• Title/Summary/Keyword: 간접측정법

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Use of Real-Time PCR and Internal Standard Addition Method for Identifying Mixed Ratio of Chicken Meat in Sausages (Real-Time PCR과 Internal Standard Addition법을 이용한 돼지고기 소시지에 혼합된 닭고기의 정량)

  • Lee, Namrye;Joo, Jae-Young;Yeo, Yong-Heon
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.46 no.9
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    • pp.1097-1105
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    • 2017
  • This study examined how much chicken meat was in sausage made with pork. Both real-time polymerase chain reaction (PCR) and internal standard addition were used. Fifty ng of chicken DNA was added to the sausages as an internal standard. The addition of standard DNA increased the amplification efficiency of PCR and confirmed the possibility of quantitative analysis. A QIAamp DNA Micro Kit was used to improve the DNA recovery and amplification efficiency. The density of template DNA and primer were suitable for $3.0{\sim}5.0{\mu}L$ and $0.5{\mu}L$, respectively. Each DNA of pig and chicken was diluted in 10-fold from steps 50 ng to 0.05 ng. The detection limit of both pig and chicken meat was more than 0.05 ng and the correlation coefficient of the standard curve was at least 0.98. The result of the quantitative analysis after heat treatment of 3 samples of pigs and chickens mixed at 70:30 showed a 5.7% difference (64.3:35.7) between the expected value and measured value. The quantitative value was changed by affecting the DNA according to the heat treatment ($70^{\circ}C$, 10 min). An analysis of the pork and chicken content in sausages showed that it was difficult to detect chicken meat and the quantitative value of DNA according to the Ct value was very low. On the other hand, when adding standard material (50 ng of chicken DNA) to the sausages, the Ct value decreased gradually with increasing chicken mixing ratio. Thus, the mixing ratio of chicken in sausages could be estimated.

Cellular Mechanism of Nicotine-mediated Intracellular Calcium Homeostasis in Primary Culture of Mouse Cerebellar Granule Cells (니코틴의 마우스 소뇌과립세포내 칼슘의 항상성 조절기전)

  • Kim, Won-Ki;Pae, Young-Sook
    • The Korean Journal of Pharmacology
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    • v.32 no.1
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    • pp.13-21
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    • 1996
  • Intracellular calcium concentration ($[Ca^{2+}]_i$) may play a crucial role in a variety of neuronal functions. Here we report that in primary culture of mouse cerebellar granule cells nicotinic acetylcholine receptors (nAChRs) are expressed in a specific developmental stage and involved in the regulation of intracellular calcium homeostasis. Nicotine-mediated calcium responses were measured using $^{45}Ca^{2+}$ or fluorometrically using the calcium-sensitive fluorescent dye fura-2. Maximal uptake of $^{45}Ca^{2+}$ evoked by nicotine in mouse cerebellar granule cells were revealed $8{\sim}12$ days in culture. In contrast, nicotine did not alter the basal $^{45}Ca^{2+}$ uptake in cultured glial cells. In cerebellar granule cells nicotine-evoked $^{45}Ca^{2+}$ uptake was largely blocked by the NMDA receptor antagonists. Glutamate pyruvate transaminase (GPT). which removes endogenous glutamate, also prevented nicotine effects, implying the indirect involvement of glutamate in nicotine-mediated calcium responses. Fluorometric studies using fura-2 showed two phases of nicotine-evoked $[Ca^{2+}]_i$ rises: the initial rising phase and the later plateau phase. Interestingly, the NMDA receptor antagonists and GPT appeared to inhibit only the later plateau phase of nicotine-evoked $[Ca^{2+}]_i$ rises. The present results imply that nicotine mediated $^{45}Ca^{2+}$ uptake and $[Ca^{2+}]_i$ rises are attributed to the calcium fluxes through both nAchRs and NMDA receptors in a time-dependent manner. Consequently, nAChRs may play an important role in neuronal development by being expressed in a specific developmental stage and regulating the intracellular calcium homeostasis.

