Molecular Events in Response to Triclosan-Induced Oxidative Stress in CRISPR/Cas9-Mediated p53-Targeted Mutants in Daphnia magna.

Triclosan (TCS), a widely used antimicrobial agent, has been implicated in the oxidative stress induction and disruption of cellular processes in aquatic organisms. As TCS is ubiquitous in the aquatic environment, many previous studies have documented the effects of exposure to TCS on aquatic organisms. Nevertheless, most of the research has concentrated on the molecular and physiological responses of TCS, but there are still limited studies on the function of specific genes and the consequences of their absence. In this study, we focused on p53, a gene that is crucial for molecular responses such as autophagy and apoptosis as a result of TCS exposure. In order to ascertain the role and impact of the p53 gene in TCS-induced molecular responses, we examined the molecular responses to TCS-induced oxidative stress in wild-type (WT) and CRISPR/Cas9-mediated p53 mutant (MT) water fleas. The result has been accomplished by examining changes in molecular mechanisms, including in vivo end points, enzyme activities, adenosine triphosphate release rate, and apoptosis, to determine the role and impact of the p53 gene on TCS-induced molecular responses. The results indicated that the sensitivity of MT water fleas to TCS was greater than that of WT water fleas; however, the difference in sensitivity was significant at short exposures within 48 h and decreased toward 48 h. Accordingly, when we confirmed the oxidative stress after 24 h of exposure, the oxidative stress to TCS exposure was stronger in the MT group, with an imbalance of redox. To identify the mechanisms of tolerance to TCS in WT and MT Daphnia magna, we checked mitochondrial and ER-stress-related biomarkers and found an increase in apoptosis and greater sensitivity to TCS exposure in the MT group than in the WT. Our results suggest that the absence of p53 caused alterations in molecular processes in response to TCS exposure, resulting in increased sensitivity to TCS, and that p53 plays a critical role in response to TCS exposure.

Dendropanoxide Attenuates High Glucose-induced Oxidative Damage in NRK-52E Cells via AKT/mTOR Signaling Pathway.

Hyperglycemia is a potent risk factor for the development and progression of diabetes-induced nephropathy. Dendropanoxide (DPx) is a natural compound isolated from Dendropanax morbifera (Araliaceae) that exerts various biological effects. However, the role of DPx in hyperglycemia-induced renal tubular cell injury remains unclear. The present study explored the protective mechanism of DPx on high glucose (HG)-induced cytotoxicity in kidney tubular epithelial NRK-52E cells. The cells were cultured with normal glucose (5.6 mM), HG (30 mM), HG + metformin (10 µM), or HG + DPx (10 µM) for 48 h, and cell cycle and apoptosis were analyzed. Malondialdehyde (MDA), advanced glycation end products (AGEs), and reactive oxygen species (ROS) were measured. Protein-based nephrotoxicity biomarkers were measured in both the culture media and cell lysates. MDA and AGEs were significantly increased in NRK-52E cells cultured with HG, and these levels were markedly reduced by pretreatment with DPx or metformin. DPx significantly reduced the levels of kidney injury molecule-1 (KIM-1), pyruvate kinase M2 (PKM2), selenium-binding protein 1 (SBP1), or neutrophil gelatinase-associated lipocalin (NGAL) in NRK-52E cells cultured under HG conditions. Furthermore, treatment with DPx significantly increased antioxidant enzyme activity. DPx protects against HG-induced renal tubular cell damage, which may be mediated by its ability to inhibit oxidative stress through the protein kinase B/mammalian target of the rapamycin (AKT/mTOR) signaling pathway. These findings suggest that DPx can be used as a new drug for the treatment of high glucose-induced diabetic nephropathy.

Comparison of Speech Outcomes Between Speech Therapy Only and Double-Opposing Z-Plasty Combined With Speech Therapy in Patients With Submucous Cleft Palate.

