The follicle loss of transplanted ovarian tissue (OT) is caused by ischemia and slow revascularization. To shorten the ischemic period and promote angiogenesis, some angiogenic factors have been treated for transplanted tissues. Angiopoietin-2 (ANG-2) is one of the major angiogenic factors and has been reported to promote blood vessels and increase vascular permeability in the ischemic and/or hypoxic environment. So, this study was designed to assess the impact of ANG-2 on follicle integrity and revascularization of mouse OT grafts. The 5-week-old B6D2F1 female mice were divided into 3 groups (a control and 2 ANG-2 groups) followed by ovary collection and vitrification. After warming, the ovaries were autotransplanted into kidney capsules with/without ANG-2 injection (50 or 500 ng/kg), and then killed at day(D)2, 7, 21 and 42 after transplantation. Total 2,437 follicles in OT grafts were assessed for the follicular density, integrity and classification by H&E staining. Apoptosis, revascularization, and serum FSH levels were evaluated by TUNEL assay, CD31 immunohistochemistry, and ELISA respectively. All the ANG-2 groups showed remarkable increase of morphologically intact follicle ratio across all the grafting duration except D21 (no statistical difference). The numbers of CD31(+) vessels (the sum of 3 fields at ×400 magnification) were significantly increased in both ANG-2 groups compared with the control group at all the grafting duration. Especially at D42, the 500ng ANG-2 group showed significantly more vessels than the 50 ng ANG-2 group as well as the control group. However the mean follicle numbers of grafts, apoptosis ratio and serum FSH levels showed no significant difference among the groups. In this study, remarkably well preserved follicles and larger amount of vessels were appeared in ANG-2 treated groups. So we thought that ANG-2 treatment is effective for OT transplantation and improve transplantation outcomes.

Polyethyleneglycol-adsorbed–superoxide dismutase (PEG-SOD), has been proposed as an effective agent for reducing free radical-mediated injury. The objective of this study was to investigate a protective effect of PEG-SOD supplementation on ovarian tissue during transplantation. Ovaries from F-1 mice were collected and vitrified. After warming, ovaries were autotransplanted under kidney capsule. Mice were randomly divided into four groups according to dose of PEG-SOD, (0 U/ml, 100 U/ml, 1,000 U/ml and 10,000 U/ml respectively). Grafted ovaries were retrieved 2, 7 and 21 days later. PEG-SOD was treated by intraperitoneal injection once every 48 hours and especially for 21 days group, after first week treatment, PEG-SOD was treated once every 4 days. Morphology of ovaries was assessed histological analysis and ELISA for FSH was performed to evaluate restoration of ovarian function. In 2 days groups, morphologically intact follicle ratio of 10,000 U/ml group was significantly higher than other groups. In 7 days groups, morphologically intact follicle ratio was significantly higher in all treatment groups. In 21 days groups, there was no significant difference of intact follicle ratio in total follicles in all groups but intact primordial, primary and secondary follicles ratio was higher in 10,000 U/ml group. FSH levels in blood serum were decrease as time goes on, but there is no statistical difference in each groups. In conclusion, the data of the present study show that PEG-SOD has a beneficial effect on preservation of the morphologically intact follicle.

Ovarian tissue cryopreservation and transplantation causes follicle depletion. To overcome this problem, we investigate the effect of Anti-Müllerian hormone (AMH), a follicle recruitment control hormone, supplementation before and/or after mouse ovarian transplantation. A total of 120 5-week-aged BD F-1 female mice were used. The mice were randomly divided into four groups according to AMH doses (0, 5, 25, 125 μg/mL, respectively). AMH was injected intraperitoneally on every other day for a week before, after, or before and after transplantation of ovaries under kidney capsules was performed. One week after transplantation, follicular normality was evaluated by histological analysis and TUNEL assay. In Group A and C, morphologically intact follicle (G1) ratios of AMH treated groups showed no statistically significant difference. In Group B, G1 ratios of 25 and 125 μg/mL of AMH treated groups were higher than those of 5 μg/mL treated group, but there was no improvement in G1 ratio after AMH treatment. In every group, apoptotic follicle ratios did not show any trend according to AMH treatment. Proportions of primordial follicle were not significantly different according to AMH treatment in all groups. The result of the present study demonstrated that AMH treatment during on transplantation of cryopreserved ovaries has no significant effect on follicle survival and prevention of follicle depletion.

