• Title/Summary/Keyword: enzyme assay

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Anti-inflammatory effect of Sinhyowoldo-san Extract with regard to Pro-inflammatory Mediators in PMA plus A23187-induced Human Mast Cells (인간 비만세포에서 PMA와 A23187에 의해 유도된 전염증 매개체에 대한 신효월도산 추출물의 항염증 효과)

  • Wi, Gyeong;Yang, Da-Wun;Kang, Ok-Hwa;Kim, Sung-Bae;Mun, Su-Hyun;Seo, Yun-Soo;Kang, Da-Hye;Lim, Jae-Soo;Kim, Ma-Ryong;Kwak, Nam-Won;Kong, Ryong;Kwon, Dong-Yeul
    • The Korea Journal of Herbology
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    • v.29 no.6
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    • pp.117-123
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    • 2014
  • Objectives : Sinhyowoldo-san (SHWDS) is said to be a traditional medicine used for shigellosis, abdominal pain, diarrhea. But mechanism of SHWDS mediated-modulation of immune function is not sufficiently understood. To ascertain the molecular mechanisms of SHWDS 70% EtOH extract on pharmacological and biochemical actions in inflammation, we researched the effect of pro-inflammatory mediators in phorbol-12-myristate-13-acetate (PMA)+ A23187-activated human mast cell line (HMC-1). Methods : In the present research, cell viability was measured by MTS assay. pro-inflammatory cytokine production was measured by performing enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis to analyze the activation of mitogen-activated protein kinases (MAPKs), nuclear factor kappa-light-chain-enhancer of activated B cells ($NF-{\kappa}B$). The investigation focused on whether SHWDS inhibited the expressions of interleukin-6 (IL-6), interleukin-8 (IL-8), MAPKs and $NF-{\kappa}B$ in PMA+A23187-activated HMC-1 cells. Results : SHWDS has no cytotoxicity at measured concentration (50, 100, and $250{\mu}g/ml$). SHWDS ($250{\mu}g/ml$) inhibits pro-inflammatory cytokine expression in PMA+ A23187-activated HMC-1 cells. Moreover, SHWDS inhibited cyclooxygenase (COX)-2 expression. In activated HMC-1 cells, SHWDS suppressed phosphorylation of extracellular signal-regulated kinase (ERK 1/2) and c-jun N-terminal Kinase (JNK 1/2). Then, SHWDS suppressed activation of nuclear factor $NF-{\kappa}B$ in nuclear, degradation of IkB ${\alpha}$ in cytoplasm. Conclusions : We propose that SHWDS has an anti-inflammatory therapeutic potential, which may result from inhibition of ERK 1/2, JNK 1/2 phosphorylation and $NF-{\kappa}B$ activation, thereby decreasing the expression of pro-inflammatory genes.

Changes in Lymphocyte DNA Damage and Antioxidant Status after Supplementing Propolis to Korean Smokers: A Placebo-Controlled, Double-Blind Cross-Over Trial (프로폴리스 섭취 후 흡연자의 임파구 DNA 손상도 및 항산화 상태의 변화: 이중맹검 교차 인체시험)

  • Kang, Myung-Hee;Lee, Hye-Jin;Kim, Mi-Kyung;Sung, Mi-Kyung;Kwon, O-Ran;Park, Yoo-Kyoung
    • Journal of Nutrition and Health
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    • v.42 no.5
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    • pp.442-452
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    • 2009
  • Smoking has been known to exacerbate the initiation and propagation of oxidative stresses. Efforts have been made to reduce the smoking-induced oxidative stresses using commercial dietary supplements. Propolis is the resinous substance collected by bees from the leaf buds and bark of trees, especially poplar and conifer trees. In this trial, we examined whether a daily supplementation of 800 mg propolis can protect endogenous lymphocytic DNA damage and modulate antioxidative enzyme activities and the level of antioxidant vitamin in smokers using a placebo-controlled, doubleblinded cross-over trial. After two weeks of running-in period, 29 smokers (mean age 34.38 ${\pm}$ 1.73) received 6 tablets/day of either propolis or placebo pills for 4 weeks. After 2 weeks of washout period the subjects switched they pills for cross-over study. The degree of DNA damage (assessed by tail DNA, tail length and tail moment) was not significantly changed with propolis intake or placebo intake. Similarly, total antioxidant status (TAS) remained at the same level regardless of the treatment. Erythrocyte catalase, glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), plasma vitamin C and tocopherol level did not differ before and after propolis treatment, and did not differ between treatments. Putting all these results together, we would suggest that it is still too early to claim that propolis possess antioxidative activities.

