of cancer stem cells, chemoresistance, and high proliferation ability. Further characterization of cancer stem-like SP cells may provide new insights for novel therapeutic targets