Earticle

Cisplatin, a platinum-complex, is currently one of the most effective compounds used in the treat-ment of solid tumors. However, its use is limited by severe side effects such as renal toxicity. Our platinum-based drug discovery program is aimed at developing drugs capable of diminishing toxicity and improving selective cytotoxicity. We synthesized new Pt (II) complex analogue containing 1,2-diaminocyclohexane (DACH) as carrier ligand and 1,2-bis (diphenylphosphino) ethane (DPPE) as a leaving group. Furthermore, nitrate was added to improve the solubility. A new series of [Pt(cia-DACH)(DPPE)] . $2NO_3$ (PC) was synthes-ized and characterized by their elemental analysis and by various spectroscopic techniques [infrared (IR), $_{13}$carbon nuclear magnetic resonance (NMR)] .PC demonstrated acceptable and significant antitumor activity against SKOV-3 and OVCAR-3 human ovarian carcinoma cell lines as compared with that of cisplatin. The cytotoxicity of PC in normal cells was found quite less than that of cisplatin using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), ($^3$H)thymidine uptake and glucose consumption tests in rabbit renal proximal tubular cells, human renal cortical cells and tissues. In conclusion, PC is considered to be more selective cytotoxicity toward human ovarian cancer cells than normal human/rabbit kidney cells.

The adverse effects of ethanol on cell proliferation have been described for a variety of tissues and cells. In the present study, we investigated whether chronic ethanol intoxication impairs the cell proliferation and DNA synthesis induced by oncogenic $H-ras^{V12}$ - and $v-K-ras^{V12}$-transformed cells. Ethanol treatment inhibited the cell proliferation and the DNA synthesis of control parental fibroblasts in a time- and dose-dependent manner. In contrast, ethanol did not suppress the proliferation of either oncogenic $H-ras^{V12}$ - or $v-K-ras^{V12}$ -transformed fibroblasts. Microinjection of oncogenic $H-Ras^{V12}$ protein induces DNA synthesis and ethanol treatment did not interfere with the DNA synthesis. The antiproliferative toxicity of ethanol was rescued by antioxidants, such as N-acetylcysteine and 4-methlpyrazole. These results indicate that the antiproliferative action site of ethanol toxicity lies upstream or is independent of Ras and ethanol exerts its toxicity through a free radical formation.

Nanomedicine constitutes the emerging field of medical applications for nanotechnology such as nanomaterial-based drug delivery systems. This technology may hold exceptional potential for novel therapeutic approaches to liver diseases. Activ ation of apoptosis pathway is a key mechanism by which cytotoxic drugs kill tumor cells. Delivering drugs sele ctively to cancer cells but not to nearby normal cells is a major obstacle in drug therapy. In this study, lithoch olic acid (LCA), a potent anti-cancer drug, is converted to two forms of poly(ethyleneglycol) (PEG) conjugates, viz., PEG-LCA (PL) and lactobionic acid (LBA) conjugated PEG-LCA (LPL). The latter form contains a galacto se ligand in LBA to target the hepatocytes. Both forms are self-assembled to form nanoparticle formulation, and they have high potency than LCA to kill HepG2 cancer cells, sparing normal LO2 cells. Besides, LPL has high specificity to mouse liver cells in vivo after intravenous (IV) injection. Western blot results confirm that the cel l death is occurred through apoptosis induced by LPL nanoparticles. In conclusion, the induction of apoptosis a nd cell death is much more efficient with LPL nanoparticles than LCA molecules as well as good hepatocyte specificity in vitro and in vivo.

In this study, we analyzed expression patterns of apoptotic and autophagic gene products in culture follicular cells of normal and miniature pigs to assess the effect of hormones on the choice for programmed cell death. Autophagic activity progressively increased from control cultures to luteinizing hormone (LH)-treated cultures of follicular cells of normal pigs, but decreased from the LH to follicle stimulating hormone (FHS) +LH-treated cultures. Expression of Casp-3 protein in follicular cells was highest in LHtreated cultures, but the activity of Casp-3 decreased in the control, FSH-treated, and FSH+LH-treated cultures. The activity of the apoptosis protein was highly expressed in the control, LH-treated, and FSH+LH-treated follicular cells of miniature pigs, but autophagy- associated proteins were expressed at low levels in all treatments groups of the miniature pig. The expression of autophagy and apoptosis proteins appeared similar in control and rapamycin-treated cells. In addition, stimulation with FSH triggered the activation of autophagy in the follicular cells of normal pigs, but induced apoptosis in the follicular cells of miniature pigs. A similar effect was obtained when LH was applied. These results suggest that the autophagy process and FSH stimulation is more effective for stable and innovative follicular cell development.

