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The quality of embryos produced by assisted reproductive techniques should be advanced by the improvement of in vitro culture conditions for successful implantation and pregnancy maintenance. We investigated the anti-oxidative effect of human adipose stem cell (ASC) conditioned medium with its optimal basal medium, Dulbecco′s modified Eagle′s medium (DMEM-CM), or keratinocyte serum-free medium (KSFM-CM) as supplements during in vitro culture (IVC) of in vitro fertilized mouse embryo. At first, preimplantation embryo development was evaluated in KSFM-CM and DMEM-CM supplemented cultures at various concentrations. The blastocyst (BL) and hatched BL formation rates were significantly increased in 5% DMEM-CM, while no difference was observed from KSFM-CM. Next, comparing the efficacy of KSFM-CM and DMEM-CM at the same concentration, DMEM-CM enhanced the developmental rate of 16 cells, morula, BL, and hatched BL. The expression level of reactive oxygen species decreased and that of glutathione increased in BL cultured with DMEM-CM, which confirms its anti-oxidative effect. Furthermore, apoptosis in BL cultured with DMEM-CM was reduced compared with that in KSFM-CM. This study demonstrated that the comparative effect of human ASC-CM made of two different basal media during mouse embryo IVC and anti-oxidative effect of 5% DMEM-CM was optimal to improve preimplantation embryo development.

Kihae Ra, Hyun Ju Oh, Eun Young Kim, Sung Keun Kang, Jeong Chan Ra, Eui Hyun Kim, Byeong Chun Lee. Animals, 10 (8), 1414, 2020.

In the mammalian testes, Sertoli cells are the only somatic cells in the seminiferous tubules that provide structural, nutritional and regulatory support for developing spermatogenic cells. Sertoli cells only proliferate during the fetal and neonatal periods and enter a quiescent state after puberty. Functional evidences suggest that the size of Sertoli cell population determines sperm production and fertility. However, factors that direct Sertoli cell proliferation and maturation are not fully understood. Transcription factor E4F1 is a multifunctional protein that serves essential roles in cell fate decisions and because it interacts with pRB, a master regulator of Sertoli cell function, we hypothesized that E4F1 may have a functional role in Sertoli cells. E4f1 mRNA was present in murine testis and immunohistochemical staining confirmed that E4F1 was enriched in mature Sertoli cells. We generated a conditional knockout mouse model using Amh-cre and E4f1flox/flox lines to study E4F1 fucntion in Sertoli cells and the results showed that E4f1 deletion caused a significant reduction in testis size and fertility. Further analyses revealed that meiosis progression and spermiogenesis were normal, however, Sertoli cell proliferation was impaired and germ cell apoptosis was elevated in the testis of E4f1 conditional knockout mice. On the basis of these findings, we concluded that E4F1 was expressed in murine Sertoli cells and served important functions in regulating Sertoli cell proliferation and fertility.

Rong-Ge Yan, Qi-Lin Yang, Qi-En Yang. Animals, 10 (9), 1691, 2020.

Eucommia ulmoides flavones (EUF) have been demonstrated to attenuate the inflammation and oxidative stress of piglets. This study aimed to test whether EUF could be used as an alternative antibiotic growth promoter to support growth performance and maintain intestinal health in weanling piglets. Weaned piglets (n = 480) were assigned into three groups and fed with a low-protein basal diet (NC), or supplementation with antibiotics (PC) or 0.01% EUF (EUF). Blood, intestinal contents, and intestine were collected on days 15 and 35 after weaning. The results showed the PC and EUF supplementations increased (p < 0.05) body weight on day 35, average daily gain and gain: feed ratio from day 15 to day 35 and day 0 to day 35, whereas decreased (p < 0.05) the diarrhea index of weanling piglets. EUF treatment increased (p < 0.05) jejunal villus height: crypt depth ratio, jejunal and ileal villus height, and population of ileal lactic acid bacteria on day 15 but decreased (p < 0.05) the population of ileal coliform bacteria on day 15 and day 35. These findings indicated the EUF, as the potential alternative to in-feed antibiotic growth promoter, could improve growth performance and intestinal morphology, and decrease colonization of coliform bacteria and diarrhea index in weanling piglets.

