• 대한수의학회지
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• 제27권1호
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• pp.53-60
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• 1987

This experiment was performed on mice to investigate the effects of an immunopotentiator, picibanil(PC), on the immune responses such as phagocytic activity of reticuloen-dothelial(RE) system, E rosette formation rate of splenic lmphocytes and morphological changes of lymph node tissue. Groups of mice were treated with a single(1KE/kg BW) or sequential(0.1, 0.25 and 0.5KE/kg BW for successive 3 days) intravenous injections of PC. PC treated and untreated control mice were sensitized with 50% sheep erythrocyte suspension(0.2ml/mouse) at 1, 3, 5, 7 and 10 days after PC treatment. Functional and morphological examinations were carried out 5 days after sensitization. The following results were obtained: The phagocytic activity of RE system and the weight of liver and spleen were increased significantly at 3rd, 5th and 7th day. The peripheral polymorphonuclear leukocyte and percent of lymphocyte and monocyte were slightly increased. The rates of E rosette formation of splenic lymphotytes, sequential PC treated groups were more increased at 3rd and 5th day in sequential PC treated groups than in single treated groups. Thereafter it returned gradually to the control level by the time of 10th day. Microscopically primary lymph follicles with indistinct germinal center (GC) were partially disrupted and the parafollicular areas were consisted of the pyroninophilic cells in control group. In PC treated group, the parafollicular areas were markedly proliferated and developments of secondary lymph follicles with enlarged and prominent GC were more pronounced in the sequential injected groups compared to single injected groups. These results indicate that PC affected not only parafollicular area of the T-cell area, but also GC of the B-cell area. It suggests that PC may potentiate both cell mediated immunity and humoral immunity.

• 약학회지
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• 제42권3호
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• pp.248-256
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• 1998

We had already reported that we purified N-${\beta}$-alanyl-5-S-glutathionyl-3,4-dihydroxyphenylalanine (5-S-GAD), a novel antibacterial substance from the immunized adult Sarcoph aga peregrina (Flesh fly). We found that the antibacterial activity of synthetic 5-S-GAD is equal to that of authentic 5-S-GAD without a specificity of antibacterial activity against Gram positive and Gram negative. Significant synergism was detected between 5-S-GAD and streptomycin against streptomycin resistant strain E.coli K12 594. It has an antitumor activity against several tumor cell lines at a concentration of $100{\mu}M$. However, no cytotoxic activity against murine macrophage was detected at a concentration of $500{\mu}M$. Furthermore, haemolytic activity against sheep erythrocytes was not detected at the same concentration. We suggest that the S-conjugation of glutathion with dihydroxyphenylalanine might be important to increase antibacterial activity of dihydroxyphenylalanme.

• Biomolecules & Therapeutics
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• 제2권3호
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• pp.236-243
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• 1994

A splenocyte culture system supplemented with liver microsomes was developed to detect immunotoxic chemicals which require metabolic activation using cyclophosphamide as a positive standard. When liver microsomes were added to splenocyte cultures isolated from female B6C3Fl mice, the proliferation of splenocytes by lipopolysaccharide (LPS) was increased and the proliferation by concanavalin A (Con A) was decreased. However, when compared with each corresponding control, cyclophophamide was successfully activated to metabolites capable of suppressing Iymphoproliferative responses. This suppression was clearly dependent upon the amounts of microsomes added and/or the concentration of cyclophosphamide exposed. In these cultures, the proliferation of splenocytes was suppressed when the cells were exposed to cyclophosphamide on the day of culture initiation. On the other hand, microsome was responsible for the increase in LPS mitogenicity and NADPH was responsible for the decrease in Con A mitogenicity. Finally, our present culture system was compared with the hepatocyte-splenocyte coculture system which we had developed earlier. We found that the hepatocyte-splenocyte coculture was better able to activate cyclophosphamide to metabolites capable of suppressing the antibody response to sheep erythrocytes. Although our present culture system was relatively poor to activate cyclophosphamide in cultures for antibody response, it will be useful as a simple screening method to detect suppression of certain in vitro immunotoxic parameters like LPS mitogenicity by chemicals which require metabolism.