The purpose of this study was to compare speech outcomes in patients with submucous cleft palate (SMCP) between speech therapy alone and double-opposing Z-plasty (DOZ) combined with speech therapy. The subjects were 67 patients with SMCP (overt type, 45 males, 22 females), who were divided into the observation group (n=18), the speech therapy group (n=24; duration, 17.8 mo), and the DOZ and speech therapy (DOZ-speech therapy) group (n=25; median age at DOZ, 5.3 years, duration, 18.6 mo). The median age at initial and final speech assessments were 3 and 5 years. After age, sex, syndromic status, duration of speech therapy, surgery timing, and speech outcomes were investigated, statistical analysis was performed. After tailored interventions, both isolated and non-isolated SMCP patients experienced significant improvements in speech outcomes, including nasal emission, hypernasality, compensatory articulation, and unintelligible speech. Since comparable improvements were observed, there were no significant differences in the final assessments regardless of initial speech issues between the speech therapy group and the DOZ-speech therapy group (all P>0.05). In the DOZ-speech therapy group, the rate of achieving "socially acceptable" speech was 92.3% in isolated cases and 90% in non-isolated cases. Multivariate analysis revealed that DOZ showed a tendency to reduce hypernasality, compensatory articulation, and "unintelligible" speech; syndromic or developmental conditions influenced outcomes in nasal emission and hypernasality; and initial hypernasality and compensatory articulation were correlated with outcomes. Therefore, DOZ surgery could be recommended to resolve hypernasality and compensatory articulation in SMCP patients before speech issues worsen.

Protective Effect of Castanopsis sieboldii Extract against UVB-Induced Photodamage in Keratinocytes.

Ultraviolet B (UVB) rays disrupt the skin by causing photodamage via processes such as reactive oxygen species (ROS) production, endoplasmic reticulum (ER) stress, DNA damage, and/or collagen degradation. Castanopsis sieboldii is an evergreen tree native to the southern Korean peninsula. Although it is known to have antioxidant and anti-inflammatory effects, its protective effect against photodamage in keratinocytes has not been investigated. Thus, in the present study, we investigated the effect of 70% ethanol extract of C. sieboldii leaf (CSL3) on UVB-mediated skin injuries and elucidated the underlying molecular mechanisms. CSL3 treatment restored the cell viability decreased by UVB irradiation. Moreover, CSL3 significantly inhibited UVB- or tert-butyl hydroperoxide-mediated ROS generation in HaCaT cells. ER stress was inhibited, whereas autophagy was upregulated by CSL3 treatment against UVB irradiation. Additionally, CSL3 increased collagen accumulation and cell migration, which were decreased by UVB exposure. Notably, epigallocatechin gallate, the major component of CSL3, improved the cell viability decreased by UVB irradiation through regulation of ER stress and autophagy. Conclusively, CSL3 may represent a promising therapeutic candidate for the treatment of UVB-induced skin damage.

JAK2 Inhibitor, Fedratinib, Inhibits P-gp Activity and Co-Treatment Induces Cytotoxicity in Antimitotic Drug-Treated P-gp Overexpressing Resistant KBV20C Cancer Cells.

P-glycoprotein (P-gp) overexpression is one of the major mechanisms of multidrug resistance (MDR). Previously, co-treatment with Janus kinase 2 (JAK2) inhibitors sensitized P-gp-overexpressing drug-resistant cancer cells. In this study, we assessed the cytotoxic effects of JAK2 inhibitor, fedratinib, on drug-resistant KBV20C cancer cells. We found that co-treatment with fedratinib at low doses induced cytotoxicity in KBV20C cells treated with vincristine (VIC). However, fedratinib-induced cytotoxicity was little effect on VIC-treated sensitive KB parent cells, suggesting that these effects are specific to resistant cancer cells. Fluorescence-activated cell sorting (FACS), Western blotting, and annexin V analyses were used to further investigate fedratinib's mechanism of action in VIC-treated KBV20C cells. We found that fedratinib reduced cell viability, increased G2 arrest, and upregulated apoptosis when used as a co-treatment with VIC. G2 phase arrest and apoptosis in VIC-fedratinib-co-treated cells resulted from the upregulation of p21 and the DNA damaging marker pH2AX. Compared with dimethyl sulfoxide (DMSO)-treated cells, fedratinib-treated KBV20C cells showed two-fold higher P-gp-inhibitory activity, indicating that VIC-fedratinib sensitization is dependent on the activity of fedratinib. Similar to VIC, fedratinib co-treatment with other antimitotic drugs (i.e., eribulin, vinorelbine, and vinblastine) showed increased cytotoxicity in KBV20C cells. Furthermore, VIC-fedratinib had similar cytotoxic effects to co-treatment with other JAK2 inhibitors (i.e., VIC-CEP-33779 or VIC-NVP-BSK805) at the same dose; similar cytotoxic mechanisms (i.e., early apoptosis) were observed between treatments, suggesting that co-treatment with JAK2 inhibitors is generally cytotoxic to P-gp-overexpressing resistant cancer cells. Given that fedratinib is FDA-approved, our findings support its application in the co-treatment of P-gp-overexpressing cancer patients showing MDR.

