This study was carried out to elucidate the protective effect of zinc chloride(ZnCl$_2$) and its mechanism against the immuno-cytotoxicity of methylmercury chloide($CH_3$HgCl). This study was observed in the culture of EMT-6 cells which are originated from mammary adenocarcinoma of Balb/c mouse. Cytotoxicity of metals was measured by cell viability and NO$_2$$^{[-10]}$ , and mitochondrial function was evaluated by adenosine triphosohate (ATP) production. $CH_3$HgCl significantly decreased the sythesis of nitric oxide(NO), ATP and glutathione(GSH) in a dose-dependent manner. ZnCl$_2$ significantly increased the synthesis of GSH in a dose-dependent manner, but synthesis of NO and ATP were not changed. The immuno-cytotoxicity of $CH_3$HgCl was not fully protected when combined addition of ZnCl$_2$, whereas ZnCl$_2$ prior to addition of $CH_3$HgCl completly protected the Hg-induced immuno-cytotoxicity. Similarly, intracellular accumulation of mercury significantly decreased by ZnCl$_2$. Degree of diminution of intracellular mercury was larger in ZnCl$_2$ prior to addition of $CH_3$HgCl than in combined addition of ZnCl$_2$ and $CH_3$HgCl.. Dithiothreitol(DTT) or buthionine sulfoximine(BSO) addition at 50$\mu$M or less, which was not toxic to the cells, did not affect synthesis of NO and ATP. DTT increased intracellular GSH level and DTT pretreatment protected toxicity induced by $CH_3$HgCl as shown complete recover in the NO and ATP values. BSO decreased intracellular GSH level and BSO pretreatment exaggerated toxicity induced by $CH_3$HgCl as shown synergistic reduction in the NO and ATP values. These results indicated that the protective effects of zinc against immuno-cytotoxicity of methylmercury associated with increasing cellular level of GSH. Increased intracellular GSH transports methylmercury to out of cells. In accordance with intracellular level of mercury decreased, immuno-cytotoxicity of methylmercury decreased. These result also suggest that the protective mechanism of zinc against the mercury toxicity would be exerted in the immune system in vivo.
This study was performed to investigate the effects of aluminium treatment on renal damage in rats. Seventy five male Sprague-Dawley strains were divided into five groups; the control, 250 ppm $AlCl_3$, group, 500 ppm AlCl3 group, 250 ppm $Al_2(SO_4)_3$, group, 500 ppm $Al_2(SO_4)_3$ group and kept on the diet for 2 weeks. The body weight gain was more increased in $AlCl_3$, treated of rats but decreased in $Al_2(SO_4)_3$, treated rats than the control, on based in histophysiological examination, in $AlCl_3$, treated group, thyrodization of kidney tubular lumen and interstitial inflammatory cell infiltration, abscess formation were demonstrated and in $Al_2(SO_4)_3$, treated animals destruction of glomeruli and necrosis in kidney cortex by inflammatory cells were shownd. On the other hand, blue-pigmented cast was noted in tubules of $AlCl_3$ administration group whereas focal aluminium accumulation was noted at interstitial tissue in kidney of $Al_2(SO_4)_3$, administration group.