• 대한수의학회지
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• 제7권2호
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• pp.8-12
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• 1967

Formalin 처리(處理) 면양적혈구(緬羊赤血球)를 용혈소(溶血素) 생산용(生産用) 항원(抗原)으로 가토(家兎)의 이정맥(耳靜脈)에 접종(接種)하였던바 고가(高價)의 양성혈청(陽性血淸)을 얻을수 있었음으로 보고(報告)하는 바이다. Formalin 0.5% 식염수(食鹽水)에 하룻밤동안 $37^{\circ}C$ 항온수조(恒溫水槽)에서 처리(處理)한 면양적혈구(緬羊赤血球)를 정유액(淨遊液) 1.0ml. 2.0ml. 3.0ml. 4.0ml.을 4일간격(日間隔)으로 4회(回) 가토(家兎)의 이정맥(耳靜脈)에 접종(接種)하였으며 최종항원접종후(最終抗原接種後) 5일(日)만에 채혈(採血)하므로서 10,000단위(單位) 이상(以上)의 용혈소력가(溶血素力價)를 갖는 가토혈청(家兎血淸)을 생산(生産)할 수 있었다. 이 방법(方法)에 의(依)하면 항원(抗原)의 제조(製造)나 면역방법(免疫方法)이 지금까지 알려져온 어떤 방법(方法)보다 더 간편(簡便)하며 가토(家兎)에 대(對)한 예민성(銳敏性) 반응(反應)이 거의 없으며 개체차이(個體差異)없이 용혈소(溶血素)를 생산(生産)하는 것이 특징이다.

• 대한수의학회지
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• 제8권2호
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• pp.125-146
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• 1968

Throughout the studies the following experimental results were summarized. 1. It was impossible to infect and kill the mice, weighing 10 to 12 gm, by inoculating 0.2ml of virulent Cl. chauvoei, diluted 1 to 10 with physiological saline, via subcutaneous, intramuscular, intraperitoneal or intraveonus, route. 2. The mice which were inoculated in brain with 0.03ml of Cl. chauvoei diluted 1 : 5120 with physiological saline were resulted in all death after infection, but not in case of attenuated strain even in dilution of one to five. 3. Virulent Cl. chauvoei were diluted with each of those of whole blood, erythrocytes and serum of horse, calf, swine, sheep, rabbit, guinea pig, chicken and duck, human plasma and 2% CaCl solution, and inoculated subcutaneously 0.25 to 0.5ml in mice, weighing 12 to 15gm. It was resulted in significant increase in virulence as comparing with the case of physiological saline solution except when horse and pig sera were used. Such a phenomena were not seen in attenuated strain. 4. Virulence of virulent Cl. Chauvoei could be increased significantly in rat, as the procedures used in mice, by suspending in whole blood, erythrocytes, serum, or plasma of various animals, or 2% $CaCl_2$ solution and by inoculating subcutaneously 0.5 to 10ml in rat, weighing 30 to 60 gm, as compared with those of control group which used physiological saline solutionos diluent. 5. Mice resisted 100 and 80 percent against challenge of $10^3$ and $10^4$ M.L.D.. respectively, 24 hours after inoculation of 0.5ml black leg antiserum. 6. Immune response to the black leg living vaccine in mice could be obtained more favorably in the group of respected vaccination rather than those of single inoculation and the most profitable inoculm size of the vacine was 0.5 to 1.0ml. 7. Challenge for the immunized mice could be carried out effectively 3 weeks after first vaccination. 8. Satisfactory results could be obtained by inoculating subcutaneously for the immunization and intracerebrally or subcutaneously for the challenge. 9. Mice which were inoculated with 0.5ml of black leg living vaccine via subtaneucously two times at seven days interval and 21 days after first inoculation and challenged with 5 and 10 M.L.D. of virulent strain, resited 100 and 70 to 80 percent respectively. Same results were obtainable in black leg killed vaccine as the procedures used in living vaccine. 10. There were significantly different resistances against the definite challenge does between the mice groups which were immnuized with the living vaccine diluted five or 10 times and the undiluted. 11. For the biological assay of black leg living vaccine and antiserum, satisfactory results could be obtained using mice.

