Multiple heterologous M2 extracellular domains presented on virus-like particles confer broader and stronger M2 immunity than live influenza A virus infection

Kim, M.C.,Lee, J.S.,Kwon, Y.M.,O, E.,Lee, Y.J.,Choi, J.G.,Wang, B.Z.,Compans, R.W.,Kang, S.M. Elsevier/North-Holland 2013 Antiviral research Vol.99 No.3

The influenza M2 ectodomain (M2e) is poorly immunogenic and has some amino acid changes among isolates from different host species. We expressed a tandem repeat construct of heterologous M2e sequences (M2e5x) derived from human, swine, and avian origin influenza A viruses on virus-like particles (M2e5x VLPs) in a membrane-anchored form. Immunization of mice with M2e5x VLPs induced protective antibodies cross-reactive to antigenically different influenza A viruses and conferred cross protection. Anti-M2e antibodies induced by heterologous M2e5x VLPs showed a wider range of cross reactivity to influenza A viruses at higher levels than those by live virus infection, homologous M2e VLPs, or M2e monoclonal antibody 14C2. Fc receptors were found to be important for mediating protection by immune sera from M2e5x VLP vaccination. The present study provides evidence that heterologous recombinant M2e5x VLPs can be more effective in inducing protective M2e immunity than natural virus infection and further supports an approach for developing an effective universal influenza vaccine.

Analysis of 22 Y chromosomal STR haplotypes and Y haplogroup distribution in Pathans of Pakistan

Lee, E.Y.,Shin, K.J.,Rakha, A.,Sim, J.E.,Park, M.J.,Kim, N.Y.,Yang, W.I.,Lee, H.Y. Elsevier Science 2014 FORENSIC SCIENCE INTERNATIONAL GENETICS Vol.11 No.-

We analyzed haplotypes for 22 Y chromosomal STRs (Y-STRs), including 17 Yfiler loci (DYS19, DYS385a/b, DYS389I/II, DYS390, DYS391, DYS392, DYS393, DYS437, DY438, DYS439, DYS448, DYS456, DYS458, DYS635 and Y-GATA-H4) and five additional STRs (DYS388, DYS446, DYS447, DYS449 and DYS464), and Y chromosomal haplogroup distribution in 270 unrelated individuals from the Pathans residing in the Federally Administered Tribal Areas and the North-West Frontier Province of Pakistan using in-house multiplex PCR systems. Each Y-STR showed diversities ranging from 0.2506 to 0.8538, and the discriminatory capacity (DC) was 73.7% with 199 observed haplotypes using 17 Yfiler loci. By the addition of 5 Y-STRs to the Yfiler system, the DC was increased to 85.2% while showing 230 observed haplotypes. Among the additional 5 Y-STRs, DYS446, DYS447 and DYS449 were major contributors to enhancing discrimination. In the analysis of molecular variance, the Pathans of this study showed significant differences from other Pathan populations as well as neighboring population sets. In Y-SNP analysis, a total of 12 Y chromosomal haplogroups were observed and the most frequent haplogroup was R1a1a with 49.3% frequency. To obtain insights on the origin of Pathans, the network analysis was performed for the haplogroups G and Q observed from the Pathans and the Jewish population groups including Ashkenazim and Sephardim, but little support for a Jewish origin could be found. In the present study, we report Y-STR population data in Pathans of Pakistan, and we emphasize the need for adding additional markers to the commonly used 17 Yfiler loci to achieve more improved discriminatory capacity in a population with low genetic diversity.