Objective : To investigate the effects of Simvastatin and Methylprednisolone on ovarian tissue cryopreservation and transplantation using mouse models. Methods : The mice were randomly distributed into 1 control and 3 experimental groups. The B6D2F1 mice were given oral Simvastatin (5 mg/kg), intravenous Methylprednisolone (15 mg/kg), or a combination of both at 2 hours before ovariectomy. Same volume of normal saline was given perorally in the control group at 2 hours before ovariectomy. The ovarian tissues were vitrified accrording to our protocols. The vitrified ovaries were warmed 1 week later and auto-transplanted under bilateral kidney capsules. The ovaries and blood sera were collected at 2, 7 or 21 days after transplantation. Histological analysis, TUNEL assay, immuno-histochemistry for CD31, serum AMH level and embryonic development after in vitro fertilization were assessed for evaluation. Results : With regard to the total grade 1 follicle rate, both Simvastatin or Methylprednisolone treated groups were significantly increased at 2, 7 or 21 days after transplantation (except Simvastatin treated group at 7 days). A combination of Simvastatin and Methylprednisolone group was significantly improved in terms of the total G1 follicle rate, apoptotic follicle rate, CD31 positive area and serum AMH after ovarian tissue transplantation. However, there were no statistically difference with respect to the oocyte maturation rate, blastulation rate, and the other embryonic development parameters after in vitro fertilization procedure among the four groups. Conclusion : Our results suggest that combined donor Simvastatin and Methylprednisolone have beneficial effects on the quality and function of transplanted ovarian tissues.

Study question: What is the optimal vitrification protocol according to the cryoprotective agent (CPA) for ovarian tissue (OT) cryopreservation? Summary answer: The two-step protocol with 7.5% ethylene glycol (EG) and 7.5% dimethyl sulfoxide (DMSO) for 10 min then 20% EG, 20% DMSO and 0.5 M sucrose for 5 min showed the best results in mouse OT vitrification. What is known already: Establishing the optimal cryopreservation protocol is one of the most important steps to improve OT survival. However, only a few studies have compared vitrification protocols with different CPAs and investigated the effect of in vitro culture (IVC) on vitrified–.warmed OT survival. Some recent papers proposed that a combination of CPAs has less toxicity than one type of CPA. However, the efficacy of different types and concentrations of CPA are not yet well documented. Study design, size, duration: A total of 644 ovaries were collected from 4-week-old BDF1 mice, of which 571 ovaries were randomly assigned to 8 groups and vitrified using different protocols according to CPA composition and the remaining 73 ovaries were used as controls. After warming, each of the eight groups of ovaries was further randomly divided into four subgroups and in vitro cultured for 0, 0.5, 2 and 4 h, respectively. Ovaries of the best two groups among the eight groups were autotransplanted after IVC. Participants/materials, setting, methods: The CPA solutions for the eight groups were composed of EDS, ES, ED, EPS, EF, EFS, E and EP, respectively (E, EG; D, DMSO; P, propanediol; S, sucrose; F, Ficoll). The IVC medium was composed of a-minimal essential medium, 10% fetal bovine serum and 10 mIU/ml follicle-stimulating hormone (FSH). Autotransplantation of vitrified–.warmed OTs after IVC (0 to 4 h) using the EDS or ES protocol was performed, and the grafts were recovered after 3 weeks. Ovarian follicles were assessed for morphology, apoptosis, proliferation and FSH level. Main results and the role of chance: The percentages of the morphologically intact (G1) and apoptotic follicles in each group at 0, 0.5, 2 and 4 h of IVC were compared. For G1 follicles at 0 and 4 h of IVC, the EDS group showed the best results at 63.8 and 46.6%, respectively, whereas the EP group showed the worst results at 42.2 and 12.8%, respectively. The apoptotic follicle ratio was lowest in the EDS group at 0 h (8.1%) and 0.5 h (12.7%) of IVC. All of the eight groups showed significant decreases in G1 follicles and increases in apoptotic follicles as IVC duration progressed. After autotransplantation, the EDS 0 h group showed a significantly higher G1 percentage (84.9%) than did the other groups (42.4–.58.8%), while only the ES 4 h group showed a significant decrease in the number of proliferative cells (80.6%, 87.6–.92.9%). However, no significant differences in apoptotic rates and FSH levels were observed between the groups after autotransplantation. Limitations, reasons for caution: The limitation of this study was the absence of in vitro fertilization using oocytes obtained from OT grafts, which should be performed to confirm the outcomes of ovarian cryopreservation and transplantation. Wider implications of the findings: We compared eight vitrification protocols according to CPA composition and found the EDS protocol to be the optimal method among them. The data presented herein will help improve OT cryopreservation protocols for humans or other animals.

