• Molecular & Cellular Toxicology
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• v.5 no.1
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• pp.32-43
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• 2009

Acetaminophen (APAP) overdose is known to cause severe hepatotoxicity mainly through the depletion of glutathione. In this study, we compared the cytotoxic effects of APAP on both a normal murine hepatic cell line, BNL CL.2, and its SV40-transformed cell line, BNL SV A.8. Gene expression profiles for APAP-treated cells were also obtained using microarray and analyzed to identify differences in genes or profiles that may explain the differences of susceptibility to APAP in these cell lines. These two cell lines exhibited different susceptibilities to APAP (0-$5,000{\mu}M$); BNL SV A.8 cells were more susceptible to APAP treatment compared to BNL CL.2 cells. A dose of $625{\mu}M$ APAP, which produced significant differences in cytotoxicity in these cell lines, was tested. Microarray analysis was performed to identify significant differentially expressed genes (DEGs) irrespective of APAP treatment. Genes up-regulated in BNL SV A.8 cells were associated with immune response, defense response, and apoptosis, while down-regulated genes were associated with catalytic activity, cell adhesion and the cytochrome P450 family. Consistent with the cytotoxicity data, no significant DEGs were found in BNL CL.2 cells after treatment with $625{\mu}M$ APAP, while cell cycle arrest and apoptosis-related genes were up-regulated in BNL SV A.8 cells. Based on the significant fold-changes in their expression, a genes were selected and their expressions were confirmed by quantitative real-time RT-PCR; there was a high correlation between them. These results suggest that gene expression profiles may provide a useful method for evaluating drug sensitivity of cell lines and eliciting the underlying molecular mechanism. We further compared the genes identified from our current in vitro studies to the genes previously identified in our lab as regulated by APAP in both C57BL/6 and ICR mice in vivo. We found that a few genes are regulated in a similar pattern both in vivo and in vitro. These genes might be useful to develop as in vitro biomarkers for predicting in vivo hepatotoxicity. Based on our results, we suggest that gene expression profiles may provide useful information for elucidating the underlying molecular mechanisms of drug susceptibility and for evaluating drug sensitivity in vitro for extrapolation to in vivo.

• Journal of Ginseng Research
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• v.42 no.4
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• pp.512-523
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• 2018

Background: 20(S)-Protopanaxadiol (20S-PPD) is a fully deglycosylated ginsenoside metabolite and has potent dermal antiaging activity. However, because of its low aqueous solubility and large molecular size, a suitable formulation strategy is required to improve its solubility and skin permeability, thereby enhancing its skin deposition. Thus, we optimized microemulsion (ME)-based hydrogel (MEH) formulations for the topical delivery of 20S-PPD. Methods: MEs and MEHs were formulated and evaluated for their particle size distribution, morphology, drug loading capacity, and stability. Then, the deposition profiles of the selected 20S-PPD-loaded MEH formulation were studied using a hairless mouse skin model and Strat-M membrane as an artificial skin model. Results: A Carbopol-based MEH system of 20S-PPD was successfully prepared with a mean droplet size of 110 nm and narrow size distribution. The formulation was stable for 56 d, and its viscosity was high enough for its topical application. It significantly enhanced the in vitro and in vivo skin deposition of 20S-PPD with no influence on its systemic absorption in hairless mice. Notably, it was found that the Strat-M membrane provided skin deposition data well correlated to those obtained from the in vitro and in vivo mouse skin studies on 20S-PPD (correlation coefficient $r^2=0.929-0.947$). Conclusion: The MEH formulation developed in this study could serve as an effective topical delivery system for poorly soluble ginsenosides and their deglycosylated metabolites, including 20S-PPD.