The Correlation between TGF-beta 1 Blood Levels and the Formation of Bullae in Patients with Spontaneous Pneumothorax (자연 기흉 환자의 혈액 내 TGF-beta 1 Ligand 양과 폐 기포 형성과의 연관관계에 대한 연구)

  • Kim, Young-Sam;Kim, Kwang-Ho;Baek, Wan-Ki;Kim, Joung-Taek;Cha, Il-Kyu;Kim, Ji-Hye;Song, Sun-U;Choi, Mi-Sook
    • Journal of Chest Surgery
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    • v.43 no.4
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    • pp.394-398
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    • 2010
  • Background: The overexpression of transforming growth factor-beta 1 receptor II (TGF-${\beta}1$RII) and transforming growth factor-beta 1 (TGF-${\beta}1$) ligand may be involved in the formation of a bulla. In this study, we tested if serum TGF-${\beta}1$ ligand levels correlated with the expression level of TGF-${\beta}1$RII and TGF-${\beta}1$ in bullous tissues from patients with spontaneous pneumothorax. Material and Method: Bullous lung tissues and blood samples were obtained from 19 patients with spontaneous pneumothorax, 18 males and 1 female, aged 17 to 35 years old. The bullous tissues were obtained by video-assisted thoracic surgery (VATS), fixed in formalin, embedded in paraffin, and cut into $5{\sim}6{\mu}m$ thick slices. Sections were immunohistochemically stained with primary antibodies against TGF-${\beta}1$ or TGF-${\beta}1$RII, and serum levels of TGF-${\beta}1$ in patients and normal controls was measured by enzyme-linked immunosorbent assay (ELISA). Result: Of the 19 patients, 16 were TGF-${\beta}1$ positive and 10 were TGF-${\beta}1$RII positive. Among the 16 TGF-${\beta}1$ positives, 9 were also TGF-${\beta}1RII$ positive. As seen previously, strong immunohistochemical staining of TGF-${\beta}1$RII and TGF-${\beta}$ was detected in the boundary region between the bullous and normal lung tissues. Average TGF-${\beta}1$ blood levels of both TGF-${\beta}1$ and TGF-${\beta}1$RII positive patients was $38.36{\pm}16.2ng/mL$, and that of five controls was $54.06{\pm}15ng/mL$. Conclusion: These results suggest that overexpression of TGF-${\beta}1$ and TGF-${\beta}1$RII expression may be involved in the formation of bullae. TGF-${\beta}1$ blood levels in patients with primary spontaneous pneumothorax is lower than normal people, suggesting that the high level of local TGF-${\beta}1$ expression in the bullous tissue region, but not in the whole blood, may contribute more in the formation of bullae.

Telomerase Activity in Primary Lung Cancers (원발성 폐암에 있어서 Telomerase 활성도에 대한 연구)

  • Yun, Sang-Myung;Kwak, Kyung-Rok;Hwang, Jee-Yoon;Park, Sam-Seok;Jeon, Doo-Soo;Kim, Cheol-Min;Lee, Min-Ki;Park, Soon-Kew
    • Tuberculosis and Respiratory Diseases
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    • v.46 no.2
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    • pp.195-203
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    • 1999
  • Background: Telomerase enzyme activity is not detected in most normal cells, a phonomenon believed to be associated with limitations on cellular proliferation. Since this activity is detected in nearly all human tumor, including lung cancers, it has been suggested that telomerase activation may be coupled to acquisition of malignant phenotype. In this study, we determined whether telomerase activity was associated with tumor pathologic stage. Methods: Primary tumor specimens obtained by bronchoscopic biopsies from 33 patients were analyzed. Telomerase activity was measured by means of a modified Telomeric Repeat Amplication Protocol(TRAP) assay. Results: Telomerase activity was detected in 23 of the 27 non-small-cell lung cancer and 5 of 6 small-cell lung cancer. A few primary tumors did not appear to have detectable telomerase activity. Positive associations were found between the telomerase-positive rate and tumor stage(p<0.05). Conclusion: High telomerase activity is detected frequently in primary lung cancers that exhibit high tumor cell proliferation rates and advanced pathologic stage.

