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A STUDY ON THE EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR IN TENS10N SIDE OF RAT MOLAR PERIODONTAL LIGAMENT FOLLOWING EXPERIMENTAL TOOTH MOVEMENT (백서 구치의 실험적 치아이동시 견인측 치근막에서 혈관성장인자의 발현에 관한 연구)

  • Lim, Yong-Kyu;Shin, Choon-Shik;Lee, Dong-Ryul
    • The korean journal of orthodontics
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    • v.31 no.1 s.84
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    • pp.121-136
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    • 2001
  • This study was performed to analyse the expression of VEGF and it's receptor(VEGFR) in the tension side of the periodontal ligament following orthodontic tooth movement. Upper first molars of Sprague-Dawley rats were moved medially using closed coil spring for 1, 2, 24 hours and 3, 7, 14 days. H&E staining, immunohistochemical staining and in situ hybridization methods were used to analyse the change of the expression of VEGF and VEGFR. The results from this study were as follows : 1. Following tensional force, periodontal ligament showed elongation of fibers, compression and congestion of vessels and regional hemorrhage. These tissue changes were recovered within 3 days of force application. New bone formation was seen after 3 days of force application and continued for the remaining experimental periods. 2. Following tensional force, VEGF and VEGF mRNA expression was increased in the periodontal ligament cells, osteoblasts and cementoblasts. This change was followed by increased vasculature in the periodontal ligament. 3. After 3 days of tensional force, VEGF and VEGF mRNA expression was confined mainly to the osteopaths and the periodontal ligament cells adjacent to the alveolar bone. After 2 weeks of force application, VEGF and VEGF mRNA expression was reduced to the level of control sample. 4. VEGFRs(Flt-1, Flk-1) showed similar expression pattern and it's expression was mainly seen in the endothelial cells and osteoblasts. Following tensional force VEGFR expression was increased in the endothelial cells and osteoblasts. In conclusion, in the tension side of the penodontal ligament, ligament cells, osteoblast and cementoblast showed increased expression of VEGF & VEGF mRNA. It preceded the increase of vasculature and new bone formation. The increased expression of VEGF mRNA in cementoblast may induce periodontal vessels, which distribute mainly the bone side half of periodontal ligament, grow in the direction of tensional force. Increased expression of VEGFR & VEGFR mRNA not only in endothelial cell but in osteoblast, osteocyte and periodontal cells showed VEGF acts not only in paracrine manner but in autocrine one.

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Changes of CGRP immunoreactivity in rat trigeminal ganglion neurons during tooth movement (백서 삼차신경절내 신경세포체의 치아이동에 따른 CGRP 면역염색성의 변화)

  • Park, Chyo-Sang;Park, Guk-Phil;Sung, Jae-Hyun
    • The korean journal of orthodontics
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    • v.27 no.4 s.63
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    • pp.607-621
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    • 1997
  • GRP was known as the modulator of Pain transmission in central nervous system and local effector to peripheral tissue causing vasodilation, increased blood flow, modulation of immune sysem, stimulation of endothelial cell proliferation, and stimulation of bone formation. Numerous study, therefore, were done to elucidate involvement of CGRP to tooth movement. To investgate the response of CGRP immunoreactive nerve cells according to cell size in trigeminal ganglion during tooth movement, immunohistochemical study was performed using rat. Experimental rats(9 weeks old, 210 gm) were divided as six groups(normal(n=6), 3 hour group(n=5), 12 hour group(n=4), 1 day group(n=5), 3 day group(n=5), 7 day group(n=5)), and were applied orthodontic force (approximately 30 gm) to upper right maxillary molar. After frozen sections of trigeminal ganglions were immunostained using rabbit antisera, the changes of CGRP immunoreactive cells in regard to cell size distribution(small cell(upto $20{\mu}m$), medium cell($20-35{\mu}m$), large cell(above $35{\mu}m$)) were observed. The results were as follows 1. The percentage of CGRP immunoreactive cells to all nerve cells in trigeminal ganglion was 33.0% in normal control group, was decreased to 24.5% in 1 day group, and was increased to 41.8% in 7 day group. 2. The percentage of small, medium, and large cells expressing CGRP immunoreactivity in normal trigeminal ganglion to all CGRP immunoreactive cells were 51.3%, 44.0%, 4.7%, respectively. 3. The percentage of small cells with CGRP immunoreactivity to all CGRP immunopositive cells was increased in 3 hour and 12 hour groups. 4. The percentage of medium cells with CGRP immunoreactivity was increaed in 3 day and 7 day groups. 5. The percentage of large cells with CGRP immunoreactivity was increaed in 7 day group. Conclusively, the small cells with CGRP immunoreactivity in trigeminal ganglion respond to orthodontic force during initial phase of tooth movement, and later the medium and large cells with CGRP immunoreactivity respond

