Chung-Hsing Chang

EXPRESSION OF BCL-2, P53 AND KI-67 IN ARSENICAL SKIN CANCERS

To investigate the regulation of apoptosis and proliferation in arsenic‐induced skin cancers, we examined the expression of bcl‐2. p53, and Ki‐67 using immunohistochemical staining. Thirty patients with Bowens disease (BD), ten with basal cell carcinoma (BCC), eight with squamous cell carcinoma (SCC) and eleven of perilesional normal skin (PLN) of the non‐sun exposure sites from endemic area were examined. The results showed that: 1) bcl‐2 was expressed in all of the BCC homogeneously, in none of the SCC, and in 12/30 of the BD focally or homogeneously; 2) p53 was expressed in all of the arsenical skin cancers with a labelling index of 75±14% of BD, 50±17% of BCC. 61±15% of SCC, and also in all of the perilesional normal skin with a labelling index of 55±24%; 3) Ki‐67 was expressed in all of the skin cancers with labelling index of 58±17% of BD. 12±7% of BCC, 47±21% of SCC, and in 9/11 of PLN with a labelling index of 41±24%. Expression of bcl‐2 in BCC or BD is related to the phenotype of germinative basal cell. The constant expression of bcl‐2 i early dysplastic cells of BD and the earliest expression of P53 in the basal cells of perilesional normal skin indicate that the initial step of arsenic‐induced carcinogenesis is from the basal germinative cells. There is no mutual relationship between bcl‐2, p53 or Ki‐67 expression in any type of the arsenical skin cancers, but there is a positive correlation between p53 and Ki‐67 expression identified in perilesional normal skin. BD had the highest labelling index of p53 and Ki‐67.

Optical characterization of visible multiquantum-well semiconductor lasers by collection/excitation modes of scanning near-field optical microscopy

Both collection and excitation modes of scanning near-field optical microscopy were used to study a strained AlGaInP/Ga0.4In0.6P low power visible multiquantum-well laser diode. Collection and excitation modes provide the near-field optical propagating intensity distribution and local photoconductivity information, respectively, at the facet of laser diode. Results show highly localized spatial correlation of the laser diode structure and its optical performance at the facet. Defect level in the energy range of 60–380 meV below the conduction band in the highly n-doped (Al0.7Ga0.3)0.5In0.5P cladding layer was found by the excitation mode using the wavelengths of 543.5 and 632.8 nm. Spatially resolved near-field optical spectra of both stimulated and spontaneous emissions were acquired as well. Longitudinal modes of the stimulated emission of laser diode were observed locally.

Atomic-scale determination of band offsets at the Gd2O3/GaAs (100) hetero-interface using scanning tunneling spectroscopy

Direct measurements of band profile and band offsets across the Gd2O3/GaAs(100) hetero-interface have been performed using cross-sectional scanning tunneling microscopy and spectroscopy. The spatial variation of the local density of states with atomic precision revealed the interfacial band alignment in this model high-κ/III-V system. In conjunction with the theoretical modeling, the band offsets for both conduction and valence states are identified, revealing critical information about the electrostatic potential landscape of the GaAs semiconductor transistor with a Gd2O3 gate dielectric.

Neuroprotective Effects of Omega-3 Polyunsaturated Fatty Acids in a Rat Model of Anterior Ischemic Optic Neuropathy

