• Asian Pacific Journal of Cancer Prevention
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• v.14 no.12
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• pp.7339-7344
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• 2013

To analyze the effects of a new unknown peptide DEF on the growth of tumor cells, a fused polypeptide TAT-DV1-DEF was designed and synthesized. The lung adenocarcinoma cell line GLC-82 treated with TAT-DV1-DEF was analyzed with a cell counting kit 8, and the location of polypeptides in cells was observed under laser confocal microscopy. The efficiency of polypeptide transfection and changes in nuclear morphology were analyzed by flow cytometry and fluorescence microscopy, respectively. Finally, the mechanism of tumor cell growth inhibition was evaluated by Western blotting. We found that TAT-DV1-DEF could significantly inhibit the growth of the lung adenocarcinoma cell line GLC-82, but not the normal human embryonic kidney cell line HEK-293. Polypeptides were found to be mostly localized in the cytoplasm and some mitochondria. The efficiency of polypeptide transfection in the two cell types was approximately 99%. Apoptotic nuclei were observed under fluorescence microscopy upon treatment with polypeptides and DAPI staining. Western blot analyses indicated that the polypeptide inhibition of tumor cell growth was apoptosis dependent. In the present study, we demonstrated that fused polypeptides could induce apoptosis of the lung adenocarcinoma cell line GLC-82, indicating that the new unknown peptide DEF has antitumor effects.

• The Korean Journal of Physiology and Pharmacology
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• v.22 no.2
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• pp.145-153
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• 2018

The subgranular zone (SGZ) of hippocampal dentate gyrus (HDG) is a primary site of adult neurogenesis. Toll-like receptors (TLRs), are involved in neural system development of Drosophila and innate immune response of mammals. TLR2 is expressed abundantly in neurogenic niches such as adult mammalian hippocampus. It regulates adult hippocampal neurogenesis. However, the role of TLR2 in adult neurogenesis is not well studied in global or focal cerebral ischemia. Therefore, this study aimed to investigate the role of TLR2 in adult neurogenesis after photochemically induced cerebral ischemia. At 7 days after photothrombotic ischemic injury, the number of bromodeoxyuridine (BrdU)-positive cells was increased in both TLR2 knock-out (KO) mice and wild-type (WT) mice. However, the increment rate of BrdU-positive cells was lower in TLR2 KO mice compared to that in WT mice. The number of doublecortin (DCX) and neuronal nuclei (NeuN)-positive cells in HDG was decreased after photothrombotic ischemia in TLR2 KO mice compared to that in WT mice. The survival rate of cells in HDG was decreased in TLR2 KO mice compared to that in WT mice. In contrast, the number of cleaved-caspase 3 (apoptotic marker) and the number of GFAP (glia marker)/BrdU double-positive cells in TLR2 KO mice were higher than that in WT mice. These results suggest that TLR2 can promote adult neurogenesis from neural stem cell of hippocampal dentate gyrus through increasing proliferation, differentiation, and survival from neural stem cells after ischemic injury of the brain.

• Journal of Physiology & Pathology in Korean Medicine
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• v.23 no.2
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• pp.405-411
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• 2009

This study evaluated the effects of Puerariae Radix on the skeletal muscle atrophy, Muscle atrophy was induced by the sciatic nerve transection in Sprague-Dawley rats, then aqueous-extract of Puerariae Radix was administered for 12 days, Muscle wet weight was measured in soleus, plantaris, and medial gastrocnemius. Muscle fiber type was classified by MHCf immunohistochemistry. Muscle fiber type proportion and cross section area of muscle fiber also was observed in medial gastrocnemius. Bax and Bcl-2 expressions in medial gastrocnemius of the damaged hind limb were evaluated with immunohistochemistry. The results are as follows; Puerariae Radix attenuated muscle atrophy in soleus of the sciatic nerve transectioned rats, but there was statistic significance. Puerariae Radix attenuated significantly atrophy in plantaris at 12 days and in medial gastrocnemius at 8 days and 12 days. Puerariae Radix improved histology of the atrophic changes and increased significantly cross section areas of type-I and type-II muscle fibers in medial gastrocnemius of the sciatic nerve transectioned rats. Puerariae Radix did not affect to muscle fiber type proportion in medial gastrocnemius of the sciatic nerve transectioned rats. Puerariae Radix attenuated significantly Bax positive nuclei but did not affect to Bcl-2 positive muscle fibers in medial gastrocnemius of the sciatic nerve transectioned rats.According to above results, Puerariae Radix may have an anti-atrophy effect on the denervated skeletal muscle through anti-apoptotic effects on muscle fibers.