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The Role and Localization of Nitric Oxide Synthase in Neurogenic Inflammation of the Rat Airways (백서의 기도 선경성 염증에서 산화질소 합성효소(Nitric Oxide Synthase)의 역할과 분포)

  • Shim, Jae-Jeong;Lee, Sang-Yub;Lee, Sang-Hwa;Suh, Jung-Kyung;Kim, Chul-Hwan;Cho, Jae-Youn;In, Kwang-Ho;Yoo, Seo-Hwa;Kang, Kyung-Ho
    • Tuberculosis and Respiratory Diseases
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    • v.43 no.3
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    • pp.420-433
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    • 1996
  • Background : There have been many debates about the effects of nitric oxide on the neurogenic inflammation. The role of nitric oxide in the neurogenic inflammation of airways will be required a better understanding of the localization and types of nitirc oxide synthase(NOS) activity in the neurogenic inflammation of airways. Method : To investigate the role of nitric oxide in airway neurogenic inflammation, 1) the effects of neurokinin receptor antagonist (FK224) and nitric oxide synthase inhibitor, $N^{\omega}$-nitro-L-arginine (L-NNA) on plasma extravastion were evaluated in four groups of Sprague-Dawley rats ; sham operation group(sham NANC group), electrical vagal stimulation group(NANC2 group), intravenous pretreatment groups with FK224 (1mg/kg ; FK224 group), and L-NNA(1mg/kg ; L-NNA group) 15 minutes before vagal NANC stimulation. 2) NOS activity in trachea with neurogenic inflammation was localized by immunohistochemical stain. Immunohistochemical stain was performed by antibodies specific for inflammatory cells(iNOS), brain(bNOS), and endothelium (eNOS) on trachea obtained from sham NANC, NANC2, and FK224 groups. Results : The results are that plasma extravsation in neurogenic inflammation of rat airways was inhibited by FK224, but enhanced by L-NNA pretreatment(P<0.05). There was significantly increased infiltration of inflammatory cells in subepithelium of neurogenic inflammatory trachea, but the reduction of subepithelial infiltration of inflammatory cells was observed after pretreatment with FK224(P<0.05). Immunostaining with anti-iNOS antibody showed strong reactivity only in infiltrated inflammatory cells in neurogenic rat trachea, and these iNOS reactivity was reduced by pretreatment with FK224. bNOS immunoreactivity was significantly increased only in the nerves both of neurogenic inflammatory and FK224 pretreated trachea compared with sham NANC trachea(p<0.05). eNOS immunoreactivity was not significant change in endothelium in neurogenic inflammation of rat trachea. Conclusion : These results suggest that nitric oxide released from iNOS in infiltrated inflammatory cells has main role in neurogenic inflammation of rat trachea. The presence of bNOS immunoreactivity in the nerves indicates that nitric oxide may be released from the nerves in rat trachea with neurogenic inflammation.