Cellulase-amyloglucosidase와 효모의 가스생성법에 의한 사료의 에너지가 측정에 관한 연구

김영길,이광목,김욱 東亞大學校生命資源科學大學附設 農業資源硏究所 1993 農業生命資援硏究 Vol.2 No.2

粗飼料와 濃厚飼料를 混合한 飼料의 營養價를 Cellulase와 Amyloglucosidase, 酵母를 利用한 가스 生成法에 의하여 消化率과 TDN을 測定할 수 있는 方法을 確立하고자 本 硏究를 實施한 實驗 結果는 다음과 같다. 1. Cellulose 加水分解 酵素인 Trichoderma viride cellulase의 最適 活性 pH는 4.5-4.8이었으며 pH가 5.0보다 높을 때에는 活性이 急激히 저하하였다. 2. Cellulose 加水分解시 Trichoderma viride celulase의 溫度의 의한 影響은 50℃에서 最大 活性을 나타내었으며, 40℃에서는 活性이 92%정도로 나타났다. 3. Trichoderma viride-aspergillus niger 混合, cellulase 使用時의 가스 生成量은 Trichoderma viride cellulase 使用時 가스 生成量 보다 20-30% 높게 나타났으므로 cellulose 加水分解時 Trichoderma viride-aspergillus niger 混合 cellulase를 使用하는 것이 더욱 效果的일 것으로 思料된다. 4. 포도당(X)과 가스 生成量(Y)과의 關係는 Y=6.82+165X의 回歸式과 相關係數는 0.92로서 高度의 有意性(P<0.01)이 있었다. 5. Trichoderma viride cellulase 使用時 가스 生成量(X)과 乾物 消化率(Y)과의 關係는 Y=29.9+1.13X(r=78**)의 回歸式을 나타내었고, Trichoderma viride-aspergillus niger 混合 vellulase 使用時 가스 生成量(X)과 乾物 消化率(Y)과의 關係는 Y=28.9+0.88X(r=0.82**)의 回歸式을 나타내었다. Trichoderma viride cellulase 使用時 보다 Trichoderma viride-aspergillus niger 혼합 cellulase 사용시 相關係數가 더 높았다. 6. 가스 生成量(X)과 TDN(Y)과의 關係는 Y=35.8+0.585X의 回歸式과 相關係數 0.93으로서 高度의 有意性을 나타내었고, TDN(X)과 가스 生成量(Y)과의 關係는 Y=1.68X-59.6의 回歸式을 나타내었다. The experiment was conducted to establish the gas production method by cellulase, amyloglucosidase and yeast to determine the digestibility and energy value(TDN) of feedstuffs. The results obtained were summarized as follows; 1. The highest activity of Trichoderma viride cellulase was shown at pH 4.8 and the activity of the cellulase decreased rapidly at higher pH than 5.0. 2. The highest activity of Trichoderma virie cellulase was shown at 50℃ and the activity at 40℃ was approximately 92% compared with that at 50℃. 3. The mixed celllulases of Trichoderma viride and Aspergillus nigher produced more gas than Trichoderma viride cellulase when incubated with feedstuffs and were identified to be more effectively used for gas production method. 4. The regression equation of gas production(Y) on glucose production (X) was Y=6.82+165X with significant correlation coefficient of 0.92. 5. The regression equation of DMD(Y) on gas production(X) was Y=29.9+1.13X(r=0.78) with Trichoderma viride cellulase and Y=28.9+0.88X (r=0.82) with both Trichoderma viride and Aspergillus niger cellulase. 6. The regression equation of TDN(Y) on gas production(X) was Y=35.8+0.58X with significant correlation coefficients of 0.93 and in conclusion energy value of feedstuffs can be estimated by gas production method.

Role of flotillins in the endocytosis of GPCR in salivary gland epithelial cells

Park, M.Y.,Kim, N.,Wu, L.L.,Yu, G.Y.,Park, K. Academic Press 2016 Biochemical and biophysical research communication Vol.476 No.4

Endocytosis has numerous functions in cellular homeostasis. Defects in the endocytic pathway of receptors may lead to dysfunction of salivary gland secretion. Therefore, elucidating the complex mechanisms of endocytosis may facilitate solutions for disease treatment and prevention. The muscarinic type 3 receptor (M3R), a G-protein-coupled receptor (GPCR) located in the plasma membrane, is involved in numerous physiological activities such as smooth muscle contraction and saliva secretion. M3R enters cells through clathrin-mediated endocytosis (CME), while flotillins (flot-1 and -2), highly conserved proteins residing in lipid-raft microdomains, make use of clathrin-independent endocytosis (CIE) for their internalization. Since these two proteins use two discrete pathways for endocytic entry, the association of flotillins with CME is poorly understood. We examined whether flotillins play a role in CME of M3R using immunoblotting, immunocytochemistry, confocal immunofluorescence microscopy, co-immunoprecipitation, and RNA interference techniques in secretory epithelial cells. Upon stimulation with a cholinergic agonist, M3R, flot-1, and flot-2 each internalized from the plasma membrane into intracellular sites. The addition of chlorpromazine and cytochalasin D, well-known inhibitors of CME, inhibited internalization of M3R via CME. Filipin III and methyl-β-cyclodextrin (mβCD) acting as lipid raft inhibitors disrupted internalization of flot-½ via CIE. Interestingly, filipin III and mβCD slightly reduced expression level of M3R whereas chlorpromazine and cytochalasin D did not affect internalization of the flotillin isoforms. M3R and flot-½ colocalized and interacted with each other as they entered the cytosol during limited periods of incubation. Moreover, knockdown of flot-1 or -2 by flotillin-specific siRNA prevented internalization and reduced the endocytic efficiency of M3R. Our results suggest that flot-1 and -2 are partially involved in CME of M3R by facilitating its internalization.

