Neuroprotection by valproic acid in an intrastriatal rotenone model of Parkinson's disease.

Rotenone, which is used as a pesticide and insecticide, has been shown to cause systemic inhibition of mitochondrial complex I activity, with consequent degeneration of dopaminergic neurons within the substantia nigra and striatum, as observed in Parkinson's disease. A novel intrastriatal rotenone model of Parkinson's disease was used to examine the neuroprotective effects of valproic acid (VPA), which is known to upregulate neurotrophic factors and other protective proteins in the brain. Sham or lesioned rats were treated with either vehicle or VPA at a dose of 4mg/mL in drinking water. The right striatum was lesioned by infusion of rotenone at three sites (2µg/site) along its rostro-caudal axis. A forelimb asymmetry (cylinder) test indicated a significant (p<0.01) decrease in use of the contralateral forelimb in rotenone-lesioned animals, in the third week post-lesioning, which was abolished by VPA treatment. Similarly, a significant (p<0.01) and persistent increase in use of the ipsilateral forelimb in lesioned animals over the 4weeks of testing, was not seen in animals treated with VPA. Results of the asymmetry test illustrate that intrastriatal infusion of rotenone causes contralateral motor dysfunction, which is blocked by VPA. The significant increase in ipsilateral forelimb use has not been documented previously, and presumably represents a compensatory response in lesioned animals. Six weeks post-surgery, animals were sacrificed by transcardial perfusion. Subsequent immunohistochemical examination revealed a decrease in tyrosine hydroxylase immunoreactivity within the striatum and substantia nigra of rotenone-lesioned animals. VPA treatment attenuated the decrease in tyrosine hydroxylase in the striatum and abolished it in the substantia nigra. Stereological cell counting indicated a significant (p<0.05) decrease in tyrosine hydroxylase-positive dopamine neurons in the substantia nigra of rotenone-lesioned animals, which was confirmed by Nissl staining. Importantly, this loss of dopamine neurons in rotenone-lesioned animals, was blocked by chronic VPA treatment. These findings strongly support the therapeutic potential of VPA in Parkinson's disease.

Change in subfoveal choroidal thickness in central serous chorioretinopathy following spontaneous resolution and low-fluence photodynamic therapy.

PURPOSE: To assess the change in subfoveal choroidal thickness (SFCT) in central serous chorioretinopathy (CSC) following spontaneous resolution and low-fluence photodynamic therapy (PDT) using the enhanced depth imaging optical coherence tomography (EDI-OCT). METHODS: A total of 36 consecutive eyes of 36 patients were included in this retrospective study: 16 eyes with spontaneously resolved CSC and 20 eyes with PDT-treated CSC. Best-corrected visual acuity and SFCT were evaluated at each visit until complete absorption of the subretinal fluid. SFCT of 32 normal subjects were also measured, as the control group. Wilcoxon's singed-rank test was used to evaluate the effects of spontaneous resolution and PDT. To compare the SFCT of the eyes with resolved CSC with that of the normal eyes, Mann-Whitney U-test with Bonferroni correction was also employed. RESULTS: SFCT of patients was 459.16±77.50 µm at the baseline, and decreased to 419.31±54.49µm after a spontaneous resolution (P=0.015). However, SFCT was not normalized in comparison with that of the normal subjects (P<0.001). SFCT in PDT group was also reduced from 416.43±74.01 to 349.50±88.99 µm (P<0.001), with no significant difference with the normal value (P=0.087). CONCLUSIONS: SFCT in patients with CSC decreased both after spontaneous resolution and low-fluence PDT. However, only in the PDT group, after disappearance of subretinal fluid, did it decrease to that of normal subjects.

Synergistic effects of genetically engineered stem cells expressing cytosine deaminase and interferon-ß via their tumor tropism to selectively target human hepatocarcinoma cells.

Stem cells have received a great deal of attention for their clinical and therapeutic potential for treating human diseases and disorders. Recent studies have shown that it is possible to genetically engineered stem cells (GESTECs) to produce suicide enzymes that convert non-toxic prodrugs to toxic metabolites, selectively migrate toward tumor sites and reduce tumor growth. In this study, we evaluated whether these GESTECs are capable of migrating to hepatocarcinoma cells and examined the potential therapeutic efficacy of gene-directed enzyme prodrug therapy against liver cancer cells in cellular and animal models. A modified transwell migration assay was performed to determine the migratory capacity of GESTECs to Hep3B hepatocarcinoma cells. GESTECs, that is, HB1.F3.CD or HB1.F3.CD.interferon-ß (IFN-ß) cells, engineered to express a suicide gene, cytosine deaminase (CD), selectively migrated toward liver cancer cells. Treatment of Hep3B, human liver cancer cells, with the prodrug 5-fluorocytosine (5-FC) in the presence of HB1.F3.CD or HB1.F3.CD.IFN-ß cells resulted in the inhibition of Hep3B cell growth. In a xenografted mouse model injected with hepatocarcinoma, we investigated the therapeutic effect of these stem cells. For 9 weeks, the xenografted mice were treated with HB1.F3.CD or HB1.F3.CD.IFN-ß in the presence of 5-FC. A growth of tumor mass was inhibited about 40-50% in the mice treated with GESTECs and a prodrug. In addition, we further confirmed the cytotoxic effect on tumor cells by histological analysis and migratory effect of therapeutic stem cells. Taken together, GESTECs expressing a fusion gene encoding CD and IFN-ß may exert a synergistic antitumor effect on this type of tumor.

Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells.

As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.

Human amniotic fluid-derived stem cells expressing cytosine deaminase and thymidine kinase inhibits the growth of breast cancer cells in cellular and xenograft mouse models.

As human amniotic fluid-derived stem cells (hAFSCs) are capable of multiple lineage differentiation, extensive self-renewal and tumor targeting, they may be valuable for clinical anticancer therapies. In this study, we used hAFSCs as vehicles for targeted delivery of therapeutic suicide genes to breast cancer cells. hAFSCs were engineered to produce AF2.CD-TK cells in order to express two suicide genes encoding bacterial cytosine deaminase (CD) and herpes simplex virus thymidine kinase (HSV-TK) that convert non-toxic prodrugs, 5-fluorocytosine (5-FC) and mono-phosphorylate ganciclovir (GCV-MP), into cytotoxic metabolites, 5-fluorouracil (5-FU) and triphosphate ganciclovir (GCV-TP), respectively. In cell viability test in vitro, AF2.CD-TK cells inhibited the growth of MDA-MB-231 human breast cancer cells in the presence of the 5-FC or GCV prodrugs, or a combination of these two reagents. When the mixture of 5-FC and GCV was treated together, an additive cytotoxic effect was observed in the cell viability. In animal experiments using female BALB/c nude mouse xenografts, which developed by injecting MDA-MB-231 cells, treatment with AF2.CD-TK cells in the presence of 5-FC and GCV significantly reduced tumor volume and weight to the same extent seen in the mice treated with 5-FU. Histopathological and fluorescent staining assays further showed that AF2.CD-TK cells were located exactly at the site of tumor formation. Furthermore, breast tissues treated with AF2.CD-TK cells and two prodrugs maintained their normal structures (for example, the epidermis and reticular layers) while breast tissue structures in 5-FU-treated mice were almost destroyed by the potent cytotoxicity of the drug. Taken together, these results indicate that AF2.CD-TK cells can serve as excellent vehicles in a novel therapeutic cell-based gene-directed prodrug system to selectively target breast malignancies.