Purpose The purpose of this study was to investigate the therapeutic effect of omega-3 polyunsaturated fatty acid (ω-3 PUFA) administration in a rat model of anterior ischemic optic neuropathy (rAION). Methods The level of blood arachidonic acid/eicosapentaenoic acid (AA/EPA) was measured to determine the suggested dosage. The rAION-induced rats were administered fish oil (1 g/day EPA) or phosphate-buffered saline (PBS) by daily gavage for 10 consecutive days to evaluate the neuroprotective effects. Results Blood fatty acid analysis showed that the AA/EPA ratio was reduced from 17.6 to ≤1.5 after 10 days of fish oil treatment. The retinal ganglion cell (RGC) densities and the P1-N2 amplitude of flash visual-evoked potentials (FVEP) were significantly higher in the ω-3 PUFA-treated group, compared with the PBS-treated group (P < 0.05). The number of apoptotic cells in the RGC layer of the ω-3 PUFA-treated rats was significantly decreased (P < 0.05) compared with that of the PBS-treated rats. Treatment with ω-3 PUFAs reduced the macrophage recruitment at the optic nerve (ON) by 3.17-fold in the rAION model. The M2 macrophage markers, which decrease inflammation, were induced in the ω-3 PUFA-treated group in contrast to the PBS-treated group. In addition, the mRNA levels of tumor necrosis factor-alpha, interleukin-1 beta, and inducible nitric oxide synthase were significantly reduced in the ω-3 PUFA-treated group. Conclusions The administration of ω-3 PUFAs has neuroprotective effects in rAION, possibly through dual actions of the antiapoptosis of RGCs and anti-inflammation via decreasing inflammatory cell infiltration, as well as the regulation of macrophage polarization to decrease the cytokine-induced injury of the ON.

Efficacy of Intravitreal Injections of Antivascular Endothelial Growth Factor Agents in a Rat Model of Anterior Ischemic Optic Neuropathy.

PURPOSE We investigated the efficacy of intravitreal injections of anti-VEGF agents in a rat model of anterior ischemic optic neuropathy. METHODS We applied laser-induced photoactivation on the optic nerve head after intravenously administered rose bengal (RB). The rats immediately received an intravitreal injection of either ranibizumab or PBS. The density of retinal ganglion cells (RGCs) was calculated using retrograde FluoroGold labeling. Visual function was assessed by flash visual-evoked potentials (FVEP). We investigated TUNEL assays of the retinal sections and ED1 staining of the optic nerve. RESULTS After treatment, the RGC densities in the anti-VEGF-treated rats were not statistically significant different from those of the PBS-treated rats (57.0% vs. 40.0% in the central retinas; 39.8% vs. 33.6% in midperipheral retinas, both P > 0.05). Measurements of FVEP showed no statistically significant differences in preserved latency or amplitude of the P1 wave between anti-VEGF and PBS groups (latency 131 ± 15 ms versus 142 ± 14 ms, P = 0.157; amplitude 34 ± 12 μv versus 41 ± 13 μv, P = 0.423). Assays of TUNEL showed that there was no statistical difference in the number of apoptotic cells in the RGC layers between anti-VEGF and PBS groups (7.0 ± 0.8 cells/high-power field [HPF] versus 7.8 ± 1.3 cells/HPF; P = 0.275). In the optic nerves, we did not observe statistically significant differences in ED1-positive cells/HPF between anti-VEGF and PBS groups (P = 0.675). CONCLUSIONS Intravitreal injections of anti-VEGF did not have a protective effect in the rat model of anterior ischemic optic neuropathy.

Protective effects of systemic treatment with methylprednisolone in a rodent model of non-arteritic anterior ischemic optic neuropathy (rAION).

This study investigated the protective effects of the administration of steroids on optic nerves (ON) and retinal ganglion cells (RGCs) in a rodent model of non-arteritic anterior ischemic optic neuropathy (rAION). We induced rAION using rose bengal and argon laser irradiation in a photodynamic procedure on the optic discs of rats. The treated groups received methylprednisolone (MP) via peritoneal injection for 2 weeks. The control group received intraperitoneal injections of phosphate-buffered saline (PBS) post-rAION. At the 4th week post-infarct, MP treatments significantly rescued the RGCs (mm(2)) in the central retinas (1920 ± 210, p < 0.001) and mid-peripheral retinas (950 ± 240, respectively, p = 0.018) compared with those of the PBS-treated rats (central: 900 ± 210 and mid-peripheral: 440 ± 180). Functional assessment with flash visual-evoked potentials demonstrated that P1 latency (ms) was shortened in the MP group compared to the PBS group (108 ± 14 and 147 ± 9, respectively, p < 0.001). In addition, the P1 amplitude (uV) was enhanced in the MP group compared to the PBS group (55 ± 12 and 41 ± 13, respectively, p < 0.05). TUNEL assays showed a decrease in the number of apoptotic cells in the RGC layers of MP-treated retinas compared to the PBS-treated group (p < 0.05). ED1 positive cells (/HPF) were significantly decreased in the ONs of the MP group compared to the PBS group (p < 0.001). In conclusion, systemic administration of MP had neuroprotective effects on RGC survival and ON function in the rAION animal model.

