• The Journal of Korean Obstetrics and Gynecology
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• 제32권2호
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• pp.1-17
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• 2019

Objectives: This study was performed to identify the anti-inflammatory effects of Salvia miltiorrhizae radix Water extract (SMW) on lipopolysaccharide (LPS) induced inflammation. Methods: RAW 264.7 cells were treated with 500 ng/ml of LPS. SMW (0.1, 0.25, 0.5 mg/ml) was treated 1 h prior to LPS. Cell viability was measured by MTT assay. Levels of nitric oxide (NO) were measured with Griess reagent and pro-inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (PCR). We also examined molecular mechanisms such as mitogen-activated protein kinases (MAPKs) and nuclear factor-kappa B ($NF-{\kappa}B$) activation by western blot. In addition, we observed mice survival rate after LPS and examined their cytokine levels of serum and liver tissue. Results: SMW itself did not have cytotoxic effects in RAW 264.7 cells less than 0.5 mg/ml. SMW treatment inhibited the production of NO, and interleukin $(IL)-1{\beta}$ which is pro-inflammatory cytokine. And SMW treatment inhibited the LPS-induced activation of MAPKs such as extracellular signal-regulated kinase1/2 (ERK1/2), p38 kinases (p38), c-Jun NH2-terminal kinase (JNK) and $NF-{\kappa}B$. In addition, it also showed reducing the level of $IL-1{\beta}$ on the serum and liver tissue of mice. Also, death of LPS-induced mice was inhibited by SMW. Conclusions: The result suggests that treatment of SMW could reduce the LPS-induced inflammation. Thereby, SMW could be used as a protective agent against inflammation. Also, this study could give a clinical basis that SMW could be a drug or agent to prevent inflammatory diseases.

• The Korea Journal of Herbology
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• 제28권5호
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• pp.7-11
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• 2013

Objectives : Seungmagalgeun-tang (SMGGT) is traditional medicine widely used for inflammatory disease and flu. But SMGGT exhibits potent anti-inflammatory activity with an unknown mechanism. To elucidate the molecular mechanisms of SMGGT water extract on pharmacological and biochemical actions in inflammation, we examined the effect of SMGGT on pro-inflammatory mediators in Phorbol-12-myristate-13-acetate (PMA)+A23187-stimulated mast cells. Methods : In the present study, pro-inflammatory cytokine production was determined by performing enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis to measure the activation of MAPKs. Cells were treated with SMGGT 1 h prior to the addition of 50 nM of PMA and $1{\mu}M$ of A23187. Cell viability was measured by MTS assay. The investigation focused on whether SMGGT inhibited the expressions of interleukin-6 (IL-6), interleukin-8 (IL-8) and mitogen-activated protein kinases (MAPKs) in PMA+A23187-stimulated mast cells. Results : SMGGT has no cytotoxicity at examined concentration (100, 250, and $500{\mu}g/ml$). Also, gene expression of IL-6 and IL-8 in HMC-1 cells stimulated by PMA+A23187 was down regulated by SMGGT. Furthermore, SMGGT suppressed the PMA+A23187-induced phosphorylation of extracellular signal-regulated kinase (ERK) and c-jun N-terminal Kinase(JNK). But, SMGGT could not regulate phosphorylation of p38 MAPK. Conclusions : These results suggest that SMGGT has inhibitory effects on PMA+A23187-induced IL-6 and IL-8 production. These inhibitory effects occur through blockades on the phosphorylation of ERK and JNK.

• Journal of Life Science
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• 제28권7호
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• pp.835-840
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• 2018

Quantitative determination of the protein expression levels is one of the most important parts in assessment of the safety of foods derived from genetically modified (GM) crops. Overexpression of AtCYP78A7, a gene encoding cytochrome P450 protein, has been reported to improve tolerance to abiotic stress, such as drought and salt stress, in transgenic rice (Oryza sativa L.). In the present study, an enzyme-linked immunosorbent assay (ELISA) kit for diagnosing AtCYP78A7 protein including AtCYP78A7-specific monoclonal antibody was developed. GST-AtCYP78A7 recombinant protein was induced and purified by affinity column. Four monoclonal antibodies (mAb 6A7, mAb 4C2, mAb 11H6, and mAb 7E8) against recombinant protein were also produced and biotinylated with avidin-HRP. After pairing test using GST-AtCYP78A7 protein and lysate of rice samples, mAb 4C2 and mAb 7E8 were selected as a capture antibody and a detecting antibody, respectively, for ELISA kit. Product test using rice samples indicated that percentages of detected protein in total protein were greater than 0.1% in AtCYP78A7-overexpressing transgenic rice (Line 10B-5 and 18A-4), whereas those in negative control non-transgenic rice (Ilpum and Hwayoung) were less than 0.1%. The ELISA kit developed in this study can be useful for the rapid detection and safety assessment of transgenic rice overexpressing AtCYP78A7.

