• Toxicological Research
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• v.14 no.4
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• pp.475-481
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• 1998

This study was performed to investigate the modifying effect of the general (GPA) and the fermented pilose antler (FPA) on experimental hepatocarcinogenesis and Natural Killer cell activity in rats. Specific pathogen free, 5-week male Sprague-Dawley rats were divided into four groups. To induce hepatocarcinogenesis, diethylnitrosamine (DEN, 200 mg/kg, i.p.) was used as a tumor initiator and was given in a single dose at experimental onset. All rats were given a partial hepatectomy (PH) at 3 weeks after experimental onset. Sodium phenobarbital (PB, 0.05% in diet), GPA (0.075% in diet) and FPA (0. 075% in diet) were given from 2 to 8 weeks. Group I of the initiation control group was only given DEN. As initiation-promotion group, Group II was given DEN and then PB. Group III and IV were given DEN-PB-GPA and DEN-PB-FPA, respectively. In hematological analysis, as compared with Group I. the number of white blood cells were significantly increased in the GPA (p<0.01) and the FPA treated group (p<0.05), respectively. Natural killer (NK) cell activity by flow cytometer (FCM) analysis was higher in group of treated with the GPA (35%) than that of the FPA (27.5%), but not significant. Result of the immunohistochemical staining of the glutathione S-transferase placental form (GST-p) indicated that the number of and area of the pre-neoplastic lesions was not significantly changed in Group III and IV compared Group II, respectively. In conclusion, the GPA and the FPA treatment significantly increased the number od WBC in peripheral blood, but the enhancing NK activity and the modifying effect on the experimental hepatocarcinogenesis were not observed.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.191-191
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• 1994

본 연구는 D-galactosamine을 이용한 중기발암성 모델에서 세포성면역중의 하나인 natural killer (NK) 세포활성과 c-myc 종양단백의 발현을 알아보기 위하여 실시하였다. 수컷 6주령의 SPF SD랫드 50마리를 3개군으로 각 군당 20마리씩 나누어 배치하였다. 실험 0일에 제 1,2 군에서 DEN을 복강내로 체중 kg당 200mg 1회 투여하여 발암유발을 하였으며 대조군인 제 3군에는 saline을 투여하였다. 제 2주에 제 1군에는 강력한 발암촉진물질인 2-acetylaminofluorene (AAF)을 사료에 0.01% 투여하였으며, 제 2군은 미약한 발암촉진물질인 phenobarbital (PB)을 0.05%로 음수에 6주간 투여하였다 제 8, 15, 36주에 경시적으로 부검하였다. 제 8주에 부검시 GST-P$^{+}$ 병변이 잘 유도되어 전암병변의 유도가 잘 되었음을 확인하였다. 제 15주에 부검시 AAF를 투여한 군에서 glutathione S-transferase placental form (GST-p)에 대한 면역조직화학적 염색에서 AAF와 PB를 투여한 제 1군 및 제 2군의 간장에서는 주위조직과 한계가 명료한 GST-P$^{+}$ 증식성 결절과 병소를 관찰할 수 있었으나 기초 사료만을 투여한 제 3군은 어떠한 GST-P$^{+}$ 증식성 결절 및 병소를 관찰할 수 없었다. 랫드에서 natural killer세포는 사람이나 마우스에서 주로 자연살생능 (natural killing activity)을 보이는 LGL(lure granular lymphocyte)의 형태를 띄고 있었으며 LGL 이라고 부르는 것처럼 특징적으로 세포질 대 핵의 비율이 높고 azurophilic 과립을 세포질내에 함유하고 있으며 일반적으로 신장 형태의 핵을 가지고 있었다. 또한 세포의 크기는 small lymphocyte와 대식세포 (macrophage)의 중간 크기였다. 15주와 시험종료시 정상대조군인 제 3군의 랫드로 부터 분리한 NK 세포활성도 (% cytotoxicity)에 비하여 발암물질 투여군의 NK 활성도는 PB 투여군들의 NK활성도 보다 약간 낮았다. 랫드에서 c-myc 종양단백은 65KD 와 671KD 에서 band가 형성된 것이 관찰되었다. 시험 개시후 15주에 부검한 랫드의 간에서 c-myc 종양단백의 발현은 모든 처리군들이 대조군에 비하여 높게 발현되는. 것이 관찰되었으나 시험개시후 26주에 부검한 랫드의 간에서 c-myc 종양단백의 발현은 대조군에 비하여 차이가 거의 없었다. 따라서 랫드에서 화학적으로 유도한 간암발생 과정에서 NK 세포활성이 현저하게 억제되는 것으로 생각되며, c-myc 종양단백의 발현은 시험개시후 15주에 그 발현이 확실한 것으로 사료되어 진다.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1993.04a
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• pp.96-96
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• 1993

