Chi-Chen Lin

Terpinen-4-ol Induces Apoptosis in Human Nonsmall Cell Lung Cancer In Vitro and In Vivo

Terpinen-4-ol, a monoterpene component of the essential oils of several aromatic plants, exhibits antitumor effects. In this study, the antitumor effects of terpinen-4-ol and the cellular and molecular mechanisms responsible for it were evaluated and studied, respectively on human nonsmall cell lung cancer (NSCLC) cells. Our results indicated that terpinen-4-ol elicited a dose-dependent cytotoxic effect, as determined by MTT assay. Increased sub-G1 population and annexin-V binding, activation of caspases 9 and 3, cleavage of poly(ADPribose) polymerase (PARP), and a decrease of mitochondrial membrane potential (MMP) indicated involvement of the mitochondrial apoptotic pathway in terpinen-4-ol-treated A549 and CL1-0 cells. Elevation of the Bax/Bcl-2 ratio and a decrease in IAP family proteins XIAP and survivin were also observed following terpinen-4-ol treatment. Notably, terpinen-4-ol was able to increase p53 levels in A549 and CL1-0 cells. Diminution of p53 by RNA interference induced necrosis instead of apoptosis in A549 cells following terpinen-4-ol treatment, indicating that terpinen-4-ol-elicited apoptosis is p53-dependent. Moreover, intratumoral administration of terpinen-4-ol significantly suppressed the growth of s.c. A549 xenografts by inducing apoptosis, as confirmed by TUNEL assay. Collectively, these data provide insight into the molecular mechanisms underlying terpinen-4-ol-induced apoptosis in NSCLC cells, rendering this compound a potential anticancer drug for NSCLC.

Nitride-based LEDs with nano-scale textured sidewalls using natural lithography

This investigation describes the development of a InGaN/GaN light-emitting diode (LED) with textured sidewalls using natural lithography with polystyrene spheres (PSs) as the etching mask and dry etching the epitaxial layers of LEDs to achieve nano-scale textured sidewalls. The LED with textured sidewalls increased the output power of the InGaN?GaN multiple quantum well (MQW) LEDs by a factor of 1.3, indicating that the LED with nano-scale textured sidewalls had larger light extraction efficiency. The wall-plug efficiency of nitride-based LEDs was increased by 30% using textured sidewalls.

The Adjuvant Effects of High-Molecule-Weight Polysaccharides Purified from Antrodia cinnamomea on Dendritic Cell Function and DNA Vaccines

The biological activity of the edible basidiomycete Antrodia cinnamomea (AC) has been studied extensively. Many effects, such as anti-cancer, anti-inflammatory, and antioxidant activities, have been reported from either crude extracts or compounds isolated from AC. However, research addressing the function of AC in enhancing immunity is rare. The aim of the present study is to investigate the active components and the mechanism involved in the immunostimulatory effect of AC. We found that polysaccharides (PS) in the water extract of AC played a major role in dendritic cell (DC) activation, which is a critical leukocyte in initiating immune responses. We further size purified and identified that the high-molecular weight PS fraction (greater than 100 kDa) exhibited the activating effect. The AC high-molecular weight PSs (AC hmwPSs) promoted pro-inflammatory cytokine production by DCs and the maturation of DCs. In addition, DC-induced antigen-specific T cell activation and Th1 differentiation were increased by AC hmwPSs. In studying the molecular mechanism, we confirmed the activation of the MAPK and NF-κB pathways in DCs after AC hmwPSs treatment. Furthermore, we demonstrated that TLR2 and TLR4 are required for the stimulatory activity of AC hmwPSs on DCs. In a mouse tumor model, we demonstrated that AC hmwPSs enhanced the anti-tumor efficacy of the HER-2/neu DNA vaccine by facilitating specific Th1 responses. Thus, we conclude that hmwPSs are the major components of AC that stimulate DCs via the TLR2/TLR4 and NF-κB/MAPK signaling pathways. The AC hmwPSs have potential to be applied as adjuvants.

Formulated extract from multiple citrus peels impairs dendritic cell functions and attenuates allergic contact hypersensitivity

It has been reported that gold lotion (GL), a formulated product made from the peels of six citrus fruits, has many pharmacological properties, such as anti-tumor, antioxidant, and anti-inflammatory activities. In this study, we investigated the immunomodulatory effect of GL on lipopolysaccharide (LPS) stimulated mouse bone marrow-derived DC maturation and function. Our experimental results have shown that GL significantly impaired the pro-inflammatory cytokine and chemokine secretion, suppressed the expression of major histocompatibility complex class I/II and costimulatory molecules (CD40, CD80 and CD86), increased phagocytic capacity, and reduced propensity to stimulate the autologous CD4(+) and CD8(+) T cell proliferation of LPS-induced DCs. Furthermore, we found that oral administration of GL attenuated the 2,4-Dinitro-1-fluorobenzene induced contact hypersensitivity (CHS) in animal models. Subsequently, our molecular mechanism studies showed that GL interfered with LPS-induced MAPK-JNK, p38 phosphorylation and nuclear translocation of NF-κB p65. In an essence, these findings are the first report to provide new insight in the immunopharmacological role of GL in terms of its effects on DC.

p53 modulates the AMPK inhibitor compound C induced apoptosis in human skin cancer cells.