In an effort to elucidate the differential actions of blueberry(BB) in both normal and cancer cells, we utilized human breast cell lines to assess the accumulation of radical oxygen species(ROS) and ROS-associated apoptosis in both human normal breast epithelial(MCF10A) and breast cancer(MCF7) cells. BB extract was added to the cultures at a final concentration of $20{\mu}g/m{\ell}$ for 0(control), 6, 12, and 24 hr intervals. The MCF10A cells evidenced no marked ROS accumulation in the presence of BB, whereas the MCF7 cells evidenced clear ROS accumulation upon BB treatment from 12 hours forward. The number of dying or dead cells did not increase in the BB-treated MCF10A cell groups, whereas that number increased profoundly from 12 hr forward. Furthermore, the expression levels of certain stress-related, and pro- and antiapoptotic gene products evidenced differential responses to BB treatment between the MCF10A and MCF7 cell groups. These results indicate that the components of BB extract differentiate cancer cells by not preventing ROS accumulation within cells and by inducing ROS-associated cell death in cancer cells. However, no marked ROS accumulation or induction of cell death was noted in the normal breast epithelial cells. The fact that BB extract exerted a differential effect on cancer cells opens further directions of research regarding the specific components that exert the differential BB-mediated effects in the selective prevention of normal cells and therapy for cancer tissues in the physiological body.

Glycosphingolipids including gangliosides play important regulatory roles in cell prolifera-tion and differentiation. UDP-glucose:ceramide glucosyltransferase(Ugcg) catalyze the initial step in glycosphingolipids biosynthesis pathway. In this study, Ugcg expression was reduced to approximately 80% by short hairpin RNAs(shRNAs) to evaluate the roles of glycosphingolipids in proliferation and gangliosdies differentiation of PK15. HPTLC/immunofluorescence analyses of shRNA transfected PK15 revealed that treatment with Ugcg-shRNA decreased expression of major gangliosides, GM1 and GD1b. Furthermore, MTT and western blot/immunofluorescence analyses demonstreated that inhibition of the Ugcg expression in PK15 resulted in decrease of cell proliferation. In addition to we showed that inflammatory respon-ses in PK15 and gangliosides knock-down PK15 cells stimulated with Escherichia coli lipopolysaccharide(LPS). The gangliosides GM3, GM2, GD3, GD1a and GD1b were detected in normal PK15 cell. However almost gangliosides undetected in gangliosides knock-down PK15 cell. In addition we found that the expression levels of TNF-α, indu-cible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and we measured the production of NO. The expression of inflammatory proteins, TNF- α, iNOS and COX-2 the elevated concentration in gangliosides knock-down PK15 cell. These results suggest that gangliosides interact with components of the proinflammatory response pathway and might, therefore, be relevant for designing future therapeutic strategies intended to prolong xenograft survival.

Glycosphingolipids including gangliosides play important regulatory roles in cell prolifera-tion and differentiation. UDP-glucose:ceramide glucosyltransferase(Ugcg) catalyze the initial step in glycosphingolipids biosynthesis pathway. In this study, Ugcg expression was reduced to approximately 80% by short hairpin RNAs(shRNAs) to evaluate the roles of glycosphingolipids in proliferation and gangliosdies differentiation of PK15. HPTLC/immunofluorescence analyses of shRNA transfected PK15 revealed that treatment with Ugcg-shRNA decreased expression of major gangliosides, GM1 and GD1b. Furthermore, MTT and western blot/immunofluorescence analyses demonstreated that inhibition of the Ugcg expression in PK15 resulted in decrease of cell proliferation. In addition to we showed that inflammatory respon-ses in PK15 and gangliosides knock-down PK15 cells stimulated with Escherichia coli lipopolysaccharide(LPS). The gangliosides GM3, GM2, GD3, GD1a and GD1b were detected in normal PK15 cell. However almost gangliosides undetected in gangliosides knock-down PK15 cell. In addition we found that the expression levels of TNF-α, indu-cible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX -2) and we measured the production of NO. The expression of inflammatory proteins, TNF- α, iNOS and COX-2 the elevated concentration in gangliosides knock-down PK15 cell. These results suggest that gangliosides interact with components of the proinflammatory response pathway and might, therefore, be relevant for designing future therapeutic strategies intended to prolong xenograft survival.