Serum glucose excretion after Roux-en-Y gastric bypass: a potential target for diabetes treatment.

OBJECTIVE: The mechanisms underlying type 2 diabetes resolution after Roux-en-Y gastric bypass (RYGB) are unclear. We suspected that glucose excretion may occur in the small bowel based on observations in humans. The aim of this study was to evaluate the mechanisms underlying serum glucose excretion in the small intestine and its contribution to glucose homeostasis after bariatric surgery. DESIGN: 2-Deoxy-2-[18F]-fluoro-D-glucose (FDG) was measured in RYGB-operated or sham-operated obese diabetic rats. Altered glucose metabolism was targeted and RNA sequencing was performed in areas of high or low FDG uptake in the ileum or common limb. Intestinal glucose metabolism and excretion were confirmed using 14C-glucose and FDG. Increased glucose metabolism was evaluated in IEC-18 cells and mouse intestinal organoids. Obese or ob/ob mice were treated with amphiregulin (AREG) to correlate intestinal glycolysis changes with changes in serum glucose homeostasis. RESULTS: The AREG/EGFR/mTOR/AKT/GLUT1 signal transduction pathway was activated in areas of increased glycolysis and intestinal glucose excretion in RYGB-operated rats. Intraluminal GLUT1 inhibitor administration offset improved glucose homeostasis in RYGB-operated rats. AREG-induced signal transduction pathway was confirmed using IEC-18 cells and mouse organoids, resulting in a greater capacity for glucose uptake via GLUT1 overexpression and sequestration in apical and basolateral membranes. Systemic and local AREG administration increased GLUT1 expression and small intestinal membrane translocation and prevented hyperglycaemic exacerbation. CONCLUSION: Bariatric surgery or AREG administration induces apical and basolateral membrane GLUT1 expression in the small intestinal enterocytes, resulting in increased serum glucose excretion in the gut lumen. Our findings suggest a novel, potentially targetable glucose homeostatic mechanism in the small intestine.

Gap Junction-Mediated Intercellular Communication of cAMP Prevents CDDP-Induced Ototoxicity via cAMP/PKA/CREB Pathway.

In the cochlea, non-sensory supporting cells are directly connected to adjacent supporting cells via gap junctions that allow the exchange of small molecules. We have previously shown that the pharmacological regulation of gap junctions alleviates cisplatin (CDDP)-induced ototoxicity in animal models. In this study, we aimed to identify specific small molecules that pass through gap junctions in the process of CDDP-induced auditory cell death and suggest new mechanisms to prevent hearing loss. We found that the cyclic adenosine monophosphate (cAMP) inducer forskolin (FSK) significantly attenuated CDDP-induced auditory cell death in vitro and ex vivo. The activation of cAMP/PKA/CREB signaling was observed in organ of Corti primary cells treated with FSK, especially in supporting cells. Co-treatment with gap junction enhancers such as all-trans retinoic acid (ATRA) and quinoline showed potentiating effects with FSK on cell survival via activation of cAMP/PKA/CREB. In vivo, the combination of FSK and ATRA was more effective for preventing ototoxicity compared to either single treatment. Our study provides the new insight that gap junction-mediated intercellular communication of cAMP may prevent CDDP-induced ototoxicity.

Multigenerational Mitigating Effects of Ocean Acidification on In Vivo Endpoints, Antioxidant Defense, DNA Damage Response, and Epigenetic Modification in an Asexual Monogonont Rotifer.