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Anti-Inflammatory Effect of Ethanol Extract from Grateloupia crispata on Lipopolysaccharide-Induced Inflammatory Responses in RAW 264.7 Cells and Mice Ears (LPS로 유도된 RAW 264.7 세포와 마우스 귀 조직에 대한 주름까막살 에탄올 추출물의 항염증 효과)

  • Bae, Nan-Young;Kim, Min-Ji;Kim, Koth-Bong Woo-Ri;Park, Sun-Hee;Jang, Mi-Ran;Ahn, Dong-Hyun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.45 no.8
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    • pp.1090-1098
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    • 2016
  • The anti-inflammatory effects of ethanol extract from Grateloupia crispata (GCEE) were investigated in lipopolysaccharide (LPS)-stimulated murine macrophages. Anti-inflammatory effects were detected by enzyme-linked immunosorbent assay, Western blotting, and immunohistochemistry. There was no cytotoxic effect on proliferation of macrophages treated with GCEE compared to the control. GCEE significantly inhibited production of pro-inflammatory cytokines [interleukin (IL)-6, tumor necrosis $factor-{\alpha}$, and $IL-1{\beta}$] as well as nitric oxide in LPS-stimulated RAW 264.7 cells. In addition, GCEE suppressed expression of inducible nitric oxide synthase, cyclooxygenase-2, and nuclear $factor-{\kappa}B$ in a dose-dependent manner. GCEE significantly reduced activation of mitogen-activated protein kinases. In the in vivo test, evaluation of anti-inflammatory activity of GCEE was performed using croton oil-induced ear edema in ICR mice. Oral administration of 10 mg/kg to 250 mg/kg of GCEE significantly reduced ear edema in a dose-dependent manner compared to croton oil-induced mice. Moreover, GCEE reduced ear thickness and the number of mast cells compared to croton oil-induced mice in the histological analysis. These data suggest that GCEE could be used as a potential source for anti-inflammatory agents.

Evaluation of Antibodies Against Haemophilus influenzae Type b in Korean Adults (우리나라 성인에서 Haemophilus influenzae type b에 대한 항체 평가)

  • Lee, Ji Hyen;Kim, Han Wool;Lee, Soyoung;Kim, Kyung-Hyo
    • Pediatric Infection and Vaccine
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    • v.24 no.3
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    • pp.125-133
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    • 2017
  • Purpose: After the introduction of Haemophilus influenzae type b (Hib) vaccine in 1995 in Korea, it was included in the national immunization program in 2013. In the post-Hib vaccine era, some studies in other countries reported that invasive Hib disease affects adults, especially the elderly and immunocompromised persons, more often than it affects children. To evaluate disease susceptibility, quantitative and qualitative analysis of anti-polyribosylribitol phosphate (PRP) antibodies were carried out in Korean adults aged 20 to 85 years. Methods: Sera were collected from 39 healthy adults (20 to 50 years of age) and from 30 elderly adults (75 to 85 years of age) who did not have immune-compromising conditions. The concentration of anti-PRP immunoglobulin G (IgG) and serum bactericidal indices (SBIs) were measured by enzyme-linked immunosorbent assay and serum bactericidal assay. Results: Geometric mean concentrations of anti-PRP IgG and geometric mean SBIs were $0.88{\mu}g/mL$ (95% confidence interval [CI], 0.17 to 3.85) and 354 (95% CI, 50 to 2,499) in young adults and $1.67{\mu}g/mL$ (95% CI, 0.53 to 5.24) and 449 (95% CI, 146 to 1,376) in elderly adults, respectively. When the threshold of seropositivity for anti-PRP IgG was applied as 0.15 or $1.0{\mu}g/mL$, which is the protective antibody level in children, seropositive rates were 87.2% or 53.8% in young adults and 100% or 60% in elderly adults. The seropositivity rates of the SBI ($SBI{\geq}4$) were 82.1% and 100% in the groups, respectively. Conclusions: Most subjects in the adult and elderly adult groups display immunity to Hib based on quantitative and qualitative antibody levels, but not all. Because high immunization and low Hib circulation rates may reduce the natural Hib immunity in the population, monitoring Hib immunity as well as disease are needed continuously.

