Dissolved organic matter (DOM) is a key component in the biogeochemical cycling in freshwater ecosystem. However, it has been rarely explored, particularly complex river watershed dominated by natural and anthropogenic sources, such as various effluent facility and livestock. The current research developed a new analytical method for TOC/TN (Total Organic Carbon/Total Nitrogen) stable isotope ratio, and distinguish DOM source using stable isotope value (δ13C-DOC) and spectroscopic indices (fluorescence index [FI] and biological index [BIX]). The TOC/TN-IR/MS analytical system was optimized and precision and accuracy were secured using two international standards (IAEA-600 Caffein, IAEA-CH-6 Sucrose). As a result of controlling the instrumental conditions to enable TOC stable isotope analysis even in low-concentration environmental samples (<1 mgC L-1), the minimum detection limit was improved. The 12 potential DOM source were collected from watershed, which includes top-soils, groundwater, plant group (fallen leaves, riparian plants, suspended algae) and effluent group (pig and cow livestock, agricultural land, urban, industry facility, swine facility and wastewater treatment facilities). As a result of comparing characteristics between 12 sources using spectroscopic indices and δ13C-DOC values, it were divided into four groups according to their characteristics as a respective DOM sources. The current study established the TOC/TN stable isotope analyses system for the first time in Korea, and found that spectroscopic indices and δ13C-DOC are very useful tool to trace the origin of organic matter in the aquatic environments through library database.

Background: The regulation of maternal immunity is critical for the establishment and maintenance of successful pregnancy. Among many cell types regulating the immune system, innate lymphoid cells (ILCs) are known to play an important role in innate immunity. Although some reports show that ILCs are present at the maternalconceptus interface in humans and mice, the expression and function of ILCs in the endometrium have not been studied in pigs. Methods: Thus, we determined the expression, localization, and regulation of ILC markers, CD127 (a common marker for ILCs), BCL11B (a ILC2 marker), and RORC (a ILC3 marker) at the maternal-conceptus interface in pigs. Results: The expression of BCL11B and RORC, but not CD127, in the endometrium changed during pregnancy in a stage-specific manner and the expression of CD127, BCL11B, and RORC was greatest on Day 15 during pregnancy. CD127, BCL11B, and RORC were also expressed in conceptus tissues during early pregnancy and in chorioallantoic tissues during the later stage of pregnancy. BCL11B and RORC proteins were localized to specific cells in endometrial stroma. The expression of CD127 and BCL11B, but not RORC, was increased by the increasing doses of interferon-γ (IFNG) in endometrial explants. Conclusions: These results suggest that ILCs present at the maternal-conceptus interface may play a role in the establishment and maintenance of pregnancy by regulating the innate immunity in pigs.

The cellular communication network factor (CCN) family proteins regulate many biological events such as angiogenesis, tumor growth, placentation, implantation, and embryogenesis. The expression and function of CCN1, CCN2, and CCN3 at the maternal-conceptus interface are established in humans and rodents, but little is known about the role of CCN4 to CCN6 in the reproductive organs in any other species. Several studies in transcriptome analysis in pigs have shown that the expression of CCN4 and CCN6 increases in the endometrium during early pregnancy. However, their expression, regulation, and function in the endometrium throughout the estrous cycle and pregnancy have not been fully understood in pigs. Thus, we determined the expression, localization, and regulation of CCN4 and CCN6 during the estrous cycle and at the maternal-conceptus interface in pigs. We found that the levels of CCN4, but not CCN6, changed during the estrous cycle. The levels of CCN4 were greater during mid- to late pregnancy than in the early stage, and the levels of CCN6 were greatest on Day 15 of pregnancy. CCN4 and CCN6 were detected in conceptus tissues during early pregnancy and in chorioallantoic tissues during the later stage of pregnancy. CCN4 mRNA was mainly localized to epithelial cells, CCN6 mRNAs to epithelial and stromal cells in the endometrium. In endometrial explant cultures, CCN4 expression was increased by progesterone, and CCN6 expression by interferon-γ. These results suggest that CCN4 and CCN6 may play roles in the establishment and maintenance of pregnancy by regulating the endometrial epithelial cell functions in pigs.