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Evaluate Utility of Thyroid Incidentaloma Discrimination by $^{18}F$-FDG PET/CT Delay Scan Images ($^{18}F$-FDG PET/CT검사에서 지연영상을 이용한 갑상선 우연종 감별의 유용성 평가)

  • Lee, Hyun-Kuk;Yang, Seoung-Oh;Song, Gi-Deok;Song, Chi-Ock;Lee, Gi-Heun
    • The Korean Journal of Nuclear Medicine Technology
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    • v.12 no.3
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    • pp.184-191
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    • 2008
  • Purpose: To evaluate the degree of malignancy of incident thyroid lesion found in $^{18}F$-FDG PET/CT findings and the usefulness of the method suggested in this study, we applicate the Delay Scan Method that differentiate a false positive benign tumor, inflammation and malignancy, as well as make the criteria of SUV. Materials and Methods: A retrograde study was conducted of 800 patients who were admitted in E hospital to receive $^{18}F$-FDG PET/CT examination. One patient who was diagnosed as primary thyroid cancer and received $^{18}F$-FDG PET/CT examination was excluded. The number of final patients of this study was 799, the reasons of $^{18}F$-FDG PET/CT examination of these patients were follow-up of old cancer or suspicious tumorous lesion in 696 and disease screening in 103. $^{18}F$-FDG PET/CT image photographing was taken in Biograph-Duo made by SIEMENS, after taking normal $^{18}F$-FDG PET/CT image (1 hr) and then 1 hr later we took the thyroid 1 bed-delayed image for the patients who showed abnormal thyroid $^{18}F$-FDG uptake and above 2.0 SUV for 2 minutes every 1 bed. For the patients who showed abnormal thyroid uptake and above 2.0 SUV, 1 hr later, we took a 1 bed-delayed image and then made a comparative study between measured $SUV_{max}$ of 1 hr-abnormal uptake image and that of 2 hr-delayed image. Results and Conclusion: In this $^{18}F$-FDG PET/CT study among the patients who showed incidental $^{18}F$-FDG thyroidal uptake the number of thyroid incidentaloma was 5 (0.63%), all of then showed benign findings. And in the case of incidental $^{18}F$-FDG uptake in thyroid, $SUV_{max}$ variance obtained from 2 hr delayed image can be a indirect criteria in differentiating benign tumor from malignancy and decrease finding error. In the cases found thyroid incidentaloma when 1) $SUV_{max}$ of focal thyroid lesion is above 5.0 and 2) $SUV_{max}$ variance between normal $^{18}F$-FDG PET/CT exam and 2 hr delayed is $1.0{\pm}0.5$, they are suspected as malignancy and confirming biopsy is to be followed. Otherwise, I also think that distinct follow-up PET or CT image study is a reasonable diagnostic method.

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Lung Uptake of $^{99m}Tc-sestamibi$ during Routine Gated Exercise SPECT Imaging : Comparison with Left Ventricular Ejection Fraction and Severity of Perfusion Defect (일상적인 운동 부하 게이트 심근 관류 SPECT에서 $^{99m}Tc-sestamibi$ 폐섭취 : 좌심실 구혈률과 관류 결손 정도와의 비교)