• Archives of Pharmacal Research
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• v.27 no.3
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• pp.305-313
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• 2004

The effect of diazoxide, a $K^{+}$channel opener, on apoptotic cell death was investigated in HepG2 human hepatoblastoma cells. Diazoxide induced apoptosis in a dose-dependent manner and this was evaluated by flow cytometric assays of annexin-V binding and hypodiploid nuclei stained with propidium iodide. Diazoxide did not alter intracellular $K^{+}$concentration, and various inhibitors of $K^{+}$channels had no influence on the diazoxide-induced apoptosis; this implies that $K^{+}$channels activated by diazoxide may be absent in the HepG2 cells. However, diazoxide induced a rapid and sustained increase in intracellular $Ca^{2+}$ concentration, and this was completely inhibited by the extracellular $Ca^{2+}$ chelation with EGTA, but not by blockers of intracellular $Ca^{2+}$ release (dantrolene and TMB-8). This result indicated that the diazoxide-induced increase of intracellular $Ca^{2+}$ might be due to the activation of a Ca2+ influx pathway. Diazoxide-induced $Ca^{2+}$ influx was not significantly inhibited by either voltage-operative $Ca^{2+}$ channel blockers (nifedipinen or verapamil), or by inhibitors of $Na^{+}$, $Ca^{2+}$-exchanger (bepridil and benzamil), but it was inhibited by flufenamic acid (FA), a $Ca^{2+}$-permeable nonselective cation channel blocker. A quantitative analysis of apoptosis by flow cytometry revealed that a treatment with either FA or BAPTA, an intracellular $Ca^{2+}$ chelator, significantly inhibited the diazoxide-induced apoptosis. Taken together, these results suggest that the observed diazoxide-induced apoptosis in the HepG2 cells may result from a $Ca^{2+}$ influx through the activation of $Ca^{2+}$-permeable non-selective cation channels. These results are very significant, and they lead us to further suggest that diazoxide may be valuable for the therapeutic intervention of human hepatomas.

• BMB Reports
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• v.36 no.4
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• pp.337-343
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• 2003

Rhus verniciflua Stokes (RVS) has been used as a traditional herbal medicine. Several earlier studies indicated that an ethanol extract of RVS has both anti-oxidant and anti-tumor properties, although the mechanism for the activity remains to be elucidated. In this report, we prepared a highly purified ethanol extract from RVS, named REEE-1 ($\underline{R}$hus $\underline{e}$thanol $\underline{e}$luted $\underline{e}$xtract-1), and investigated the mechanism involved in its growth-inhibitory effect on the human B and T lymphoma cell lines, BJAB and Jurkat, respectively. Results from tritium uptake proliferation assays showed that the proliferative capacities of both BJAB and Jurkat cells were strongly suppressed in the presence of REEE-1. This was further confirmed through trypan blue exclusion experiments that revealed a dose-dependent decrease in viable cell numbers after REEE-1 treatment. REEE-1-mediated suppression of cell growth was verified to be apoptotic, based on the increase in DNA fragmentation, low fluorescence intensity in nuclei after propidium iodide staining, and the appearance of DNA laddering. In particular, REEE-1 exerted its anti-oxidant activity through the inhibition of hydroxyl radical-mediated degradation by iron ion chelation rather than direct scavenging of hydroxyl radicals. Furthermore, REEE-1 was revealed to be a potential scavenger of superoxide anions. Collectively, our findings suggest that REEE-1 is a natural anti-oxidant that could be used as a cancer chemo-preventive and therapeutic agent.