• Korean Journal of Food Science and Technology
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• v.30 no.5
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• pp.1229-1235
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• 1998

To see the correlation between the rate of in vitro starch hydrolysis and the glycemic index, an in vitro digestion was carried out by incubating the cereal samples for 2 hours with ${\alpha}-amylase$ in dialysis tubing. Also the levels of blood glucose were measured over 2 hours after feeding healthy volunteers with 50 g carbohydrate portions. Hydrolysis area, hydrolysis index (HI) and the dialysate content of carbohydrate throughout the digestion time for barley was significantly below those for other cereals (p<0.05), and unpolished glutinous rice was significantly above (p<0.05). The GI-glucose of barley $(57%{\pm}7)$ to glucose as standard was significantly (p<0.05) lower than those of other cereals whereas the GI-glucose of glutinous rice $(110%{\pm}8)$ was significantly higher (p<0.05) than other cereals. The GI-rice values to rice as standard were $122%{\pm}4$ for glutinous sorghum, $116%{\pm}13$ for job's tear, $115%{\pm}13$ for glutinous millet, $106%{\pm}6$ for unpolished glutinous rice, $102%{\pm}7$ for glutinous rice, $100%{\pm}0$ for rice, $90%{\pm}12$ for unpolished rice, $85%{\pm}6$ for foxtail millet, $79%{\pm}5$ for buckwheat and $63%{\pm}6$ for barley. The GI-rice was significantly correlated to hydrolysis area and HI (r=0.75, p<0.01). It suggests that the in vitro starch hydrolysis offers good potential to predict the in vivo glycemic response of starch foods.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.7
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• pp.3065-3071
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• 2012

Aim: The present investigation was to evaluate the effects of essential oils of Wedelia chinensis (Osbeck) on free radicals and in vivo antioxidant properties. Methods: Essential oils were extracted using hydro-distillation and compound analysis was performed by GC-MS analysis. Screening for inhibitory activity was conducted by DPPH and OH-scavenging assays. In addition an in vivo study was carried out in cell line implanted cancer bearing mice with assessment of levels of catalase, superoxide dismutase, glutathione peroxidase, lipid peroxidation, nitric oxide and reduced glutathione. Finally, lungs were dissected out for histopathology study of metastasis. Results: GC-MS analysis revealed the presence of carvocrol and trans-caryophyllene as the major compounds with 96% comparison with the Wilily and NBS libraries. The essential oil exhibited significant inhibition in DPPH free radical formation. Whereas reducing power and hydroxyl radical scavenging activity are dose dependent. When compared with the standard, it was found that the essential oil has more or less equal activity in scavenging free radicals produced. In the animal studies, the level of antioxidant enzymes catalase, superoxide dismutase and glutathione peroxidase, as well as glutathione, were found to be increased in treated groups whereas lipid peroxidation and nitric oxide were reduced. Histopathology report also shows that the essential oil has a significant combating effect against cancer development. Conclusion: In all the in vitro assays, a significant correlation existed between the concentrations of the essential oil and percentage inhibition of free radicals. The in vivo studies also has shown a very good antioxidant property for the essential oil during cancer development. From, these results the essential oil can be recommended for treating disease related to free radicals and to prevent cancer development.

• Ocean and Polar Research
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• v.33 no.2
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• pp.127-133
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• 2011

The comet assay, also called single-cell electrophoresis (SCGE) assay, is a potential sensitive monitoring tool for DNA damage in cells. The primary objective of this study was to use comet assay to ascertain if the blood cells of flounder (Pleuronichthys olivaceus) and muscle cells of clam (Saxidomus purpurata) are suitable for genotoxicity screening. This was achieved by initially exposing blood and muscle cells under in vitro conditions to the reference genotoxin hydrogen peroxide ($H_2O_2$); strong correlation between $H_2O_2$ concentration and comet values were found. Subsequently, the identification of DNA damage in isolated cells from flounder and clam was performed under in vivo exposure to benzo(a)pyrene (BaP) and tributyltin (TBT). Flounder and clam were exposed to different concentrations (1, 10, 50, 100 ${\mu}g/L$) of BaP or TBT for 4 days. Regardless of treated chemicals, blood cells of flounder were more prone to DNA breakage compared to muscle cells of clam. In conclusion, in vivo genotoxicity of BaP and TBT can be biomonitored using the comet assay. This study suggests that flounder and clam do show potential as mediums for monitoring genotoxic damage by comet assay.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.7
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• pp.1049-1056
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• 2007

