• Journal of the Society of Cosmetic Scientists of Korea
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• v.42 no.1
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• pp.75-85
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• 2016

Pectin, a naturally occurring polysaccharide, has in recent years attracted considerable attention. Its benefits are increasingly appreciated by scientists and consumers due to its safety and usefulness. The chemistry and gel-forming characteristics of pectin have enabled to be used in pharmaceutical industry, health promotion and treatment. Yet, it has been rarely used in cosmetics because of its incompatibility with many cosmetic ingredients, including alcohols, and unstable viscosity of pectin gels under various pH and salt conditions. However, low-molecular-weight pectin oligomers have excellent biological activities, and depolymerization of pectin to produce cosmetic ingredients would be very useful. In this study, we attempted the development of cosmetic ingredients using pectin with an excellent effect on human skin. We developed a bio-conversion process that uses enzymatic hydrolysis to produce pectin hydrolysates containing mainly low-molecular-weight pectin oligomers. Gel permeation chromatography was used to determined the ratio of hydrolysis. The molecular weight of the pectin hydrolysates obtained varied between 200 and 2,700 Da. The two newly developed low-molecular-weight pectin hydrolysates, LMPH A and B, had higher anti-oxidative activities than pectin or D-galacturonic. Exposure to UVB radiation induces apoptotic cell death in epidermal cells. Annexin V binding and propidium iodide uptake were measured by flow cytometry to evaluate UVB-induced cell death in HaCaT cells. Both LMPH A and B reduced UVB-induced cell death and increased cell proliferation by 22% and 30% at 0.5% concentration respectively, while pectin had no significant activity. In conclusion, this study suggests that the newly developed low-molecular-weight pectin hydrolysates can be used as safe and biologically active cosmetic ingredients.

• Journal of Ginseng Research
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• v.44 no.6
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• pp.815-822
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• 2020

Background: Recently, beneficial roles of ginsenoside F2 (GF2), a minor constituent of Panax ginseng, have been demonstrated in diverse inflammatory diseases. However, its roles in alcoholic liver inflammation and injury have not been clearly understood. Here, we investigated the underlying mechanism by which GF2 ameliorated alcoholic liver injury. Methods: To induce alcoholic liver injury, C57BL/6J wild type (WT) or interleukin (IL)-10 knockout (KO) mice were orally administered with ethanol (3 g/kg) or ethanol-containing GF2 (50 mg/kg) for 2 wk. Liver injury and infiltration of macrophages and neutrophils were evaluated by serum biochemistry and immunohistochemistry, respectively. The changes of hepatic immune cells were assessed by flow cytometry and polymerase chain reaction analysis. In vitro differentiation of naïve T cells was performed. Results: GF2 treatment significantly attenuated alcoholic liver injury, in which infiltrations of inflammatory macrophages and neutrophils were decreased. Moreover, the frequencies of Foxp3⁺ regulatory T cells (Tregs) increased but IL-17-producing T (Th17) cells decreased in GF2-treated mice compared to controls. Furthermore, the mRNA expression of IL-10 and Foxp3 was significantly increased, whereas IL-17 mRNA expression was suppressed in GF2-treated mice. However, these beneficial roles of GF2 were not observed in GF2-treated IL-10 KO mice, suggesting a critical role of IL-10. Similarly, GF2 treatment suppressed differentiation of naïve T cells into Th17 cells by inhibiting RORgt expression and stimulating Foxp3 expression. Conclusion: The present study suggests that GF2 treatment attenuates alcoholic liver injury by increasing IL-10 expression and Tregs and decreasing IL-17 expression and Th17 cells.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.4
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• pp.299-305
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• 2010