• International Journal of Oral Biology
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• v.43 no.2
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• pp.69-76
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• 2018

Oral squamous cell carcinoma (OSCC) is the most common type of oral malignancy. Numerous therapies have been proposed for its cure. Research is continually being conducted to develop new forms of treatment as current therapies are associated with numerous side-effects. Luteolin, a common dietary flavonoid, has been demonstrated to possess strong anti-cancer activity against various human cancer cell lines. Nevertheless, research into luteolin-based anticancer activity against oral cancer remains scarce. Thus, the objective of this study was to assess the effect of luteolin as an anti-cancer agent. After treatment with luteolin, Ca9-22 and CAL-27 oral cancer cells showed condensed nuclei and enhanced apoptotic rate with evidence of mitochondria-mediated apoptosis. Epithelialmesenchymal transition (EMT) is closely related to tumor migration and invasion. Luteolin suppressed cancer cell invasion and migration in the current study. Elevated expression of E-cadherin, an adherens junction protein, was evident in both cell lines after luteolin treatment. Luteolin also significantly inhibited transcription factors (i.e., N-cadherin, Slug, Snail, Twist, and ZEB-1) that regulated expression of tumor suppressors such as E-cadherin based on Western blot analysis and quantitative PCR. Thus, luteolin could induce mitochondrial apoptosis and inhibit cancer cell invasion and migration by suppressing EMT-induced transcription factors.

• The Journal of Internal Korean Medicine
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• v.31 no.2
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• pp.273-289
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• 2010

Objectives : To investigate the anti-cancer effect of Baekduong-tang(BDOT) against cancer cells, the signaling pathway of apoptosis was explored in human colon cancer cells. Materials and Methods : Human colon cancer cell lines, including HT-29 and HCT-116 cells, were used. Cell viability was measured by MTT assay. Apoptosis was determined by DAPI nuclei staining and flow cytometry in HCT-116 cells treated with 0.25 mg/$m{\ell}$ Baekduong-tang for 48 hrs. Results : Baekduong-tang induced the apoptosis of p53 positive HCT-116 cells with G2/M phase arrest. Treatment with Baekduong-tang led to increased expression and phosphorylation of p53 and decreased expression of CDK2 and CDK6 in HCT-116 cells. It also activated caspase-3 through caspase-10 and caspase-9 activation. Finally, Baekduong-tang induced production $H_2O_2$, superoxide anion ($O_2^-$) and NO and modulated proteins expression including SOD, NOS, Bax and Bcl-2. Conclusions : These results indicate Baekduong-tang induces apoptotic death of HCT-116 cells through G2/M phase arrest and disturbance of intracellular redox status in a p53-dependent manner.

• Korean Journal of Medicinal Crop Science
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• v.13 no.2
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• pp.95-102
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• 2005

Caulis Bambusae in Taenia is widely used in Korea and China due to its various pharmacological activity. The present study aims to investigate the effect of the methanol extract of Caulis Bambusae in Taenia (CB) from Phyllostachys nigra Munro var. henonis Stapf (Gramineae) on amyloid ${\beta}$ protein (25-35) $(A{\beta}\;(25-35))$, a synthetic 25-35 amyloid peptide, -induced neurotoxicity using cultured rat cortical neurons. CB, over a concentration range of $10-50{\mu}g/{\mu}l$, inhibited the $A{\beta}\;(25-35)\;(10\;{\mu}M)$-induced neuronal cell death, as assessed by a 3-[4,5-dimethyIthiazole-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) assay and the number of apoptotic nuclei, evidenced by Hoechst 33342 staining. CB $(50\;{\mu}g/{\mu}l)$ inhibited glutamate release into medium induced by $10\;{\mu}M\;A{\beta}$, (25-35) which was measured by HPLC. Pretreatment of CB $(50\;{\mu}g/{\mu}l)$ inhibited $10{\mu}M\;A{\beta}$ (25-35)-induced elevation of cytosolic calcium concentration $([Ca^{2+}]_c)$, which was measured by a fluorescent dye, fluo-4 AM, and generation of reactive oxygen species. These results suggest that CB prevents $A{\beta}$ (25-35)-induced neuronal ell damage in vitro.