  • Jeong, Shin-Young;Lee, Jae-Tae;Bae, Jin-Ho;Ahn, Byeong-Cheol;Lee, Kyu-Bo
    • The Korean Journal of Nuclear Medicine
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    • v.37 no.2
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    • pp.83-93
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    • 2003
  • Background: Lung-to-heart uptake ratio (LHR) in $^{201}Tl-chloride$ myocardial perfusion scan is believed to be a reliable marker for left ventricular (LV) dysfunction, but the clinical value of LHR is controversial for $^{99m}Tc-MIBI$ imaging. Furthermore, most of results suggesting lung uptake of $^{99m}Tc-MIBI$ as a potential marker for LV dysfunction used immediate post-stress images, instead of routine images acquired 1 hour after tracer injection. The goal of our study was to investigate whether LHR evaluated with routine gated $^{99m}Tc-MIBI$ imaging can reflect the degree of perfusion defect or left ventricular performance. Subjects and Methods: 241 patients underwent exercise $^{99m}Tc-MIBI$ myocardial SPECT were classified into normal myocardial perfusion (NP, n=135) and abnormal myocardial perfusion (AP, n=106) group according to the presence of perfusion defect. LHR was calculated from anterior projection image taken at 1-hour after injection. Two legions of interest (ROIs) were placed on left lung above LV and on myocardium showing the highest radioactivity. Subjects were classified by left ventricular ejection fraction (LVEF), as Gr-I: >50%, Gr-II: 36-50%, Gr-III: <36% and by summed stress score (SSS), as Gr-A: <4, Gr-B: 4-8, Gr-C: 9-13, Gr-D: >13, LHR was compared among these groups. Results: In NP group(n=135), LHR, were higher in men than women ($men:\;0.311{\pm}0.03,\;women:\;0.296{\pm}0.03,\;p<0.05$). Significant difference, in LHR were found between NP and AP groups both for men and women ($men:\;0.311{\pm}0.03\;vs\;.\;0.331{\pm}0.06,\;women:\;0.296{\pm}0.03\;vs.\;0.321{\pm}0.07.\;p<0.05$). There were weak negative correlation between LHR and LVEF (r=-0.342, p<0.05) and weak positive correlation between LHR and SSS (r=0.478, p<0.05) in men, but not in women (LVEF: r=-0.279, p=0.100, SSS: r=0.276, p=0.103). Increased LHR was defined when for more than mean + 2SD value ($men{\geq}0.38,\;women{\geq}0.37$) of the LHR of the subject with normal perfusion. Increased LHR were observed more frequently in subjects with lower LVEF (Gr-I: 11.1%, Gr-II: 27.0%, Gr-III: 35.4%, p<0.05) and higher SSS(Gr-A: 14.0%, Gr-B: 5.7%, Gr-C: 18.2%, Gr-D: 40.7%, p<0.05). Conclusions: LHRs obtained from routine $^{99m}Tc-MIBI$ gated SPECT images were weakly correlated with LVEF and perfusion defect. Although significant overlaps were observed between normal and abnormal perfusion group, LHRs could be used as an indirect marker of severe perfusion defect or reduced left ventricular function.

Toll-like receptor 2 expression on monocytes from patients with BCG vaccine-associated suppurative lymphadenitis (BCG 접종 후 발생한 화농성 림프절염 환자의 단핵구에서 Toll-like receptor 2의 발현)

  • Oh, Hyun Joo;Shin, Kyung-Sue
    • Clinical and Experimental Pediatrics
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    • v.52 no.6
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    • pp.667-673
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    • 2009
  • Purpose : Toll-like receptor 2 (TLR2) is critical in the immune response to mycobacterial infections. The purpose of this study was to analyze TLR2 surface expressions and TLR2-mediated tumor necrosis factor-alpha ($TNF-{\alpha}$) and interleukin-6 (IL-6) production in patients with BCG vaccine-associated suppurative lymphadenitis. Methods : Peripheral monocytes were separated from 16 patients with BCG vaccine-associated suppurative lymphadenitis and 10 healthy controls using a magnet cell isolation kit. Monocytes ($1{\times}10^6$ cells/well) were incubated with a constant amount of $Pam_3CSK_4$ ($100{\mu}g/mL$) for 24 hours. TLR2 surface expression on monocytes was analyzed by FACS analysis and TLR-2 mRNA expression was determined by RT-PCR. TLR2-mediated $TNF-{\alpha}$ and IL-6 production were measured by ELISA. Results : In patients with BCG vaccine-associated suppurative lymphadenitis, low TLR2 expression on monocytes ($3.39{\pm}$1.2% versus $4.64{\pm}2.6%$) together with significantly lower TLR2 mRNA expression than in the healthy controls was seen after $Pam_3CSK_4$ stimulation. TLR2-mediated $TNF-{\alpha}$ and IL-6 production in patients with BCG vaccine-associated suppurative lymphadenitis ($TNF-{\alpha}$, $775.5{\pm}60.8pg/mL$; IL-6, $4,645.8{\pm}583.9pg/mL$) were also lesser than that in healthy controls ($TNF-{\alpha}$, $1,098.5{\pm}94.3pg/mL$; IL-6, $6,696.3{\pm}544.3pg/mL$). Conclusion : These findings suggest that low TLR2 expression on monocytes might be associated with increased susceptibility to BCG vaccine-associated suppurative lymphadenitis.