Purpose: Adipose tissue is located beneath the skin, around internal organs, and in the bone marrow in humans. Its main role is to store energy in the form of fat, although it also cushions and insulates the body. Adipose tissue also has the ability to dynamically expand and shrink throughout the life of an adult. Recently, it has been shown that adipose tissue contains a population of adult multipotent mesenchymal stem cells and endothelial progenitor cells that, in cell culture conditions, have extensive proliferative capacity and are able to differentiate into several lineages, including, osteogenic, chondrogenic, endothelial cells, and myogenic lineages. Materials and Methods: This study focused on endothelial cell culture from the adipose tissue. Adipose tissues were harvested from buccal fat pad during bilateral sagittal split ramus osteotomy for surgical correction of mandibular prognathism. The tissues were treated with 0.075% type I collagenase. The samples were neutralized with DMEM/and centrifuged for 10 min at 2,400 rpm. The pellet was treated with 3 volume of RBC lysis buffer and filtered through a 100 ${\mu}m$ nylon cell strainer. The filtered cells were centrifuged for 10 min at 2,400 rpm. The cells were further cultured in the endothelial cell culture medium (EGM-2, Cambrex, Walkersville, Md., USA) supplemented with 10% fetal bovine serum, human EGF, human VEGF, human insulin-like growth factor-1, human FGF-$\beta$, heparin, ascorbic acid and hydrocortisone at a density of $1{\times}10^5$ cells/well in a 24-well plate. Low positivity of endothelial cell markers, such as CD31 and CD146, was observed during early passage of cells. Results: Increase of CD146 positivity was observed in passage 5 to 7 adipose tissue-derived cells. However, CD44, representative mesenchymal stem cell marker, was also strongly expressed. CD146 sorted adipose tissue-derived cells was cultured using immuno-magnetic beads. Magnetic labeling with 100 ${\mu}l$ microbeads per 108 cells was performed for 30 minutes at $4^{\circ}C$ a using CD146 direct cell isolation kit. Magnetic separation was carried out and a separator under a biological hood. Aliquous of CD146+ sorted cells were evaluated for purity by flow cytometry. Sorted cells were 96.04% positivity for CD146. And then tube formation was examined. These CD146 sorted adipose tissue-derived cells formed tube-like structures on Matrigel. Conclusion: These results suggest that adipose tissue-derived cells are endothelial cells. With the fabrication of the vascularized scaffold construct, novel approaches could be developed to enhance the engineered scaffold by the addition of adipose tissue-derived endothelial cells and periosteal-derived osteoblastic cells to promote bone growth.

• Biomedical Science Letters
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• v.2 no.2
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• pp.167-174
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• 1996

Alcoholics increased susceptibility to microbial infection that is associated with decreased immunity. but there has been little experimental evidence to support alcoholics-induced increase of microbial infection directly in non-specific immunity. Therefore, we were used the method of phagocytic-plaque including all the stimulating factors for the phagocytosis, subtypes of lymphocytes and T-lymphocyte proliferation. The experimental groups were divided into 3 groups: (1) alcoholics who were hospitalized less than 1 week (newly hospitalized alcoholics), (2) alcoholics who were hospitalized more than 2 weeks (old hospitalized alcoholics), (3) healthy blood donors. We have studied 98 alcoholics and 35 healthy blood donors and control groups. A physician has checked the biological markers and diagnosed the body-condition alcoholics. The immunity and non-specific immunity on the alcoholics were analyzed by using the simultest kit and flow cytometry. Proliferation of the lymphocytes was analyzed by the phytohemmagglutinine mitogen. Phagocytosis and migration properties of leukocytes were identified on the layer formed by Staphylococcus aureus Cowan I strain. Biological markers of alcoholics and control groups, by such as blood glucose, ${\gamma}$-glutamyl transpeptidase and mean corpuscular volumes of red blood cells, were determined by biochemical and hematological methods. Compared with control groups, cluster of differentiation (CD)3+, CD8+ and CD19+ in alcoholic were more decreased except CD4+/CD8+ ratio. Proliferation of the T-lymphocytes, phagocytosis and migration properties of the leukocytes in alcoholics were decreased compared with those of control groups. According to the results observed in our experiment, they can be summerized as follows: 1, Cellular, humoral and non-specific immunities, are markedly decreased in alcoholics than those in control groups. 2. It is inferred that Phagocytic plaque formation is a very useful method to evaluate phagocytosis and migration properties of the alcoholic leukocytes 3. It is thought that the subtypes of lymphocytes, especially CD4+/CD8+ ratio, are essential methods to analyzed the alcoholic immunity. 4. Specific and non-specific immunity on the old hospitalized alcoholics was slightly increased, which depends upon the alcoholic medication.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.18
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• pp.7849-7855
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• 2014

