Enhanced NIR radiation-triggered hyperthermia by mitochondrial targeting.

Mitochondria are organelles that are readily susceptible to temperature elevation. We selectively delivered a coumarin-based fluorescent iron oxide nanoparticle, Mito-CIO, to the mitochondria. Upon 740 nm laser irradiation, the intracellular temperature of HeLa cells was elevated by 2.1 °C within 5 min when using Mito-CIO, and the treatment resulted in better hyperthermia and a more elevated cytotoxicity than HeLa cells treated with coumarin iron oxide (CIO), which was missing the mitochondrial targeting unit. We further confirmed these results in a tumor xenograft mouse model. To our knowledge, this is the first report of a near-infrared laser irradiation-induced hyperthermic particle targeted to mitochondria, enhancing the cytotoxicity in cancer cells. Our present work therefore may open a new direction in the development of photothermal therapeutics.

Effect of Achyranthes bidentata Blume on 3T3-L1 Adipogenesis and Rats Fed with a High-Fat Diet.

The present study investigated the antiobesity effect of Achyranthes bidentata Blume root water extract in a 3T3-L1 adipocyte differentiation model and rats fed with a high-fat diet. To investigate the effect of Achyranthes bidentata Blume on adipogenesis in vitro, differentiating 3T3-L1 cells in adipocyte-induction media were treated every two days with Achyranthes bidentata Blume at various concentrations (1 to 25 µ g/mL) for eight days. We found that Achyranthes bidentata Blume root inhibited 3T3-L1 adipocyte differentiation without affecting cell viability, and Western blot analysis revealed that phospho-Akt expression was markedly decreased, whereas there was no significant change in perilipin expression. Furthermore, administration of Achyranthes bidentata Blume root (0.5 g/kg body weight for six weeks) to rats fed with a high-fat diet significantly reduced body weight gain without affecting food intake, and the level of triglyceride was significantly decreased when compared to those in rats fed with only a high-fat diet. These results suggest that Achyranthes bidentata Blume root water extract could have a beneficial effect on inhibition of adipogenesis and controlling body weight in rats fed with a high-fat diet.

Neuroprotective effect of Chunghyuldan (Qing Xue Dan) on hypoxia-reoxygenation induced damage of neuroblastoma 2a cell lines.

OBJECTIVES: Chunghyuldan (CHD), a combinatorial drug that has anti-hyperlipidemic and antiinflammatory activities, has been shown to reduce infarct volume in a focal ischemia-reperfusion rat model. To explore the molecular basis of CHD's neuroprotective effect, we examined whether CHD shows a cell-protective activity and has a regulatory effect on Bax and/or B-cell leukemia/lymphoma 2 (Bcl-2) expression in mouse neuroblastoma 2a (N2a) cells subjected to hypoxia-reoxygenation (H/R). METHODS: In order to evaluate the effects of CHD on the cytotoxicity induced from hypoxia or H/R condition, lactate dehydrogenase (LDH) assay was performed. To explore whether the suppression of neural damage when pre-treated with CHD is associated with its anti-apoptotic effect, the CHD effect on the expression of Bcl-2 and Bax was analyzed by Western blotting analysis. RESULTS: Cytotoxicity of N2a cell line was slightly increased in 42 h hypoxia condition and dramatically increased under the H/R condition. CHD treatment markedly decreased the cytotoxicity in both conditions (P<0.01, P<0.05). H/R markedly increased the expression of the pro-apoptotic protein, Bax, but slightly increased the expression of the anti-apoptotic protein, Bcl-2, compared with the normoxia or hypoxia group. CHD significantly decreased Bax expression (P<0.01) and slightly decreased Bcl-2 expression (P>0.05), resulted in a reduction of Bax/Bcl-2 ratio in N2a cells subjected to H/R. CONCLUSION: CHD has neuroprotective effect in N2a cells subjected to H/R, which might be derived at least in part from its ability to decrease the expression of the pro-apoptotic protein, Bax.

Effects of tetramethylpyrazine on microglia activation in spinal cord compression injury of mice.