• Applied Biological Chemistry
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• 제39권2호
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• pp.147-152
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• 1996

천연으로부터 보체 저해활성 물질을 선발할 목적으로 용혈성 보체측정법(hemolytic complement assay, $CH_{50}$)을 이용한 미량 탐색법을 확립하였다. 보체결합 반응은 microplate에서 수행하였으며, 표준 용혈(50% hemolysis)을 설정하기 위한 최적조건은 classical pathway (CP)의 경우 양 적혈구 농도를 $5.0{\times}10^8\;cells/ml$로 하였을때 hemolysin 희석농도 $1/75{\sim}1/100$ 및 보체 희석농도 $1/80{\sim}1/120$이었으며, alternative pathway (AP)의 경우는 토끼 적혈구 농도 $4.0{\times}10^8\;cells/ml$에서 보체 희석농도 l/5, EGTA 4mM 및 $Mg^{2+}\;4{\sim}8mM$ 이었다. 비수용성 화합물의 검정에는 dimethyl sulfoxide를 1% 수준이 되도록 시료의 농도를 조절하여 첨가하였다. 본 실험 조건에서 3종의 phenylpropanoid는 모두 농도에 비례하여 항보체 활성을 나타내었는데, 그중 rosmarinic acid의 항보체 활성은 CP의 경우 $0.063{\sim}0.5mM$에서 $5.4{\pm}3.6%{\sim}95.8{\pm}0.2%$, AP의 경우 $0.063{\sim}1\;mM$에서 $35.1{\pm}0.9%{\sim}95.6{\pm}1.1%$으로 측정되었다.

• Restorative Dentistry and Endodontics
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• 제30권4호
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• pp.335-343
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• 2005

세균의 용혈활성은 세균이 숙주 내에서 생존하기 위해 필요한 철을 획득하기 위한 특성이며 기능 면에서 볼 때 숙주에 대한 중요한 독력인자로 간주될 수 있다. 본 연구에서는 괴사치수 및 치근단 치주염으로 진단된 환자의 근관에서 분리한 Prevotella nigrescens의 용혈활성을 다양한 조건 하에서 측정하여 그 특성을 규명하는 것을 목적으로 한다. 1. 임상에서 분리한 P. nigrescens와 표준균주인 P. nigrescens ATCC 33563에서 모두 용혈활성이 나타났다. 2. 사람, 면양 및 말 세 가지 종에 대해 용혈활성을 비교한 결과 사람의 적혈구에서 가장 강한 용혈활성을 나타내었다. 3. 용혈소 억제제인 $NaN_3$와 dithiothreitol(DTT)는 농도의존적으로 P. nigrescens의 용혈활성을 감소시켰다 (p<0.05) 4. P. nigrescens가 최대 용혈활성을 나타내는 최적의 pH는 4이었으며, $50^{\circ}C$이하의 온도에서는 용혈활성을 보였으나 $95^{\circ}C$에서 급격히 감소하였다. 5.배양조건에 따른 P. nigrescens의 용혈활성을 비교한 결과 $10\%\;CO_2$배양기에 배양한 경우 혐기성 조건에서 배양한 것보다 더 높은 용혈활성을 보였다.

• Journal of Chest Surgery
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• 제32권11호
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• pp.978-983
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• 1999

