A new compound, phomaherbarine A, induces cytolytic reactivation in epstein-barr virus-positive B cell lines.

Epstein-Barr virus (EBV), the first virus found to induce cancer in humans, has been frequently detected in various types of B cell lymphomas. During its latent phase, EBV expresses a limited set of proteins crucial for its persistence. Induction of the lytic phase of EBV has shown promise in the treatment of EBV-associated malignancies. The present study assessed the ability of phomaherbarine A, a novel compound derived from the endophytic fungus Phoma herbarum DBE-M1, to stimulate lytic replication of EBV in B95-8 cells. Phomaherbarine A was found to efficiently initiate the expression of both early and late EBV lytic genes in B95-8 cells, with this initiation being further heightened by the addition of phorbol myristate acetate and sodium butyrate. Moreover, phomaherbarine A demonstrated notable cytotoxicity against the EBV-associated B cell lymphoma cell lines B95-8 and Raji. Mechanistically, phomaherbarine A induces apoptosis in these cells through the activation of caspase-3/7. When combined with ganciclovir, phomaherbarine A does not interfere with the reduction of viral replication by ganciclovir and sustains its apoptosis induction. In conclusion, these findings indicate that phomaherbarine A may be a promising candidate for therapeutic intervention in patients with EBV-associated B cell lymphomas.

Identification of novel dimers and chemical profiling of acid amide alkaloids in Piper nigrum.

Peppers (Piper nigrum L.) are distinguished by their pungent flavor and aroma. Piperine is a major acid-amide alkaloid with a piperidine ring that gives pepper its flavor and scent. In plant metabolomics research, the accessibility of the chemical standards is critical for scientific credibility. We isolated and identified 10 novel dimers of acid amide alkaloids (9-15 and 20-22), along with 12 known monomers (1-6) and dimers (7, 8, 16-19) from black pepper. Subsequently, we found the distribution of monomers and dimers of acid amide alkaloids in black and white peppers by twenty-two acid amide alkaloids which we obtained using the molecular networking technique and multivariate analysis to reveal the molecular relationships between the acid amide alkaloids in black and white peppers. Our research delved into the chemical diversity of acid amide alkaloids in black and white peppers, which could help inform future culinary and potential medicinal utilization of pepper.

Horizontal ridge augmentation with porcine bone-derived grafting material: a long-term retrospective clinical study with more than 5 years of follow-up.

Objectives: The purpose of this study was to evaluate the outcomes of implants placed in horizontally augmented alveolar ridges using porcine bone grafts and to investigate the long-term stability of the porcine bone grafts. Materials and Methods: A retrospective analysis was conducted on 49 sites that underwent horizontal ridge augmentation using porcine bone grafts and implant placement with a follow-up period longer than 5 years. Furthermore, additional analysis was conducted on 24 sites where porcine bone grafts were used exclusively for horizontal ridge augmentation and implant placement. Results: The mean follow-up period after prosthesis loading was 67.5 months, with a mean marginal bone loss of 0.23 mm at 1 year and a cumulative mean marginal bone loss of 0.40 mm over the entire follow-up period. Of the 49 implants, 2 were lost and 3 did not meet the success criteria, resulting in a survival rate of 95.9% and a success rate of 89.8%. In 24 sites, the mean marginal bone loss was 0.23 mm at 1 year and 0.41 mm at 65.8 months, with 100% survival and success rates. Conclusion: Porcine bone grafts can be successfully used in horizontal ridge augmentation for implant placement in cases of ridges with insufficient horizontal width.

Targeted Isolation of N-Acetylcysteine-Containing Angucycline Derivatives from Streptomyces sp. MC16 and Their Antiproliferative Effects.

Liquid chromatography-mass spectrometry (LC-MS/MS)-based molecular networking analysis was applied to Streptomyces sp. MC16. The automatic classification of the MolNetEnhancer module revealed that its major constituent was an angucycline derivative. By targeted isolation of unique clusters in the molecular network, which showed different patterns from typical angucycline compounds, two new N-acetylcysteine-attached angucycline derivatives (1 and 2) were isolated. The structures were elucidated based on intensive NMR analysis and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS). All isolated compounds (1-4) were tested for their inhibitory effects on the proliferation of A431, A549, and HeLa cell lines. Antibiotics 100-1 (3) and vineomycinone B2 (4) showed moderate inhibitory effects on these three cell lines with IC50 values ranging from 18.5 to 59.0 µM, while compounds 1 and 2 with an additional N-acetylcysteine residue showed weak inhibitory effects only on the HeLa cell line with IC50 values of 54.7 and 65.2 µM, respectively.

