• Asian Pacific Journal of Cancer Prevention
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• v.15 no.2
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• pp.629-635
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• 2014

Background: Acute myeloid leukemia (AML) is a fatal hematological malignancy which is resistant to a variety of chemotherapy drugs. Myeloid cell leukemia-1 (Mcl-1), a death-inhibiting protein that regulates apoptosis, has been shown to be overexpressed in numerous malignancies. In addition, it has been demonstrated that the expression level of the Mcl-1 gene increases at the time of leukemic relapse following chemotherapy. The aim of this study was to target Mcl-1 by small interference RNA (siRNA) and analyze its effects on survival and chemosensitivity of acute myeloid leukemia cell line HL-60. Materials and Methods: siRNA transfection was performed with a liposome approach. The expression levels of mRNA and protein were measured by real-time quantitative PCR and Western blot analysis, respectively. Trypan blue assays were performed to evaluate tumor cell growth after siRNA transfection. The cytotoxic effects of Mcl-1 siRNA (siMcl-1) and etoposide were determined using MTT assay on their own and in combination. Apoptosis was quantified using a DNA-histone ELISA assay. Results: Transfection with siMcl-1 significantly suppressed the expression of Mcl-1 mRNA and protein in a time-dependent manner, resulting in strong growth inhibition and spontaneous apoptosis. Surprisingly, pretreatment with siMcl-1 synergistically enhanced the cytotoxic effect of etoposide. Furthermore, Mcl-1 down-regulation significantly increased apoptosis sensitivity to etoposide. No significant biological effects were observed with negative control siRNA treatment. Conclusions: Our results suggest that specific suppression of Mcl-1 by siRNA can effectively induce apoptosis and overcome chemoresistance of leukemic cells. Therefore, siMcl-1 may be a potent adjuvant in leukemia chemotherapy.

• The Journal of Korean Medicine
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• v.29 no.4
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• pp.146-160
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• 2008

Objectives: The present study was designated to evaluate the direct effects of Fructus Liquidambaris on suppression of eosinophil activity and on suppression of chemokines such as eotaxin, IL-8, ICAM-1, and VCAM-1, in vitro. Methods: A549 cells were stimulated with TNF-$\alpha$ (100 ng/ml), IL-4 (100 ng/ml) or IL-$1{\beta}$(10 ng/ml) to induce chemokines and adhesion molecules involved in eosinophil chemotaxis. Then after treatment of Fructus Liquidambaris, inhibition effect assay such as ELISA, real-time RT-PCR, and chemotaxis assay was performed. Results: Eotaxin level was suppressed in both protein secretion and mRNA expression. Eosinophil recruitment to lung epithelial cells was also reduced by Fructus Liquidambaris, implying the role of eotaxin in eosinophil recruitment. In addition, expression of IL-8 was also suppressed by Fructus Liquidambaris (p<0.05). However, expression of adhesion molecules (ICAM-1, VCAM-1) related to eosinophil was not affected. The eosinophil migration was inhibited at high concentration of Fructus Liquidambaris (p<0.05). Conclusion: These results suggest that Fructus Liquidambaris may regulate a common signaling pathway of eotaxin and IL-8. FS might be of therapeutic value in diseases such as asthma.

• Macromolecular Research
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• v.12 no.1
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• pp.46-52
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• 2004

In a continuation of our previous studies on blood compatibility profiles of anion-substituted poly(vinyl alcohol) (PVA) membranes, in which hydroxyl groups have been replaced with carboxymethyl (C-PVA) and sulfonyl groups (S-PVA), we have studied the activation of complement components and the changes in white cell and platelet count in vitro and compared them with those of unmodified PVA, Cuprophane, and low-density polyethylene. Complement activation of fluid phase components, C3a, Bb, iC3b, and SC5b-9, and of bound phases, C3c, C3d, and SC5b-9, were assessed by enzyme-linked immunosorbent assay (ELISA) and immunoblot, respectively. The changes in the number of white cells and platelets following complement activation were counted using a Coulter counter. C-PVA and S-PVA activated C3 to a lesser extent than did PVA, which we attribute to the diminished level of surface nucleophiles of the samples. In addition, C- and S-PVA exhibit increased inhibition of Bb production, resulting in a decrease in the extent of C5 activation. Consequently, because of the reduced activation of C3 and C5, C- and S-PVA samples cause marked decreases in the SC5b-9 levels in plasma. We also found that the negatively charged sulfonate and carboxylate groups of the samples cause a greater extent of adsorbtion of the positively charged anaphylatoxins, C3a and C5a, because of strong electrostatic attraction, which in turn provides an inhibition of chemotaxis and activation of leukocytes. The ability to inhibit complement production, together with the binding ability of anaphylatoxins of the C- and S-PVA samples, leads to a prominent decrease in lysis of leukocytes as well as activation of platelets.