Clinical perfusion data on 16 cases of cardiopulmonary bypass using Sigmamotor pump and RyggKyvsgaard Oxygenator which performed at Seoul National University Hospital during the period of Aug. 1968 to Aug. 1970 was analized. AIl cases were hemodiluted and the perfusion was carried out under the normothermic condition. The age of the patients ranged between 6 and 43 years. The b:dy weight varied between 18.3 and 54.0 kg and the body surface area between 0.78 and 1. 59$M^2$. The priming solution was consiste:I with fresh ACD blood. Hartmann solution and Mannitol. The average amount of priming was approximately 2242 ml. The average hemodilution rate was 17%. The flow rate ranged from 1.7L to 3.5L/Min/$M^2$ and averaged 2.4L/Min/$M^2$ or 78mI/Min/kg. The duration of perfusion varied from 22 to 110 min with average of 56.9 minutes. Some hemodynamic responses were observed. The arterial pressure dropped immediately after the initiation of partial perfusion and was more marked after the total perfusion foIlowed by gradual increase to the safety level. The central venous pressure reflected the reduced blood volume especially in the cases of prolonged perfusion which lasted over 60 min. In most of the cases, red blood cell count decreased and white blood ceIl count increased after the perfusion. Hemoglobin level was decreased, averaging of 12.5mg%, Hct 3.3% and platelets count of 18% postoperatively. Plasma hemoglobin increased mildly, from pre-perfusion average value of 4. 06mg% to postperfusion value of 22.5mg%. Serum potassium was 4.4mEq/L pre-operatively and was decreased to 3.7mEq/L postoperatively. Five cases showed definite hypopotassemia immediately after the operation. Sodium and chloride decreased mildly. These electrolyte changes are thought to be related with hemodilution. diuretics and reduced blood volume during and after the perfusion. Arterial blood pH value revealed minimal to moderate elevation from preperfusion average value of 7.376 to 7.461 during perfusion and then 7.365 after perfusion. The pC02 and hicarbonate showed minimal to moderately lowered values. The total CO2 was decreased. Buffer base decreased during perfusion (Av. 42.6mEq/L) and further decreased after the perfusion (Av. 40.8mEq/L). These arterial blood acid base changes suggested that the metabolic acidosis was accompanied by respiratory alkalosis during and immediately after the perfusion. Authors belived that the acidosis could more effectively be corrected with the more additional dose of bicarbonate than we used by this study. The chest tune drainage during the first 24 hours following operation was 1158 ml in average. One case (Case No. 15) showd definite bleeding tendency and it was believed that the cause might be due to the defect of heparin and protamine titration. The average urinary out put during 24 hours post-perfusion was 1291ml. One case (Case No. ]) showed definite post perfusion oliguria. As conclusion hemodilution using fresh ACD blood. Hartmann and Mannitol solution added with Bivon and high flow rate unler normothermia. was thought to amelioratc the severity of mctabolic acidosis during and after perfusion with relatively satisfactory effect on the diuresis and bleeding tendency.
Ozonization for rearing seawater in land-based culture system has recently been utilized for disinfection of pathogenic microorganisms and improvement of water quality. This study was conducted to examine the effects of total residual oxidants (TRO) in ozone-treated seawater on survival, blood parameters, osmolality and oxygen consumption, and gill tissue of the flounder. Paralichthys olivaceus. Experiments were carried out with the starved flounder of 12~19 cm in total length at $20^{\circ}C$. The 48-hr and 96-hr $LC_{50}$ for the flounder amounts to 26.4 ppb and 22.3 ppb, respectively. With increase of TRO concentration from 24 to 39 ppb, the values of hematocrit, hemoglobin concentration, red blood cell count and osmolality of the flounder with respect to exposure time were significantly elevated, however, the oxygen consumption rates decreased. In the case of the fish exposed to 13 ppb for 96 hrs, blood glucose increased with an elapse of exposure time, while survival rate was 100%. Death apparently resulted by massive destruction of gill lamellar epithelium, severe osmotic imbalance and the lack of oxygen uptake. The results of this experiment indicated that to protect aquaculture organisms, the ozone-treated seawater should not contain any residual oxidants, and that relatively long-term exposure to TRO of low concentration can impact on survival and physiological conditions of the flounder.
The purpose of this study was to investigate the inhibitory effect of antibacterial substances produced by probiotic lactic acid bacteria (LAB) against biogenic amines-producing bacteria and the influence of culture conditions on the antibacterial activity of bacteriocin and organic acid. The bacteriocin solutions of Lactobacillus plantarum FIL20 (64 AU/ml) and Lactobacillus paracasei FIL31 (128 AU/ml) showed strong antibacterial activity against Serratia marcescens CIH09 and Aeromonas hydrophilia RIH28, respectively. And the lactic acid contents in the cell-free culture supernatants (CFCS) obtained from FIL20 and FIL31 strains were $107.3{\pm}2.7mM$ and $129.5{\pm}4.6mM$, respectively. Therefore, the bacteriocin solution (200 AU/ml) and the CFCS ($200{\mu}l/ml$) produced by L. plantarum FIL20 and L. paracasei FIL31 significantly (P < 0.05) decreased the bacterial numbers and histamine and tyramine production ability of S. marcescens CIH09 and A. hydrophilia RIH28. The amounts of histamine and tyramine produced by the CIH09 strain under conditions of low initial pH (5.0) and incubation temperature ($15^{\circ}C$) was significantly reduced by treatment with bacteriocin solution and CFCS obtained from L. plantarum FIL20. In addition, the bacterial counts and biogenic amines contents of CIH09 strain were significantly decreased (P < 0.05) when sodium chloride (5%) or potassium nitrite (200 mg/g) were mixed with the antibacterial substances of L. plantarum FIL20. Consequently, the bacteriocin and organic acid solution of L. plantarum FIL20 and L. paracasei FIL31 can be used as a biological preservation to effectively control the production of biogenic amines by the application of hurdle technology.