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Comparative Quantitative Study of Surfactant Protein C mRNA by Filter Hybridization and Solution Hybridization in Rats (Filter Hybridization과 Solution Hybridization 방법에 의한 백서 Surfactant Protein C mRNA 정량측정의 비교)

  • Kim, Jin-Ho;Sohn, Jang-Won;Yang, Seok-Chul;Yoon, Ho-Joo;Shin, Dong-Ho;Park, Sung-Soo
    • Tuberculosis and Respiratory Diseases
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    • v.51 no.6
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    • pp.517-529
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    • 2001
  • Background : Surfactant protein C(SP-C) is a hydrophobic 5,000 dalton molecule. SP-C has the primary roles in accelerating surface spreading of a surfactant phospholipid. The filter hybridization and solution hybridization assays are both rapid and sensitive and can be used to measure the RNAs complementary to any cloned DNA sequence. Methods : The authors measured the SP-C mRNA levels quantitatively using solution hybridization and filter hybridization assays to obtain a standard curve equation to quantify the mRNA of unknown samples comparatively. Results : 1. The minimum level of the specimens by solution hybridization was 3 pg for SP-C mRNA. 2. The standard curve equation of the solution hybridization assay between the counts per minute(Y) and the SP-C mRNA transcript input(X) was Y=6.46 X+244. The correlation coefficient was 0.99. 3. The minimum detection level of specimens by filter hybridization was 0.1 ng for SP-C mRNA. 4. The standard curve equation of the filter hybridization assay between the counts per minute(Y) and SP-C mRNA transcript input(X) is Y=2541.6 X+252.7. The correlation coefficient was 0.99. Conclusions : A comparison of CPM/filter in the linear range allowed an accurate and reproducible estimation of the SP-C mRNA copy number. Filter hybridization and solution hybridization assays are both rapid and sensitive and can be used to measure the RNAs complementary to any cloned DNA sequence. It is ideally suited to situations where accurate quantitation of multiple samples is required.

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Inhibition of Viability and Genetic Change in Hypoxia-treated Lung Pericytes (허파혈관주위세포에서 저산소증에 의한 생존능의 억제와 유전자 발현의 변화)

  • Shin, Jong Wook;Kim, Kae-Young;Lee, Young Woo;Jung, Jae Woo;Lee, Byoung Jun;Kim, Jae-Yeol;Jo, Inho;Park, In Won;Choi, Byoung Whui
    • Tuberculosis and Respiratory Diseases
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    • v.57 no.1
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    • pp.37-46
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    • 2004
  • Background : Lung pericytes are important constituent cells of blood-air barrier in pulmonary microvasculature. These cells take part in the control of vascular contractility and permeability. In this study, it was hypothesized that change of lung pericytes might be attributable to pathologic change in microvasculature in acute lung injury. The purpose of this study was how hypoxia change proliferation and genetic expression in lung pericytes. Methods : From the lungs of several Sprague-Dawley rats, performed the primary culture of lung pericytes and subculture. Characteristics of lung pericytes were confirmed with stellate shape in light microscopy and immunocytochemistry. 2% concentration of oxygen and $200{\mu}M$ $CoCl_2$ were treated to cells. Tryphan blue method and reverse transcription-polymerase chain reaction were done. Results : 1. We established methodology for primary culture of lung pericytes. 2. Hypoxia inhibited cellular proliferation in pericytes. 3. Hypoxia could markedly induce vascular endothelial growth factor(VEGF) and smad-2. 4. Hypoxia-inducible factor-$1{\alpha}$(HIF-$1{\alpha}$) was also induced by 2% oxygen. Conclusion : Viability of lung pericytes are inhibited by hypoxia. Hypoxia can stimulate expression of hypoxia-responsive genes. Pericytic change may be contributed to dysfunction of alveolar-capillary barrier in various pulmonary disorders.