Feed Intake Patterns and Growth Performance of Purebred and Crossbred Meishan and Yorkshire Pigs

Hyun, Y.,Wolter, B.F.,Ellis, M. Asian Australasian Association of Animal Productio 2001 Animal Bioscience Vol.14 No.6

Two experiments were conducted to compare the feed intake patterns and growth performance of Meishan and Yorkshire growing pigs. Experiment 1 was carried out over a 6-wk period and used 48 barrows with equal numbers of purebred Meishan (M) and Yorkshire (Y). Pigs were allocated to four groups of 12 pigs consisting of equal numbers of M and Y. Initial BW were $36.4{\pm}0.32kg$ and $42.1{\pm}1.41kg$ for M and Y, respectively. Experiment 2 was carried out over a 5-week period and used 48 pigs consisting of equal numbers of both barrows and gilts and of crossbred Meishan$\times$Yorkshire (MY) and purebred Yorkshire (Y) animals. Pigs were allotted to 6 pens of 8 pigs, with 4 single- and 2 mixed-genotype groups (initial $BW=28.5{\pm}0.99kg$). In both experiments, pigs were given ad libitum access to a grower diet (17% crude protein, 0.9% lysine, 3365 kcal/kg ME) via feed intake recording equipment (F.I.R.E.). Pigs carried an ear-tag transponder with an unique identification which allowed the time, duration, and size of individual meals to be recorded. In Exp. 1, Y had higher ADG (721 vs 353 g, p<0.01), daily feed intake (DFI; 2.338 vs 1.363 kg, p<0.01), made more frequent visits to the feeder per day (NFV; 18.5 vs 7.7, p<0.01), had a shorter feeder occupation time per visit (FOV; 7.4 vs 12.9 min, p<0.01), and ate less feed per visit (FIV; 130 vs 177 g, p<0.01) than M pigs. Feed consumption rates (CR) were greater for Y compared to M (19.3 vs 14.8 g/min, p<0.01). Feeder occupation time per day (FOD) was longer for Y than M (114.3 vs 82.8 min/pig, p<0.01). Yorkshire pigs visited the feeder more frequently between 0800 and 1100 h. Meishan pigs showed more frequent feeder visits between 0600 and 0800 h, and between 1600 and 2100 h when feeding competition with Y was reduced. In Exp. 2, there was no effect of genotype or group composition on DFI, ADG or gain:feed ratio. Crossbred pigs (MY) made fewer feeder visits (12.6 vs 17.7, p<0.01), and had greater FIV (124 vs 98 g/visit, p<0.01), and longer FOV (8.11 vs 7.24 min/visit, p<0.01) and FOD (112 vs 100 min, p<0.05) than Y pigs. Results of this study suggest substantial genetic variation in feeding patterns as well as in growth performance.

Supplemented vaccination with tandem repeat M2e virus-like particles enhances protection against homologous and heterologous HPAI H5 viruses in chickens

Song, B.M.,Kang, H.M.,Lee, E.K.,Jung, S.C.,Kim, M.C.,Lee, Y.N.,Kang, S.M.,Lee, Y.J. Butterworths ; Elsevier Science Ltd 2016 Vaccine Vol.34 No.5

Highly pathogenic avian influenza (HPAI) H5 viruses derived from A/Goose/Guangdong/1/96 have been continuously circulating globally, severely affecting the public health and poultry industries. The matrix 2 protein ectodomain (M2e) is considered a promising candidate for a universal cross-protective influenza vaccine that provides more effective control over HPAI H5 viruses harboring variant hemagglutinin (HA)-antigens. Here, we evaluated the protective efficacy of a tandem repeat construct of heterologous M2e presented on virus-like particles (M2e5x VLPs) either alone or as a supplement against HPAI H5 viruses in a chicken model. Chickens immunized with M2e5x VLPs alone induced M2e-specific antibodies but were not protected against HPAI H5. The homo- and cross-protective efficacy of M2e5x VLP-supplemented vaccination of chickens was also examined. Importantly, supplementation with M2e5x VLPs induced significantly higher levels of antibodies specific for M2e and different viruses as well as provided improved protection against homologous and heterologous HPAI H5 viruses. Considering the limited efficacy of inactivated vaccines, supplement vaccination with M2e5x VLPs may be an effective measure for preventing outbreaks of HPAI viruses that have the ability to constantly change their antigenic properties in poultry.