Daixiu Yuan, Jing Wang, Dingfu Xiao, Jiefeng Li, Yanhong Liu, Bie Tan, Yulong Yin. Animals, 10 (11), 1998, 2020.

Endoplasmic reticulum (ER) stress can be triggered during in vitro embryo production and is a major obstacle to embryo survival. MicroRNA (miR)-210 is associated with cellular adaptation to cellular stress and inflammation. An experiment was conducted to understand the effects of miR-210 on in vitro embryo development, ER stress, and apoptosis; to achieve this, miR-210 was microinjected into parthenogenetically activated embryos. Our results revealed that miR-210 inhibition significantly enhanced the cleavage rate, blastocyst formation rate, and total cell number (TCN) of blastocysts, and reduced expression levels of XBP1 (p < 0.05). miR-210 inhibition greatly reduced the expression of ER stress-related genes (uXBP1, sXBP1, ATF4, and PTPN1) and Caspase 3 and increased the levels of NANOG and SOX2 (p < 0.05). A miR-210-mimic significantly decreased the cleavage, blastocyst rate, TCN, and expression levels of XBP1 compared with other groups (p < 0.05). The miR-210-mimic impaired the expression levels of uXBP1, sXBP1, ATF4, PTPN1, and Caspase 3 and decreased the expression of NANOG and SOX2 (p < 0.05). In conclusion, miR-210 plays an essential role in porcine in vitro embryo development. Therefore, we suggest that miR-210 inhibition could alleviate ER stress and reduce apoptosis to support the enhancement of in vitro embryo production.

Muhammad Rosyid Ridlo, Eui Hyun Kim, Geon A. Kim. Animals, 11 (1), 221, 2021.

The main factor of embryonic demise is endoplasmic reticulum (ER) stress. Successful attenuation of ER stress results in an improvement in embryo development. We studied the impact of adiponectin in the in vitro culture (IVC) of porcine embryos derived from parthenogenetic activation and somatic cell nuclear transfer (SCNT). The first experiment revealed that 15 and 30 μg/mL adiponectin treatments improved cleavage, blastocyst rates, and total cell number (TCN) of parthenogenetic embryos and reduced the expression of XBP1 compared to the 5 μg/mL adiponectin treatment and control groups (p < 0.05). The second experiment showed that cleavage rate, blastocyst formation rate, and TCN of blastocysts were improved in the 15 μg/mL adiponectin treatment group compared with the control group, with significantly reduced XBP1 expression in ≥4-cell stage SCNT embryos and blastocysts (p < 0.05). Treatment with 15 μg/mL adiponectin significantly improved the expression of XBP1 and reduced the expression of ER stress-related genes (uXBP1, sXBP1, PTPN1, and ATF4), increased the expression levels of pluripotency-related genes (Nanog and SOX2), and decreased apoptosis-related gene expression (Caspase-3). These results suggest that 15 μg/mL adiponectin enhanced the in vitro developmental capacity of early-stage SCNT porcine embryos by reducing ER stress and apoptosis.

Muhammad Rosyid Ridlo, Eui Hyun Kim, Anukul Taweechaipaisankul, Byeong Chun Lee, Geon A. Kim. Animals, 11 (2), 473, 2021.