Ocean acidification (OA) is caused by changes in ocean carbon chemistry due to increased atmospheric pCO2 and is predicted to have deleterious effects on marine ecosystems. While the potential impacts of OA on many marine species have been studied, the multigenerational effects on asexual organisms remain unknown. We found that low seawater pH induced oxidative stress and DNA damage, decreasing growth rates, fecundity, and lifespans in the parental generation, whereas deleterious effects on in vivo endpoints in F1 and F2 offspring were less evident. The findings suggest that multigenerational adaptive effects play a role in antioxidant abilities and other defense mechanisms. OA-induced DNA damage, including double-strand breaks (DSBs), was fully repaired in F1 offspring of parents exposed to OA for 7 days, indicating that an adaptation mechanism may be the major driving force behind multigenerational adaptive effects. Analysis of epigenetic modification in response to OA involved examination of histone modification of DNA repair genes and a chromatin immunoprecipitation assay, as Bombus koreanus has no methylation pattern for CpG in its genome. We conclude that DSBs, DNA repair, and histone modification play important roles in multigenerational plasticity in response to OA in an asexual monogonont rotifer.

Disclosure at #SAGES2018: An analysis of physician-industry relationships of invited speakers at the 2018 SAGES national meeting.

BACKGROUND: Financial conflicts of interest (COI) have been shown to affect the interpretation of scientific findings. Publications with unreported COI tend to be more favorable to industry. Since 2014 industry payments to United States (US) physicians are publicly reported in the Open Payments Database (OPD). Several studies show high levels of unreported COI in medical literature; however, there is no research examining COI reporting at surgical conferences. We hypothesized that compliance with the COI disclosure requirement would be high at the 2018 SAGES meeting. However, we expected to find significant discrepancy between speaker-reported and OPD-reported COI. A secondary aim was to characterize the amount, source, and variation in industry payments to invited speakers. METHODS: We reviewed all available presentations from SAGES 2018 as recorded and publicly available on YouTube™ for the presence of COI disclosure and the disclosed industry relationships. For US physicians we searched the OPD and recorded all industry payments > $500. We compared the self-disclosed COI for each speaker with OPD records. Presentation topics were divided into ten groups to determine which topics received the most funding. RESULTS: Of the 526 invited presentations, 479 (91%) videos were available. Disclosures were reported by 414 presenters (86.4%). There were 420 unique presenters of which 315 were listed in the OPD. Speaker-reported disclosures were fully concordant with the OPD in 38.3% (121/315) of cases with 39% (123/315) under-reporting disclosures. Of presenters listed in OPD, the median payment was $992 ($0-$374,502) with a total of $6,389,097 paid in 2017. SAGES speakers failed to disclose $2,049,535 worth of industry payments with an average undisclosed payment of $16,662.88 (± $40,733.19). The largest financial contributor was Intuitive Surgical with $1,981,169 paid. Among topics, robotics and hernia received the most funding with $2,593,925 (40.6%) and $2,591,671 (40.5%) paid, respectively. CONCLUSIONS: Overall compliance with SAGES disclosure rules is high. There remains a discrepancy between speaker- and industry-reported disclosures, including a number of undisclosed payments, some of which are substantial. Adjustments to disclosure rules to include the relative amount of compensation may be warranted.

Characterization of gene expression profiles and functional analysis of peptidoglycan recognition protein 2 from rock bream (Oplegnathus fasciatus).

Peptidoglycan recognition protein 2 (PGRP2) is a Zn2+-dependent peptidase that plays important roles in binding to microbial components of the cell membrane, inducing phagocytosis and antimicrobial activity. Rock bream (Oplegnathus fasciatus) PGRP2 (RbPGRP2) was identified in the intestine by next generation sequencing (NGS) analysis. The open reading frame (ORF) the RbPGRP2 cDNA (470 amino acid residues) contains a peptidoglycan recognition protein domain (residues 300 to 446). Alignment analysis revealed that RbPGRP2 shares 37.6-53.5% overall sequence identity with the PGRP2s of other species. Phylogenetic analysis revealed that RbPGRP2 clustered together with PGRP2s from teleosts. In healthy rock bream, RbPGRP2 was found to be ubiquitously expressed in all of the examined tissues, especially in the liver. RbPGRP2 expression was significantly upregulated in all of the examined tissues of rock bream after infection with Edwardsiella piscicida, Streptococcus iniae and red sea bream iridovirus (RSIV) compared with the control. Purified rRbPGRP2 interactions with bacteria and inhibited the growth of bacteria in the presence of Zn2+. These results indicate that RbPGRP2 plays an important role in the innate immune response against bacterial infection.

Protective Role of Freshwater and Marine Rotifer Glutathione S-Transferase Sigma and Omega Isoforms Transformed into Heavy Metal-Exposed Escherichia coli.