Daldipyrenones A-C: Caged [6,6,6,6,6] Polyketides Derived from an Endolichenic Fungus Daldinia pyrenaica 047188.

Daldipyrenones A-C (1-3), three unprecedented caged xanthone [6,6,6,6,6] polyketides featuring a spiro-azaphilone unit, were discovered from an endolichenic fungus, Daldinia pyrenaica 047188. The structures of 1-3 were determined by using spectroscopic analysis and chemical derivatization. Daldipyrenones are likely derived by combining a chromane biosynthesis intermediate, 1-(2,6-dihydroxyphenyl)but-2-en-2-one, and a spiro-azaphilone, pestafolide A, via radical coupling or Michael addition to form a bicyclo[2.2.2]octane ring. Genome sequencing of the strain revealed two separate biosynthetic gene clusters responsible for forming two biosynthetic intermediates, suggesting a proposed biosynthetic pathway. Daldipyrenone A (1) exhibited significant antimelanogenic activity with lower EC50's than positive controls and moderate adiponectin-secretion promoting activity.

Discovery of Bioactive Metabolites by Acidic Stress to a Geldanamycin Producer, Streptomyces samsunensis.

In an effort to activate silent biosynthetic gene clusters, Streptomyces samsunensis DSM42010, a producer of geldanamycin, was cultured at four different pHs (4.5, 5.4, 6.6, and 7.4). An acidic culture condition (pH 5.4) was selected for a chemical investigation since S. samsunensis showed a different metabolic profile compared to when it was cultured under other conditions. Seven new (1-7) and four known (8-11) compounds were isolated from these cultures. The structures of the isolated compounds were determined by spectroscopic techniques and chemical derivatization. Relative and absolute configurations of the new compounds (1-5) were established using JBCA, PGME method, advanced Marfey's method, modified Mosher's method, and comparison of observed and calculated ECD data. Interestingly, compounds 1-3 were truncated versions of geldanamycin, and compound 4 was also deduced to originate from geldanamycin. Compound 5 was composed of 3-methyltyrosine and 6-hydroxy-2,4-hexadienoic acid connected through an amide bond. Compounds 6 and 7 were dihydrogenated forms of geldanamycin with a hydroxy substitution. It is possible that culturing this strain under acidic conditions interfered to some degree with the geldanamycin polyketide synthase, leading to production of truncated versions as well as analogues of geldanamycin. Compounds 1, 8, and 9 showed significant antivirulence activity, inhibiting production of α-toxin by methicillin-resistant Staphylococcus aureus without growth attenuation and global regulatory inhibition; compounds 1, 8, and 9 may become promising α-toxin-specific antivirulence leads with less risk of resistance development.

Anti-Obesity Potential through Regulation of Carbohydrate Uptake and Gene Expression in Intestinal Epithelial Cells by the Probiotic Lactiplantibacillus plantarum MGEL20154 from Fermented Food.

We investigated the probiotic characteristics and anti-obesity effect of Lactiplantibacillus plantarum MGEL20154, a strain that possesses excellent intestinal adhesion and viability. The in vitro properties, e.g., gastrointestinal (GI) resistance, adhesion, and enzyme activity, demonstrated that MGEL20154 is a potential probiotic candidate. Oral administration of MGEL20154 to diet-induced obese C57BL/6J mice for 8 weeks resulted in a feed efficacy decrease by 44.7% compared to that of the high-fat diet (HFD) group. The reduction rate of weight gain was about 48.5% in the HFD+MGEL20154 group compared to that of the HFD group after 8 weeks, and the epididymal fat pad was also reduced in size by 25.2%. In addition, the upregulation of the zo-1, pparα, and erk2, and downregulation of the nf-κb and glut2 genes in Caco-2 cells by MGEL20154 were observed. Therefore, we propose that the anti-obesity effect of the strain is exerted by inhibiting carbohydrate absorption and regulating gene expression in the intestine.

The Mediating Effects of Social Support on the Relationship between Uncertainty and Quality of Life among Patients with Chronic Low Back Pain: A Cross-Sectional Survey.

Background: This study aimed to investigate the mediating effects of social support on the relationship between uncertainty and quality of life (QOL) in patients with chronic low back pain (LBP). Methods: From 1 July 2019 to 25 March 2020, data were collected using a structured questionnaire from inpatients and outpatients > 20 years of age with chronic LBP lasting > 3 months. Inpatients included patients waiting for surgery and those recovering after surgery. The exclusion criteria were cancer and other serious pathological diseases. The relationships between uncertainty, social support, and QOL were analyzed using Pearson's correlation coefficients. Results: Uncertainty, the independent variable, exerted a significant effect on social support, the mediator (B = 0.33, p < 0.001). In addition, both uncertainty (B = 0.37, p < 0.001) and social support (B = 0.45, p < 0.001) exerted statistically significant effects on QOL, the dependent variable. Conclusions: Disease-related uncertainty can reduce QOL in patients with chronic LBP, and this relationship is mediated by the level of social support. To develop strategies for strengthening social support from healthcare providers, family, and friends, future studies should examine the experiences of patients with chronic LBP from various perspectives, including pain intensity and duration.