• Journal of Microbiology and Biotechnology
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• v.16 no.10
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• pp.1491-1498
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• 2006

The feasibility of pullulan acetate nanoparticles (PAN) with ionic strength (IS) sensitivity as a radioisotope carrier to inhibit tumor growth is demonstrated. PAN was radiolabeled with rhenium 188 (Re-188) without any chelating agents. The labeling efficiency of Re-188 into PAN (Re-188PAN) was $49.3{\pm}4.0%$ as determined by TLC. The tumor volumes of mice treated with 0.45 mCi of Re-188-PAN were measured and compared with that of free Re-188 after 5 days of intratumoral injection. For the histological evaluation of apoptotic nuclei of tumor cells, hematoxylin and eosin (H&E), and terminal deoxynucleotidyl transferase biotinylated deoxyuridine triphosphate nick end labeling (TUNEL) staining were performed. The mean tumor volume of the Re-188-PAN-treated group was decreased by 36% after 5 days, whereas that the free Re-188-treated group was decreased by only 15% (P<0.05). The mean number of TUNEL-positive cells in Re-188-PAN-treated tumors at $144.3{\pm}79.9$ cells/section was significantly greater than the control ($26.7{\pm}7.9$ cells/section, P=0.03). The numbers of leukocyte and lymphocyte were decreased in both free Re-188- and Re-188-PAN-treated mice. These results indicated that the intratumoral injection of Re-188-PAN effectively inhibits the tumor growth by prolonging Re-188 retention time in tumor site induced by the IS sensitivity.

• Clinical and Experimental Pediatrics
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• v.51 no.10
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• pp.1112-1117
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• 2008

Purpose : We intended to observe cell death and apoptotic changes in neurons in organotypic hippocampal slice cultures following oxygen-glucose deprivation (OGD), using propidium iodide (PI) uptake, Fluoro-Jade (FJ) staining, TUNEL staining and immunofluorescent staining for caspase-3. Methods : The hippocampus of 7-day-old rats was cut into $350{\mu}m$ slices. The slices were cultured for 10 d (date in vitro, DIV 10) and and exposed to OGD for 60 min at DIV 10. They were then incubated for reperfusion under normoxic conditions for an additional 48 h. Fluorescence of PI uptake was observed at predetermined intervals, and the cell death percentage was recorded. At 24 h following OGD, the slices were Cryo-cut into $15{\mu}m$ thicknesses, and Fluoro-Jade staining, TUNEL staining, and immunofluorescence staining for caspase-3 were performed. Results : 1) PI uptake was restricted to the pyramidal cell layer and DG in the slices after OGD. The fluorescent intensities of PI increased from 6 to 48 h during the reperfusion stage. The cell death percentage significantly increased time-dependently in CA1 and DG following OGD (P<0.05). 2) At 24 h after OGD, many FJ positive cells were detected in CA1 and DG. Some neurons had distinct nuclei and processes while others had fragmented nuclei and disrupted processes in CA1. TUNEL and immunofluorescent staining for caspase-3 showed increased expression of TUNEL labeling and caspase-3 in CA1 and DG at 24 h after OGD. Conclusion : The numerous dead cells in the slice cultures after OGD tended to display apoptotic changes mediated by the activation of caspase-3.