• Parasites, Hosts and Diseases
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• 제24권1호
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• pp.25-41
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• 1986

The applicability of micro-ELISA was evaluated in human neuro-cysticercosis using paired samples of serum and CSF. A total of 355 cases who were mostly neurologic patients was subjected. Cystic fluid of C. cellulosae was used as antigen in protein concentration of $2.5{\;}{\mu}g/ml$. Serum was diluted to 1 : 100 and CSF was undiluted in the assay for the specific IgG antibody level. The differential criterion of the positive reaction was the abs. of o. 18 in both samples. The results were summarized as follows: 1. The overall sensitivity of the micro-ELISA in 71 confirmed neurocysticercosis was 90.1% ; the sensitivity by serum was 77.5% and that by CSF was 83.1%. CSF was a more sensitive and valuable material. Most of the false negative cases of neuro-cysticercosis showed far lower level of abs. rather than marginal. 2. The overall specificity of the micro-ELISA in 52 confirmed other neurologic diseases was 88.5%; the specificities by serum and by CSF were 94.2% respectively. Cases of other neurologic diseases did not show false positive reactions in both samples. 3. When serum was assayed, taeniasis(2/18), sparganosis(2/20), paragonimiasis(1/56), clonorchiasis(1/15) and fascioliasis(1/1) cases showed cross reactions. When CSF was assayed, 2 ot 10 neuro-sparganosis showed cross reactions while none of 9 neuro-paragonimiasis showed it. Out of 71 confirmed neuro-cysticercosis cases, 6 and 11 showed cross reactions by serum and CSF to crude extract antigen of sparganum; but no case did show it to crude extract antigen of Paragonimus westermani. 4. Ventricular CSF showed low or negative levels of IgG antibody than lumbar CSF unless the lesion was at the lateral ventricle itself. 5. Out of 4 racemose cysticercosis cases, 3 showed positive reaction in serum while all of 3 examined CSF were positive. The above results indicated that the serological test for detecting the specific IgG antibody by micro-ELISA using paired samples of serum and CSF was very helpful for clinical differentiation of neuro-cysticercosis from neurologic diseases of other causes.

• Parasites, Hosts and Diseases
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• 제24권2호
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• pp.187-193
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• 1986

Enzyme-linked immunosorbent assay(ELISA) using crude and affinity-purified antigens of adult worms of Paragonimus westermani was performed for infected cat sera with different worm burden, from preinfection to 18th week after infection. Crude antigen was used with supernatant of homogenated worms by freezing-thawing method, and the supernate was centrifuged for 1 hour at 10,000 rpm at $4^{\circ}C$. Affinity-purified antigen(antibody-bound antigen) was prepared from fractions(bound and unbound) of crude antigen by affinity chromatography on CNBr-activated sepharose 4B, and IgG as a ligand was prepared from paragonimiasis cat serum(6 months infected) obtained by ammonium sulfate ($40%{\sim}45%$ saturated) precipitation method. By SDS-PAGE, crude antigen showed 22 polypeptide fractions while purified antigen showed 4 fractions: 36, 400, 34, 700, 27, 600 and 11, 500 in molecular weights. All cats were divided into five groups($G_1-G_5$) by different worm burdens. The mean of recovered worms(${\pm}SD$) and the number of cats in each group are as follows: $G_1$, 2 worms(0) and 4 cats; $G_2$, 4.75 (${\pm}0.66$) and eight; $G_3$, 10.75(${\pm}1.92$) and four; $G_4$, 23.20(${\pm}3.43$) and five; $G_5$, 48(${\pm}12.63$) and five cats. The results were summarized as follows: 1. The antibody levels(OD value) increased by worm burden in $G_1$ to $G_4$ generally. However, individual antibody levels were not exactly related with worm burden in all groups, especially there was a wide difference in $G_4$ and $G_5$. These results suggested that the worm burden in $G_4$ (about $20{\sim}30$ worms) is enough to produce antibody maximum in cats of $2{\sim}3kg$ weight. 2. The antibody levels increased significantly(p<0.05) compared to control sera at the 3rd week in $G_1$ and $G_2$, at the 2nd week in $G_3$, and at the 1st week in $G_4$ and $G_5$. Especially in the 4th week, OD value increased more in $G_1$(p<0.01) and in $G_2$ to $G_5$(p<0.001). In the pattern of antibody levels by ELISA in each group, OD in $G_1$ increased to the 18th week continuously, in $G_2$ OD was maintained same after the 16th week, but in $G_3$ it decreased after the 16th week, and it was maintained same in $G_4$ and $G_5$ after the 14th week. 3. The antibody levels by ELISA with the affinity-purified antigen were higher than those with crude antigen in all groups generally. Especially, the difference of OD values between two antigens was larger from the 4th to the 10th week. In $G_1$ and $G_2$ OD with purified antigen was higher than that with crude one to the 18th week. It was also higher in $G_3$ than that with crude antigen to the 16th week and OD of $G_4$ and $G_5$ were higher before the 14th week than that with crude antigen, however became lower at the 16th week. Consequently, the antibody level in ELISA with affinity-purified antigen was more sensitive at the early weeks after infection and in light infection groups than that with crude antigen.