• The Korea Journal of Herbology
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• v.21 no.2
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• pp.135-142
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• 2006

Objectives : On-Bi-Tang(OB) has been prescribed Chinese traditional medicine for the treatment of inflammatory disease such as chronic renal failure. In this study, we investigated the anti-inflammatory effect of OB extract in the BV2 murine microglial cells. Methods : After the water extract of OB was treated in BV2 cells, murine microglial line, the cell viability was measured by MTT assay. The production of nitric oxide (NO) and $TNF-{\alpha}$ was determined based on Griess reagent and enzyme linked immunosorbant assay (ELISA). mRNA expression of inducible nitric oxide synthase (iNOS) and $TNF-{\alpha}$ was analyzed by RT-PCR. Results : OB extract significantly inhibited the LPS-induced production of NO and TNF-a in BV2 cells. OB extract also suppressed the mRNA expression of iNOS and $TNF-{\alpha}$ in BV2 cells activated with LPS. Conclusion : These data suggests that OB extract may have the anti-inflammatory effect through the modulation of NO production and inflammatory cytokine such as $TNF-{\alpha}$.

• The Journal of Internal Korean Medicine
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• v.29 no.1
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• pp.243-257
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• 2008

Myostatin, a negative regulator of myogenesis, is shown to function by controlling the proliferation of myoblasts. In this study we show that myostatin is an inhibitor of myoblast differentiation and that this inhibition is mediated through Smad 3. To determine MyoD expression by Dodamtang treatment, we compared the expression pattern of $C_{2}C_{12}$ mouse myoblasts that constitutively express myostatin with control cells. In vitro, increasing concentrations of Dodamtang reversibly prevented the myogenic blockage of myoblasts by myostatin expression. ELISA assay, Western and confocal analysis indicated that treatment of Dodamtang to the low serum culture media increased the levels of MyoD leading to the inhibition of myogenic differentiation by myostatin. The stable transfection of $C_{2}C_{12}$ myoblasts with myostatin expressing constructs did rescue MyoD-induced myogenic differentiation. Consistent with this, the treatment of Dodamtang rescued the expression of a MyoD in $C_{2}C_{12}$ myoblasts treated with myostatin. Taken together, these results suggest that induction of MyoD by Dodamtang inhibits myostatin activity and expression via SMAD3 resulting in the rescue of the myoblasts to differentiate into myotubes. Thus we propose that myostatin action by Dodamtang plays a critical role in myogenic differentiation and that the muscular hyperplasia and hypertrophy seen in animals that blockage of functional myostatin is because of deregulated proliferation and differentiation of myoblasts.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.12
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• pp.7719-7724
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• 2013

Background: Overexpression of survivin, a known inhibitor of apoptosis, is associated with tumor progression and drug resistance in numerous malignancies, including leukemias. The aim of this study was to investigate the effect of a specific survivin small interference RNA (siRNA) on proliferation and the sensitivity of HL-60 acute myeloid leukemia (AML) cells to the chemotherapeutic drug etoposide. Materials and Methods: The cells were transfected with siRNAs using Lipofectamine $^{TM}2000$ transfection reagent. Relative survivin mRNA and protein levels were measured by quantitative real-time PCR and Western blotting, respectively. Trypan blue exclusion assays were performed to monitor tumor cell proliferation after siRNA transfection. The cytotoxic effects of etoposide and survivin siRNA, alone and in combination, on leukemic cells were determined using MTT assay. Apoptosis was assessed by ELISA cell death assay. Results: Survivin siRNA markedly reduced both mRNA and protein expression levels in a time-dependent manner, leading to distinct inhibition of cell proliferation and increased spontaneous apoptosis. Surprisingly, survivin siRNA synergistically increased the cell toxic effects of etoposide. Moreover, survivin down-regulation significantly enhanced its induction of apoptosis. Conclusions: Our study suggests that down-regulation of survivin by siRNA can trigger apoptosis and overcome drug resistance of leukemia cells. Therefore, survivin siRNA may be an effective adjuvant in AML chemotherapy.

• Journal of Pharmacopuncture
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• v.11 no.1
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• pp.119-125
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• 2008

Objective : This experimental study was performed to investigate the effects of Houttuyniae Herba extract on anti-inflammation and anti-oxidation. Methods : The cytotoxicity of Houttuyniae Herba water extract and ethanol extract about viability of Raw 264.7 cell were tested using a colormetric tetrazolium assay(MTT assay). We investigated the inhibitory effects of Houttuyniae Herba water extract and ethanol extract on Propionibactrium acnes using paper disk diffusion method. To investigate the anti-inflammation effects of Houttuyniae Herba water extract and ethanol extract on LPS-induced macrophage Raw 264.7 cell, we used ELISA kit. We evaluated anti-oxidation effects of Houttuyniae Herba water extract and ethanol extract on HaCaT cell by Enzyme recycling method. Results : 1. In Houttuyniae Herba water extract and ethanol extract, cell toxicity depended on the density and wasn't difference between two extracts. 2. Houttuyniae Herba water extract and ethanol extract has not the significant inhibition effect of Propionibactrium acnes. 3. Concentration of 50, $100{\mu}g/m{\ell}$ Houttuyniae Herba water extract inhibited the production of NO in the Raw 264.7 cell stimulated with LPS. 4. All extracts except for $20{\mu}g/m{\ell}$ Houttuyniae Herba water extract showed anti-oxidation effect by decreasing the DPPH radicals. Conclusion : These results indicate that Houttuyniae Herba extract has anti-inflammation and anti-oxidation effects. If further study is performed, the use of Houttuyniae Herba extract will be valuable and benificial in the therapy of Propionibactrium acnes.