간부분 절제술을 하지 않는 비수술적 방법으로서 D-galactosamine을 이용한 중기발암성 시험의 개발을 목적으로 F344 수괵 랫드를 이용하여 본 실험을 수행하였다. 실험 I 에서는 실험방법에 따라 3가지 모델로 구분하고, 각 모델에 처치군과 대조군을 두었다. 모델 1 에서는 실험개시시에 diethylinitrosamine (DEN)을 200 mg/kg body weight로 복강내로 1회 투여하고, 실험개시후 2및 5주에 D-galactosamine을 300 mg/kg body weight로 복강내로 각각 1회 투여하였다. 처치군에는 실험개시후 2주부터 6주간 2-acetylaminofluorene을 0.01%로 혼합한 사료를 급여하였으며, 대조군에는 기초사료를 계속 급여하였다. 모델 2에서는 모델 1의 4주차까지의 처치를 2회 반복하였다. 모델 3은 간부분 절제술을 하는 DEN-PH (diethylnitrosamine-partial hepatectomy) 모델과 같은 방법으로 처치하였다. 사육기간 중 매주 체중 및 사료소비량을 측정하였고, DEN 투여후 8주에 전동물을 부검하여 적출한 간의 중량을 측정하고, glutathione S-transferase placental form (GST-P) 양성 foci에 대한 면역조직화학적 염색표본을 만들어 GST-P 양성 foci의 수 및 면적을 측정하였다. 실험 II에서는 모델 1의 방법으로 phenobarbital(PB), 3-methylcholanthrene (3-MC), n-ethyl-n'-nitro-n-nitrosoguanidine 및 3,3'-diaminobenzidine외 GST-P 양성 foci의 발현정도를 조사하였다. 실험 I의 결과, 모델 1이 정상적인 체중 증가를 보여주었으며, 간조직의 GST-P 양성 foci 의 발현율이 가장 좋았다. GST-P 양성 foci의 면적은 큰것 부터 미상엽, 내측우엽, 외측우엽의 순으로 나타났으나 foci의 수는 모델별로 다르게 나타났다. 실험 II의 PB 투여군과 3-MC 투여군에서 GST-P 양성 foci의 수 및 면적의 유의성 있는 증가가 관찰되었다. 이와 같은 결과로 볼때, 비수술적 방법인 D-galactosamine 을 이용한 중기 발암성 검색법은 간부분 절제술을 이용한 중기발암성 검색법에 비하여 GST-P 양성foci의 발현능력이 동등하거나 더 우수하였으며, 간 및 간이외 장기의 발암물질에 대한 발암성 검색에 보다 유용할 것으로 생각된다.

• Korean Journal of Veterinary Research
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• v.44 no.4
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• pp.507-513
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• 2004

Aflatoxins are produced mainly by Aspergillus flavus and Aspergillus parasiticus that grow in improperly stored cereals. Aflatoxin B1 ($AFB_1$) is a potent hepatocarcinogen in a variety of experimental animals including human beings. In spite of a high attention to the hepatocarcinogenecity of $AFB_1$, the relative toxicity of aflatoxins ($AFB_2$ and $AFG_1$) is not fully clarified. Sprague-Dawley male rats were orally administered with $AFB_1$, $AFB_2$, and $AFG_1$ at the dose of 250 ${\mu}g/kg$ (additionally including a dose of $1250{\mu}g/kg $ for $AFB_1$) body weight. Animals were then killed at 12, 24 or 48 hrs following aflatoxin exposure. Subsequently the immunohistochemical examination of p53, cytochrome p450 1A1 (CYP450 1A1), and glutathione-S-transferase placental form (GST-P) were performed. The level of the 8-OxodG in the liver was determined. Expressions of CYP450 1A1 and p53 were high in the liver of rats through 48 hrs after treatment of $AFB_1$ at the single dose of $250{\mu}g/kg $. This pattern was more clear as increasing doses. The treatment of $AFB_2$ and $AFG_1$ did not affect the expression of CYP450 1A1 but it caused weak expression of p53. The activity of GST were not found in the liver of rats treated with aflatoxins. The formation of 8-OxodG by $AFB_1$ increased in a dose-dependent manner up to 24 hrs after a single treatment of $AFB_1$ thereafter decreased to the level of control. The treatment of $AFB_2$ and $AFG_1$ did not affect the levels of 8-OxodG in the liver of rats with increasing time. These results in the present study indicate that $AFB_1$ among aflatoxins with low comparable levels is the most toxic as determined by early biomarkers such as CYP450 1A1, p53, GST-P, and 8-OxodG.