The PepMoV has been considered the most frequently detected potyvirus. When it co-infects with CMV or PMMoV, it gives severe impact to total pepper harvest in Korea. Since F1 hybrid that resistant to PepMoV has not been developed, we have developed transgenic peppers using Agrobacterium-mediated transformation with a Hc-Pro gene of the PepMoV. A large number of GM peppers were tested for resistance to the PepMoV, and after consequent self-crossing up to T4 generation, a highly tolerant pepper to PepMoV called T20 was selected. So far, BC4F1 lines have been selected by back-crossing with 4 elite lines through a breeding program. Very recently, based on molecular analysis, we have selected another event, #10-2, which is also resistant to PepMoV. Horticultural difference was investigated for both GM lines, #T20 and 10-2, and no significance was found comparing to non-GM lines.

Genome sequencing researches for considerable numbers of crops and wild plants are being developed. Cytogenetic researches according to chromosome number and size are essential to confirm and comprehend ploidy level and genome size before genome sequencing project is actually conducted. Cytogenetic researches on six food crop plants were carried out by DAPI staining and fluorescence in situ hybridization (FISH) method. Fagopyrum esculentum Moench showed 2n=2x=16, each chromosome length of 1.42㎛ to 1.77㎛, total chromosome length of 13.31㎛, and karyotypic formula of 2n=8m; Phaseolus angularis W.F. Wight, 2n=2x=22, 2.01㎛ to 3.84㎛, total 28.03㎛, 2n=9m+2sm, Perilla frutescens var. japonica Hara, 2n=2x=40, 1.73㎛ to 2.76㎛, total 44.36㎛, 2n=5m+13sm+2st. Chromosome sizes of the other three species such as, Panicum miliaceum L., 2n=2x=36, total chromosome length of 30.83㎛, Sesamum indicum L., 2n=2x=26, 27.39㎛, lpomoea batatas L., 2n=2x=30, total 33.51㎛ were too small for each chromosome type to be identified and analyzed. The result of FISH analysis using 5S and 45S rDNA probe showed species-specific chromosome locations in the genome. These preliminary analyses were carried out to decide which food crop to prioritize for genome sequencing. This work was supported by the “Cooperative Research Program for Agriculture Science & Technology Development (No.PJ009837), Rural Development Administration, Republic of Korea.

The fruit shape is an important character in tomato. OVATE is one of genes controlling fruit elongation in tomato. Two loci suppress the ovate mutation, sov1 and sov2, on chromosome 10 and chromosome 11 respectively. sov1 appears to control neck constriction in the fruits (Rodriguez et al, 2013). We sequenced the genomes of Gold Ball Livingston and Yellow Pear using the Illumina Hiseq 2000 generating 101 PE reads and developed molecular markers tightly linked to sov1. The locus was confirmed by fruit shape index analysis, marker genotyping and progeny testing of recombinants. We find mapped sov1 to a 145 kb interval corresponding to a region comprising two candidate genes. One of the candidate genes for sov1 is SlOFP20 another member of the Ovate Family Protein class. Although there is no difference expression of SlOFP20 in the parents at anthesis, when the gene is expressed very high, the mutation appears to be a 34 kb promoter deletion of SlOFP20 in Yellow Pear, conferring a pear shaped and neck-constricted fruit.