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 $H_2O_2$.

The Heterotrimeric Kinesin-2 Family Member KIF3A Directly Binds to Creatine Kinase B (Heterotrimeric kinesin-2의 KIF3A와 creatine kinase B의 결합)

  • Jeong, Young Joo;Park, Sung Woo;Seo, Mi Kyoung;Kim, Sang-Jin;Lee, Won Hee;Kim, Mooseong;Urm, Sang-Hwa;Lee, Jung Goo;Seog, Dae-Hyun
    • Journal of Life Science
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    • v.31 no.3
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    • pp.257-265
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    • 2021
  • Heterotrimeric kinesin-2 is a molecular motor protein of the kinesin superfamily (KIF) that moves along a microtubule plus-end directed motor protein. It consists of three different motor subunits (KIF3A, KIF3B, and KIF3C) and a kinesin-associated protein 3 (KAP3) that form a heterotrimeric complex. Heterotrimeric kinesin-2 interacts with many different binding proteins through the cargo-binding domain of the KIF3s. The activity of heterotrimeric kinesin-2 is regulated to ensure that the cargo is directed to the right place at the right time. How this regulation occurs, however, remains in question. To identify the regulatory proteins for heterotrimeric kinesin-2, we performed yeast two-hybrid screening and found a specific interaction with creatine kinase B (CKB), which is the brain isoform of cytosolic creatine kinase enzyme. CKB bound to the cargo-binding domain of KIF3A but did not interact with the KIF3B, KIF5B, or KAP3 in the yeast two-hybrid assay. The carboxyl (C)-terminal region of CKB is essential for the interaction with KIF3A. Another protein kinase, CaMKIIa, interacted with KIF3A, but GSK3a did not interact with KIF3A in the yeast two-hybrid assay. KIF3A interacted with GST-CKB-C but not with GSK-CKB-N or GST alone. When co-expressed in HEK-293T cells, CKB co-localized with KIF3A and co-immunoprecipitated with KIF3A and KIF3B but not KIF5B. These results suggest that the CKB-KIF3A interaction may regulate the cargo transport of heterotrimeric kinesin-2 under energy-compromised conditions in cells.

Mycoplasma pneumoniae-induced production of proasthmatic mediators in airway epithelium (인체 기관지 상피세포에서 Mycoplasma pneumoniae 감염에 의한 천식 매개물질의 발현)

  • Kim, Kyung Won;Lee, Byung Chul;Lee, Kyung Eun;Kim, Eun Soo;Song, Tae Won;Park, Mi Yeoun;Sohn, Myung Hyun;Kim, Kyu-Earn
    • Clinical and Experimental Pediatrics
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    • v.49 no.9
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    • pp.977-982
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    • 2006
  • Purpose : There has been an increasing amount of literature concerning the association between Mycoplasma pneumoniae and asthma pathogenesis. Interleukin(IL)-6 stimulates the differentiation of monocytes, and can promote Th2 differentiation and simultaneously inhibit Th1 polarization. IL-8 is a potent chemoattractant and, it has been suggested, has a role in asthma pathogenesis. Nitric oxide (NO) synthesized by airway epithelium may be important in the regulation of airway inflammation and reactivity. Vascular endothelial growth factor(VEGF) has been reported to be a mediator of airway remodeling in asthma. We investigated the effects of M. pneumoniae on IL-6, IL-8, NO and VEGF production in human respiratory epithelial cells. Methods : A549 cells were cultured and inoculated with M. pneumoniae at a dose of 20 cfu/cell. After infection, the presence of M. pneumoniae in epithelial cell cultures was monitored by immunofluorescence and confirmed by polymerase chain reaction(PCR) detection. IL-6, IL-8 and VEGF were determined by an enzyme-linked immunosorbent assay and reverse transcriptase-polymerase chain reaction. NO was measured using the standard Griess reaction. Results : In A549 cells, M. pneumoniaeinduced IL-6, IL-8, NO and VEGF release in time-dependent manners. It also induced mRNA expression of IL-6, IL-8 and VEGF in similar manners. Conclusion : These observations suggest that M. pneumoniae might have a role in the pathogenesis of the allergic inflammation of bronchial asthma.