Lipase-Inhibitory and Anti-Oxidative Activity of the Methanol Extract and the Powder of Phellinus linteus (상황버섯 자실체 메탄올 추출물과 분말의 지방소화효소 억제 및 항산화 활성)

  • Kim, Ji-Hyun;Son, In-Suk;Kim, Jong-Sang;Kim, Ki-Hoon;Kwon, Chong-Suk
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.37 no.2
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    • pp.154-161
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    • 2008
  • Phellinus linteus (PL) has been known to exhibit potent biological activity. The present study was designed to investigate lipase-inhibitory and anti-oxidative activity of the methanol extract and the powder of PL fruiting body. The methanol extract of PL appeared to have the inhibitory activity against pancreatic lipase with an $IC_{50}$ value of $36.3\;{\mu}g/mL$, and the scavenging activity of DPPH radical with an $IC_{50}$ value of $20.1\;{\mu}g/mL$, which was similar to that of vitamin C ($IC_{50}\;18.3\;{\mu}g/mL$). To investigate the lipase-inhibitory and anti-oxidative effect of PL on animal, Sprague-Dawley rats were fed with high-fat diet supplemented with either 2% or 5% PL powder for 8 weeks. Total food intake was significantly increased, but body weight was not changed by PL powder supplementation. However, fecal fat excretion of the experimental groups fed with the PL powder were higher than that of the control group. PL powder showed a decrease in the plasma total cholesterol, LDL-cholesterol, and the hepatic total cholesterol levels. The anti-oxidative enzyme activities were also affected by PL supplementation. Glutathione peroxidase (GSH-Px) in the plasma and liver were significantly increased by 98% and 46% in the 2% PL group, and 99% and 32% in the 5% PL group, respectively. Total superoxide dismutase (T-SOD) activity was not affected by PL supplementation. DNA damage was measured by the comet assay in the lymphocytes collected after 2 weeks, 4 weeks, and 8 weeks of feeding PL supplemented diet. Lymphocyte DNA damage was decreased in the PL supplemented group. Furthermore, PL feeding enhanced the resistance to lymphocyte DNA damage caused by an oxidant challenge with $H_2O_2$.

STUDY OF RAT EPIGASTRIC VESSELS ACCORDING TO THE FREEZING TIME : HISTOLOGIC, HISTOMORPHOMETRIC, IMMUNOHISTOCHEMICAL & SCANNING ELECTRON MICROSCOPIC STUDY (백서 상복부 혈관의 동결시간에 따른 변화에 대한 연구)

  • Kim, Woo-Chan;Lee, Chong-Heon;Kim, Kyung-Wook;Kim, Chang-Jin
    • Maxillofacial Plastic and Reconstructive Surgery
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    • v.21 no.2
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    • pp.89-109
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    • 1999
  • Vascular spasm which has been reported to occur in 25% of clinical cases continues to be a problem in microvascular surgery; When prolonged and not corrected, it can lead to low flow, thrombosis, and replant or free flap failure. Ischemia, intimal damage, acidosis and hypovolemia have been implicated as contributors to the vascular spasm. Although much work has been done on the etiology and prevention of vasospasm, a spasmolytic agent capable of firmly protecting against or reversing vasospasm has not been found. Therefore vascular freezing was introduced as a new safe method that immediately and permanently relieves the vasospasm and can be applied to microsurgical transfers. Cryosurgery can be defined as the deliberate destruction of diseased tissue or relief the vascular spasm in microvascular surgery by freezing in a controlled manner. 96 Sprague Dawley rats each weighing within 250g were used and divided into 2 group, experimental 1 and 2 group. In the experimental 1 group, right epigastric vessels (artery and vein) were freezed with a cryoprobe using $N_2O$ gas for 1 min. In the experimental 2 group, after freezing for 1 min, thawing for 30 secs and repeat freezing for 30 secs. Left side was chosen as control group in both group. We sacrified the experimental animals by 1 day, 3 days, 1 week, 2 weeks, 4 weeks & 5 months and observed the sequential change that occur during regeneration of epigastric vessels using a histologic, histomorphometric, immunohistochemical and SEM study after the vascular freezing. The results were as follows1. In epigastric arteries, internal diameters had statistically significant enlargement in 1 day, 3 days of Exp-1 group and 1 day, 3 days, 1 week & 2 weeks of Exp-2 group. Wall thickness had statistically significant thinning in 2 weeks of Exp-2 group. 2. In epigastric veins, internal diameters had enlargement of statistical significance in 1 day of Exp-1 and Exp-2 group. 3. The positive PCNA reactions in smooth muscle appeared in 1 week and increased until 2 weeks, decreased in 4 weeks. There was no statistical significance between Exp-1 and Exp-2 group. 4. The positive ${\alpha}$-SMA reaction in smooth muscles showed weak responses until 1 week and slowly increased in 2 weeks and showed almost control level in 4 weeks. 5. The positive S-100 reactions in the perivascular nerve bundles showed markedly decrease in 1 day, 3 days and increased after 1 week and showed almost control level in 4 weeks. Exp-1 group had stronger response than Exp-2 group. 6. In SEM, we observed defoliation of endothelial cell and flattening of vessel wall. Exp-2 group is more destroyed and healing was slower than Exp-1 group. To sum up, relief of vasospasm (vasodilatation) by freezing with cryoprobe was originated from the damage of smooth muscle layer and perivascular nerve bundle and the enlargement of internal diameter in vessels was similar to expeimental groups, but Exp-2 group had slower healing course and therefore vessel freezing in microsurgery can be clinically used, but repeat freezing time needs to be studied further.

