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

• Journal of Korean Biological Nursing Science
• /
• 제3권2호
• /
• pp.21-33
• /
• 2001

The objective of this study was to examine the effects of antioxidant vitamins on the cellular oxidant damage by observing the mitogenicity in the mouse spleen and the strand breaks of DNA in mouse blood induced by $AFB_2$. Intraperitoneal(i.p.) injections of vitamin C(VC) of 10 mg/kg and vitamin E(VE) of 63.8 mg/kg were repeatedly administered to male ICR mice of 6 weeks old at intervals of 4 times every 2 days. After one hour vitamin treatments, $AFB_1$ of 0.4 mg/kg was injected into the $AFB_2$ plus vitamin treated groups in the same way. On the other hands, into the $AFB_2$ only treated group, only $AFB_2$ was injected without vitamins in the same method as above. The results of the experiment are as follows ; as regard to comet assay, DNA strand breaks were clearly present and they formatted a typical comet tail in the mice blood of the $AFB_2$ only treated groups. However, comet tails apparently disappeared in $AFB_2$ plus antioxidant vitamins treated groups since oxidant damage was controlled in an almost similar level to the control group. Mitogenicity of the spleen also showed a similar tendency as before, and these differences were more remarkably observed in the reaction against Con-A, which is a T-cell mitogen. In these data, the statistical significance was p<0.01. The LDL and VLDL levels were 408.72, 504.47 mg/dl respectively in the $AFB_2$ only treated groups. Compared with the $AFB_1$ only treated groups, those of $AFB_2$ plus antioxidant vitamin treated groups decreased to 272.06(VC), 305.28 mg/dl(VE), respectively. On the other hand, HDL levels were diminished to 32.60, 29.60 mg/dl in $AFB_2$ only treated groups, compared to 42.23, 41.14 mg/dl in the $AFB_2$ plus antioxidant vitamins treated groups. But, blood glucose levels were not statistically significant.

• Archives of Pharmacal Research
• /
• 제21권2호
• /
• pp.120-127
• /
• 1998

Peroxisome proliferators induce hepatic peroxisome proliferation and hepatic tumors in rodents. These chemicals increase the expression of the peroxisomal $\beta$-oxidation pathway and the cytochrome P-450 4A family, which metabolizes lipids, including eicosanoids. Peroxisome proliferators transiently induce increased cell proliferation in vivo. However, peroxisome proliferators are weakly mitogenic and are not co-mitogenic with epidermal growth factor (EGF) in cultured hepatocytes. Earlier study found that the peroxisome proliferator ciprofibrate is cornitogenic with eicosanoids. In order to study possible mechanisms of the comitogenicity of peroxisome proliferator ciprofibrate and eicosanoids' we hypothesized that the co-mitogenicity may result from synergistic or additive increases of second messengers in mitogenic signal pathways. We therefore examined the effect of the peroxisome proliferator ciprofibrate, prostaglandin $F_2_{\alpha}$($PGF_2{\alpha}$) and the combination of ciprofibrate and $PGF_2{\alpha}$ with or without growth factors on the protein kinase C (PKC) activity, and inositol-1, 4, 5-triphosphate ($IP_{3-}$) and intracellular calcium ($[Ca^{2+}]_i$) concentrations in cultured rat hepatocytes. The combination of ciprofibrate and $PGF_2{\alpha}$ significantly increased particulate PKC activity. The combination of ciprofibrate and $PGF_2{\alpha}$ also significantly increased EGF, transforming growth factor-$\alpha$ ($TGF_2{\alpha}$) and hepatic growth factor (HGF)-induced particulate PKC activity. The combination of ciprofibrate and $PGF_2_\alpha$greatly increased $[Ca^{2+}]_i$. However, the increases of PKC activity and $[Ca^{2+}]_i$ by ciprofibrate and $PGF_2{\alpha}$ alone were much smaller. Neither ciprofibrate or $PGF_2{\alpha}$ alone nor the combination of ciprofibrate and $PGF_2{\alpha}$ significantly increased the formation of $IP_3$. The combination of ciprofibrate and $PGF_2{\alpha}$, however, blocked the inhibitory effect of $TGF-{\beta}$ on particulate PKC activity and formation of $IP_3$ induced by EGF. These results show that co-mitogenicity of the peroxisome proliferator ciprofibrate and eicosanoids may result from the increase in particulate PKC activity and intracellular calcium concentration but not from the formation of $IP_3$.