This study was conducted to compare the methane ($CH_4$) production estimated by in vivo (sulfur hexafluoride tracer technique ($SF_6$)) with that of two in vitro rumen simulation (RUSITEC) and gas production (IVGPT)) techniques. Four adult dry Holstein cows, aged $7.4{\pm}3.0$ years and weighing $697{\pm}70$ kg, were used for measuring methane production from five diets by the $SF_6$ technique. The experimental diets were alfalfa hay ($D_1$), corn silage + soybean meal (SBM) (910: 90, $D_2$), Italian rye grass hay +SBM (920: 80, $D_3$), rice straw +SBM (910: 90, $D_4$) and Sudan grass hay +SBM (920: 80, $D_5$). Each diet was individually fed to all 4 cows and 5 feeding studies of 17 d each were conducted to measure the methane production. In the RUSITEC, methane production was measured from triplicate vessels for each diet .In vitro gas production was measured for each of the diets in triplicate syringes. The gas produced after 24 and 48 h was recorded and gas samples were collected in vacuum vials and the methane production was calculated after correction for standard temperature and pressure (STP). Compared to the $SF_6$ technique, estimates of methane production using the RUSITEC were lower for all diets. Methane production estimated from 24 h in vitro gas production was higher (p<0.001) on $D_1$ as compared to that measured by $SF_6$, whereas on $D_2$ to $D_5$ it was lower. Compared to $SF_6$, methane production estimated from 48 h in vitro gas production was higher on all diets. However, methane estimated from the mean of the two measurement intervals (24+48 h/2) in IVGPT was very close to that of $SF_6$ (correlation 0.98), except on $D_1$. The results of our study confirmed that IVGPT is reflective of in vivo conditions, so that it could be used to generate a database on methane production potential of various ruminant diets and to examine strategies to modify methane emissions by ruminants.

• Journal of Biomedical Engineering Research
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• v.19 no.4
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• pp.379-384
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• 1998

To measure precisely the blood velocity in the skin microcirculation, we have used time domain correlation (called Cross-Correlation) based on the processing of the backscattered RF signal obtained with a wideband echographic imaging transducer, although it is difficulties of adaptation of the pulsed wave system, because of the data processing in real time and the hardware problem. This dedicated technology based on a 20MHz echographic imaging system has been developed. We present how the experimental data, i.e. the backscattered RF signal, have to be analyzed. After RF lines realignment, stationary echo canceling procedure and correlation level control, a velocity profile has been obtained. In-vitro result show that velocity measurements as low as 0.1mm/sec attainable with a 80${\mu}m$ in axial resolution. We have also validated with in-vivo experimentation on the external ear of a rabbit using B-mode sector scanning image and M-mode image of a custom made 20MHz skin image system. The flow of the "auriculares caudales" vein, a microvessel of 600 m diameter, has been detected and studied. This technique will allow a more precise exploration of circulatory troubles in cutaneous pathologies.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.11
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• pp.1721-1728
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• 2007

An in vitro and a feeding trial were conducted to investigate the effect of xylanase supplementation on the feeding value of growing pig diets containing high proportions of Chinese double-low rapeseed meals (DLRM). Seven diets were formulated to meet NRC (1998) nutrient requirements. Diet 1 based on corn-soybean meal was used as positive control 1, and diet 2, a practical diet which incorporated a conventional level of Chinese DLRM (60 g/kg diet), as positive control 2. Diet 3 contained a higher level of DLRM (100 g/kg diet) as the negative control. Diet 3 plus xylanase at 0.10, 0.25, 0.50 and 0.70 g/kg diet created diets 4, 5, 6 and 7, respectively. The seven diets were incubated in triplicate with the in vitro two-stage enzyme incubation method to predict responses of diets to xylanase in terms of digestibility of dry matter (DM), crude protein (CP) and neutral detergent fibre (NDF). In vitro, the negative control had the lowest CP and NDF digestibility. Both DM and CP digestibility were increased (p<0.05) owing to xylanase supplementation either at 0.50 or 0.70 g/kg diet, and NDF digestibility was improved following xylanase addition at all of the test levels. There was a high linear correlation ($r^2>90$, p<0.05) between the activity concentration of the enzyme when transformed into its logarithmic value and in vitro digestibility coefficients of DM, CP or NDF. In the feeding trial, 112 crossbred pigs were randomly assigned to seven dietary treatments with 16 replicate pens of one pig each. An obvious dose effect on growth rate was observed ($r^2=0.79$, p<0.05) within the inclusion levels of xylanase. Compared with the negative control, xylanase addition at 0.70 g/kg diet resulted in significantly increased ADG (878 g/d vs. 828 g/d, p<0.05), and a tendency towards improved growth rate (868 g/d vs. 828 g/d, p = 0.10) was also observed following the inclusion of xylanase at 0.50 g/kg diet. It would appear that the nutrient utilization of corn and Chinese DLRM diets by pigs could be enhanced by an appropriate amount of xylanase addition. The in vitro and in vivo results suggested that the in vitro incubation method is feasible for predicting responses of pigs to exogenous enzymes and identifying those preparations that possess potential for improvement of the nutritive values of feedstuffs.