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Effect of Dietary Protein and Calcium Levels on Calcium Metabolism of the Rat (식이단백질과 칼슘 수준이 흰쥐의 칼슘대사에 미치는 영향)

  • Yoon, Gun-Ae;Hwang, Hye-Jin
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.34 no.2
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    • pp.176-180
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    • 2005
  • This study was conducted to examine effect dietary protein and calcium levels on calcium metabolism of the rat. Weaned 6-week old male rats were divided into 4 groups and were fed experimental diets for six weeks. Experimental groups were HPNC group-high protein normal calcium (protein: 400 g/kg diet, calcium: 0.5%), HPLC group-high protein low calcium (protein: 400 g/kg diet, calcium: 0.1%), NPNC group-normal protein normal calcium (protein: 200 g/diet, calcium: 0.5%), NPLC group-normal protein low calcium (protein: 200 g/diet, calcium: 0.1%). The calcium excretion in urine was higher in high protein group than in normal protein group, and it was highest in HPLC group. The activation of alkaline phophatase had a tendency to low in normal calcium group, and the concentration of parathyroid hormone (PTH) was the lowest in HPLC group. The deoxypyridinoline (DPD) concentration of urine was investigated as the highest in HPLC group and it was significantly lower in HPNC group that consumed normal calcium. The bone density of the femur was the highest in NPNC group and the lowest in NPLC group. As the results of this study, calcium excretion in urine and DPD density were the highest and the bone density was the lowest in HPLC group. It may suggest that the deficiency of calcium causes adversely effect in calcium metabolism upon consuming high protein diet. Therefore, it should be emphasized to consume enough calcium to prevent the hindrance of skeletal metabolism caused by deficiency of calcium upon consuming high protein diet.

The Role of Pulmonary Capillary Pressure in the Oxygen Free Radical-Induced Acute Lung Injury (산소기에 의한 급성 폐손상에서 폐모세혈관압의 역할에 관한 연구)