Background: Complement activation with transpulmonary leukocyte sequestration is considered a main mediator leading to ischemia-reperfusion lung(I-R) injury. We studied the role of leukocytes in the formation of I-R injury in ovine cardiopulmonary bypass(CPB) model with a membrane oxygenator. Material and Method: Five sheep were used. CPB circuitry consisted of a roller pump(American Optical Corp., Greenwich, CT, USA) and a membrane oxygenator(UNIVOX-IC, Bentley, Baxter Health Corp, Irvine, CA, USA). The CPB time was fixed at 120 min. Ten minutes after the start of CPB, total CPB was established. Thereafter a total CPB of 100 min was performed, followed by another 10 min of partial CPB. The CPB was discontinued and the animals were fully recovered. For measuring left and right atrial leukocyte counts, blood samples were taken before thoracotomy, 5 min and 109 in after the start of CPB, and 30 min and 120 min after weaning. C3a was measured before thoracotomy, 109 min after the start of CPB, and 30 min and 120 min after weaning. Plasma malondialdehyde(MDA) was checked before thoracotomy, 109 min after the start of CPB, and 30 min after weaning. One to two grams of lung tissue were taken for water content measurement before thoracotomy, 109 min after the start of CPB, and 30 min after weaning. Lung biopsy specimens were examined by light and electron microscopy. Result: Of 5 animals, 4 survived the experimental procedures. Of these, 3 animals survived on a long-term basis. No significant differences in transpulmonary gradients of leukocyte were found and no significant complement activation was expressed by C3a levels. MDA level did not show significant changes related to lung reperfusion despite an increase after the start of CPB. On both light and electron microscopic examinations, mild to moderate acute lung change was observed. Interstitial edema, leakage of erythrocytes into the alveolar space and endothelial cell swelling were the main findings. Water content of the lung showed a slight increase after the start of CPB, but there was no statistical significance. Conclusion: These findings indicate that ischemia-repersusion lung injury may not be from complement activation-leukocyte sequestration but from another source of oxygen free radicals related to CPB.

• Biomolecules & Therapeutics
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• 제22권2호
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• pp.149-154
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• 2014

Effects of diallyl sulfide (DAS) on thioacetamide-induced hepatotoxicity and immunotoxicity were investigated. When male Sprague-Dawley rats were treated orally with 100, 200 and 400 mg/kg of DAS in corn oil for three consecutive days, the activity of cytochrome P450 (CYP) 2E1-selective p-nitrophenol hydroxylase was dose-dependently suppressed. In addition, the activities of CYP 2B-selective benzyloxyresorufin O-debenzylase and pentoxyresorufin O-depentylase were significantly induced by the treatment with DAS. Western immunoblotting analyses also indicated the suppression of CYP 2E1 protein and/or the induction of CYP 2B protein by DAS. To investigate a possible role of metabolic activation by CYP enzymes in thioacetamide-induced hepatotoxicity, rats were pre-treated with 400 mg/kg of DAS for 3 days, followed by a single intraperitoneal treatment with 100 and 200 mg/kg of thioacetamide in saline for 24 hr. The activities of serum alanine aminotransferase and aspartate aminotransferase significantly elevated by thioacetamide were protected in DAS-pretreated animals. Likewise, the suppressed antibody response to sheep erythrocytes by thioacetamide was protected by DAS pretreatment in female BALB/c mice. Taken together, our present results indicated that thioacetamide might be activated to its toxic metabolite(s) by CYP 2E1, not by CYP 2B, in rats and mice.

• BMB Reports
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• 제41권2호
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• pp.119-125
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• 2008

Our previous work confirmed that Sph2/LA1029 was a sphigomyelinase-like hemolyisn of Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai. Characteristics of both hemolytic and cytotoxic activities of Sph2 were reported in this paper. Sph2 was a heat-labile neutral hemolysin and had similar hemolytic behavior as the typical sphingomyelinase C of Staphylococcus aureus upon sheep erythrocytes. The cytotoxic activity of Sph2 was shown in mammalian cells such as BALB/C mouse lymphocytes and macrophages, as well as human L-02 liver cells. Transmission electron microscopic observation showed that the Sph2 treated BALB/C mouse lymphocytes were swollen and ruptured with membrane breakage. They also demonstrated condensed chromatin as a high-density area. Cytoskeleton changes were observed via fluorescence confocal microscope in Sph2 treated BALB/C mouse lymphocytes and macrophages, where both cytokine IL-$1{\beta}$ and IL-6 were induced. In addition, typical apoptotic morphological features were observed in Sph2 treated L-02 cells via transmission electron microscope and the percentage of apoptotic cells did increase after the Sph2 treatment detected by flow cytometry. Therefore, Sph2 was likely an apoptosis-inducing factor of human L-02 liver cells.