Secretory leukocyte protease inhibitor (SLPI), also known as neutrophil elastase and cathepsin-G protease inhibitor, functions in protection of epithelial cells from proteases. SLPI is expressed and secreted by many mucosal tissues, including lungs, seminal vesicles and cervix in women. SLPI plays an important role in protection of endometrial epithelial cells during pregnancy from degradation by degradation by proteases derived from trophoblast at the maternal-conceptus interface. In pigs, SLPI mRNA is known to be expressed in endometrial tissues, but the expression of SLPI in the endometrium throughout the estrous cycle and pregnancy has not been determined. Therefore, we analyzed the expression and regulation of SLPI mRNA in the endometrium throughout the whole stages of the estrous cycle and pregnancy in pigs. We obtained endometrial tissues from gilts on Days 0 (day of estrus), 3, 6, 9, 12, 15, and 18 of the estrous cycle and on Days 10, 12, 15, 30, 60, 90, and 114 of pregnancy. Real-time RT-PCR analysis showed that the expression of SLPI mRNA in the endometrium increases during midt-o late pregnancy. During the estrous cycle, levels of SLPPI mRNA in estrus and proestrus were higher than those in diestrus and metestrus. In situ hybridization analysis showed that SLPI mRNA was specifically localized to the glandular epithelial cells in the endometrium during pregnancy with strong signal intensity during mid-to late pregnancy. SLPI mRNA was not detectable in conceptus tissues on Days 12 and 15 of pregnancy, but SLPI mRNA was expressed in chorioallantoic tissues during mid-to term pregnancy with increasing levels toward term pregnancy. To determine the effects of steroid hormones, estrogen and progesterone, on the expression of SLPI mRNA, endometrial explant tissues from immature pigs were treated with increasing doses of estradiol-17β (E2) and progesterone (P4). Increasing doses of E2 and P4 increased the expression of SLPI mRNA in endometrial tissues. These results showed that SLPI was expressed in the endometrium in a pregnancy stage-and cell type-specific manner and the expression of SLPI was regulated by E2 and P4 in endometrial tissues, suggesting that SLPI may play an important role in regulating the endometrial epithelial cell function during mid-to late pregnancy in pigs. Further analysis to determine the roles of SLPI at the maternal-conceptus interface is still needed.

Caspases are a family of cysteine protease enzymes composed of more than 10 members that play essential roles in apoptosis and inflammation. It has been reported that caspases play a critical role in regulating apoptosis at the maternal-conceptus interface in many species. However, the expression and regulation of caspases have not been determined in the endometrium in pigs. Therefore, we analyzed the expression, localization, and regulation of caspases in the endometrium during the estrous cycle and pregnancy in pigs. Real-time RT-PCR analysis showed that caspases were expressed in the endometrium during the estrous cycle and pregnancy. The expression of CASP6, CASP7, and CASP8 during the estrous cycle and CASP3, CASP6, CASP7, and CASP10 changed during pregnancy. Levels of CASP3 mRNA in the endometrium were higher on Day 12 of pregnancy than the estrous cycle and levels of CASP7 mRNAs were highest on Day 15 of estrous cycle and pregnancy. Immunohistochemistry analysis showed that CASP3 protein was localized to endometrial epithelial cells on Days 12 and 15 the estrous cycle and pregnancy, but cleaved CASP3 was localized only to luminal epithelial (LE) cells on Days 12 and 15 of pregnancy in the endometrium. CASP7 protein was localized to endometrial LE cells only on Day 15 of pregnancy. CASP3, CASP6, CASP7, CASP8, and CASP10 mRNAs were detectable in conceptus on D12 and D15 of pregnancy, and chorioallantoic tissues expressed CASP3, CASP6, CASP7, CASP8, CASP8, and CASP10 with increasing levels toward term pregnancy, except CASP3 mRNA. The effect of steroid hormones and interleukin-1βß (IL1B) on CASP3 expression and the effect of interferon-γ(IFNG) on CASP7 expression was determined by endometrial explant cultures and we found that CASP3 expression was increased by IL1B and CASP7 expression was increased by IFNG in a dose-dependent manner. These results showed that caspases were expressed in the endometrium during the estrous cycle and pregnancy in a stage- and/or pregnancy-specific dependent manner and some caspases were regulated by IL1B or IFNG in the endometrial tissues, suggesting that caspase may play an important role in regulating apoptosis for the establishment and maintenance of pregnancy at the maternal-conceptus interface in pigs.