• Food Science of Animal Resources
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• 제29권2호
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• pp.238-244
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• 2009

Food allergy is a serious nutritional problem in both children and adults. Therefore, food allergenicity reduction methods are greatly needed. The allergenicity is altered by various manufacturing processes, and the digestibility of food proteins can be affected by food processing. This study was conducted to investigate the effect of in-vitro digestibility on the allergenicity of autoclaved market pork sausages using pepsin (30min) and trypsin (5, 30, 60, 90, and 120min). The binding ability of the porcine serum albumin (PSA) from sausages A and B significantly decreased by about 30 and 23%, respectively, after autoclave treatment (121; 5, 10, and 30 min). After the pepsin and trypsin treatments, the binding ability of products A and B at 30 min decreased. These competitive indirect enzyme-linked immunosorbent assay (ci-ELISA) results corresponded well with the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting results. The results demonstrated that the allergenicity of pork sausages considerably decreased after autoclave treatment, and were also maintained or decreased after enzyme treatment. Accordingly, autoclave treatment represents a promising processing technology for the reduction of the allergenicity of diverse food products.

• Clinical and Experimental Pediatrics
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• 제48권4호
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• pp.411-415
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• 2005

Purpose : This study was performed to determine the developmental expression pattern of nerve growth factor(NGF) in the urine of healthy children. It was hypothesized that NGF may contribute to the development of the spinobulbospinal micturition reflex that represents the adult micturition pattern. Methods : Voided urine was collected in 60 healthy children during the first 5 years of life(0-1 month, n=10; 1 month-1 year, n=10; 2 years, n=10; 3 years, n=10; 4 years, n=10). The urinary NGF was analyzed by using an enzyme linked immunosorbent assay. Results were normalized based on creatinine or total protein in urine. Results : NGF was significantly greater among neonates compared to other age groups(P<0.05). NGF levels declined during 1 month to 1 year and increased by age 2 years. NGF levels at age 3 years were less than in neonates examined. When comparing the NGF levels according to continence in children over 1 year old, NGF levels were significantly lower in children with continence than in children with incontinence(P<0.05). Conclusion : These data suggest that NGF is involved in the achievement of continence and in mechanisms of bladder nerve growth and in the reorganization of bladder reflex pathway.

• IMMUNE NETWORK
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• 제4권1호
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• pp.23-30
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• 2004

Background: A human orthologue of mouse S100A6-binding protein (CacyBP), Siah-1-interacting protein (SIP) had been shown to be a component of novel ubiquitinylation pathway regulating $\beta$-catenin degradation. The role of the protein seems to be important in cell proliferation and cancer evolution but the expression pattern of SIP in actively dividing cancer tissues has not been known. For the elucidation of the role of SIP protein in carcinogenesis, it is essential to produce monoclonal antibodies specific to the protein. Methods: cDNA sequence coding for ORF region of human SIP gene was amplified and cloned into an expression vector to produce His-tag fusion protein. Recombinant SIP protein and monoclonal antibody to the protein were produced. The N-terminal specificity of anti-SIP monoclonal antibody was conformed by immunoblot analysis and enzyme linked immunosorbent assay (ELISA). To study the relation between SIP and colon carcinogenesis, the presence of SIP protein in colon carcinoma tissues was visualized by immunostaining using the monoclonal antibody produced in this study. Results: His-tag-SIP (NSIP) recombinant protein was produced and purified. A monoclonal antibody (Korea patent pending; #2003-45296) to the protein was produced and employed to analyze the expression pattern of SIP in colon carcinoma tissues. Conclusion: The data suggested that anti-SIP monoclonal antibody produced here was valuable for the diagnosis of colon carcinoma and elucidation of the mechanism of colon carcinogenesis.