Mungbean (Vigna radiata) is a fast-growing, warm-season legume crop that is primarily cultivated in developing countries of Asia. We constructed a draft genome sequence of mungbean to facilitate genome research into the subgenus Ceratotropis and to enable a better understanding of the evolution of leguminous species. The draft genome sequence covers 80% of the estimated genome, of which 50.1% consists of repetitive sequences. In total, 22,427 high confidence protein-coding genes were predicted. Based on the de novo assembly of additional wild mungbean species, the divergence of what was eventually domesticated and the sampled wild mungbean species appears to have predated domestication. Moreover, the de novo assembly of a tetraploid Vigna species (Vigna reflexo-pilosa var. glabra) provided genomic evidence of a recent allopolyploid event. To further study speciation, we compared de novo RNA-seq assemblies of 22 accessions of 18 Vigna species and protein sets of Glycine max and Cajanus cajan. The species tree was constructed by a Bayesian Markov chain Monte Carlo method using highly confident orthologs shared by all 24 accessions. The present assembly of V. radiata var. radiata will facilitate genome research and accelerate molecular breeding of the subgenus Ceratotropis.

Neutropenic enterocolitis (NE), the most serious gastrointestinal complication, has been reported as a clinical syndrome that occurs in the setting of disease- or chemotherapy-induced neutropenia. Complications of NE include bowel necrosis with perforation, fistula, stenosis, massive bleeding, abscess formation, and pneumatosis intestinalis (PI). Most physicians recommend initial conservative management with bowel rest, intravenous fluids, total parenteral nutrition, broad-spectrum antibiotics, and normalization of neutrophil counts. Surgical intervention is recommended in the event of obstruction, perforation, persistent gastrointestinal bleeding despite correction of thrombocytopenia and coagulopathy, or clinical deterioration. We experienced a patient whose abdominal computed tomography scan showed pneumoretroperitoneum, intramural gas in the colon, and inferior vena caval gas. Her condition improved after treatment with granulocyte colony-stimulating factor and broad-spectrum antibiotics. We report on this case along with a review of the literature.

It has been known that adenomyoma is a rare non-neoplastic lesion of the biliary tract. In literature, adenomyoma was managed by extensive surgical procedures for its diagnosis and treatment because adenomyoma showed symptoms like obstructive jaundice and epigastric pain, which made the decision for the lesion difficult to distinguish from malignant neoplasm.We successfully treated a patient who had 12 mm sized adenomyoma of the major papilla with incomplete pancreas divisum by using an endoscopic papillectomy in the asymptomatic stage. The final diagnosis with an immunohistochemical stain revealed a lesion for adenomyoma of the major papilla.

Soybean mosaic virus (SMV), a member of Potyviridae family, is one of the most typical viral diseases and results in yield and quality loss of cultivated soybean. Due to the depletion of genetic resources for resistance breeding, a trial of genetic transformation to improve disease resistance has been performed by introducing SMV-CP and HC-Pro gene by RNA interference (RNAi) method via Agrobacterium-mediated transformation. Transgenic plants were infected with SMV strain G5 and investigated the viral response. As a result, two lines (3 and 4) of SMV-CP(RNAi) transgenic plants and three lines (2, 5 and 6) of HC-Pro(RNAi) transgenic plants showed viral resistance. In genomic Southern blot analysis, most of lines contained at least one T-DNA insertion in both SMV-CP(RNAi) and HC-Pro(RNAi) transgenic plants. Subsequent investigation confirmed that no viral CP and HC-Pro gene expression was detected in two SMV-resistant lines of SMV-CP(RNAi) and three lines of HC-Pro(RNAi) transgenic plants, respectively. On the other hand, non-transgenic plants and other lines showed viral RNA expression. Viral symptoms affected seed morphology, and clean seeds were harvested from SMV-resistant line of SMV-CP(RNAi) and HC-Pro(RNAi) transgenic plants. In addition, strong viral gene expression was detected from seeds of SMV-susceptible non-transgenic plants and SMV-susceptible transgenic lines. When compared the viral resistance between SMV-CP(RNAi) and HC-Pro(RNAi) transgenic plants, soybean transgenic plants with the HC-Pro gene using RNAi strategy showed much stronger and higher frequency of viral resistance.