Dietary glutamic acid (GLU) is used as a feed additive because of its functional characteristics that may affect the growth performance and health of pigs. This study was carried out to determine the effects of dietary GLU on growth performance, nutrient digestibility, immune responses, and intestinal health of weaned pigs. A total of ninety-six weaned pigs (8.07 ± 1.17 kg of body weight; 28 days of age) were assigned to two dietary treatments (8 pigs/pen; 6 replicates/treatment) in a randomized complete block design (block: body weight): (1) a typical weaner diet (CON) and (2) CON supplemented with 0.5% GLU. The experimental period was for 4 weeks. All data and sample collections were performed at the specific time points during the experimental period. Pigs fed GLU had higher average daily gain and average daily feed intake for the first two weeks and nutrient digestibility than pigs fed CON. In addition, dietary GLU increased villus height to crypt depth ratio, number of goblet cells, and ileal gene expression of claudin family and occludin compared with CON, but decreased serum TNF-α and IL-6 and ileal gene expression of TNF-α. Moreover, pigs fed GLU had increased relative composition of bacterial communities of genus Prevotella and Anaerovibrio and decreased genus Clostridium and Terrisporobacter compared with those fed CON. This study suggests that dietary GLU influences growth performance and health of weaned pigs by modulating nutrient digestibility, intestinal morphology, ileal gene expression of tight junction proteins and cytokines, immune responses, and microbial community in the gut.

Hyunjin Kyoung, Jeong Jae Lee, Jin Ho Cho, Jeehwan Choe, Joowon Kang, Hanbae Lee, Yanhong Liu, Younghoon Kim, Hyeun Bum Kim, Minho Song. Animals, 11 (2), 504, 2021.

Effects of marine microalga Chlorella pyrenoidosa 55% ethanol extract (CPE55) on lipid metabolism, gut microbiota and regulation mechanism in high fat diet-fed induced hyperlipidaemia rats were investigated. Structure characterizations of major compounds in CPE55 were determined by ultra-performance liquid chromatography-quadrupole/time of flight mass spectrometry (UPLC-Q-TOF-MS/MS). The compositions of gut microbiota in rats were analyzed by high-throughput next-generation 16S rRNA gene sequencing. Oral administration with CPE55 markedly alleviated dyslipidemia through improving adverse blood lipid profile and inhibiting hepatic lipid accumulation and steatosis. CPE55 has downregulated the gene expression levels of acetyl CoA carboxylase, sterol regulatory element-binding transcription factor-1c, and 3-hydroxy-3-methyl glutaryl coenzyme A reductase and upregulated adenosine 5′-monophosphate-activated protein kinase-α. It has also improved the abundance of bacteria Alistipes, Prevotella, Alloprevotella, and Ruminococcus1 and decreased the abundances of Turicibacter and Lachnospira. Turicibacter and Lachnospira were both positive correlations of metabolic phenotypes. The findings above illustrated that CPE55 might be developed as food ingredients to ameliorate lipid metabolic disorders and hyperlipidaemia.

Xuzhi Wan, Tiantian Li, Dan Liu, Yihan Chen, Yuanyuan Liu, Bin Liu, Huiying Zhang, Chao Zhao. Marine Drugs, 16 (12), 498, 2018.

Human high-density lipoproteins (HDL) show a broad spectrum of antiviral activity in terms of anti-infection. Although many reports have pointed out a correlation between a lower serum HDL-C and a higher risk of COVID-19 infection and progression, the in vitro antiviral activity of HDL against SARS-CoV-2 has not been reported. HDL functionality, such as antioxidant and anti-infection, can be impaired by oxidation and glycation and a change to pro-inflammatory properties. This study compared the antiviral activity of native HDL with glycated HDL via fructosylation and native low-density lipoproteins (LDL). After 72 h of fructosylation, glycated HDL showed a typical multimerized protein pattern with an elevation of yellowish fluorescence. Glycated HDL showed a smaller particle size with an ambiguous shape and a loss of paraoxonase activity up to 51% compared to native HDL. The phagocytosis of acetylated LDL was accelerated 1.3-fold by glycated HDL than native HDL. Native HDL showed 1.7 times higher cell viability and 3.6 times higher cytopathic effect (CPE) inhibition activity against SARS-CoV-2 than that of glycated HDL under 60 μg/mL (approximately final 2.2 μM) in a Vero E6 cell. Native HDL showed EC50 = 52.1 ± 1.1 μg/mL (approximately final 1.8 μM) for the CPE and CC50 = 79.4 ± 1.5 μg/mL (around 2.8 μM). The selective index (SI) of native HDL was calculated to be 1.52. In conclusion, native HDL shows potent antiviral activity against SARS-CoV-2 without cytotoxicity, while the glycation of HDL impairs its antiviral activity. These results may explain why patients with diabetes mellitus or hypertension are more sensitive to a COVID-19 infection and have a higher risk of mortality.