붉은곰팡이는 곰팡이독소를 생성하는 식물병원균으로 수확된 보리 알곡에 잔존하여 저장 중 적절한 환경이 조성 되면 곰팡이독소를 생성할 수 있다. 저장 중 겉보리의 저장온도와 곡물수분함량이 붉은곰팡이와 곰팡이독소 오염에 미치는 영향을 알아보기 위해 전라도에서 수집한 겉보리 시료 3점의 수분함량을 14%와 20%로 조절한 후 온도 조절이 되지 않는 상온창고와 12oC 이하로 온도를 조절한 저온창고에 각각 저장하였다. 창고의 온도와 습도를 실시간으로 측정하면서 1, 3, 6, 12개월 후 시료의 수분함량, 붉은곰팡이와 곰팡이독소의 오염 정도를 조사하였다. 창고의 온·습도 조사결과 상온창고는 월별 평균온도가 최 저 3oC에서 최고 29oC였으며, 평균습도는 58~70% 인 반면 저온창고는 평균온도가 3~13oC 였으며, 평균습도는 62~74% 였다. 시료의 수분함량은 상온창고에서 감소하였으나 저온창고에서는 초기수분함량 14% 시료는 수분함량이 증가하였고, 초기수분함량 20% 시료는 감소하였다. 붉은곰팡이 오염립은 상온창고에서 저장기간이 경과할수록 감소하였으나 저온창고에서는 시료의 수분함량이 높아감 소폭이 적었다. 곰팡이독소는 대부분의 시료에서 저장 12 개월 후에는 상온창고에서 니발레놀이 더 많이 검출되었 으나 1, 3, 6개월에는 뚜렷한 차이가 없었다. 따라서 겉보리의 건조가 덜 된 경우 저온창고에 보관하는 것이 붉은 곰팡이의 오염을 줄이는데 효과적이며, 겉보리를 1년이상 장기 저장할 경우 저온창고에 보관해야 니발레놀의 오염을 줄일 수 있다.

The objective of this study was to optimize analytical methods for ochratoxin A (OTA) and zearalenone (ZEA) in rice straw silage and winter forage crops using ultra-high performance liquid chromatography (UHPLC). Samples free of mycotoxins were spiked with 50 μg/kg, 250 μg/kg, or 500 μg/kg of OTA and 300 μg/kg, 1500 μg/kg, or 3000 μg/kg of ZEA. OTA and ZEA were extracted by acetonitrile and cleaned-up using an immunoaffinity column. They were then subjected to analysis with UHPLC equipped with a fluorescence detector. The correlation coefficients of calibration curves showed high linearity (R2 ≧ 0.9999 for OTA and R2≧0.9995 for ZEA). The limit of detection and quantification were 0.1 μg/kg and 0.3 μg/kg, respectively, for OTA and 5 μg/kg and 16.7 μg/kg, respectively, for ZEA. The recovery and relative standard deviation (RSD) of OTA were as follows: rice straw = 84.23~95.33%, 2.59~4.77%; Italian ryegrass = 79.02~95%, 0.86~5.83%; barley = 74.93~97%, 0.85~9.19%; rye = 77.99~ 96.67%, 0.33~6.26%. The recovery and RSD of ZEA were: rice straw = 109.6~114.22%, 0.67~7.15%; Italian ryegrass = 98.01~109.44%, 1.65~4.81%; barley = 98~113.53%, 0.25~5.85%; rye = 90.44~108.56%, 2.5~4.66%. They both satisfied the standards of European Commission criteria (EC 401-2006) for quantitative analysis. These results showed that the optimized methods could be used for mycotoxin analysis of forages.

