Myo-Kyoung Kim

Incorporation of Bloom’s Taxonomy into Multiple-Choice Examination Questions for a Pharmacotherapeutics Course

Objective. To incorporate Bloom’s taxonomy into multiple-choice examination questions in a pharmacotherapeutics course and assess its effectiveness in detecting areas of improvement in learning. Design. Bloom’s taxonomy was incorporated into examination questions through a multi-step process: Sample questions representing each learning domain within Bloom’s taxonomy (knowledge, comprehension, application, analysis, synthesis, and evaluation) were introduced to students during lecture presentations and discussions. Quiz and examination containing questions categorized according to Bloom’s taxonomy were administered to students. During review sessions following each quiz or examination, the categorization of each question was provided to students and feedback from students was gathered. Assessment. The effect of the 5 types of test questions on the correct response fraction and discrimination index was determined after combining synthesis and evaluation. Correct response fractions for knowledge, comprehension, and application questions were significantly higher than those for analysis and synthesis/evaluation questions (p<0.05). However, discrimination index for application and synthesis/evaluation questions were significantly higher than those for knowledge and comprehension questions (p<0.05). In interviews with students who had requested learning assistance, the majority realized the importance of critical-thinking skills in the learning process. Conclusion. Well-designed multiple-choice questions incorporating different learning domains of Bloom’s taxonomy may be a potential method of assessing critical-thinking skills in large classes of students.

ERK Activation by Fucoidan Leads to Inhibition of Melanogenesis in Mel-Ab Cells

Fucoidan, a fucose-rich sulfated polysaccharide derived from brown seaweed in the class Phaeophyceae, has been widely studied for its possible health benefits. However, the potential of fucoidan as a possible treatment for hyperpigmentation is not fully understood. This study investigated the effects of fucoidan on melanogenesis and related signaling pathways using Mel-Ab cells. Fucoidan significantly decreased melanin content. While fucoidan treatment decreased tyrosinase activity, it did not do so directly. Western blot analysis indicated that fucoidan downregulated microphthalmia-associated transcription factor and reduced tyrosinase protein expression. Further investigation showed that fucoidan activated the extracellular signal-regulated kinase (ERK) pathway, suggesting a possible mechanism for the inhibition of melanin synthesis. Treatment with PD98059, a specific ERK inhibitor, resulted in the recovery of melanin production. Taken together, these findings suggest that fucoidan inhibits melanogenesis via ERK phosphorylation.

Tumor apoptosis by indole-3-acetic acid/light in B16F10 melanoma-implanted nude mice

Recently, we reported that UVB-activated indole-3-acetic acid (IAA) induces the apoptosis of G361 human melanoma cells. In the present study, we used IAA and visible light combinations to treat B16F10 melanoma-implanted nude mice using an experimental intense pulsed light (IPL) therapy model. We first investigated whether activated IAA by horseradish peroxidase (HRP) or UVB causes apoptosis of B16F10 melanoma cells. IAA/HRP or IAA/UVB combination lead to apoptosis of B16F10 cells, as reported in other cell lines. Interestingly, IAA alone was not cytotoxic. These findings suggested the potential use of IAA in the treatment of melanoma. For the future clinical use, we also tested whether visible light has the same effects like UVB and found that visible light also activates IAA to produce free radicals and that IAA/visible light decreased cell viability significantly. Based on these results, IAA/IPL combination was tried whether it can induce apoptosis in vivo status. TUNEL staining showed that IAA/IPL treatment induced apoptosis of tumor cells. In addition, the expressions of p53, Fas, and PARP were upregulated in the IAA/IPL-treated group than in untreated control, demonstrating that IAA/IPL treatment caused apoptosis in melanoma-implanted nude mice. In conclusion, we showed that IAA/IPL induces melanoma regression in B16F10 melanoma-implanted nude mice. These results suggest the potential use of IAA/IPL in the treatment of malignant melanoma.

Scopoletin from Cirsium setidens Increases Melanin Synthesis via CREB Phosphorylation in B16F10 Cells.

In this study, we isolated scopoletin from Cirsium setidens Nakai (Compositae) and tested its effects on melanogenesis. Scopoletin was not toxic to cells at concentrations less than 50 µM and increased melanin synthesis in a dose-dependent manner. As melanin synthesis increased, scopoletin stimulated the total tyrosinase activity, the rate-limiting enzyme of melanogenesis. In a cell-free system, however, scopoletin did not increase tyrosinase activity, indicating that scopoletin is not a direct activator of tyrosinase. Furthermore, Western blot analysis showed that scopoletin stimulated the production of microphthalmia-associated transcription factor (MITF) and tyrosinase expression via cAMP response element-binding protein (CREB) phosphorylation in a dose-dependent manner. Based on these results, preclinical and clinical studies are needed to assess the use of scopoletin for the treatment of vitiligo.