Secondary mechanisms, including inflammation and microglia activation, serve as targets for the development and application of pharmacological strategies in the management of spinal cord injury (SCI). Tetramethylpyrazine (TMP), an active ingredient of Ligusticum wallichii (chuanxiong), has shown anti-inflammatory and neuroprotective effects against SCI. However, it remains uncertain whether the inflammation-suppressive effects of TMP play a modulatory role over microglia activation in SCI. The present study investigated the effects of TMP on microglia activation and pro-inflammatory cytokines in spinal cord compression injury in mice. For a real-time PCR measurement of pro-inflammatory cytokines, SCI was induced in mice by the clip compression method (30 g force, 1 min) and TMP (15 or 30 mg/kg, i.p.) was administered once, 30 minutes before the SCI induction. For immunohistochemistry, TMP (30 mg/kg, i.p.) treatment was given three times during the first 48 hours after the SCI. 30 mg/kg of TMP treatment reduced the up-regulation of TNF-α, IL-1ß and COX-2 mRNA in the spinal tissue at four hours after the SCI induction. TMP also significantly attenuated microglia activation and neutrophil infiltration at 48 hours after the SCI induction. In addition, iNOS expression in the spinal tissue was attenuated with TMP treatment. These results suggest that TMP plays a modulatory role in microglia activation and may protect the spinal cord from or potentially delay secondary spinal cord injury.

Effects of Atractylodes macrocephala Koidzumi rhizome on 3T3-L1 adipogenesis and an animal model of obesity.

ETHNOPHARMACOLOGICAL RELEVANCE: Atractylodes macrocephala Koidzumi (AMK) is an herbal medicine traditionally used for treatment of abdominal pain, gastrointestinal disease, obesity, and related complications. AIM OF THE STUDY: We investigated the effects and molecular mechanism of AMK rhizome water extract on 3T3-L1 adipogenesis and an animal model of obesity. MATERIALS AND METHODS: To study the effect of AMK on adipogenesis in vitro, differentiating 3T3-L1 cells were treated every two days with AMK at various concentrations (1-25µg/ml) for eight days. Oil Red O staining was performed to determine the lipid accumulation in 3T3-L1 cells. To elucidate the inhibitory mechanism of AMK on adipogenesis, phosphorylation levels of Akt and expression of perilipin, were analyzed by Western blotting. AMK was administered orally to high fat diet (HFD)-induced obese rats to confirm its effect in vivo. RESULTS: AMK inhibited 3T3-L1 adipocyte differentiation in a dose-dependent manner without cellular toxicity. Phospho-Akt expression was highly decreased by AMK treatment, whereas there was no significant change in perilipin expression. AMK administration significantly reduced the body weight of rats fed a HFD. Plasma triglyceride levels were significantly lower in the AMK-treated HFD group than those in the HFD control group or normal diet (ND) group, although serum total, HDL- and LDL-cholesterol levels did not differ between the groups. CONCLUSION: These results demonstrate an inhibitory effect of AMK on adipogenesis through reduction of an adipogenic factor, phospho-Akt. AMK had a beneficial effect, reducing body weight gain in a HFD-induced animal model of obesity.

Zingiber officinale protects HaCaT cells and C57BL/6 mice from ultraviolet B-induced inflammation.

Ultraviolet (UV) light is a physical carcinogen, and UV irradiation from sunlight has negative effects on human skin. UVB-induced inflammation is linked to excessive induction of various inflammatory cytokines/chemokines in many types of cells, including keratinocytes. The purpose of this study was to investigate the anti-inflammatory effect of water extract of Zingiber officinale, gingerol, and shogaol on UVB-induced skin damage in the human keratinocyte cell line HaCaT and C57BL/6 mice. To test for an effective compound to protect against inflammation in UV-damaged skin, we prepared a water extract of ginger rhizomes and examined the effects of Z. officinale, gingerol, and shogaol on cell viability and cytokine/chemokine production in UV-irradiated HaCaT cells. We also investigated the in vivo relevance of these findings in C57BL/6 mice using hematoxylin and eosin staining and cytokine measurements. A water extract of Z. officinale, gingerol, and shogaol inhibited production of cytokines in UVB-irradiated HaCaT cells effectively. Treatment with Z. officinale attenuated UVB-induced hyperplasia, infiltration of leukocytes, and dilation of blood vessels in the dermis of mice. Z. officinale, gingerol, and shogaol show potential as anti-inflammatory agents to protect skin against UVB irradiation damage.