Undomesticated soybeans (Glycine soja) are an important source of genetic variation for introducing useful traits to domesticated soybeans (Glycine max). Although Korea is known as the origin of the soybean, a little is known about genetic diversity and structure analysis of G.soja. The objectives of this study were to investigate the genetic diversity and the structure analysis of wild soybeans, and to construct a core collection of G. soja accessions in Korea. To evaluate the genetic diversity and structure analysis of G.soja, we analyzed allelic profiles at 21 SSR loci of 1028 accessions using POWERMARKER V3.25. These markers generated a total of 581 alleles over all loci. The number of alleles per locus ranged from 21 to 40, with a mean of 28 alleles per locus and a mean gene diversity of 0.886 in this accessions tested. Polymorphic information content value ranged from 0.737 to 0.946, with an average of 0.877. Using STRUCTURE V2.34, wild soybean originated from Korea was divided into two distinct populations, largely corresponding to two geographic regions. Population 1 consisted of eight sub-groups corresponds to mountains; population 2 to entire regions in Korea. Based on theses 21 SSR markers, a core collection development was performed by POWERCORE V1.0. A G. soja core collection consisted of 148 accessions which were established from 1028 accessions in Korea. Most accessions of the core collection were belonged to population 2 and only four were belonged to population 1. These results of this study would provide valuable information for future breeding programs using the G. soja core collection.

MicroRNAs (miRNAs) are a class of non-coding RNAs of approximately 21-nt which play important roles in regulating gene expression in plants. Although many miRNA studies have focused on a few model plants, the miRNAs and their target genes remain largely unknown in pepper, one of the most important crops cultivated worldwide. Here we employed high-throughput small RNA sequencing to extensively identify miRNAs in pepper from 10 different libraries, including leaf, stem, root, flower, and six developmental stage fruits. Based on bioinformatics pipeline, we successfully identified 29 and 35 families of conserved and novel miRNAs, respectively. We noticed that some miRNAs, whose targets were validated experimentally in this study, exhibited prominent changes in expression levels during fruit development stages. From the qRT-PCR analysis of the target mRNAs, including the SBP-transcription factor and F-box protein, we found that expression of these two target mRNAs gradually decreased in general during fruit development and was negatively correlated with the expression of their corresponding miRNAs. The validation of miRNA-directed cleavage of these target mRNAs, combined with the results of qRT-PCR analysis, likely suggests that some miRNAs in pepper may play a role in fruit development. Conclusively, our study first utilizes high-throughput sequencing to identify and characterize conserved and novel miRNAs and their targets in pepper, providing a basis for understanding the functional roles of miRNAs in pepper.

Recently, detection of esophageal diverticula by neck ultrasonography has been increasing. In particular, esophageal diverticula near the thyroid gland can be diagnosed as thyroid nodule on ultrasonography. To date, 13 cases of ultrasonographic features of Killian-Jamieson (K-J) diverticula have been reported. We report on four cases of K-J diverticula and discuss clinical characteristics, including ultrasonographic findings, in comparison with previously reported cases. Awareness of ultrasonographic finding of esophageal diverticula, a hypoechoic nodule containing echogenic foci found in the posterolateral aspect of the thyroid, is most important for making a differential diagnosis of K-J diverticulum from a thyroid nodule.

Acinetobacter baumannii is usually considered an opportunistic pathogen that it responsible for a variety of nosocomial infections, such as, pneumonia, tracheobronchitis, meningitis, endocarditis, peritonitis, skin and soft tissue infections, and urinary tract infections. However, over the years, the organism has developed substantial antimicrobial resistance, and thus, the management of infections has become more difficult. The less common infective endocarditis is one of the more serious consequences of nosocomial bacteremia. Here, we report the successful treatment of the first case of multidrug-resistant A. baumannii endocarditis in a 33-year-old patient.