Leucine-rich glioma inactivated 3 promotes HaCaT keratinocyte migration

Our finding that human skin expresses leucine‐rich glioma inactivated 3 (LGI3) raises the question of the function of this cytokine in keratinocytes. We have shown that LGI3 stimulates human HaCaT keratinocyte migration without affecting viability or proliferation. Western blot analysis showed that LGI3 induced focal adhesion kinase activation, Akt phosphorylation, and glycogen synthase kinase 3β (GSK3β) phosphorylation in these cells. Using the scratch wound assay and a modified Boyden chamber, we found that LY294002, a selective phosphatidylinositol 3‐kinase inhibitor, and LiCl, a selective GSK3β inhibitor, abolished LGI3‐induced cell migration. We tested β‐catenin levels after LGI3 treatment because the Akt‐GSK3β pathway regulates β‐catenin accumulation, and β‐catenin promotes cell migration. LGI3 treatment increased β‐catenin protein and nuclear localization, whereas LY294002 prevented LGI3‐induced focal adhesion kinase and Akt activation as well as β‐catenin accumulation. Overall, these data suggest that LGI3 stimulates HaCaT cell migration following β‐catenin accumulation through the Akt pathway.

Geranylgeranylacetone induces apoptosis via the intrinsic pathway in human melanoma cells

The aim of this study was to test the anti-cancer effects of geranylgeranylacetone (GGA), an isoprenoid compound, on human melanoma cells. Human melanoma cell lines G361, SK-MEL-2, and SK-MEL-5 were treated with GGA at various doses (1-100μM). Cell viability was measured by crystal violet assay. Western blot analysis was adopted to detect marker proteins of apoptosis. GGA significantly reduced the viability of G361, SK-MEL-2, and SK-MEL-5 human melanoma cells at concentrations above 10μM. Western blot analysis showed the phosphorylation of p38 MAPK and c-Jun N-terminal kinase (JNK) after GGA treatment, as well as activation of caspase-9, caspase-3, and poly(ADP-ribose) polymerase (PARP) cleavage. GGA also induced p53 and Bax expression, but did not affect expression of Bcl-2 and MITF. These findings suggest that GGA induces apoptosis through the intrinsic pathway. Accordingly, GGA should be considered for further development as a potential agent for melanoma.

Dual hypopigmentary effects of punicalagin via the ERK and Akt pathways

Punicalagin is a phenolic compound with antioxidant properties. However, the effects of punicalagin on melanin synthesis have been poorly evaluated. Therefore, we investigated the effects of punicalagin on melanogenesis in Mel-Ab cells. Punicalagin significantly inhibited melanin synthesis in a dose-dependent manner. In accordance with the melanin content, punicalagin also dose-dependently decreased tyrosinase activity. Punicalagin did not directly inhibit tyrosinase in a cell-free system but did downregulate the expression of microphthalmia-associated transcription factor (MITF) and tyrosinase. Therefore, we examined the effects of punicalagin on melanogenesis-related signaling pathways. Punicalagin induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation but had no effect on β-catenin level. We measured melanin content and MITF expression in the presence of the ERK pathway inhibitor PD98059 and/or the Akt pathway inhibitor LY294002. Cotreatment with PD98059 and LY294002 almost completely restored punicalagin-induced hypopigmentation. These data indicate that punicalagin inhibits melanin synthesis through ERK and Akt phosphorylation, with subsequent downregulation of MITF and tyrosinase.

Menadione (Vitamin K3) decreases melanin synthesis through ERK activation in Mel-Ab cells

Menadione is a synthetic vitamin K3 derivative. Here, we examined the effects of menadione on melanogenesis and its related signaling pathways. Our results showed that melanin content was significantly reduced after menadione treatment in a dose-dependent manner. However, menadione treatment did not reduce tyrosinase activity directly. Wnt signaling is known to play a major role in the control of melanin synthesis. Thus, we tested the effects of menadione treatment on GSK3β and β-catenin signaling, but found that menadione did not influence either of these signaling pathways. We also investigated changes in the phosphorylation of ERK, which is related to melanin regulation. These results indicated that menadione treatment led to the phosphorylation of ERK. Additionally, menadione treatment reduced both MITF and tyrosinase protein levels. Treatment with PD98059, a specific ERK pathway inhibitor, restored menadione-induced melanin reduction and also prevented MITF and tyrosinase downregulation by menadione. These results suggest that the hypopigmentary action of menadione is due to MITF and tyrosinase downregulation by ERK activation.

UVB-irradiated indole-3-acetic acid induces apoptosis via caspase activation

Abstract Objective: Indole-3-acetic acid (IAA) activation has been suggested as a new strategy for cancer therapy. It has been reported that ultraviolet B (UVB) radiation can activate IAA. In the present study, we investigated whether UVB-irradiated IAA (IAAUVB) can induce apoptosis of G361 human melanoma cells and examined the apoptotic pathway involved. Methods: DNA fragmentation was measured to examine apoptosis. IAAUVB-induced signaling pathways were investigated by Western blot analysis. Results: Our results show that IAAUVB reduced cell viability of G361 human melanoma cells, and induced DNA fragmentation, a hallmark of apoptosis. We also found that c-Jun NH2-terminal kinase (JNK) and p38, which are activated by IAAUVB, are not associated with this cell death. We further investigated the IAAUVB-mediated apoptotic pathway after pretreatment with NS398, vitamin C, and N-acetylcysteine (NAC). Although NS398, an inhibitor of cyclooxygenase-2, was not protective, vitamin C and NAC ameliorated IAAUVB-mediated cell death. In addition, when cells were pretreated with a caspase inhibitor, IAAUVB-induced apoptosis was inhibited. Conclusions: These results suggest that free radicals generated from IAA by UV irradiation may cause apoptosis, and IAAUVB induces apoptosis of G361 human melanoma cells by activating caspases.