• 약학회지
• /
• 제41권4호
• /
• pp.421-432
• /
• 1997

A new lectin was partially purified from starfish,Asterina pectinifera by means of physiological saline extraction, salt fractionation, ion exchange chromatography and hy droxyapatite chromatography, and it was named APL. The biochemical properties of the APL were characterized. In addition, its effects on lymphocyte mitogenicity and cancer cell agglutinability were tested. The APL agglutinated nonspecifically human erythrocytes and rabbit blood cells. Agglutinability was decreased to 30% of control activity below pH 5 and above pH 9 and was relatively unstable at increasing temperatures above 60$^{\circ}C$. The activity was reduced by addition of two kinds of metal ions, $Ba^{2+},\;Mn^{2+}$ and chelating agent, EDTA. APL was proved to be glycoproteins containing 9% sugars. For carbohydrate specificity, it was found that the activity of APL was inhibited by D(+)-glucosamine, D(+)-galactosamine, stachyose, N-acetyl-galactosamine and methyl-${\alpha}$-D-galactopyranoside among 35 sugars tested. In amino acid composition, the contents of acidic amino acids such as aspartic acid and glutamic acid were relatively high. This result suggest that the isoelectric point would be in a lower range. APL was found that it promotes the division of human lymphocytes. APL was proved to be a potent agglutinin for cancer cells such as HeLa, L929 and L1210 cells. Significant changes on the HeLa cell surfaces affected by APL were observed under the electron microscope.

• 약학회지
• /
• 제37권3호
• /
• pp.254-261
• /
• 1993

Developing new substance for immunosuppressor or immunomodulator from natural products is extremely important in the present biomedicine. In this paper, we focused our efforts on the mitogenicity and immunochemical properties of the two lectins (LCC-I, LCC-II) obtained from marine animal Lunella coronata coreensis. Immunochemical techniques were employed to elucidate the structural and/or functional similarities between the LCC lectins. Molecular weight of the LCC lectins, LCC-I and LCC-II were estimated to be around 60 KD and 66-70 KD, respectively. LCC lectins were mitogens for murine splenic lymphocytes, and the optimum mitogenic doses were 31.25 $\mu\textrm{g}$/ml and 3.91 $\mu\textrm{g}$/ml, respectively. LCC-II lectin was a good mitogen toward human peripheral lymphocytes at a concentration about 31.25 $\mu\textrm{g}$/ml.

• Nutrition Research and Practice
• /
• 제1권2호
• /
• pp.94-99
• /
• 2007

The immunostimulating activities of mucilage fraction from yam were investigated. The proliferation of BSA-primed lymph node cells was enhanced between 4.1- to 10.9-fold compare to control, when cultured with 1 to $25{\mu}g/mL$ of yam-mucilage fraction. It showed strong immunostimulating activity than ginseng extract and as remarkable as Bifidobacterium adolescentis M101-4 known as a positive immunostimulator. Mitogenicity to lymph node cells was fully induced by concanavalin A and lipopolysaccharide. The proliferation of splenocytes and Peyer's patch cells was enhanced between 5.0- to 14.1-fold and 2.4- to 6.4-fold, respectively, when cultured with 1 to $25{\mu}g/mL$ of yam-mucilage fraction. It enhanced the production of cytokines such as tumor necrosis $factor-{\alpha}$ and IL-6 in the culture of RAW 264.7 macrophage cells. In the culture of lipopolysaccharide-stimulated RAW 264.7 cells, production of cytokines was as similar as compared to controls. In unstimulated RAW 264.7 cells, both tumor necrosis $factor-{\alpha}$ and IL-6 production were enhanced between 15.6- to 60.1-fold and 2.3- to 9.1-fold, respectively. Mucilage fraction from yam is expected to be a safe immunopotentiator to maintain the host immunity and develop a physiologically functional food.

• Journal of Periodontal and Implant Science
• /
• 제34권2호
• /
• pp.461-474
• /
• 2004

The purpose of this study was to assess some biological activities of lipopolysaccharides (LPSs) from P. intermedia and P. nigrescens. LPS was prepared by the standard hot phenol-water method. NO production was assayed by measuring the accumulation of nitrite in culture supernatants. $TNF-{\alpha}$ production was determined by enzyme-linked immunosorbent assay. Western blot analysis of iNOS and analysis of reverse transcription (RT)-PCR products were carried out. LPS from P. intermedia demonstrated higher KDO content than those from two stains of P. nigrescens. LPSs from P. intermedia and P. nigrescens were mitogenic for spleen cells of BALB/C mouse. The present study clearly shows that LPSs from P. intermedia and P. nigrescens fully induced iNOS expression and NO production in RAW264.7 cells in the absence of other stimuli. Moreover, LPSs from P. intermedia and P. nigrescens clearly induced $TNF-{\alpha}$ production in RAW264.7 cells. The biological activities of LPS from P. intermedia was found to be comparable to those of P. nigrescens LPS. The ability of LPSs from P. intermedia and P. nigrescens to promote the production of NO and $TNF-{\alpha}$ may be important in the pathogenesis of inflammatory periodontal disease.