형광을 이용한 암 진단을 위해 배양된 정상 및 암세포주에 광민감제인 5-ALA를 투여하고 세포 내ㆍ외에서 생성된 Protoporphyrin IX (PpIX)의 형광을 측정하여 5-ALA 투여의 최적농도를 조사하였다. 정상 간세포주 (Chang) 및 자궁경부암 세포주 (HeLa)에 5-ALA를 농도별로 투여하여 ALA에 의해 유도된 PpIX의 생성을 확인하고, MTT assay로 세포생존율을 측정하였다. 배양된 cell에 5-ALA를 투여한 후 24시간 동안 배양함으로써 생성되는 PpIX의 양은 형광의 강도로 측정하였다. HeLa 세포주에 대한 5-ALA의 최적농도는 50 μg/mL이며, 이 때의 형광 (emission) 스펙트럼은 여기 파장이 410 nm일 때 602.3 nm, 659.9 nm에서 형광 봉우리가 관찰되었다. PpIX의 형광 강도를 측정한 결과, PpIX는 정상세포에서는 낮은 농도로 축적이 되는 반면에 암세포에서 더 높은 농도로 축적되었으며, 세포 외보다는 세포 내에서 더 높은 농도로 축적됨을 알 수 있었다.

To clarify the usefulness of fluorescent diagnosis for cancer, We investigated the optimal method of administrating 5-aminolevulinic acid (5-ALA) by analyzing fluorescence signal of Protoporphyrin IX (PpIX) in the cultured normal and cancer cells. 5-ALA was injected as a photosensitizer to the cervico-uterine cancer cell line (HeLa) and in normal liver cells (Chang). Chang and HeLa cells were incubated with various concentrations of 5-ALA (0-800 ㎍/㎖). The accumulation of PpIX induced by 5-ALA was in HeLa and Chang cells. The cell viability was measured by MTT assay. The optimal concentration of ALA that induced maximum levels of PpIX was 50 ㎍/㎖ in HeLa cell cultured for 24 hours after 5-ALA injection. Fluorescence of PpIX in HeLa cell was excited at a wavelength (λ=410 ㎚) and showed an emission spectrum at 602.3 ㎚, 659.9 ㎚ which could be related to the PpIX generation induced by the applied 5-ALA. The experimental results showed that fluorescence signal of PpIX was proportional to the concentration of 5-ALA in cancer cells, but measured with low concentration in normal cells.

The flowers of Buddleja officinalis are used to treat sore and damaged eyes, a condition which is similar to skin wounds. However, whether it has any protective effect on oxidative DNA damage and cell death induced by hydroxyl radical remains unclear. In this study, we evaluated the protective effects of the extracts against oxidative DNA and cell damage caused by hydroxyl radical. DPPH radical, hydroxyl radical, hydrogen peroxide and intracellular ROS scavenging assay, and $Fe^{2+}$ chelating assay were used to evaluate the antioxidant properties. phi X 174 RF I plasmid DNA and intracellular DNA migration assay were used to evaluate the protective effect against oxidative DNA damage. Lastly, MTT assay and lipid peroxidation assay were used to evaluate the protective effect against oxidative cell damage. It was found to prevent intracellular DNA and the normal cells from oxidative damage caused by hydroxyl radical via antioxidant activities. These results suggest that Buddleja officinalis may exert the inhibitory effect on ROS-induced carcinogenesis by blocking oxidative DNA damage and cell death.

Cancer treatment remains a serious problem worldwide. Analysis of the relationship between cancer cells and normal cells reveals that these two share characteristics in contradiction, thus could be analyzed by using contradictory principles. Under the theory of contradictory principles, induction of a dormant state or reversal of cancer cells is an important treatment strategy beyond traditional cytotoxic therapy. Normal cells are also the targets and under the influence of anti-cancer treatments and should be considered during therapy. Findings based on crosstalk between these two cell types may offer opportunities for the development of new biomarkers and therapies.

Piperlongumine, a natural alkaloid isolated from the long pepper, selectively increases reactive oxygen species production and apoptotic cell death in cancer cells but not in normal cells. However, the molecular mechanism underlying piperlongumine-induced selective killing of cancer cells remains unclear. In the present study, we observed that human breast cancer MCF-7 cells are sensitive to piperlongumine-induced apoptosis relative to human MCF-10A breast epithelial cells. Interestingly, this opposing effect of piperlongumine appears to be mediated by heme oxygenase-1 (HO-1). Piperlongumine upregulated HO-1 expression through the activation of nuclear factor-erythroid-2-related factor-2 (Nrf2) signaling in both MCF-7 and MCF-10A cells. However, knockdown of HO-1 expression and pharmacological inhibition of its activity abolished the ability of piperlongumine to induce apoptosis in MCF-7 cells, whereas those promoted apoptosis in MCF-10A cells, indicating that HO-1 has anti-tumor functions in cancer cells but cytoprotective functions in normal cells. Moreover, it was found that piperlongumine-induced Nrf2 activation, HO-1 expression and cancer cell apoptosis are not dependent on the generation of reactive oxygen species. Instead, piperlongumine, which bears electrophilic ${\alpha},{\beta}$-unsaturated carbonyl groups, appears to inactivate Kelch-like ECH-associated protein-1 (Keap1) through thiol modification, thereby activating the Nrf2/HO-1 pathway and subsequently upregulating HO-1 expression, which accounts for piperlongumine-induced apoptosis in cancer cells. Taken together, these findings suggest that direct interaction of piperlongumine with Keap1 leads to the upregulation of Nrf2-mediated HO-1 expression, and HO-1 determines the differential response of breast normal cells and cancer cells to piperlongumine.