The Separate and Combined Effects of Insulin, Dexamethasone and Growth Hormone on the OB Gene Expression and Leptin Secretion from Cultured Human Visceral Adipose Tissue (인체의 복강 내 지방조직 배양을 통한 OB 유전자 발현과 Leptin 분비에 미치는 인슐린, Dexamethasone과 성장호르몬의 단독 또는 복합적 영향에 관한 연구)

  • Hwang, Il Tae;Kim, Kyung Hee;Hwang, Jin Soon;Shin, Choong Ho;Yang, Sei Won
    • Clinical and Experimental Pediatrics
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    • v.46 no.8
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    • pp.795-802
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    • 2003
  • Purpose : We investigated the hormonal control of OB gene expression and leptin secretion in cultured human visceral adipose tissue. Methods : Visceral adipose tissues were cultured for up to 48 hrs in modified Eagle's medium with varying concentration of hormones : Control(no hormone), bovine insulin(100 nM), Dexamethasone(DEX, 100 nM), growth hormone(GH, 40 ng/mL), insulin+DEX(100 nM each), insulin+DEX+GH(100 nM insulin and DEX, 40 ng/mL GH). Quantitative analysis of leptin mRNA was performed by competitive reverse transcription polymerase chain reaction, and leptin secretion in culture medium was measured by IRMA using a commercial kit. Results : The addition of dexamethasone to the medium significantly increased OB gene expression and leptin secretion(P<0.05). Unlike dexamethasone, insulin did not affect OB gene expression and leptin secretion. Both insulin and dexamethasone, at high concentration, significantly stimulated leptin secretion compared with basal values(P<0.05). Leptin gene expression was not significantly increased by GH treatment alone, however GH, in combination with high concentrations of insulin and dexamethasone, attenuated the stimulatory effects of high concentrations of insulin and dexamethasone. Conclusion : Insulin cannot increase leptin secretion without the presence of dexamethasone. The mechanism suggested is that insulin may increase leptin secretion in cytoplasm only after dexamethasone increases the expression of OB gene. Further studies are necessary to elucidate the mechanism of the action of insulin on leptin secretion after increasing OB gene expression by dexamethasone.

Relationship of serum IL-13 and eotaxin level with airway hyperresponsiveness in children with asthma (소아 천식 환아에서 혈청 IL-13 및 eotaxin 농도와 기관지과민성의 관계)

  • Park, Eun Young;Shim, Jung Yeon;Yoo, Myung Hwan;Kim, Deok Soo;Shim, Jae Won;Jung, Hye Lim;Park, Moon Soo
    • Clinical and Experimental Pediatrics
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    • v.49 no.8
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    • pp.889-894
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    • 2006
  • Purpose : Asthma is characterized by the presence of airway hyperresponsiveness(AHR) and inflammation. The extensive eosinophil infiltration into the lung is the hallmark of asthma and contributes to the damage of respiratory epithelium during late phase airway responses. Eotaxin is the major eosinophil chemoattractant found in bronchoalveolar lavage(BAL) fluid of allergic inflammation. IL-13 has been known to induce the expression of exotaxin and eosinophilia. IL-13 also induces airway inflammation, mucus production and leads to marked fibrosis, airway remodeling and AHR. We investigated whether serum IL-13 levels can reflect the presence of airway hyperresponsiveness in children with asthma, and the relationship between serum IL-13 and eotaxin levels. Methods : Using sandwich enzyme-linked immunosorbent assay, the serum IL-13 and eotaxin levels were measured in 13 atopic asthmatics, 5 atopic non-asthmatics and 12 control subjects. Metacholine challenge tests were performed in all subjects. Airway hyperresponsiveness to metacholine was expressed as provocative concentration of metacholine causing a 20% fall in FEV1[$PC_{20}mg/mL$]. $PC_{20}$ value of 25 mg/mL was used as a cut-off for defining a AHR. Results : Serum IL-13 levels showed positive correlation with eotaxin levels. Serum IL-13 and eotaxin levels showed no differences among atopic asthmatics, atopic non-asthmatics and control subjects. And there were no differences serum IL-13 and eotaxin levels in children with and without AHR and atopy. Serum IL-13 and eotaxin levels did not correlate with $logPC_{20}$ levels. Conclusion : IL-13 is closely related to the eotaxin release. But serum IL-13 and eotaxin per se can't predict the severity of airway hyperresponsiveness. IL-13 and eotaxin may have local effect on respiratory epithelium or there can be some factors to induce airway hyperresponsiveness other than serum IL-13 in asthmatic airways.