Factors influencing the retrograde labeling of retinal ganglion cells with fluorogold in an animal optic nerve crush model.

Purpose: To investigate whether different crush durations or a different fluorogold (FG) injection timing can affect the efficiency of FG retrograde labeling of retinal ganglion cells (RGCs) in the optic nerve (ON) crush model. Methods: We performed the ON crush in rats with a clip at different durations or a jewel forceps to compare the effects of different crush methods with FG staining. RGC density was compared between the FG injection 1 week before the sacrifice of the animals (group A) and the injection before the crush experiment (group B). Double staining with CD11b and FG in the retinal sections was conducted to investigate the relationship between the overcounting of RGCs and microglia. Results: The FG-stained particles were significantly decreased at the distal part of the crush site compared to the proximal site of the ON with a crush duration of over 30 s or when crushed with the jewel forceps. Two weeks after ON crush, the RGC count was higher both in the central and mid-peripheral retinas in group B. The percentage of CD11b-stained cells among the FG-stained cells in the RGC layer of retinas in group B was higher than that of group A (34% in group B vs. 4% in group A, p = 0.0001). Overcounting of RGC density in group B was due to additional microglia with FG engulfing. Conclusions: Our results suggest that each laboratory should test its setting conditions to avoid factors influencing the RGC density measurement before conducting ON crush experiments.

Efficacy of Intravitreal Injections of Triamcinolone Acetonide in a Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy.

PURPOSE To investigate effects of intravitreal injections of triamcinolone acetonide (IVI-TA) at different times in a rodent model of nonarteritic anterior ischemic optic neuropathy (rAION). METHODS After inducing ischemic optic neuropathy, the rats received either IVI-TA (0.32 mg/2 μL) at 1 day (1d-TA), 1 week (7d-TA), 2 weeks (14d-TA), or PBS. The density of retinal ganglion cells (RGCs) was calculated using retrograde Fluorogold labeling. Electrophysiological visual function was assessed by flash visual evoked potentials (FVEPs). Apoptosis assays of the retinal sections and immunohistochemistry of ED1 staining of the optic nerves were performed. RESULTS Four weeks postinfarct, the 1d- and 7d-TA groups had significantly rescued RGCs in the central (2160 ± 250 mm(2), P = 0.004; 2050 ± 660, P = 0.008, respectively) and midperipheral retinas (1240 ± 130; 1250 ± 220, respectively, both P = 0.004) compared with those of the PBS-treated rats. Flash visual evoked potentials demonstrated improvements in P1 amplitude (μV) in the 1d- and 7d-TA groups (both P < 0.05). Assays of TUNEL showed a decrease in the number of apoptotic cells in the RGC layers of 1d- and 7d-TA-treated retinas compared with the PBS-treated group (both P = 0.004). Cells ED1-positive were significantly decreased in the optic nerve (ON) of the 1d- and 7d-TA groups compared with the PBS group (P = 0.004 and 0.02, respectively). CONCLUSIONS Within 1 week postinfarct of rAION, IVI-TA had neuroprotective effects on RGC survival with an increase in the electrophysiological amplitude of VEPs and a decrease in microglial infiltration in the ONs.