Purpose: To investigate the effect of deacetylase inhibitory trichostatin A (TSA) on anti HepG2 liver carcinoma cells and explore the underlying mechanisms. Materials and Methods: HepG2 cells exposed to different concentrations of TSA for 24, 48, or 72h were examined for cell growth inhibition using CCK8, changes in cell cycle distribution with flow cytometry, cell apoptosis with annexin V-FTIC/PI double staining, and cell morphology changes under an inverted microscope. Expression of ${\beta}$-catenin, HDAC1, HDAC3, H3K9, CyclinD1 and Bax proteins was tested by Western blotting. Gene expression for ${\beta}$-catenin, HDAC1and HDAC3 was tested by q-PCR. ${\beta}$-catenin and H3K9 proteins were also tested by immunofluorescence. Activity of Renilla luciferase (pTCF/LEF-luc) was assessed using the Luciferase Reporter Assay system reagent. The activity of total HDACs was detected with a HDACs colorimetric kit. Results: Exposure to TSA caused significant dose-and time-dependent inhibition of HepG2 cell proliferation (p<0.05) and resulted in increased cell percentages in G0/G1 and G2/M phases and decrease in the S phase. The apoptotic index in the control group was $6.22{\pm}0.25%$, which increased to $7.17{\pm}0.20%$ and $18.1{\pm}0.42%$ in the treatment group. Exposure to 250 and 500nmol/L TSA also caused cell morphology changes with numerous floating cells. Expression of ${\beta}$-catenin, H3K9and Bax proteins was significantly increased, expression levels of CyclinD1, HDAC1, HDAC3 were decreased. Expression of ${\beta}$-catenin at the genetic level was significantly increased, with no significant difference in HDAC1and HDAC3 genes. In the cytoplasm, expression of ${\beta}$-catenin fluorescence protein was not obvious changed and in the nucleus, small amounts of green fluorescence were observed. H3K9 fluorescence protein were increased. Expression levels of the transcription factor TCF werealso increased in HepG2 cells following induction by TSA, whikle the activity of total HDACs was decreased. Conclusions: TSA inhibits HDAC activity, promotes histone acetylation, and activates Wnt/${\beta}$-catenin signaling to inhibit proliferation of HepG2 cell, arrest cell cycling and induce apoptosis.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.7
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• pp.4421-4426
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• 2013