• Journal of Environmental Health Sciences
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• 제38권3호
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• pp.204-212
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• 2012

Objectives: The purpose of this study was to determine house dust mite concentrations in living rooms and bedding materials in atopic case-control groups. Methods: Fifty four homes with children suffering atopic diseases as cases and fifty one homes without atopic diseases as controls were selected after diagnosis at three elementary schools in Ulsan City. Dust samples were collected from the living rooms and bedding materials in each home during summer and winter with a vacuum cleaner and analyzed for house dust mites using enzyme-linked immunosorbent assay (ELISA). Environmental factors and family history were surveyed during sampling. Results: Both species of house dust mite Dermatophagoides farina (Der f 1) and Dermatophagoides pteronyssinus (Der p 1) were found in all homes. Derf1 concentrations were much higher than Der P1 concentrations, both in atopic homes (1518.9 vs. 27.0 ng/g. dust, respectively) and non-atopic homes (810.8 vs. 44.3 ng/g. dust, respectively). Der f 1 concentrations were significantly higher in the atopic group than in the non-atopic group (atopic: 1518.9 ng/g.dust, non-atopic: 810.8 ng/g.dust, p = 0.035). However, total house dust mite Der p 1 concentrations were significantly higher in the non-atopic group than in the atopic group (atopic: 27.0 ng/g.dust, non-atopic: 44.3 ng/g.dust, p = 0.035). Multiple regression implied that mothers with family history of atopic diseases (OR = 4.79, 95% CI = 1.81-12.69), Der f 1 concentrations (OR = 1.74, 95% CI = 1.07-2.81), and air freshener use (OR = 4.60, 95% CI = 1.72-12.34) had significant associations with atopic children. Conclusion: House dust mite Der f 1 concentrations were associated with atopic children. This study suggests that parents should reduce house dust mite Der f 1 concentrations through environmental controls.

• Korean Journal of Clinical Laboratory Science
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• 제37권1호
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• pp.56-60
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• 2005

Human chorionic gonadotrophin (hCG) and unconjugated estriol (uE3) were added to AFP to make what is commonly known as the Triple test. The Triple test combines results from these three tests and has been a standard screening procedure for several years. Recent studies have demonstrated the usefulness of adding inhibin-A to Down's syndrome risk assessment. The Quad test adds dimeric Inhibin-A (DIA) to the three other markers and uses the same computer program to calculate risk factors. Testing was performed between 14 and 21 weeks of gestation. Sample size were 648 samples and period of study was from 1, July, 2004 to 30, September, 2004. Used analytical methods for AFP, hCG and uE3 were radioimmunoassay (RIA) and dimeric inhibin A was enzyme-linked immunosorbent assay (ELISA). Adding dimeric inhibin-A as a fourth marker to the standard triple test increases the detection rate from 62 % to 75 % with a false-positive rate of 5%. The DIA based Quad test has been shown to be the most effective second trimester screening test for Down's syndrome suitable for routine use. Increased DIA values are observed during normal pregnancy where a bimodal pattern response is seen. Values increase during the first trimester, decline after 14 weeks, and re-ascend between 17-25 weeks. Values for DIA may be additionally elevated during a Down's syndrome pregnancy. Dimeric inhibin A is a glycoprotein hormone made by the ovary and placenta. DIA levels are twice as high in Down's syndrome pregnancies. AFP, hCG, and uE3 levels vary with gestational age, and incorrect gestational dating will influence results. DIA levels do not vary substantially with gestational age, resulting in greater screening accuracy. Although the Quad test is an improvement over the Triple test, it is important to underscore the fact that a positive test on both should be done. Most women who initially screen positive will be found to be carrying normal babies when amniocentesis and definitive diagnostic chromosome analysis are done.