Glutathione S-transferases (GSTs) play an important role in phase II of detoxification to protect cells in response to oxidative stress generated by exogenous toxicants. Despite their important role in defense, studies on invertebrate GSTs have mainly focused on identification and characterization. Here, we isolated omega and sigma classes of GSTs from the freshwater rotifer Brachionus calyciflorus and the marine rotifer Brachionus koreanus and explored their antioxidant function in response to metal-induced oxidative stress. The recombinant Bc- and Bk-GSTs were successfully transformed and expressed in Escherichia coli. Their antioxidant potential was characterized by measuring kinetic properties and enzymatic activity in response to pH, temperature, and chemical inhibitor. In addition, a disk diffusion assay, reactive oxygen species assay, and morphological analysis revealed that GST transformed into E. coli significantly protected cells from oxidative stress induced by H2O2 and metals (Hg, Cd, Cu, and Zn). Stronger antioxidant activity was exhibited by GST-S compared to GST-O in both rotifers, suggesting that GST-S plays a prominent function as an antioxidant defense mechanism in Brachionus spp. Overall, our study clearly shows the antioxidant role of Bk- and Bc-GSTs in E. coli and provides a greater understanding of GST class-specific and interspecific detoxification in rotifer Brachionus spp.

Molecular characterization, expression and functional analysis of peptidoglycan recognition protein-SC2 from rock bream, Oplegnathus fasciatus.

Peptidoglycan recognition proteins are members of the family of pattern recognition receptors (PRRs), that play important roles in the recognition of peptidoglycan and various biological processes. In this study, we have characterized peptidoglycan recognition protein-SC2 (PGRP-SC2) in rock bream (Oplegnathus fasciatus) (RbPGRP-SC2) and analysed its expression in various tissues after pathogen challenge. A sequence alignment revealed that the residues essential to zinc binding of the deduced protein were highly conserved among all the organisms. Phylogenetic analysis revealed that RbPGRP-SC2 is most closely related to the large yellow croaker PGRP-SC2. RbPGRP-SC2 was ubiquitously expressed in all tissues analysed, predominantly distributed in muscle and skin. After challenge with microbial pathogens (Edwardsiella piscicida), Streptococcus iniae or red seabream iridovirus [RSIV]), RbPGRP-SC2 was up-regulated in all the tissues examined, especially in liver. We produced recombinant RbPGRP-SC2 (rRbPGRP-SC2) using an Escherichia coli expression system. The rRbPGRP-SC2 had agglutination activity towards both Gram-negative (E. piscicida) and Gram-positive bacteria (S. iniae). In addition, rRbPGRP-SC2 induced leukocyte apoptosis and promoted leukocyte phagocytosis. These results suggest that the RbPGRP-SC2 plays an important role in the immune system and in maintaining cellular homeostasis of rock bream.

Olive flounder CD276 (B7-H3) a coinhibitory molecule for T cells: Responses during viral hemorrhagic septicemia virus (VHSV) stimulation.

Coinhibitory pathways in the B7-CD28 family provide critical inhibitory signals that regulate immune homeostasis, defense and protect tissue integrity. CD276 (B7-H3) is an important immune checkpoint member of this family, which is induced on antigen-presenting cells (APCs), and plays an important role in the inhibition of T-cell function. We have characterized the CD276 gene of olive flounder, Paralichthys olivaceus. OfCD276 has an ORF of 912 bp that codes for 303 amino acids with a predicted molecular mass of 33 kDa. It is a type I transmembrane protein with a single extracellular V- and C-like Ig domains, a transmembrane region, and a highly diverse cytoplasmic tail. This gene was distinctly expressed in gill, spleen, and skin, and sparsely expressed in other tissues. Pathogen stimulation by VHSV revealed that transcription of OfCD276 was induced on early hours in liver and expressed late in head kidney, spleen, intestine and gill tissues. Flow cytometry analysis of leukocytes revealed the percentage of granulocytes and lymphocytes that expressed OfCD276 molecules on their cell surface was 85.1% and 3.1%, respectively. Our study shows a significant role played by this coinhibitory molecule that participate in the regulation of the cell mediated immune response.