• KSBB Journal
• /
• 제16권6호
• /
• pp.547-553
• /
• 2001

본 연구에서는 민방이나 한방에서 천연 약재로 많이 사용하고 있는 느릅나무 뿌리껍질(유근피)로부터 수용액 추출물을 분리하여 이 중 단백다당체를 분리하여 성분분석 및 단백다당체가 갖고 있는 면역활성에 의한 항암효과를 실험적으로 제시하였다. 느릅나무 추출물 당단백질의 화학적 조성을 보면 총당함량은 55.8∼72.1%였고, 총산성당 함량은 30.0∼30.5%, 총단백질 함량은 5.0∼6.1%으로 수용성 고분자이며, 50 내지 500 kDa 범위의 분자량을 나타내었다. 그리고, 단백다당체를 구성하는 주요 당성분으로는 글루크론산, 람노스아라비노스, 글루코스, 갈락토스 등이었다. 단백다당체의 면역활성 검정에 따르면 면역세포의 증식을 자극하는 mitogen 역할을 하며, 특히 T세포에 의한 면역증강 효과를 나타내었다 B16 흑색종 이식 마우스를 이용한 생체내 면역활성에 의한 항암효과를 조사한 결과 유의성 있는 항암효과를 나타내었으며, 느릅나무 추출물인 단백다당체가 3mg/kg 군에서 가장 강한 항암효과를 나타내어 용매대조군에 대해 약 140% 생존연장효과를 나타내었다.

• 한국식품영양과학회지
• /
• 제33권9호
• /
• pp.1552-1559
• /
• 2004

새로운 probiotic 유산균으로써의 잠재적 이용가능성을 평가할 목적으로, 김치로부터 분리한 Lactobacillus brevis FSB-1을 대상으로 각종 면역증진활성의 특성이 조사되었다. L. brevis FSB-1을 전균체, 세포벽, 세포질 및 균체외 획분으로 각각 분리하고 장관면역 활성을 측정한 결과, Peyer's patch 세포를 매개로 한 골수세포 증식활성의 경우, 세포벽 및 세포질 획분에서 상대적으로 높은 활성을 농도 의존적으로 보인 반면, 직접적인 골수세포 증식활성은 나타내지 않았다. 마크로파지의 활성화능은 전균체, 세포벽 및 세포질 획분에서 상대적으로 높은 활성을 보였으며, splenocyte mitogen 활성의 경우, 이들 획분에서 공히 대조군의 약 200%이상의 활성 증가가 관찰되었다. 그러나 양성대조군인 LPS의 활성에는 다소 미치지 못하였다. 한편 보체계 활성화능을 검토한 결과, 균체외 획분을 제외한 모든 획분에서 높은 활성을 보였으며, 특히 세포질 획분에서 농도 의존적으로 매우 강력한 활성을 나타냈다. 또한 세포질 획분에 의한 보체계 활성화는 anti-human C3를 이용한 2차원 면역전기영동에 의해 classical 및 alternative pathway 양 경로를 경유함을 확인할 수 있었다.

• 한국식품과학회지
• /
• 제44권3호
• /
• pp.343-349
• /
• 2012

한국 전통발효음료 중에 존재하는 특이 다당류의 화학적 특성 및 생물활성을 규명할 목적으로, 농가에서 직접 발효한 복분자주로부터 다당류를 분리하고 이들의 각종 면역증강활성에 대해 검토하였다. 복분자주에 80% ethanol 침전을 행하여 얻어진 조다당획분 RB-0를 이용, 연속적인 3회의 column chromatography를 행하여, 복분자주의 주요 다당인 RB-1b-I으로 정제할 수 있었다. RB-1b-I획분은 HPLC상에서 대칭을 유지하는 단일 peak로 검출되었으며, 분자량은 약 180 kDa으로 평가되었다. 정제다당인 RB-1b-I의 구성당 조성을 확인한 결과, mannose가 76.8%로 주를 이루고 있었으며, 그 외에 galactose(15.8%), arabinose(3.8%), glucose(2.6%) 및 rhamnose(1.2%)의 순으로 함유되어 있었다. 이러한 결과는 복분자 중에 존재하는 주 다당이 발효 효모의 세포벽에서 기원한 mannan임을 추정하게 하였다. 한편 복분자주에서 정제한 다당 RB-1b-I은 in vitro상에서 macrophage의 활성화를 높은 비율로 유도하였으며, 비장세포에 대해 높은 증식능을 보였다. 또한 RB-1b-I을 정맥 투여한 경우, NK cell을 활성화하여 YAC-1 종양세포에 대한 세포독성을 증가시킴이 관찰되었다. 한편 고분자 정제 다당인 RV-1b-I은 비특이적 면역계에 있어 중요 역할을 담당하고 있는 보체계에 대하여 농도 의존적인 활성화 경향을 보였다. 이들은 Ca++ 이온이 제거된 상태에서의 항보체 활성과 antihuman C3를 이용한 2차원 면역전기영동에 의하여 C3 산물을 동정한 결과로부터 보체계의 classical pathway와 alternative pathway양 경로를 모두 경유하여 활성을 나타냄을 확인할 수 있었다. 이상의 결과로부터 복분자주에는 인체 건강에 유익한 면역증진 활성을 제공하는 다당류가 함유되어 있음을 알 수 있었다.