Atmospheric plasma's electron temperature is less than thermal plasma, so it is useful at bio experiment. We have investigated the optical emission spectroscopy (OES) lines by spectrometer during Atmospheric plasma bombardment onto the PBS surface by using an Ar gas flow. Also we have measured the OH radical density inside the solution induced by the Atmospheric plasma bombardment. OH radical species are appeared at 308 nm and 309 nm. Densities of OH radical species has been found to be significantly decreased versus depth of the solution from 2 mm to 6 mm. OH radical density inside the PBS is measured to be about $1.87{\times}1016cm-3$ downstream at 2 mm from the surface under optimized Ar gas flow of 200 sccm in Atmospheric plasma. Also we have investigated cell viability of lung cancer and normal cell after Atmospheric plasma treatment for fixed exposure time in 60 seconds, but different depths. We used SEM, we observed change of cell morphorogy, did experiment about FDA & PI Staining method. It is found that there is selectivity between the lung cancer and lung normal cell, in which cancer cell definitely has higher cell death ratio more than normal cell. We have investigated change of bond structure in FT-IR spectroscopy, the following peaks were observed: and intense O-H peak at 3422 cm-1 and at 2925 cm-1 corresponds to C-H stretch vibrations of methylene group.

N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) is a carcinogen made of modified guanine on which alkyl group is added on 6th oxygen. It has been used for inducing different types of cancers experimentally in vivo and in vitro. Stomach cancer might be the best well established particular cancer induced with MNNG. Comparative analysis of gene expression between normal stomach cell and MNNG-treated stomach cell could give much information to understand cancer formation in stomach. To this end, normal human stomach cells HS738 were treated with DMSO or MNNG. Genetic comparison was conducted with purified RNA from the treated cells for 6 hours or 24 hours. Total 13 genes were selected based on their high induction folds and comprehensible function to cancer formation. Some of the genes were cancer-promoting whereas the others were anti-cancer genes. These results could give important information of genetic changes in stomach cells during MNNG-induced stomach cancer formation.

The cancer stem cell (CSC) hypothesis has captured the attention of many scientists. It is believed that elimination of CSCs could possibly eradicate the whole cancer. CSC surface markers provide molecular targeted therapies for various cancers, using therapeutic antibodies specific for the CSC surface markers. Various CSC surface markers have been identified and published. Interestingly, most of the markers used to identify CSCs are derived from surface markers present on human embryonic stem cells (hESCs) or adult stem cells. In this review, we classify the currently known 40 CSC surface markers into 3 different categories, in terms of their expression in hESCs, adult stem cells, and normal tissue cells. Approximately 73% of current CSC surface markers appear to be present on embryonic or adult stem cells, and they are rarely expressed on normal tissue cells. The remaining CSC surface markers are considerably expressed even in normal tissue cells, and some of them have been extensively validated as CSC surface markers by various research groups. We discuss the significance of the categorized CSC surface markers, and provide insight into why surface markers on hESCs are an attractive source to find novel surface markers on CSCs.

This study was designed to assess, whether a new chemotherapeutic microtubule inhibitor, Epothilone B (EpoB, Patupilone), can induce DNA damage in normal ovarian cells (MM14.Ov), and to evaluate if such damage could be repaired. The changes were compared with the effect of paclitaxel (PTX) commonly employed in the clinic. The alkaline comet assay technique and TUNEL assay were used. The kinetics of DNA damage formation and the level of apoptotic cells were determined after treatment with IC50 concentrations of EpoB and PTX. It was observed that PTX generated significantly higher apoptotic and genotoxic changes than EpoB. The peak was observed after 48 h of treatment when the DNA damage had a maximal level. The DNA damage induced by both tested drugs was almost completely repaired. As EpoB in normal cells causes less damage to DNA it might be a promising anticancer drug with potential for the treatment of ovarian tumors.