Objective: The present study employed 5-aza-2'-deoxycytidine (5-Aza-CdR) to treat non-small cell lung cancer (NSCLC) cell line A549 to investigate the effects on proliferation and expression of the TFPI-2 gene. Methods: Proliferation was assessed by MTT assay after A549 cells were treated with 0, 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR, a specific demethylating agent, for 24, 48 and 72h. At the last time point cells were also analyzed by flow cytometry (FCM) to identify any change in their cell cycle profiles. Methylation-specific polymerase chain reaction (MSPCR), real time polymerase chain reaction(real-time PCR) and western blotting were carried out to determine TFPI-2 gene methylation status, mRNA expression and protein expression. Results: MTT assay showed that the growth of A549 cells which were treated with 5-Aza-CdR was significantly suppressed as compared with the control group (0 ${\mu}mol/L$ 5-Aza-CdR). After treatment with 0, 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR for 72h, FCM showed their proportion in G0/G1 was $69.7{\pm}0.99%$, $76.1{\pm}0.83%$, $83.8{\pm}0.35%$, $95.5{\pm}0.55%$ respectively (P<0.05), and the proportion in S was $29.8{\pm}0.43%$, $23.7{\pm}0.96%$, $15.7{\pm}0.75%$, $1.73{\pm}0.45%$, respectively (P<0.05), suggesting 5-Aza-CdR treatment induced G0/G1 phase arrest. MSPCR showed that hypermethylation in the promoter region of TFPI-2 gene was detected in control group (0 ${\mu}mol/L$ 5-Aza-CdR), and demethylation appeared after treatment with 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR for 72h. Real-time PCR showed that the expression levels of TFPI-2 gene mRNA were $1{\pm}0$, $1.49{\pm}0.14$, $1.86{\pm}0.09$ and $5.80{\pm}0.15$ (P<0.05) respectively. Western blotting analysis showed the relative expression levels of TFPI-2 protein were $0.12{\pm}0.01$, $0.23{\pm}0.02$, $0.31{\pm}0.02$, $0.62{\pm}0.03$ (P<0.05). TFPI-2 protein expression in A549 cells was gradually increased significantly with increase in the 5-Aza-CdR concentration. Conclusions: TFPI-2 gene promoter methylation results in the loss of TFPI-2 mRNA and protein expression in the non-small cell lung cancer cell line A549, and 5-Aza-CdR treatment could induce the demethylation of TFPI-2 gene promoter and restore TFPI-2 gene expression. These findings provide theoretic evidence for clinical treatment of advanced non-small cell lung cancer with the demethylation agent 5-Aza-CdR. TFPI-2 may be one molecular marker for effective treatment of advanced non-small cell lung cancer with 5-Aza-CdR.

• Journal of Chest Surgery
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• v.43 no.5
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• pp.499-505
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• 2010

Background: There have been several reports using animal experiments that CD1-restricted T-cells have a key role in tumor immunity. To address this issue, we studied the expression of markers for CD1c+ myeloid dendritic cells (DCs) isolated from peripheral blood in the clinical setting. Material and Method: A total of 24 patients with radiologically suspected or histologically confirmed lung cancer who underwent pulmonary resection were enrolled in this study. The patients were divided according to histology findings into three groups: primary adenocarcinoma of lung (PACL), primary squamous cell carcinoma of lung (PSqCL) and benign lung disease (BLD). We obtained 20 mL of peripheral venous blood from patients using heparin-coated syringes. Using flow-cytometry after labeling with monoclonal antibodies, data acquisition and analysis were done. Result: The ratio of CD1c+CD19- dendritic cells to CD1c+ dendritic cells were not significantly different between the three groups. CD40 (p=0.171), CD86 (p=0.037) and HLA-DR (p=0.036) were less expressed in the PACL than the BLD group. Expression of CD40 (p=0.319), CD86 (p=0.036) and HLA-DR (p=0.085) were less expressed in the PACL than the PSqCL group, but the differences were only significant for CD86. Expression of co-stimulatory markers was not different between the PSqCL and BLD groups. Expression of markers for activated DCs were dramatically lower in the PACL group than in groups with other histology (CD40 (p=0.005), CD86 (p=0.013) HLA-DR (p=0.004). Conclusion: These results suggest the possibility that CD1c+ myeloid DCs participate in control of the tumor immunity system and that low expression of markers results in lack of an immune response triggered by dendritic cells in adenocarcinoma of the lung.

• Tuberculosis and Respiratory Diseases
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• v.41 no.4
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• pp.354-362
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• 1994