Early applications of granulocyte colony-stimulating factor (G-CSF) can stabilize the blood–optic-nerve barrier and ameliorate inflammation in a rat model of anterior ischemic optic neuropathy (rAION)

ABSTRACT Granulocyte colony-stimulating factor (G-CSF) was reported to have a neuroprotective effect in a rat model of anterior ischemic optic neuropathy (rAION model). However, the therapeutic window and anti-inflammatory effects of G-CSF in a rAION model have yet to be elucidated. Thus, this study aimed to determine the therapeutic window of G-CSF and investigate the mechanisms of G-CSF via regulation of optic nerve (ON) inflammation in a rAION model. Rats were treated with G-CSF on day 0, 1, 2 or 7 post-rAION induction for 5 consecutive days, and a control group were treated with phosphate-buffered saline (PBS). Visual function was assessed by flash visual evoked potentials at 4 weeks post-rAION induction. The survival rate and apoptosis of retinal ganglion cells were determined by FluoroGold labeling and TUNEL assay, respectively. ON inflammation was evaluated by staining of ED1 and Iba1, and ON vascular permeability was determined by Evans Blue extravasation. The type of macrophage polarization was evaluated using quantitative real-time PCR (qRT-PCR). The protein levels of TNF-α and IL-1β were analyzed by western blotting. A therapeutic window during which G-CSF could rescue visual function and retinal ganglion cell survival was demonstrated at day 0 and day 1 post-infarct. Macrophage infiltration was reduced by 3.1- and 1.6-fold by G-CSF treatment starting on day 0 and 1 post-rAION induction, respectively, compared with the PBS-treated group (P<0.05). This was compatible with 3.3- and 1.7-fold reductions in ON vascular permeability after G-CSF treatment compared with PBS treatment (P<0.05). Microglial activation was increased by 3.8- and 3.2-fold in the early (beginning treatment at day 0 or 1) G-CSF-treated group compared with the PBS-treated group (P<0.05). Immediate (within 30 mins of infarct) treatment with G-CSF also induced M2 microglia/macrophage activation. The cytokine levels were lower in the group that received immediate G-CSF treatment compared to those in the later G-CSF treatment group (P<0.05). Early treatment with G-CSF stabilized the blood–ON barrier to reduce macrophage infiltration and induced M2 microglia/macrophage polarization to decrease the expressions of pro-inflammatory cytokines in this rAION model. Summary: Early G-CSF treatment (within 24 hours of a rAION infarct) can stabilize optic nerve (ON) vascular permeability to reduce macrophage infiltration into the ON and induce M2 microglia/macrophage polarization at the ON, resulting in decreased inflammation.

Real-time vascular imaging and photodynamic therapy efficacy with micelle-nanocarrier delivery of chlorin e6 to the microenvironment of melanoma

BACKGROUND Strategies combining anti-vascular therapy and vascular imaging may facilitate the prediction of early response and outcome in cancer treatment. OBJECTIVE The aim of this study was to investigate the relationship between the tumor-associated vasculature in melanoma and to develop an approach for melanoma treatment by utilizing the free form and micelle form of the photosensitizer (PS) chlorin e6 in photodynamic therapy (PDT). METHODS Green fluorescence protein (GFP) expressing B16-F10 melanoma cells were implanted into the mouse ear dermis. Ce6 in free form or in micelle form was administered via the tail vein. An OV100 imaging system was used to record the red fluorescence of Ce6 to obtain real-time vascular images in the GFP tumor. RESULTS Compared to free Ce6, Ce6 linked to the micelle-nanocarrier depicted a much clearer vascular image and had an effective vascular destruction by PDT. Micelle Ce6 was localized in lysosomes and in the endoplasmic reticulum of cultured endothelial cells, implying an active endocytosis of the nano-carrier. CONCLUSION Micelle Ce6 can serve as a bifunctional PS for vascular imaging and PDT, which facilitates its delivery in the tumor microenvironment.