Kim, Seung-Yeon;Kim, Ga-Yeon;You, Hyeong-Ju;Kang, Man-Jong
Animal Bioscience
/
v.35
no.1
/
pp.126-137
/
2022
Objective: Efficient gene editing technology is critical for successful knock-in in domestic animals. RAD51 recombinase (RAD51) gene plays an important role in strand invasion during homologous recombination (HR) in mammals, and is regulated by checkpoint kinase 1 (CHK1) and CHK2 genes, which are upstream elements of RAD51 recombinase (RAD51). In addition, mismatch repair (MMR) system is inextricably linked to HR-related pathways and regulates HR via heteroduplex rejection. Thus, the aim of this study was to investigate whether clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9)-mediated knock-in efficiency of human lactoferrin (hLF) knock-in vector in the bovine β-casein gene locus can be increased by suppressing DNA MMR-related genes (MSH2, MSH3, MSH6, MLH1, and PMS2) and overexpressing DNA double-strand break (DSB) repair-related genes (RAD51, CHK1, CHK2). Methods: Bovine mammary epithelial (MAC-T) cells were transfected with a knock-in vector, RAD51, CHK1, or CHK2 overexpression vector and CRISPR/sgRNA expression vector to target the bovine β-casein gene locus, followed by treatment of the cells with CdCl2 for 24 hours. After 3 days of CdCl2 treatment, the knock-in efficiency was confirmed by polymerase chain reaction (PCR). The mRNA expression levels of DNA MMR-related and DNA DSB repair-related genes were assessed by quantitative real-time PCR (RT-qPCR). Results: Treatment with CdCl2 decreased the mRNA expression of RAD51 and MMRrelated genes but did not increase the knock-in efficiency in MAC-T cells. Also, the overexpression of DNA DSB repair-related genes in MAC-T cells did not significantly affect the mRNA expression of MMR-related genes and failed to increase the knock-in efficiency. Conclusion: Treatment with CdCl2 inhibited the mRNA levels of RAD51 and DNA MMR-related genes in MAC-T cells. However, the function of MMR pathway in relation to HR may differ in various cell types or species.
Transmembrane protein TMEM16A, which encodes calcium-activated chloride channel has been implicated in tumorigenesis. Overexpression of TMEM16A is associated with poor prognosis and low overall survival in multiple cancers including lung adenocarcinoma, making it a promising biomarker and therapeutic target. In this study, three structure-related sesquiterpene lactones (mecheliolide, costunolide and dehydrocostus lactone) were extracted from the traditional Chinese medicine Aucklandiae Radix and identified as novel TMEM16A inhibitors with comparable inhibitory effects. Their effects on the proliferation and migration of lung adenocarcinoma cells were examined. Whole-cell patch clamp experiments showed that these sesquiterpene lactones potently inhibited recombinant TMEM16A currents in a concentration-dependent manner. The half-maximal concentration (IC50) values for three tested sesquiterpene lactones were 29.9 ± 1.1 µM, 19.7 ± 0.4 µM, and 24.5 ± 2.1 µM, while the maximal effect (Emax) values were 100.0% ± 2.8%, 85.8% ± 0.9%, and 88.3% ± 4.6%, respectively. These sesquiterpene lactones also significantly inhibited the endogenous TMEM16A currents and proliferation, and migration of LA795 lung cancer cells. These results demonstrate that mecheliolide, costunolide and dehydrocostus lactone are novel TMEM16A inhibitors and potential candidates for lung adenocarcinoma therapy.