Metabolic characterization of (1-(5-fluoropentyl)-1H-indol-3-yl)(4-methyl-1-naphthalenyl)-methanone (MAM-2201) using human liver microsomes and cDNA-overexpressed cytochrome P450 enzymes

Kong, T. Y.,Kim, J. H.,Choi, W. G.,Lee, J. Y.,Kim, H. S.,Kim, J. Y.,In, M. K.,Lee, H. S. Springer Science + Business Media 2017 Analytical and bioanalytical chemistry Vol.409 No.6

<P>MAM-2201 is a synthetic cannabinoid that is increasingly found in recreational drug abusers and cases of severe intoxication. Thus, characterization of the metabolic pathways of MAM-2201 is necessary to predict individual pharmacokinetics and toxicity differences, and to avoid toxic drug-drug interactions. Collectively, 19 phase 1 metabolites of MAM-2201 were identified using liquid chromatography-Orbitrap mass spectrometry following human liver microsomal incubations in the presence of NADPH: 7 hydroxy-MAM-2201 (M1-M7), 4 dihydroxy-MAM-2201 (M8-M11), dihydrodiol-MAM-2201 (M12), N-(5-hydroxypentyl)-MAM-2201 (M13), hydroxy-M13 (M14), N-dealkyl-MAM-2201 (M15), 2 hydroxy-M15 (M16, M17), MAM-2201 N-pentanoic acid (M18), and hydroxy-M18 (M19). On the basis of intrinsic clearance values in human liver microsomes, hydroxy-MAM-2201 (M1), N-(5-hydroxypentyl)-MAM-2201 (M13), and hydroxy-M13 (M14) were the major metabolites. Based on an enzyme kinetics study using human cDNA-expressed cytochrome P450 (CYP) enzymes and an immunoinhibition study using selective CYP antibodies in human liver microsomes, CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 enzymes were responsible for MAM-2201 metabolism. The CYP3A4 enzyme played a prominent role in MAM-2201 metabolism, and CYP1A2, CYP2B6, CYP2C8, and CYP2C9 enzymes played major roles in the formation of some metabolites. MAM-2201 is extensively metabolized by multiple CYP enzymes, indicating that MAM-2201 and its metabolites should be used as markers of MAM-2201 abuse and toxicity.</P>

早期破膜에 關한 臨床的 考察

許湘,吳文愛,安東源,申吳永 대한산부인과학회 1972 Obstetrics & Gynecology Science Vol.15 No.7

Effect of Antimuscarinic Autoantibodies in Primary Sjögren’s Syndrome

Kim, N.,Shin, Y.,Choi, S.,Namkoong, E.,Kim, M.,Lee, J.,Song, Y.,Park, K. SAGE Publications 2015 Journal of dental research Vol.94 No.5

<P>The presence of functional autoantibodies against the muscarinic type 3 receptor (M3R) has been reported in primary Sjogren's syndrome (pSS). However, the pathogenic role of these autoantibodies in pSS development remains to be elucidated. In this experiment, we investigated a pathologic role of pSS autoantibodies (pSS IgG) associated with downregulation of the major histocompatibility complex I (MHC I) molecule with M3R through internalization. Anti-M3R autoantibodies in purified control and pSS IgG were detected using 4 synthesized cyclic M3R peptides by enzyme-linked immunosorbent assay. The binding reactivity of pSS IgG to M3R in situ was analyzed by a dual immunostaining method. Surface expression, interaction, and internalization of M3R with MHC I were analyzed by immunofluorescence confocal microscopy and biochemical assays. Synthetic cyclic peptides M3RP(205-221) and M3RP(520-527) showed significantly high reactivity with pSS IgG compared to the control IgG or the other 3 peptides (P < 0.05). Significantly high reactivity of pSS IgG to M3R in situ was observed. PSS IgG increased the interaction of membrane M3R with MHC I and induced their internalization in primary human submandibular gland cells. The pSS IgG-induced internalization of M3R with MHC I was significantly inhibited by the cholesterol-sequestering drug filipin. Our novel findingnamely, strong downregulation of the membrane MHC I with M3R through internalization of the cholesterol-rich microdomain associating with anti-M3R autoantibodiescould be an important mechanism contributing to the impaired salivation seen in pSS and linking secretory hypofunction to autoimmune pathogenesis.</P>