Totally, 26 collections, 17 from Korea and 9 from China, were investigated for their sequences of 5S rDNA, especially the non-transcribed spacers (NTSs). Sequences of 5S rDNA were isolated by PCR using the primers, 5s-rRNA1 and 5s-rRNA2. Genomic DNA PCR produced single amplification of 300, 330, or 350 base pair fragments. Sequence analysis revealed that all inserts contained the part of 5S rDNA gene sequence and the full length of the NTS region. Three different sizes of the fragments were confirmed due to different size of NTS and their length were 300bp, 330bp and 350bp, respectively. Among 17 Korean foxtail millets tested, 14 collections showed single 300bp amplification. Longest fragment amplification, 350bp, was obtained only from the foxtail millet from China origin, even though 2 of them include 300bp fragment. CLUSTALW multiple alignments of 26 foxtail millets clearly revealed 4 areas with certain degree of sequence heterogeneity (region I, II, III, IV). Among 4 boxed areas, foxtail millet genotypes from China have distinct insertion especially in region III. Five of them have extra insertion of sequence and their additional sequences were either 45 or 48 base pair. Three Korean foxtail millets have 32 bp insertion. Other 8 Korean collections have short insert sequences (6 to 8 bp), 3 with 8 bp and 5 with 6 bp. In addition to insert, deletion sequences were also confirmed as major deletion was observed in region II of Chinese collection. The size of deletion was 7 bp long. According to phylogenic tree constructed using MEGA4 program, clear grouping was not revealed. To obtain more convincing results various collections from many countries should be obtained and analyzed to distinguish different germplasm from different origin.

Twenty two common millet (Panicum miliaceum L.) varieties collected from Korea, China and Russia were investigated for their phylogenetic relationship using 5S ribosomal DNA sequences with a hope to provide the basic information on their exact origin. Sequences of 5S rDNA were isolated by PCR. The primers, 5s-rRNA1 and 5s-rRNA2, were designed to isolate the complete NTS. Genomic DNA amplification produced two fragments with different length, 900 bp and 400 bp fragments, confirming the presence of two types of 5S rDNA repeats that differed from each other in the length of the NTS region. Amplified DNAs of 400 bp fragment were subcloned and used for further investigation. The obtained NTS sequences ranged from 200 to 300 bp and homology of sequences among plant materials was much higher than long repeat. CLUSTALW multiple aligment of 5S rDNA sequences from 22 different common millets revealed the clear difference by their origin. And critically different areas with insert or deletion were also confirmed. Those sequence difference seems to be used for discrimination of cultivars from different origin and use as molecular markers for origin identification. In phylogenic tree construction, the clear classification was shown where the genotypes from China and Russia is positioned together and stay away from domestic genotypes.

Herb extracts commercially used in Korea were screened for PPAR-γ agonist test and α-glucosidase inhibition assay. Total 16 herb plants had a PPAR-γ agonist activity. Specially, Alisma orientale Juz (108.41%), Ephedra sinica (98.22%), Sasa japonica Makino var. purpurascens Nakai (140.68%), Astragalus membranaceus Bunge (106.79%) and Cnidium officinale Makino (113.00%) showed high PPAR-γ agonist activity rate compared with rosiglitazone's (167.46%). And Cornus officinalis S. et Z. (90.3%), Cinnamomum cassia Blume (89.2%), Psoralea corylifolia L. (89.8%), Paeonia japonica (Makino) Miyabe (92.4%) and Paeonia suffruticosa Andr (93.2%), showed high α-glucosidase inhibition rates. These results support previous reports of the efficacy of Oriental medicinal plants used for diabetes mellitus.

Muscle strength and endurance activities of Korean ginseng (Panax ginseng C. A. Meyer; KG) were compared with those of wild simulated cultivation ginseng (WCG) in mice. Fifty male ICR mice were divided into five groups: A (vehicle); B (WCG 100 mg/kg); C (WCG 500 mg/kg); D (KG 100 mg/kg); E (KG 500 mg/kg). Subsequently, the mice were subjected to the forced swimming test (FST) and treadmill test at the 4th and 7th weeks. The glycogen content in the muscle and blood analysis (levels of glucose, triglyceride (TG), IGF-1) were also performed immediately after the last FST and treadmill test at the 7th week. Immobility times in FST were shorter in WCG- than KG-treated groups, and the results of the treadmill tests were also significant except for KG-treated at 100 mg/kg. The glycogen content was increased in both groups with a peak at 500 mg/kg of WCG groups. Serum concentrations of TG and glucose were decreased by administration of KG and WCG and all treated groups showed increase in the level of IGF-1 in serum. These results suggest that KG and WCG supplementations are effective in escalating the muscle strength and endurance.