Kyung-Hyun Cho, Jae-Ryong Kim, In-Chul Lee, Hyung-Jun Kwon. Antioxidants, 10 (2), 209, 2021.

Oxidative stress is a major cause of damage to the quantity and quality of embryos produced in vitro. Antioxidants are usually supplemented to protect embryos from the suboptimal in vitro culture (IVC) environment. Amniotic membrane-derived mesenchymal stem cells (AMSC) have emerged as a promising regenerative therapy, and their paracrine factors with anti-oxidative effects are present in AMSC conditioned medium (CM). We examined the anti-oxidative potential of human AMSC-CM treatment during IVC on mouse preimplantation embryo development and antioxidant gene expression in the forkhead box O (FoxO) pathway. AMSC-CM (10%) was optimal for overall preimplantation embryo developmental processes and upregulated the expression of FoxOs and their downstream antioxidants in blastocysts (BL). Subsequently, compared to adipose-derived mesenchymal stem cell (ASC)-CM, AMSC-CM enhanced antioxidant gene expression and intracellular GSH levels in the BL. Total antioxidant capacity and SOD activity were greater in AMSC-CM than in ASC-CM. Furthermore, SOD and catalase were more active in culture medium supplemented with AMSC-CM than in ASC-CM. Lastly, the anti-apoptotic effect of AMSC-CM was observed with the regulation of apoptosis-related genes and mitochondrial membrane potential in BL. In conclusion, the present study established AMSC-CM treatment at an optimal concentration as a novel antioxidant intervention for assisted reproduction.

Kihae Ra, Hyun Ju Oh, Eun Young Kim, Sung Keun Kang, Jeong Chan Ra, Eui Hyun Kim, Se Chang Park, Byeong Chun Lee. Antioxidants, 10 (2), 268, 2021.

Melatonin and phytanic acid (PA) are known to be involved in lipid metabolism and β-oxidation, in which peroxisomal activities also significantly participate. In addition, other studies have reported that the nuclear factor-erythroid-derived 2-like 2 (Nrf2 or NFE2L2) signaling pathway mediates lipid metabolism and its subsequent cascades. As these mechanisms are partially involved in porcine oocytes or embryonic development, we hypothesized that the factors governing these mechanisms could be interconnected. Therefore, we aimed to investigate possible crosstalk between peroxisomal activities and Nrf2 signaling in porcine embryos following melatonin and PA treatment. Porcine embryos were cultured for seven days after parthenogenetic activation, and subsequently treated with melatonin and PA, or injected with Pex19-targeted siRNAs. Real-time PCR, immunocytochemistry, and BODIPY staining were used to evaluate peroxisomal activities, Nrf2 signaling, and subsequent lipid metabolism. We found that melatonin/PA treatment enhanced embryonic development, whereas injection with Pex19-targeted siRNAs had the opposite effect. Moreover, melatonin/PA treatment upregulated peroxisomal activities, Nrf2 signaling, lipid metabolism, and mitochondrial membrane potentials, whereas most of these mechanisms were downregulated by Pex19-targeted siRNAs. Therefore, we suggest that there is a connection between the action of melatonin and PA and the Nrf2 signaling pathway and peroxisomal activities, which positively influences porcine embryonic development.

Eui-Hyun Kim, Muhammad-Rosyid Ridlo, Byeong-Chun Lee, Geon A. Kim. Antioxidants, 10 (5), 771, 2021.