Monitoring the Rate of Frozen Denaturation of Bovine Myosin by Competitive Indirect ELISA Method (Competitive Indirect ELISA를 이용한 Bovine Myosin의 동결 변성도 측정)

  • Kim, Seong-Bae;Lee, Ju-Woon;Park, Jong-Heum;Do, Hyung-Ki;Hyun, Chang-Kee;Shin, Heuyn-Kil
    • Korean Journal of Food Science and Technology
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    • v.30 no.4
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    • pp.862-870
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    • 1998
  • This study shows the application of Ci-ELISA method for monitoring the denaturation of myosin by the frozen treatment in order to differentiate thawed beef from chilled. Hanwoo M.Semitendinosus (n=25) was treated under the two different frozen process as follows; simple frozen treatment (Exp-1) at 4 different temperatures, -10, -20, -50 and $-80^{\circ}C$, respectively, and repeated thawing-refreezing treatment (Exp-2) stored at 4 different temperatures, -10, -20, -50 and $-80^{\circ}C$, respectively. Antibodies (Abs) were produced from rabbits immunized with myosin whole molecule (MWM) isolated from beef round, heavy meromyosin S-1 (S-1) and light meromyosin (LMM) prepared by digestion of MWM. Each immunoglobulin G (IgG) was separated from antiserum. At 6 month storage, IA of anti-MWM IgG for myosin was decreased to 32.67, 32. 23, 51.52 and 34.27% in Exp-1 and to 14.82, 15.61, 25.3 and 23.7% in Exp-2 at -10, -20, -50 and $-80^{\circ}C$, respectively (P<0.05). In Exp-1, the reactivities of anti-LMM IgG were decreased to 25.12, 21.42, 49.05 and 28.96%, and those of Exp-2 were to 11.88, 9.56, 20.63 and 12.64% at -10, -20, -50 and $-80^{\circ}C$, respectively, at 6 times thawing (P<0.05). Conclusively, myosin was denaturated by freezing treatment and LMM or myosin rod part might have suffered from more extreme demage than HMM S-1, and samples at $-50^{\circ}C$ were slightly injured less than others by freezing treatment.

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The Increased Expression of Gelatinolytic Proteases Due to Cigarette Smoking Exposure in the Lung of Guinea Pig (기니픽에서 흡연 노출에 의한 젤라틴 분해 단백 효소의 발현 양상에 관한 연구)