Nanoplastic Ingestion Enhances Toxicity of Persistent Organic Pollutants (POPs) in the Monogonont Rotifer Brachionus koreanus via Multixenobiotic Resistance (MXR) Disruption.

Among the various materials found inside microplastic pollution, nanosized microplastics are of particular concern due to difficulties in quantification and detection; moreover, they are predicted to be abundant in aquatic environments with stronger toxicity than microsized microplastics. Here, we demonstrated a stronger accumulation of nanosized microbeads in the marine rotifer Brachionus koreanus compared to microsized ones, which was associated with oxidative stress-induced damages on lipid membranes. In addition, multixenobiotic resistance conferred by P-glycoproteins and multidrug resistance proteins, as a first line of membrane defense, was inhibited by nanoplastic pre-exposure, leading to enhanced toxicity of 2,2',4,4'-tetrabromodiphenyl ether and triclosan in B. koreanus. Our study provides a molecular mechanistic insight into the toxicity of nanosized microplastics toward aquatic invertebrates and further implies the significance of synergetic effects of microplastics with other environmental persistent organic pollutants.

Reproducibility of an intraoral scanner: A comparison between in-vivo and ex-vivo scans.

INTRODUCTION: The purpose of this study was to assess the reproducibility of in-vivo and ex-vivo scans using an intraoral scanner. METHODS: Twenty adults with no missing teeth except for third molars were included in the study. Alginate impressions were taken, and plaster models were made from the impressions. Each subject underwent full-arch intraoral scanning twice with a TRIOS scanner (3Shape, Copenhagen, Denmark) at an interval of 2 weeks, and, the plaster models were scanned at the same interval with the same scanner. The first images of each scan were superimposed on the second scanned images using surface-based registration. In each case, the differences between the 2 scanned images were evaluated with color mapping. The reproducibility between the in-vivo and ex-vivo scans was compared using independent t tests and Bland-Altman analysis. RESULTS: The discrepancies between the first and second images were greater in the posterior than in the anterior regions for both the in-vivo and ex-vivo scans. Average surface differences between the first and second images were greater for the in-vivo scans (0.04 mm) than for the ex-vivo scans (0.02 mm). The Bland-Altman plots showed that the reproducibility of both scans was within the limits of agreement. CONCLUSIONS: The reproducibility of in-vivo scanning was comparable with ex-vivo scanning, although it showed a slight difference (0.02 mm) compared with ex-vivo scanning.

Genetic, management, and nutritional factors affecting intramuscular fat deposition in beef cattle - A review.

Intramuscular fat (IMF) content in skeletal muscle including the longissimus dorsi muscle (LM), also known as marbling fat, is one of the most important factors determining beef quality in several countries including Korea, Japan, Australia, and the United States. Genetics and breed, management, and nutrition affect IMF deposition. Japanese Black cattle breed has the highest IMF content in the world, and Korean cattle (also called Hanwoo) the second highest. Here, we review results of research on genetic factors (breed and sex differences and heritability) that affect IMF deposition. Cattle management factors are also important for IMF deposition. Castration of bulls increases IMF deposition in most cattle breeds. The effects of several management factors, including weaning age, castration, slaughter weight and age, and environmental conditions on IMF deposition are also reviewed. Nutritional factors, including fat metabolism, digestion and absorption of feed, glucose/starch availability, and vitamin A, D, and C levels are important for IMF deposition. Manipulating IMF deposition through developmental programming via metabolic imprinting is a recently proposed nutritional method to change potential IMF deposition during the fetal and neonatal periods in rodents and domestic animals. Application of fetal nutritional programming to increase IMF deposition of progeny in later life is reviewed. The coordination of several factors affects IMF deposition. Thus, a combination of several strategies may be needed to manipulate IMF deposition, depending on the consumer's beef preference. In particular, stage-specific feeding programs with concentrate-based diets developed by Japan and Korea are described in this article.

MCM7 polymorphisms associated with the AML relapse and overall survival.