We investigated the inhibitory effects of solvent extracts from dried tuna on the growth of cancer cell lines (HT1080 human fibrosarcoma and HT-29 human colon cancer cells) and $H_2O_2$-induced oxidative stress. Inhibitory effects of acetone with methylene chloride (A+M) and methanol (MeOH) extracts on the growth of HT1080 and HT-29 cancer cells increased in a dose dependent manner (p<0.05). The inhibitory effect was more significant on the growth of HT1080 cells, and A+M extracts had a higher inhibitory effect compared to MeOH extracts. The treatments of hexane, 85% aq. methanol, butanol and water fractions significantly inhibited the growth of both cancer cells (p<0.05). Among the fractions, hexane and 85% aq. methanol fractions showed higher inhibitory effects. In order to determine the protective effect on $H_2O_2$-induced oxidative stress, a DCHF-DA (dichlorodihydrofluorescin diacetate) assay was conducted. All fractions, including crude extracts of dried tuna, appeared to significantly reduce the levels of intracellular reactive oxygen species (ROS) with dose responses (p<0.05). Among the fractions, BuOH and 85% methanol fractions showed a higher protective effect on the production of lipid peroxides. These results indicate that the consumption of tuna may be recommended as a potent functional food for preventing cellular oxidation and cancer.
We investigated the antioxidant effects of hederagenin 3-O-b-D-glucopyranosyl($1{\rightarrow}3$)-a-L-rhamnopyranosyl($1{\rightarrow}2$)-a-L-arabinopyranoside (HDL) isolated from root bark of Ulmus davidiana on the activity of enzymes related to reactive oxygen species (ROS) in human osteosarcoma U2OS cells. Cobalt chloride ($CoCl_2$), a transition metal, was used as an inducer of oxidative stress, generating hydrogen peroxide ($H_2O_2$) via increasing xanthine oxidase (XO) activity. The increased levels of $H_2O_2$, XO, ferritin, and ferritin iron by $CoCl_2$ were diminished effectively by co-treatment with HDL in U2OS cells. Furthermore, decreased levels of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT) by $CoCl_2$ were highly increased by co-treatment with HDL in U2OS cells; however, the levels of glutathione peroxidase (GPx) did not change. The increased contents of TBARS related to lipid peroxidation were significantly reduced by HDL in U2OS cells. The concentration of GSH changed in a pattern that went against regulated TBARS by $CoCl_2$ and HDL. We examined the expression of p53, $p21^{CIP1/WAF1}$, and $p27^{KIP1}$ proteins related to oxidative stress and cell cycle regulation. As a result, the expression of $p27^{KIP1}$ modulated by $CoCl_2$ was not changed by HDL. However, the expression of p53 and $p21^{CIP1/WAF}$ increased by $CoCl_2$ was reduced by HDL in U2OS cells. Together with alteration of p53 and $p21^{CIP1/WAF1}$ proteins, the accumulated cells at G1 phase by $CoCl_2$ was decreased by HDL in U2OS cells. Our data suggests that HDL inhibits $CoCl_2$-generated ROS in U2OS cells, providing potentially new antioxidant compounds that are isolated from natural products.
In order to determine chemical components of onion flesh and peel, general nutrients, vitamin C, and total flavonoids were measured. Onion peel showed less moisture (14.3%) and no vitamin C compared to onion flesh. Onion peel contained more amounts of total flavonoids compared to onion flesh. In addition, the inhibitory effects of solvent extracts from onion flesh and peel on $H_2O_$-induced oxidative stress and growth of cancer cell lines (AGS human gastric adenocarcinoma and HT-29 human colon cancer cells) were investigated. Acetone with methylene chloride (A+M) and methanol (MeOH) extracts from onion flesh and peel appeared to significantly reduce the levels of intracellular reactive oxygen species (ROS) (p<0.05) and a greater antioxidant effect was observed in onion peel. Among fractions, 85% aq. methanol showed a higher protective activity against oxidative stress in both flesh and peel and there was no effect in the water and hexane fractions. The growth of cancer cells exposed to medium containing extracts and fractions from onion flesh and peel was inhibited dose-dependently. The growth of AGS was inhibited more in both flesh and peel compared to HT-29, and onion peel was more effective than onion flesh. Among fractions, 85% aq. methanol showed the greatest effect on growth inhibition in both flesh and peel. $IC_{50}$ values of 85% aq. methanol fraction from onion flesh and peel on AGS were 0.04 and 0.03 mg/ml, respectively, while those on HT-29 were 0.23 and 0.04 mg/ml. From our results, 85% aq. methanol fraction had an inhibitory effect against oxidative stress and growth of cancer cells, suggesting that it may contain biological active compounds.
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