  • Kang, Min-Jong;Lee, Jae-Ho;Yoo, Chul-Gyu;Lee, Choon-Taek;Chung, Hee-Soon;Seo, Jeong-Wook;Kim, Young-Whan;Han, Sung-Koo;Shim, Young-Soo
    • Tuberculosis and Respiratory Diseases
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    • v.50 no.4
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    • pp.426-436
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    • 2001
  • Background : Chronic obstructive pulmonary disease(COPD) is one of the major contributors to morbidity and mortality among the adult population. Cigarette smoking(CS) is undoubtedly the single most important factor in the pathogenesis of COPD. However, its mechanism is unclear. The current hypothesis regarding the pathogenesis of COPD postulates that an imbalance between proteases and antiproteases leads to the destructive changes in the lung parenchyma. This study had two aims. First, to evaluate the effect of CS exposure on histologic changes of the lung parenchyme, and second, to evaluate the effect of CS exposure on the expression of the gelatinolytic enzymes in BAL fluid cells in guinea pigs. Methods : Two groups of five guinea pigs were exposed to the whole smoke of 20 commercial cigarettes per day, 5 hours/day, 5 days/week, for 6weeks, and 12 weeks, respectively, using a smoking apparatus. Five age-matched guinea pigs exposed to room air were used as controls. Five or more sections were microscopically extamined(${\times}400$) and the number of cellular infiltration of the alveolar wall was measured in order to evaluate the effect of CS exposure on the histologic changes of lung parenchyme. The statistical significance was analyzed by a linear regression method. To evaluate the expression of the gelatinolytic enzymes in intraalveolar cells, BAL fluid was obtained and the intraalveolar cells were separated by centrifugation (500 g for 10 min at $4^{\circ}C$). Two sets of culture plates were loaded with $1{\times}10^6$ intraalveolar cells. One plate, contained O.1mM EDTA, a inhibitor of matrix metalloproteases(MMPs), and the other plate had no EDTA. Both plates were incubated for 48 hours at $37^{\circ}C$. After incubation, gelatinolytic protease expression in the supernatants was analyzed by gelatin zymography. Results : At the end of CS exposure, the level of blood carboxy Hb had increased significantly(4.1g/dl in control group, 24g/dl immediately after CS exposure, 18g/dl 30 min after CS exposure, 15g/dl 1 hour after CS exposure). Alveolar inflammatory cells were identified in the CS exposed guinea pigs. The number of alveolar cellular cells observed in a microscopic field ($400{\times}$) was $121.4{\pm}7.2$, $158.0{\pm}20.2$, $196.8{\pm}32.8$, in the control, the 6 weeks, and the 12 weeks group, respectively. The increased extent of inflammatory cellular infiltration of the lung parenchema showed a statistically significant linear relationship with the duration of CS exposure(p=0.001, $r^2=0.675$). Several types of gelatinolytic enzymes in the intraalveolar cells of CS exposed guinea pigs were expressed, of which some were inhibited by EDT A. However, the gelatinolytic enzymes were not expressed in the control groups. Conclusion : CS exposure increases inflammatory cellular infiltration of the alveolar wall and the expression of gelatinolytic proteases in guinea pigs. EDTA inhibits some of the gelatinolytic proteases. These findings suggest a possibility that CS exposure may increase MMP expression in the lungs of guinea pigs.

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Economic evaluation of a weekly administration of a sustained-release injection of recombinant human growth hormone for the treatment of children with growth hormone deficiency (소아 성장호르몬결핍증 치료에 사용되는 성장호르몬 서방형 주사제의 경제성 평가)

  • Kang, Hye-Young;Kim, Duk Hee;Yang, Sei-Won;Kim, Yoon-Nam;Kim, Miseon
    • Clinical and Experimental Pediatrics
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    • v.52 no.11
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    • pp.1249-1259
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    • 2009
  • Purpose:From a societal perspective, we evaluated the cost-effectiveness of a novel sustained-release injection of recombinant human growth hormone (GH) administered on a weekly basis compared with that of the present daily GH injection for the treatment of children with GH deficiency. Methods:Health-related utility for GH therapy was measured based on the visual analogue scale. During July 2008, caregivers of 149 children receiving GH therapy form 2 study sites participated in a web-based questionnaire survey. The survey required the caregivers to rate their current subjective utility with daily GH injections or expected utility of weekly GH injections. Because there was no difference in the costs of the daily and weekly therapies, for the purposes of this study, only drug acquisition costs were considered. Results:Switching from daily to weekly injection of GH increased the utility from 0.584 to 0.784 and incurred an extra cost of 4,060,811 Korean won (KW) per year. The incremental cost-utility ratio (ICUR) for a base case was 20,305,055 KW per quality-adjusted life year (QALY) gained. Scenario analyses showed that the ICUR ranged from 15,751,198 to 25,489,929 KW per QALY. Conclusion:The ICUR for a base case and worst case scenario analyses ranged from 0.85 to 1.37-times per capita gross domestic product of Korea, which is considered to be within the generally accepted willingness-to-pay threshold. Thus, it is concluded that switching from daily to weekly injection of GH would be cost-effective.