• Journal of the Korean Society of Food Science and Nutrition
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• v.31 no.5
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• pp.782-787
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• 2002

This study was done to investigate the effects of Korean mistletoe water extract and lectin on the apoptosis and preneoplastic lesion in chemically induced rat hepatocarcinogenesis. To attain the above objectives, weanling Sprague-Dawley male rats were fed modified AIN-76 diets containing 10% corn oil for 9 weeks. One week after feeding starts, rats were intraperitoneally injected twice with a dose of diethylnitrosamine (DEN, 50 mg/kg body weight (BW). Rats were provided with 0.05% phenobarbital (PB) in drinking water from one week after DEN treatment until the end of experiment. During the period of PB treatment, rats were injected with mistletoe extract (100 $\mu\textrm{g}$/kg BW) and lectin (10 $\mu\textrm{g}$/kg BW) twice a week. At the end of 9th week, rats were sacrificed and the formation of hepatic glutathione S-transferase placental form positive (GST-P$^{+}$) foci, apoptosis, DNA fragmentation and apoptosis related proteins were determined respectively. The formation of GST-P$^{+}$foci was significantly decreased by mistletoe extract or lectin treatment. Although there was no effect on apoptosis and DNA fragmentation in hepatic tissue by mistletoe extract or lectin treatment, caspase-9 and fas-L were increased. These results suggest that Korean mistletoe extract and lectin have a potential to inhibit hepatocarcinogenesis by increasing apoptosis.sis.

• Journal of Food Hygiene and Safety
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• v.16 no.3
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• pp.212-220
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• 2001

To investigate the modifying effect of Kwao Kreu, Pueraria mirifica (PM), we performed two kind of studies which are the non-surgical medium-term carcinogenicity study and the modulation of gap junctional intercellular communication study. The first study, a non-surgical medium-term carcinogenicity bioassay was done to investigate the modifying effect of Kwao Keru, Pueyaria mirifca (PH), a rejuvenating folk medicine from Thailand, on the male F344 rat liver. Specific pathogen free, male 6-week-old F3444 rats were divided into ten groups. To induce hepatocarcinogenesis, those in all groups were given a single i.p. injection of DEN (200 mg/kg) and were received two i.p. injection of DGA (300 mg/kg) at the ends of weeks 2 and 5. Rats of group 3-6 were given sodium phenobarbital (PB 0.05% in drink). A diet containing 10 mg/kg PM was given to group 2 during the post-initiation phase and to groups 4 and 5 during promotion and initiation phase, respectively. Group 6 was given the experimental diet alone throughout the experiment (8 weeks). Rats of group 7, 8, 9 and 10 were fed 1000 mg/kg PH in the same manner as group 2, 4, 5 and 6. All animals were sacrificed at 8 weeks after DEN administration. Result of the immunohistochemical staining of the glutathione S-transferase placental form (GST-p) indicated that the numbers and areas of the preneoplastic leisions were not significantly changed in all PM treatment group comparing to control group. Also the numbers and areas of GST-p positive foci among group 7, 8, 9 and 10 were not significantly changed in comparing to control group. To study the effect of PM on the modulation of gap junctional intercellular communication, the present study was performed scrape-loading dye transfer (SL/DT) assay in human keratinocytes. The results showed that PM could not modulate GJIC. These results indicate that Pueraria mirifica may have no carcinogenic effects on experimental hepatocarcinogenesis in rats and gap junctional intercellular communication in human keratinocyte.