Compound C, a well-known inhibitor of the intracellular energy sensor AMP-activated protein kinase (AMPK), has been reported to cause apoptotic cell death in myeloma, breast cancer cells and glioma cells. In this study, we have demonstrated that compound C not only induced autophagy in all tested skin cancer cell lines but also caused more apoptosis in p53 wildtype skin cancer cells than in p53-mutant skin cancer cells. Compound C can induce upregulation, phosphorylation and nuclear translocalization of the p53 protein and upregulate expression of p53 target genes in wildtype p53-expressing skin basal cell carcinoma (BCC) cells. The changes of p53 status were dependent on DNA damage which was caused by compound C induced reactive oxygen species (ROS) generation and associated with activated ataxia-telangiectasia mutated (ATM) protein. Using the wildtype p53-expressing BCC cells versus stable p53-knockdown BCC sublines, we present evidence that p53-knockdown cancer cells were much less sensitive to compound C treatment with significant G2/M cell cycle arrest and attenuated the compound C-induced apoptosis but not autophagy. The compound C induced G2/M arrest in p53-knockdown BCC cells was associated with the sustained inactive Tyr15 phosphor-Cdc2 expression. Overall, our results established that compound C-induced apoptosis in skin cancer cells was dependent on the cells p53 status.

5-demethylnobiletin promotes the formation of polymerized tubulin, leads to G2/M phase arrest and induces autophagy via JNK activation in human lung cancer cells

5-Demethylnobiletin is a hydroxylated polymethoxyflavone found in citrus plants that shows antiproliferative activities in several cancer cell lines. In this study, we investigated the effects and underlying molecular mechanisms of 5-demethylnobiletin on inhibition of cell growth, apoptosis, cell cycle and autophagy in A549 and CL1-5 lung cancer cells. The results of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay suggested that 5-demethylnobiletin inhibited cell growth in a dose- and time-dependent manner. Flow cytometry results suggested that 5-demethylnobiletin inhibited proliferation in lung cancer cells by inducing G2/M cell cycle phase arrest but predominantly not through apoptosis. Western blot results illustrated that the blockade of the cell cycle was associated with reduced levels of cdc25 and cdc2. Notably, our results indicated that 5-demethylnobiletin induced significant abnormal microtubule dynamics in A549 and CL1-5 cells, a novel finding. Studies conducted with isolated tubulin and docking models suggest that 5-demethylnobiletin promoted the polymerization of microtubules and bound to the taxol site. Additionally, 5-demethylnobiletin might also induce autophagy via activation of the JNK signaling pathway in A549 and CL1-5 cells. Pretreatment of the cells with the autophagy inhibitor 3-methyladenine significantly potentiated 5-demethylnobiletin-induced apoptosis, suggesting that 5-demethylnobiletin-induced autophagy mitigated cell apoptosis. Further investigation revealed that 5-demethylnobiletin inhibition of CL1-5 lung cancer cell growth was reproducible in a nude mouse model. Taken together, these studies suggest that 5-demethylnobiletin has anti-lung cancer efficacy both in vitro and in vivo possibly through induction of G2/M arrest, autophagy and apoptosis.

Baicalein Triggers Mitochondria-Mediated Apoptosis and Enhances the Antileukemic Effect of Vincristine in Childhood Acute Lymphoblastic Leukemia CCRF-CEM Cells

Acute lymphoblastic leukemia (ALL) accounts for approximately 75% of childhood leukemia, and chemotherapy remains the mainstay therapy. Baicalein is an active flavonoid used in traditional Chinese medicine and has recently been found to have anticancer, anti-inflammatory, and antiallergic properties. This study aims to investigate the molecular apoptotic mechanisms of baicalein in CCRF-CEM leukemic cells and to evaluate the combined therapeutic efficacy of baicalein with several commonly used chemotherapeutic drugs in CCRF-CEM cells. Our results demonstrate that baicalein induces mitochondria-dependent cleavage of caspases-9 and -3 and PARP with concomitant decreases in IAP family proteins, survivin, and XIAP. Furthermore, our results present for the first time that baicalein triggers a convergence of the intrinsic and extrinsic apoptotic pathways via the death receptor-caspase 8-tBid signaling cascade in CCRF-CEM cells. In addition, we also present for the first time that the combination of baicalein and vincristine results in a synergistic therapeutic efficacy. Overall, this combination strategy is recommended for future clinical trials in the treatment of pediatric leukemia owing to baicaleins beneficial effects in alleviating the vomiting, nausea, and skin rashes caused by chemotherapy.