수도의 유묘기 냉해에 관한 연구 II. 주요 신품종들의 제 3 엽기 내냉한계

권용웅,김주헌,오윤진,이문희,Kwon, Y.W.,Kim, J.H.,Oh, Y.J.,Lee, M.H. 한국작물학회 1979 한국작물학회지 Vol.24 No.2

Indica \times Japonica 교잡에 의해 육성된 수도신품종들의 유묘기 내냉한계와 내냉발현특성을 Janica품종들과 비교 고찰하고자 10개 품종을 유묘기 내냉성이 제일 약한 제 3 엽기에 주간 11\circ C야간 5\circ C의 저온조건에 0, 3, 4, 5, 8, 10일 처리한 결과는 다음과 같다. 1. 저온처리 전후에 주야 26\circ C / 18\circ C에서 생장한 파종 후 35일묘의 건물중이 무처리묘의 90% 이상이 되는 최장저온처리기간은 신품종들의 경우 0.8~2.8일 간으로써 특히 조생통일이 짧고 밀야 2003가 길었으며, Japonica 품종들의 경우 4.0~5.2일간으로써 신품종들보다 가역적 내냉한계기간이 배정도 길었다. 2. 저온처리에 따른 냉해발현 종료후 묘생존율이 90%이상되는 최장저온처리기간은 신품종들의 경우 3.0~5.6 일간으로써 수원 258호가 제일 짧고 수원 25001가 제일 길었으며, Japonica 품종들은 공시품종 모두 10일간 이상으로 신품종들보다 비가역적 내냉한계기간이 2~3배 이상 길었다. 3. 저온처리중 엽신변색고사율(y_1) 및 묘고사율(y_2), 저온처리 후 발현된 모신변색고사율(y_3), 묘고사율(y_4), 냉해를 받은 35일묘의 초장(y$_{5}$)과 건물중(y$_{6}$)은 모두 저온처리기간(x)과 고도로 유의한 상관성을 보였는데 특히 y_1, y_3, y$_{6}$와 높은 상관성을 보였으며, y_1~y$_{6}$들 상호간에도 모두 고도로 유의한 상관성을 보였으나 특히 y_1과 y_3, y_3와 y$_{6}$의 상관성이 컸다. 4. 냉해징상들의 발현순서와 상태들간의 상관성 분석결과로부터 가역적 및 비가역적, 직접적 및 간접적 냉해의 발생과 그들의 복합현상으로써 결과되는 관찰되는 냉해징상의 발현과정 모식도를 작성하여 그림 5에 제시하였다. 5. 수도유묘의 내냉성 평가에 있어서 가역적 피해와 비가역적 피해를 종합적으로, 그리고 정량적으로 나타내며 일반적으로 냉해를 나타내는 조사형질들 중 냉해에 가장 예민한 반응을 보이는 건물중율을 1차요인으로 취급할 것으로 제의한다. Comparative tolerance of the new rice varieties from Indicax Japonica cross to elastic and plastic chilling injuries was examined at the 3rd leaf-stage, the most sensitive stage to chilling during nursery, using a chilling chamber set 11\circ C, 13hrs. 20K lux day/5$^{\circ}C$ night. Seedlings were reared until 35 days after seeding in a 26\circ C, 13hrs. day / 18\circ C night chamber except the period of chilling treatment for 0, 3, 4, 6, 8 or 10 days. The new varieties were approximately twice susceptible to elastic injuries, and thrice susceptible to plastic injuries than the Japonica varieties in the term of chilling duration. Among the variables indicating apparant chilling injuries the reduction in dry matter responded most sensitively and differentiated well the varietal difference in chilling resistance even with shorter chilling period. Sequential aspects of the cause and effect relations involved in the development of the apparant chilling injuries are discussed and a schematic diagram is presented as Fig. 5 in the text.