Inhibitory effects of 5-chloroacetyl-2-piperidino-1,3-selenazole, a novel selenium-containing compound, on skin melanin biosynthesis.

OBJECTIVES: Increased production and accumulation of melanin leads to many hyperpigmentation disorders such as melasma, freckles and geriatric pigment spots. Thus, there is a need for the development of depigmenting agents. Based on our previous reports, selenium derivatives as anti-melanogenic lead compounds could be very important. The aim of this study was to investigate the depigmenting effect of novel selenium-containing compounds. METHODS: The inhibitory effects of 5-chloroacetyl-2-piperidino-1,3-selenazole (CS1), a novel selenium-containing compound, on melanogenesis were investigated in B16F10 melanoma cells and cultured brownish guinea pig skin tissue with alpha-melanocyte-stimulating hormone stimulation. KEY FINDINGS: We found that CS1 inhibited melanin production in B16F10 cells by suppressing tyrosinase activity and its protein expression. In addition, Western blotting analysis revealed that CS1 suppressed the expression of tyrosinase-related protein (TRP)-1 and TRP-2. Therefore, the depigmenting effect of CS1 might have been due to inhibition of tyrosinase activity and expression of melanogenic enzymes. Furthermore, CS1 had inhibitory effects on melanin biosynthesis of primary cultured skin of brownish guinea pig. CONCLUSIONS: The results suggested that CS1 could be a useful candidate for the treatment of skin hyperpigmentation.

Genipin inhibits the inflammatory response of rat brain microglial cells.

Microglia are the prime effectors in immune and inflammatory responses of the central nervous system (CNS). Under pathological conditions, the activation of these cells helps restore CNS homeostasis. However, chronic microglial activation endangers neuronal survival through the release of various proinflammatory and neurotoxic factors. Thus, negative regulators of microglial activation have been considered as potential therapeutic candidates to target neurodegeneration, such as that in Alzheimer's and Parkinson's diseases. Genipin, the aglycon of geniposide found in gardenia fruit has long been considered for treatment of various disorders in traditional oriental medicine. Genipin has recently been reported to have diverse pharmacological functions, such as antimicrobial, antitumor, and anti-inflammatory effects. The specific aim of this study was to examine whether genipin represses brain microglial activation. Genipin was effective at inhibiting LPS-induced nitric oxide (NO) release from cultured rat brain microglial cells. Genipin reduced the LPS-stimulated production of tumor necrosis factor-alpha, interleukin-1beta, prostaglandin E(2), intracellular reactive oxygen species, and NF-kappaB activation. In addition, genipin reduced NO release from microglia stimulated with interferon-gamma and amyloid-beta. Both pretreatment and post-treatment of genipin to LPS-stimulated microglia were effective at decreasing NO release. Furthermore, genipin effectively inhibited microglial activation in a mouse model of brain inflammation. These results suggest that genipin provide neuroprotection by reducing the production of various neurotoxic molecules from activated microglia.

Effects of red ginseng extract on UVB irradiation-induced skin aging in hairless mice.