Mcl-1 determines the imiquimod-induced apoptosis but not imiquimod-induced autophagy in skin cancer cells.

BACKGROUND Imiquimod had been shown to induce apoptosis and autophagy in several skin cancer cells, especially basal cell carcinoma (BCC) cells. OBJECTIVE We evaluate the molecular mechanisms of imiquimod-induced apoptosis and autophagy in skin cancer cell lines. METHODS The Mcl-1, Bcl-2 and Bcl-xL proteins were determined by immunoblotting. The Mcl-1 mRNA level was examined by RT-PCR and real-time PCR. The mechanisms of imiquimod-induced decrease in Mcl-1 protein were evaluated by addition of cycloheximide, MG132 proteasome inhibitor or pan-caspase inhibitor. The phosphorylation of eIF4E, 4E-BP1 and eEF2 in imiquimod treated cells were examined by immunoblotting. The imiquimod-induced apoptosis and autophagy were evaluated in Mcl-1-overexpressing cells by XTT test, mitochondrial membrane potential measurement, DNA content assay, LC3 immunoblotting, EGFP-LC3 puncta formation and quantification of acidic vesicular organelle with acridine orange staining. RESULTS The decrease in the Mcl-1 protein level was faster and stronger than the decrease in Bcl-2 and Bcl-xL in imiquimod-treated skin cancer cells. The imiquimod-induced decrease in Mcl-1 protein was not caused by blocked transcription or the promotion of degradation but was associated with inactivation of translation factors in BCC cells. The Mcl-1-overexpressing BCC cells were more resistant to intrinsic cellular apoptosis than control BCC cells during imiquimod treatment. Mcl-1 overexpression in BCC cells resulted in the basal activation of autophagy but did not modulate imiquimod-induced autophagy or rescue imiquimod-induced autophagic cell death in BCC cells. CONCLUSIONS Imiquimod may rapidly downregulate Mcl-1 protein levels by inhibiting translation in skin cancer cells. Mcl-1 may act to protect against apoptosis but not autophagy and autophagic cell death during imiquimod treatment in skin cancer cells.

Comparative Nephrotoxicity of Aristolochic Acid and Tetrandrine In Vitro and In Vivo

Aristolochic acid (AA) and tetrandrine (TET) are the major bioactive components in Chinese herbs used for weight loss. The nephropathy caused by the 2 Chinese herbs has not been simultaneously investigated. The aim of this study was to examine the potential nephrotoxicity of AA and TET using Madin-Darby canine kidney (MDCK) cells and mice. The results showed that TET was more potent than AA in inhibiting MDCK cell growth via inducing apoptosis, as determined by annexin-V staining, 4’, 6’-diamino-2-phenylindole (DAPI) staining, DNA fragmentation, and caspase 3 activity. Mice treated with AA (10 mg/kg) by intraperitoneal administration for 3 months showed nephrotoxicity, elevated blood urea nitrogen, and increased renal tubular injuries. In contrast, mice treated with 50 mg/kg of TET in the same time period had moderate hydropic degeneration of the distal tubules in the kidneys. These results suggest that TET is more cytotoxic than AA in MDCK cells but shows less nephrotoxic than AA in mice.

Dextromethorphan Inhibits Activations and Functions in Dendritic Cells

Dendritic cells (DCs) play an important role in connecting innate and adaptive immunity. Thus, DCs have been regarded as a major target for the development of immunomodulators. In this study, we examined the effect of dextromethorphan (DXM), a common cough suppressant with a high safety profile, on the activation and function of DCs. In the presence of DXM, the LPS-induced expression of the costimulatory molecules in murine bone marrow-derived dendritic cells (BMDCs) was significantly suppressed. In addition, DXM treatment reduced the production of reactive oxygen species (ROS), proinflammatory cytokines, and chemokines in maturing BMDCs that were activated by LPS. Therefore, DXM abrogated the ability of LPS-stimulated DCs to induce Ag-specific T-cell activation, as determined by their decreased proliferation and IFN-γ secretion in mixed leukocyte cultures. Moreover, the inhibition of LPS-induced MAPK activation and NF-κB translocation may contribute to the suppressive effect of DXM on BMDCs. Remarkably, DXM decreased the LPS-induced surface expression of CD80, CD83, and HLA-DR and the secretion of IL-6 and IL-12 in human monocyte-derived dendritic cells (MDDCs). These findings provide a new insight into the impact of DXM treatment on DCs and suggest that DXM has the potential to be used in treating DC-related acute and chronic diseases.