Objectives and Methods : The presence of aneuploidy or high proliferative activity in cytologic specimens is considered as complementary for the diagnosis of malignancy. To evaluate the diagnostic usefulness of DNA ploidy and cell cycle analysis in lung cancer, we compared the diagnostic yielding rates of DNA ploidy test by brushing specimens using flow cytometry with bronchoscopic forceps biopsy and brushing cytology. Results : Of the seventy-six cases, 55 cases proved to have malignant diseases(squamous cell cancer: 27, adenocarcinoma: 7, large cell cancer: 1, undifferentiated: 4 and small cell cancer: 16). The incidence of aneuploidy in lung cancer patients was 32.7%(18/55), as opposed to no cases in benign disease. And the proportion of high proliferative activity(S+G2M>22%) in lung cancer patients was 42.9%(15/35), but none in benign diseases. In fifty-six of 75 cases(74.7%), cytology of brushing specimens and DNA analysis(either aneuploidy or high proliferative activity vs. diploidy and low proliferative activity) were in concordance. The sensitivity with only brushing cytology was 41.8%(23/55), but with the addition of DNA analysis, it was increased to 56.4%(31/55), without decreasing the specificity(100%). And there was a case whose clue for malignancy was absent except aneuploidy, and he was confirmed to have squamous cell cancer following open thoracotomy. There were no differences in the frequency of aneuploidy or high proliferative activity between histologic subtypes of bronchogenic malignancy. Conclusions : The diagnostic detection rate of lung cancer was improved with the addition of DNA ploidy and cell cycle analysis, and the presence of aneuploidy or high proliferative activity was a relatively specific indicator of malignant disease. It would be useful to test DNA ploidy and cell cycle analysis with brushing specimen for the diagnosis of bronchogenic malignancy particularly in patients whose biopsy specimen could not be obtainable.

• Journal of Life Science
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• v.21 no.7
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• pp.961-968
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• 2011

The formation of reactive lipid aldehydes, 4-hydroxynonenal (HNE) is shown to be derived from fatty acid hydroperoxides through the oxidative process. Among its known effects in cytotoxicity, HNE has been implicated in apoptotic cell death. To delineate its putative role as a potential mediator, we investigated the mechanism by which HNE induces apoptosis of endothelial cells (ECs). The anti-proliferative effects of HNE were tested through MTT assay after exposure to various concentrations ($5\sim15\;{\mu}M$) of HNE. We observed apoptotic bodies with propidium iodide staining, and measured the HNE induction of endothelial apoptosis by flow cytometry assay. We observed that cells exposed to HNE for 24 hr resulted in increased poly(ADP-ribose) polymerase cleavage and up-regulation of Bax. Data on the HNE action strongly indicated the involvement of reactive species, namely, intracellular ROS, nitrite, and peroxynitrite. To obtain evidence on the implication of ROS and peroxynitrite in HNE-induced apoptosis, a ROS scavenger, N-acetylcysteine (NAC), and a peroxynitrite scavenger, penicillamine, were tested. Results clearly indicate that the induction of apoptosis by HNE was effectively inhibited by NAC and penicillamine. Based on the present data, we conclude that the endothelial apoptosis induced by HNE involves both ROS generation and peroxynitrite activity. Our new data could lead to a redefinition of HNE action on apoptosis in ECs.

• Journal of Life Science
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• v.27 no.2
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• pp.225-232
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• 2017

This study aimed to evaluate several biological activities of Pharbitis nil and to isolate an anticancer agent from its methanol extract. Pharbitis nil seeds were extracted with methanol (PNM). Then, PNM was fractionated into solvent layers such as ethyl acetate fraction (PNE), butanol fraction (PNB), and water fraction (PNW). The biological activities of the fractions were analyzed for tyrosinase inhibition, lipase inhibition, DPPH-free radical scavenging, and cell growth inhibition. PNM showed strong growth inhibition of prostate cancer PC-3 cells. PNM was subjected to Diaion HP-20 and eluted stepwise with 50%, 80%, and 100% methanol. Then, for activity-guided fraction, each fraction was analyzed for growth inhibition of prostate cancer PC-3 cells by using an MTT assay. Because the 100% fraction showed significantly strong inhibitory activity, the fraction was further separated in the reverse phase C18, which was eluted with 80% and 90% methanol. The 90% fraction was further subjected to Sephadex LH-20 using a mobile solvent of 100% methanol. Finally, the compound PN was partially purified for HPLC analysis. PN showed cell growth inhibitory activity and induced the apoptosis and cell cycle arrest of prostate cancer PC-3 cells, as measured by flow cytometry. The results together suggest that Pharbitis nil possesses various biological activities, especially the inhibitory activity for the proliferation of prostate cancer PC-3 cells, suggesting the possibility of its use as an anticancer agent.