아플라톡신은 옥수수 같은 작물에 Aspergillus flavus와 A. parasiticus 같은 곰팡이에서 생성되는 독소로서 발암 등 식품 안전성에 심각한 문제를 일으키는 물질이다. 이런 곰팡이의 감 염 환경에 적게 노출된 우리나라와 달리, 아프리카 동부에 위 치하고 있는 우간다의 경우, 고온 다습한 기후에 의해 곰팡이 오염이 심각하며, 이에 따른 아플라톡신의 생성 또한 많아, 대 표적인 식량 수출국으로서 심각한 피해를 보고 있다. 이를 저 감하기 위해, 곰팡이 저항성 품종 육성, 수확 후 관리 기술 개발 등의 신기술이 필요한 실정이다. 본 연구에서는 곰팡이 저항성 신품종 옥수수 육종의 일환으로 우간다 현지 재배 옥수수에 피해를 주는 곰팡이 균주 조사 및 아플라톡신 오염 현 황을 조사하고자 하였다. 우간다 각지의 옥수수 생산 농가와 곡물 상점을 대상으로 55종의 시료를 확보하고, 이들에 대한 아플라톡신 오염을 조사하였다. 그 결과 총 시료 가운데 약 11% 이상의 시료에서 아플라톡신의 오염이 확인되었고, 이들 의 함량 또한 12.7 - 123.5 μg/Kg으로 우간다 현지의 규제 기 준을 초과하는 시료의 경우도 9% 이상 발견되었다. 이 수치 는 곰팡이 감염에 대한 대책 수립의 필요성을 뒷받침하고, 아 플라톡신 등의 곰팡이 독소의 생성을 억제 할 신품종 및 기술 개발이 필요함을 의미한다.

Interleukin-12 (IL12) and IL23 are members of the IL12 family and secreted from antigen presenting cells (APCs) such as dendritic cells and macrophages. IL12 and IL23 are composed of two subunits of a sharing subunit, IL12B, and a unique subunit, IL12A for IL12 and IL23A for IL23. IL12 is involved in induction of T helper (Th) type 1 response, whereas IL23 is associated with the differentiation of naive T cells into Th17 cells. It has shown that IL12, a proinflammatory cytokine, is down-regulated during pregnancy for successful establishment and maintenance pregnancy and increases in plasma levels in women with preeclampsia. IL23 decreases IL12 expression to change the immune microenvironment at the maternal fetal interface in humans and mice. In the present study we determined the expression of IL12 and IL23 and their receptors in the endometrium and placenta during pregnancy. Real-time RT-PCR analysis showed levels of IL12A and IL12B mRNAs in the endometrium were high during early pregnancy, but maintained low during mid- to term pregnancy. During pregnancy, levels of IL12RB1 mRNA in the endometrium showed a biphasic pattern with the highest levels on Days 15 and 60 of pregnancy, while levels of IL12RB2 mRNA did not change. Levels of IL23A and IL23R mRNAs in the endometrium decreased toward term pregnancy. Immunohistochemical analysis showed that IL12A protein was localized specifically to scattered cells in endometrial stroma, but barely detected during mid- to term pregnancy. Conceptuses from early pregnancy expressed IL12, IL23, and their receptors, except IL12RB2, and corioallantoic tissues during mid- to late pregnancy expressed IL12, IL23, and their receptors, but not IL23R. These results showed that IL12 and IL23 and their receptors were expressed at the maternal-conceptus interface, suggesting that IL12 and IL23 may play a key role in regulating maternal immune environment for the establishment and maintenance of pregnancy in pigs.

S100 protein family is small calcium-binding proteins with two EF-hand motifs and comprises more than 20 proteins in human. Although S100A proteins are known to play important roles in proinflammatory responses including damage-associated molecular pattern (DAMP) signaling and in the establishment of pregnancy, the expression of S100As have not been determined in the uterine endometrium during the estrous cycle in pigs. Thus, this study was performed to investigate expression and localization of S100A8, S100A9, and S100A12 in the uterine endometrial tissues during the estrous cycle in pigs. Real-time RT-PCR analysis showed that S100A8, S100A9, and S100A12 mRNAs were expressed in the uterine endometrium during the estrous cycle with higher levels on days 15 and 18 of the estrous cycle than the other days of cycle. Immunohistochemistry analysis showed that S100A9 and S100A12 proteins were mainly localized to the immune cells in the uterine endometrium. Especially, S100A9- and S100A12-positive immune cells were detected in the uterine blood vessels on day 15 of the estrous cycle, and also localized to stroma near to luminal epithelium on days 0 and 18 of the estrous cycle. These results showed that S100As were expressed in the uterine endometrium during the estrous cycle in a cyclic stage-specific manner, and these proteins were localized to the immune cells in the endometrium. These suggest that immune cells expressing S100A proteins may be recruited into the endometrium during the estrous cycle and play an important role in regulating endometrial function in pigs.