• Herbal Formula Science
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• v.27 no.4
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• pp.245-255
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• 2019

Objective : Herbal processing is one of the traditional techniques used in Korean medicine to increase the effectiveness of herbs or reduce their toxicity. In this study, Gardeniae Fructus processed with ginger juice and alcohol was prepared to evaluate the anti-inflammatory effect on lipopolysaccharide (LPS)-induced macrophages. Methods : The processing of Gardeniae Fructus was performed by adding 40 % ginger juice or 10% alcohol to the total weight of Gardeniae Fructus and then roasting at 150℃ for 5 minutes. Cell viability was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. To detect nitric oxide (NO) production, culture media were mixed with Griess reagent and measured the absorbance at 540 nm. Prostaglandin E₂ (PGE₂) and pro-inflammatory cytokines were detected by enzyme-linked immunosorbent assay (ELISA). Western blot was applied to monitor protein expression levels. Results : LPS-induced NO, PGE₂ and inflammatory cytokines were decreased by the treatment of normal or processed Gardeniae Fructus ethanol extracts (GFE). Compared to normal GFE, the processed GFE showed a stronger inhibitory effect on the production of NO and PGE₂. These inhibitory effect of GFE was due to the suppression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) mediated from the inhibition of nuclear factor kappa B (NF-κB). Furthermore, processed GFE showed more suppressive effects on the expression of iNOS, COX-2 and IκBα proteins than normal GFE. Conclusion : From these results, it was concluded that GFE had an improved anti-inflammatory effect compared to normal GFE. These results provide an objective evidences for the use of herbal processing in Korean medicine.

• Archives of Pharmacal Research
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• v.26 no.12
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• pp.1029-1035
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• 2003

Osteoporosis is recognized as one of the major hormonal deficiency diseases, especially in menopausal women and the elderly. When estrogen is reduced in the body, local factors such as IL-1 $\beta$ and IL-6, which are known to be related with bone resorption, are increased and promote osteoclastogenesis, which is responsible for bone resorption. In the present study, we investigated whether glucosidic isoflavones (Isocal, PIII) extracted from Sophorae fructus affect the proliferation of osteoblasts and prevent osteoclastogenesis in vitro by attenuating upstream cytokines such as IL-1$\beta$ and IL-6 in a human osteoblastic cell line (MG-63) and in a primary osteoblastic culture from SD rat femurs. Interestingly, IL-1$\beta$ and IL-6 mRNA were significantly suppressed in osteoblast-like cells treated with 17$\beta$-estradiol (E2) and PIII when compared to positive control (SDB), and this suppression was more effective at $10^{-8}$% than at the highest concentration of $10^{-4}$%. In addition, these were confirmed in protein levels using ELISA assay. In the cell line, the cells showed that E2 was the most effective in osteoblastic proliferation over the whole range of concentration ($10^{-4}%-10^{-12}$%), even though PIII also showed the second greatest effectiveness at $10^{-8}$%. Nitric oxide (NO) was significantly (p<0.05) upregulated in PIII and E2 over the concentration range $10^{-6}% to 10^{-8}$% when compared to SDB, without showing any dose dependency. In bone marrow primary culture, we found by TRAP assay that PIII effectively suppressed osteoclastogenesis next to E2 in comparison with SDB and culture media (control). In conclusion, these results suggest that local bone-resorbing cytokines can be regulated by PIII at lower concentrations and that, therefore, PIII may preferentially induce anti-osteoporosis response by attenuating osteoclastic differentiation and by upregulating NO.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.36 no.1
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• pp.1-20
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• 2023

Objectives : The purpose of this study was to evaluate the anti-inflammatory effect of Tanghwajitongsan(THJTS) through inhibition of NF-κB and MAPK. Methods : We evaluated cell survival rate by MTT assay, NO production by nitrite content in the culture medium. We quantified TNF-α, IL-1β, IL-6 and PGE₂ of the cultured supernatant by ELISA. And we evaluated the effect of THJTS on protein expression of NF-κB, MAPK, iNOS and COX-2 by Western blot analysis. THJTS ameliorates LPS-activated alterations in protein expression of NF-κB, p-38, iNOS and COX-2 and production of NO, pro-inflammatory cytokines and PGE₂. Also, THJTS ameliorates LOX, PGN and FLA-activated alterations in protein expression of NO, iNOS. THJTS ameliorates only PGN-activated alterations in protein expression of COX-2. Results : THJTS ameliorates LPS-activated alterations in protein expression of NF-κB, p-38, iNOS and COX-2 and production of NO, pro-inflammatory cytokines and PGE₂. Also, THJTS ameliorates LOX, PGN and FLA-activated alterations in protein expression of NO, iNOS. THJTS ameliorates only PGN-activated alterations in protein expression of COX-2. Conclusions : This study can provide scientific evidence for the anti-inflammatory effects and underlying mechanisms of THJTS.