UNLABELLED: Panax ginseng C.A. Meyer, Korean herb medicine, has been widely used in China and Japan for fatigue and enhancement of resistance to many diseases. AIM OF THE STUDY: This study is aimed to assess the effects of Korean red ginseng extract on UVB irradiation induced skin aging in hairless mice. MATERIALS AND METHODS: Red ginseng extracts prepared with ethanol were used in this study. To standardize Korean red ginseng, it was analyzed by HPLC. And inhibitory effects of red ginseng extract on UVB irradiation-induced skin aging in hairless mice were determined by the measurement of wrinkle, expression of type I procollagen and MMP-1 and immunohistology. RESULTS: Based on the HPLC quantitative analysis, ginsenoside Rb1 content in Korean red ginseng was 43.5mg/g of extract. In the result of body weight gain and food efficiency rate, body weights of all groups were increased during experimental periods. In the wrinkle measurement and image analysis of skin replicas, the results showed that the dietary supply containing red ginseng extract significantly inhibited wrinkle formation caused by chronic UVB irradiation. In the changes of expression of procollagen type I and MMP-1 in the skin of UV irradiated hairless mice fed dietary supplement containing 2.5% red ginseng extract, level of mRNA of procollagen type I was decreased. But protein level of that was increased. And in terms of MMP-1, either mRNA or protein levels of MMP-1 were significantly decreased. These results showed anti-wrinkle effect of Korean red ginseng involved the inhibition of collagen degradation rather than increased collagen synthesis. CONCLUSION: It is shown that Korean red ginseng may be functional food candidate for skin photoaging.

Chunghyuldan attenuates brain microglial inflammatory response.

Microglial cells are the prime effectors in immune and inflammatory responses of the central nervous system (CNS). During pathological conditions, the activation of these cells helps restore CNS homeostasis. However, chronic microglial activation endangers neuronal survival through the release of various proinflammatory molecules and neurotoxins. Thus, negative regulators of microglial activation have been considered as potential therapeutic candidates to target stroke and neurodegenerative diseases. Chunghyuldan, a combinatorial drug consisting of Scutellariae Radix, Coptidis Rhizoma, Phellodendri Cortex, Gardeniae Fructus, and Rhei Rhizoma, has an inhibitory effect on stroke recurrence in patients with small-vessel disease. It has also been reported to confer antihypertensive, antihyperlipidemic, and antiinflammatory effects. The aim of this study was to examine whether Chunghyuldan suppresses microglial activation. Chunghyuldan was effective at inhibiting LPS-induced nitric oxide (NO) release from rat brain microglia. Real-time reverse transcriptase PCR analysis revealed that pretreatment of rat brain microglia with Chunghyuldan attenuated the LPS-induced expression of mRNAs encoding inducible NO synthase, tumor necrosis factor (TNF)-alpha, interleukin-1beta, and cyclooxygenase-2. In rat brain microglia, Chunghyuldan reduced the LPS-stimulated production of TNF-alpha and prostaglandin E2. In addition, Chunghyuldan significantly decreased LPS-induced phosphorylation of the ERK1/2 and p38 signaling proteins. These results suggest that Chunghyuldan provide neuroprotection by reducing the release of various proinflammatory molecules from activated microglia.

Tetrandrine cytotoxicity and its dual effect on oxidative stress-induced apoptosis through modulating cellular redox states in Neuro 2a mouse neuroblastoma cells.

Tetrandrine (TET), a plant alkaloid, is known primarily as a non-selective Ca(2+) channel blocker. On the contrary to the cytoprotective effect on ischemia/reperfusion injury, TET has also been reported to cause cytotoxicity. In this study, we wished to understand the apparently disparate effects of this potential drug and thus investigated molecular mechanisms on proliferation and apoptosis and its effect on oxidative stress-induced apoptosis in Neuro 2a mouse neuroblastoma cells. We showed that TET, at high concentrations, induced cell cycle arrest and apoptosis through oxidative stress with following observations. Firstly, 10 microM TET elevated the reactive oxygen species (ROS) level and accordingly depleted glutathione (GSH) content. Secondly, pretreatment with antioxidants (NAC or GSH) protected cells from TET-induced apoptosis. We also demonstrated that treatment with 10 microM TET caused not only induction of p53, p21(waf1), and Bax, but also nuclear translocation of p53 and hypo-phosphorylation of pRb concurrently. Our important finding is that the concentration-dependent dual effect of TET, either inhibiting or promoting cell death induced by H(2)O(2) was observed, probably through regulating redox balance, which was well reflected on the GSH content in each condition. Besides, inhibition of Ca(2+) influx protected cells from H(2)O(2)-induced apoptosis even in the presence of 10 microM TET. Taken together, our data suggest that TET regulation of cellular redox states may play a major role in its dual action of cytotoxicity and cytoprotection.