The intensity and frequency of harmful algal blooms (HABs) have increased, posing a threat to human seafood resources due to massive kills of cultured fish and toxin contamination of bivalves. In recent years, bacteria that inhibit the growth of HAB species were found to be densely populated on the biofilms of some macroalgal species, indicating the possible biological control of HABs by the artificial introduction of macroalgal beds. In this study, an artificially created Ulva pertusa bed using mobile floating cages and a natural macroalgal bed were studied to elucidate the distribution of algal growth-limiting bacteria (GLB). The density of GLB affecting fish-killing raphidophyte Chattonella antiqua, and two harmful dinoflagellates, were detected between 106 and 107 CFU g−1 wet weight on the biofilm of artificially introduced U. pertusa and 10 to 102 CFU mL−1 from adjacent seawater; however, GLB found from natural macroalgal species targeted all tested HAB species (five species), ranging between 105 and 106 CFU g−1 wet weight in density. These findings provide new ecological insights of GLB at macroalgal beds, and concurrently demonstrate the possible biological control of HABs by artificially introduced Ulva beds.

Nobuharu Inaba, Isamu Kodama, Satoshi Nagai, Tomotaka Shiraishi, Kohei Matsuno, Atsushi Yamaguchi, Ichiro Imai. Applied Sciences, 10 (16), 5658, 2020.

Arthonia ulleungdoensis Lee & Hur is described as a new lichen species from South Korea. The new species is distinguishable from Arthonia ruana A. Massal. by its large, rounded and non-punctiform apothecia, taller apothecial section, asci with fewer spores, and larger and permanently colorless spores. Molecular analyses employing mitochondrial small subunit (mtSSU) and RNA polymerase subunit II (RPB2) sequences strongly support Arthonia ulleungdoensis as a distinct species in the genus Arthonia. Overall, 22 Arthonia species are currently recorded in South Korea. A surrogate key is provided to assist in the identification of all 10 taxa of Arthonia/Arthothelium with muriform spores in Northeast Asia.

Beeyoung Gun Lee, Jae-Seoun Hur. Microorganisms, 7 (7), 205, 2019.

Ralstonia solanacearum is a major phytopathogenic bacterium that attacks many crops and other plants around the world. In this study, a novel actinomycete, designated strain NEAU-SSA 1T, which exhibited antibacterial activity against Ralstonia solanacearum, was isolated from soil collected from Mount Song and characterized using a polyphasic approach. Morphological and chemotaxonomic characteristics of the strain coincided with those of the genus Streptomyces. The 16S rRNA gene sequence analysis showed that the isolate was most closely related to Streptomyces aureoverticillatus JCM 4347T (97.9%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain formed a cluster with Streptomyces vastus JCM4524T (97.4%), S. cinereus DSM43033T (97.2%), S. xiangluensis NEAU-LA29T (97.1%) and S. flaveus JCM3035T (97.1%). The cell wall contained LL-diaminopimelic acid and the whole-cell hydrolysates were ribose, mannose and galactose. The polar lipids were diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), hydroxy-phosphatidylethanolamine (OH-PE), phosphatidylinositol (PI), two phosphatidylinositol mannosides (PIMs) and an unidentified phospholipid (PL). The menaquinones were MK-9(H4), MK-9(H6), and MK-9(H8). The major fatty acids were iso-C17:0, C16:0 and C17:1 ω9c. The DNA G+C content was 69.9 mol %. However, multilocus sequence analysis (MLSA) based on five other house-keeping genes (atpD, gyrB, recA, rpoB, and trpB), DNA–DNA relatedness, and physiological and biochemical data showed that the strain could be distinguished from its closest relatives. Therefore, it is proposed that strain NEAU-SSA 1T should be classified as representatives of a novel species of the genus Streptomyces, for which the name Streptomyces sporangiiformans sp. nov. is proposed. The type strain is NEAU-SSA 1T (=CCTCC AA 2017028T = DSM 105692T).

Junwei Zhao, Liyuan Han, Mingying Yu, Peng Cao, Dongmei Li, Xiaowei Guo, Yongqiang Liu, Xiangjing Wang, Wensheng Xiang. Microorganisms, 7 (9), 360, 2019.