The minichromosome maintenance complex component 7 (MCM7) encodes a member of MCM complex, which plays a critical role in the initiation of gene replication. Due to the importance of MCM complex, MCM7 gene has been regarded as a candidate gene for cancer development. In the present study, seven MCM7 polymorphisms were genotyped in 344 subjects composed of 103 acute myeloid leukemia (AML) patients and 241 normal controls to examine the possible associations between MCM7 polymorphisms and the risk of AML. MCM7 polymorphisms were not associated with the risk of AML (P > 0.05). However, MCM7 polymorphisms were significantly related to the relapse of AML and overall survival. The rs2070215 (N144S) showed a protective effect to the risk of AML relapse (OR = 0.37; P corr = 0.02). In haplotype analyses, the ht1 and ht2 showed significant associations with the risk of AML relapse (P corr = 0.02-0.03). In addition, rs1534309 showed an association with the overall survival of AML patients. Patients with major homozygote genotype (CC) of rs1534309 showed a higher survival rate than the patients with other genotypes (CG and GG). The results of the present study indicate that MCM7 polymorphisms may be able to predict the prognosis of AML patients.

Protective responses of two paralogs of granulocyte colony stimulating factor (GCSF) in rock bream, Oplegnathus fasciatus during bacterial and viral infection.

Granulocyte colony stimulating factor (GCSF) has a key role in the production of neutrophilic granulocytes during normal hematopoietic development and release of neutrophils into the blood circulation. In this study we have identified and characterized two paralogs of GCSF (RbGCSF) in rock bream. Although RbGCSF-1 and RbGCSF-2 share low sequence conservation, its domains and protein structure still share significant similarity. Basal levels of RbGCSF-1 gene expression was high in peripheral blood leukocytes (PBLs), spleen and intestine whereas the RbGCSF-2 was highly expressed in PBLs and kidney, of healthy animals. A significant induction of RbGCSFs were observed after the challenge with Streptococcus iniae in kidney, spleen and gills during initial hours of infection. Whereas Edwarsiella tarda infection caused a reasonable expression in kidney. Red seabream iridovirus caused induction of RbGCSF-1 transcription only in gills during initial hours. This higher expression of RbGCSF in early hours may be its response to induce emergency hematopoiesis, due to shortage of neutrophils to combat the surge in pathogens. The difference in induction of RbGCSF paralogs during infection may constitute to its different roles or overlapping functions.

The first report of CD2 associated protein gene, in a teleost (Rock bream, Oplegnathus fasciatus): An investigation of the immune response upon infection with several pathogens.

CD2 is expressed on the surfaces of virtually all T cells and natural killer (NK) cells. In mammals, the CD2 molecule is 50 kDa. The cytoplasmic tail of CD2 interacts with CD2-associated protein (CD2AP), which plays an important role in mediating the trigger signal in outer magnetic pole cells. In this study, we identified CD2AP from rock bream and investigated its gene expression. The ORF of CD2AP (1950 bp) encodes 650 amino acids (aa). CD2AP has a Src homology 3 (SH3) domain. Quantitative real-time PCR analysis revealed that CD2AP shows higher expression in the gills and skin. Under experimental challenge, CD2AP gene expression was increased as relative to the control after 7 days. This result will improve our understanding of blood vessels in teleost fish, and will provide a basis for the study of CD2-related genes.

Molecular characterization of a T cell co-stimulatory receptor, CD28 of rock bream (Oplegnathus fasciatus): Transcriptional expression during bacterial and viral stimulation.

CD28 is a co-stimulatory receptor that provides a critical second signal alongside T cell receptors for the activation of naive T cells. We characterized the CD28 gene of rock bream, which has a deduced amino acid sequence of 221 residues with an extracellular Ig-superfamily V domain, transmembrane region, and cytoplasmic tail. The conservation in domain structures and other motifs shows that it is highly likely that RbCD28 is a homologue of mammalian CD28 and may have related co-stimulatory functions. RbCD28 is constitutively expressed in most tissues that were analysed, with a relatively higher expression in teleost lymphoid organs, such as spleens, gills, trunk kidneys and skin. Unlike human CD28, RbCD28 is highly expressed in skin and gill-associated lymphoid organs. Although gills showed constitutive expression of RbCD28 in control animals, after a pathogen challenge, induction of CD28 was low, particularly in RSIV and E. tarda infection. Whereas induction of RbCD28 was observed in kidney during E. tarda and S. iniae infection, downregulation was observed during RSIV infection. In the case of the liver, E. tarda caused an initial upregulation of RbCD28. RbCD28 activation of T cells in the spleen was limited to S. iniae infection. Activation of RbCD28 observed in lymphoid organs during infection of various pathogens shows its key role as a co-stimulatory receptor of T cells.