• International Journal of Stem Cells
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• v.16 no.4
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• pp.438-447
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• 2023

Recently, ex-vivo gene therapy has emerged as a promising approach to enhance the therapeutic potential of mesenchymal stem cells (MSCs) by introducing functional genes in vitro. Here, we explored the need of using selection markers to increase the gene delivery efficiency and evaluated the potential risks associated with their use in the manufacturing process. We used MSCs/CD that carry the cytosine deaminase gene (CD) as a therapeutic gene and a puromycin resistance gene (PuroR) as a selection marker. We evaluated the correlation between the therapeutic efficacy and the purity of therapeutic MSCs/CD by examining their anti-cancer effect on co-cultured U87/GFP cells. To simulate in vivo horizontal transfer of the PuroR gene in vivo, we generated a puromycin-resistant E. coli (E. coli/PuroR) by introducing the PuroR gene and assessed its responsiveness to various antibiotics. We found that the anti-cancer effect of MSCs/CD was directly proportional to their purity, suggesting the crucial role of the PuroR gene in eliminating impure unmodified MSCs and enhancing the purity of MSCs/CD during the manufacturing process. Additionally, we found that clinically available antibiotics were effective in inhibiting the growth of hypothetical microorganism, E. coli/PuroR. In summary, our study highlights the potential benefits of using the PuroR gene as a selection marker to enhance the purity and efficacy of therapeutic cells in MSC-based gene therapy. Furthermore, our study suggests that the potential risk of horizontal transfer of antibiotics resistance genes in vivo can be effectively managed by clinically available antibiotics.

• Radiation Oncology Journal
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• v.17 no.2
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• pp.158-165
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• 1999

Purpose : To investigate the percentage of colonies wi1h16or more cells distribution of human skin fibroblast according to in vitro aging, and to evaluate the relationship between percentage of colonies with 10 or more cells and in vivo donor age in human skin fibroblast culture. Material and Method : C1, C2, C3a, and C3b human skin fibroblast samples from three breast cancer patients were used as subjects. The C1, C2, and C3a donor were 44, 54, and 55 years old, respectively. C3a and C3b cells were isolated from the same person. Single cell suspension of skin fibroblasts was prepared with primary explant technique. One hundred cells are plated into 100m1 tissue culture flask and cultured for two weeks. The colony size was defined as colonies with 16 or more cells. The cultured cell was stained with crystal violet, and number of cells in each colony was determined with stereo microscope at $\times$10 magnification. Passage number of C1, C2, C3a and C3b skin fibroblast were 12th, 17th, and 14th, respectively. Results : Percentage of colonies with 16 or more cells of skin fibroblast samples decreased with increasing in vitro passage number. In contrast, cumulative population doublings of skin fibroblast sample increased with increasing in vitro passage number. Percentage of colonies with 16 or more cells also decreased with increasing population doublings in human skin fibroblast culture. There was strong correlation with percentage of colonies with 16 or more cells and population doublings En C3a skin fibroblast sampie. At the same point of population doublings, the percentage of colonies with 16 or more cells of the young C1 donor was higher level than the old C3a donor. Conclusion : The population doublings increased with increasing in vitro passage number but percentage of colonies with 16 or more cells decreased. The results of this study imply that percentage of colonies with 16 or more cell is useful as a indicator of in vitro human skin fibroblast aging and may estimate the in vivo donor age.