Effects of Hwangryunhaedok-Tang and Geongangbuja-Tang on the Change of Interleukin-6 and $TNF-{\alpha}$ Level Induced by LPS I.C.V. Injection in Mice (황연해독탕(黃連解毒湯)과 건강부자탕(乾薑附子湯)이 LPS유도에 의한 마우스 혈중 IL-6와 $TNF-{\alpha}$ 변화에 미치는 영향)

  • Park, Su-Hyun;Kwon, Yong-Uk;Lee, Tae-Hee
    • Herbal Formula Science
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    • v.15 no.1
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    • pp.185-197
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    • 2007
  • Objective : This study was conducted to investigate the effects of Hwangryunhaedok-Tang and Geongangbuja-Tang on the change of interleukin-6 (IL-6) and tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$) level induced by LPS I.C.V. injection in mice. Method : We devided group into 6 mice and 6 mice were assingned to each group. In the normal group only saline was administered intragastrically, and in the control group LPS was injected intracerebroventricularly 1 hr after intragastric administration of saline. In the experiment groups Hwangryunhaedok-Tang(0.5g/kg, 1.0g/kg, 3.0g/kg) was administered intragastrically to mice 1 hr prior to LPS(100mg/mouse) I.C.V. injection.. Also Geongangbuja-Tang (0.5g/kg, 1.0g/kg, 3.0g/kg) was administered intragastrically to mice 1 hr prior to LPS(100mg/mouse) I.C.V. injection. To measure the plasma IL-6 and $TNF-{\alpha}$ level of mice, their blood samples were collected from retro-orbital plexus, immediately centrifuged at $4^{\circ}C$, and plasma was removed and stored frozen at $-83^{\circ}C$ for later determination of IL-6 and $TNF-{\alpha}$. The level of IL-6 and $TNF-{\alpha}$ production was measured by enzyme-linked immunosorbent assay in the plasma. Result : Regarding IL-6 level, The 0.5g/kg and the 1g/kg groups of Geongangbuja-Tang decreased IL-6 level. Especially the 3g/kg control group decreased IL-6 level significantly than the normal group(p<0.01). Regarding $TNF-{\alpha}$ level, the 3g/kg group of Geongangbuja-Tang decreased it significantly(p<0.05). Conclusion : These data revealed that Hwangryunhaedok-Tang might not have the anti imflammatory effect and Geongangbuja-Tang(3g/kg)might have the anti imflammatory effect by reducing the plasma IL-6 and $TNF-{\alpha}$ level in mice LPS Injection.EIM (Eighteen Incompatible Medicaments) is an important component in Oriental pharmacology and is directly related to clinical prescriptions. Medical practitioners argued that the definite cause and meaning of EIM was ambiguous and therefore debated the issue of clinical application of the EIM. This study conducted an in-depth literary research on the origin, meaning and contents of EIM with the purpose to contribute in its efforts to be used clinically. Even after thousands of years have past since establishment of Oriental medicine, EIM is still tabooed and was an obstacle that hindered ideologies. Modern herbal medicine texts claim that the use of EIM can reduce treatment effects and promote poisoning and side effects. However, since long ago, there has been medical practitioners who reject this as false. Recently, poisoning caused by EIM has been claimed to be from the toxicity of the drug itself, rather than the result of interaction between the drugs, and therefore they suggest that EIM is not a forbidden domain. In addition, EIM showed a difference in number depending on the era. However, this can be understood not as a definite number, but instead as a warning to be careful during combination of drugs for use as clinical medicine. Historically, there were very few cases in which EIM was used for clinical tests and thus, the clinical value is not, while others applied EIM directly to their bodies, which showed signs for the usefulness and potential of EIM for us. A more concrete and in-depth study must be made on EIM.

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