  • Yoo, Chul-Gyu;Kim, Young-Whan;Han, Sung-Koo;Shim, Young-Soo;Kim, Keun-Youl;Han, Yong-Chol
    • Tuberculosis and Respiratory Diseases
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    • v.39 no.6
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    • pp.474-483
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    • 1992
  • Background: Regardless of its causes, acute lung injury is characterized pathophysiologically by increased pulmonary arterial pressure and the protein-rich edema. Many inflammatory mediators are known to be involved in the pathogenesis of acute lung injury, including oxygen free radicals (OFR). But the changes in pulmonary capillary pressure in the OFR-induced acute lung injury is not clear. While the pulmonary edema characterized by the movement of fluid and solutes is dependent on the pressure gradient and the alveolar-capillary permeability, the role of pulmonary capillary pressure in the development of pulmonary edema is also not well understood. Method: Male Sprague-Dawley rats were divided into 5 groups: normal control (n=5), xanthine/xanthine oxidase (X/XO)-treated group (n=7), catalase-pretreated group (n=5), papaverine-pretreated group (n=7), and indomethacin-pretreated group (n=5). In isolated perfused rat lungs, the sequential changes in pulmonary arterial pressure, pulmonary capillary pressure by double occlusion method, and lung weight as a parameter of pulmonary edema were determined. Results: Pulmonary arterial pressure and pulmonary capillary pressure were increased by X/XO. This increase was significantly attenuated by catalase and papaverine, but indomethacin did not prevent the X/XO-induced increase. Lung weight gain was also observed by X/XO perfusion. It was prevented by catalase. Papaverine did not completely block the increase, but significantly delayed the onset. Indomethacin had no effect on the increase in lung weight. Conclusion: These data suggest that increased pulmonary capillary pressure by OFR may aggravate pulmonary edema in the presence of increased alveolar-capillary permeability and this may not be mediated by cyclooxygenase metabolites.

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Superoxide Dismutase Gene Expression Induced by Lipopolysaccharide in Alveolar Macrophage of Rat (폐포대식세포에서 내독소 자극에 의한 Superoxide Dismutase 유전자발현의 조절 기전)

  • Park, Kye-Young;Yoo, Chul-Gyu;Kim, Young-Whan;Han, Sung-Koo;Shim, Young-Soo;Hyun, In-Gyu
    • Tuberculosis and Respiratory Diseases
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    • v.42 no.4
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    • pp.522-534
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    • 1995
  • Background: In the pathogenesis of acute lung injury induced by lipopolysaccharide(LPS), oxygen radiclls are known to be involved in one part. Superoxide dismutase(SOD) protects oxygen radical-induced tissue damage by dismutating superoxide to hydrogen peroxide. In eukaryotic cells, two forms of SOD exist intracellularly as a cytosolic, dimeric copper/zinc-containing SOD(CuZnSOD) and a mitochondrial, tetrameric manganese-containing SOD(MnSOD). But there has been little information about SOD gene expression and its regulation in pulmonary alveolar macrophages(PAMs). The objective of this study is to evaluate the SOD gene expression induced by LPS and its regulation in PAMs of rat. Method: In Sprague-Dawley rats, PAMs obtained by broncholaveolar lavage were purified by adherence to plastic plate. To study the effect of LPS on the SOD gene expression of PAMs, they were stimulated with different doses of LPS($0.01{\mu}g/ml{\sim}10{\mu}g/ml$) and for different intervals(0, 2, 4, 8, 24hrs). Also for evaluating the level of SOD gene regulation actinomycin D(AD) or cycloheximide(CHX) were added respectively. To assess whether LPS altered SOD mRNA stability, the rate of mRNA decay was determined in control group and LPS-treated group. Total cellular RNA extraction by guanidinium thiocyanate/phenolfchlorofonn method and Northern blot analysis by using a $^{32}P$-labelled rat MnSOD and CuZnSOD cDNAs were performed. Results: The expression of mRNA in MnSOD increased dose-dependently, but not in CuZnSOD. MnSOD mRNA expression peaked at 8 hours after LPS treatment. Upregulation of MnSOD mRNA expression induced by LPS was suppressed by adding AD or CHX respectively. MnSOD mRNA stability was not altered by LPS. Conclusion: These findings show that PAMs of rat could be an important source of SOD in response to LPS, and suggest that their MnSOD mRNA expression may be regulated transcriptionally and require de novo protein synthesis without affecting mRNA stability.

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