Performance and potential appraisal of various microalgae as direct combustion fuel.

Direct combustion of biomass is considered the most effective and simple means to contribute to CO2 reduction. In this context, the life-cycle potential of microalgal solid fuel, which has been overlooked so far, was comprehensively scrutinized ranging from cultivation to direct combustion. According to the quantitative data, using the raw fuel was confirmed to offer great benefits over the conventional lipid-targeted microalgal fuel systems through exploiting all of the biomass' energy potential, thereby being able to significantly increase the energy yield from biomass. The solid fuel is shown to exhibit diverse positive aspects owing to its remarkable calorific value, productivity and CO2 fixation ability. The combustion test reveals coal-microalgae co-combustion brings beneficial consequences on combustibility and environmental impacts with no notable thermal efficiency drop. This holistic appraisal shows microalgae patently possess high potential as a direct combustion fuel, even outperforming that of extensively used woody fuels.

Multilateral approach on enhancing economic viability of lipid production from microalgae: A review.

Microalgae have been rising as a feedstock for biofuel in response to the energy crisis. Due to a high lipid content, composed of fatty acids favorable for the biodiesel production, microalgae are still being investigated as an alternative to biodiesel. Environmental factors and process conditions can alternate the quality and the quantity of lipid produced by microalgae, which can be critical for the overall production of biodiesel. To maximize both the lipid content and the biomass productivity, it is necessary to start with robust algal strains and optimal physio-chemical properties of the culture environment in combination with a novel culture system. These accumulative approaches for cost reduction can take algal process one step closer in achieving the economic feasibility.

Discovery of candidate genes and pathways regulating oviduct development in chickens.

Estrogen is a key regulator in the development of the female reproductive system. It also stimulates oviduct development in immature chicks. We identified candidate genes and pathways associated with the development of chicken oviducts. A pellet containing the synthetic estrogen analog diethylstilbestrol (DES) was implanted subcutaneously in 1-wk-old female chicks for 10 days. The pellet was removed from half the group for 10 days, and an additional dose was given for a further 10 days. Total RNA was extracted from the oviducts of DES-treated and untreated chicks and subjected to an Affymetrix chicken GeneChip analysis. We found differential expression of 2290 and 1745 transcripts from the oviducts that were treated with DES once and twice, respectively. We also found a twofold or greater change in the expression of 77 and 390 transcripts between the two control and DES-treated time points, respectively, while we found a change in the expression of 10 transcripts that were common to all groups. Analyses of real-time PCR and in situ hybridization of selected genes confirmed the validity of the gene expression patterns observed in the microarray analysis. In particular, CCRN4L, FAM26F, HAS2, NELF, and NTM were up-regulated in the DES-treated chicken oviducts. High-throughput analysis revealed that the differentially expressed genes were related to tubular formation, epithelial differentiation, hormone interactions, nerve development, and tissue remodeling in the chicken oviduct. This study provides novel insights into candidate genes regulating oviduct development and differentiation via estrogen. The identified genes may serve as biomarkers of reproductive tract development in chicks.

The distribution of neuron-specific gene family member 1 in brain and germ cells: Implications for the regulation of germ-line development by brain.

Vesicular acidification at early endosomes dissociates endocytosed receptor-ligand complexes. The ligands, receptors, or both are then directed to late endosomes for degradation or recycled back to the plasma membrane. Of neuron-specific gene (NSG) family members, early endosomal protein neuron-specific gene family member 1 (NSG1) is the most important in receptor recycling. In this study, we characterized chicken NSG1 (cNSG1). We found several functional sites related to endocytotic machinery in cNSG1 that were highly conserved with most other vertebrate NSG1 proteins. We examined the tissue and duration specificity and the temporal and spatial patterns of cNSG1 expression. cNSG1 expression was preferentially located in all regions of the brain, neuroendocrine glands, and spinal cord. Unexpectedly, cNSG1 expression was strongly detected during male and female germ-line development. Expression of NSG1 in two apparently unrelated cell types such as neurons and germ cells suggests NSG1 roles in neurons and germ-cells chemotaxis and endocytotic machinery.

Reactivation of transgene expression by alleviating CpG methylation of the Rous sarcoma virus promoter in transgenic quail cells.

In this study, we investigated the relative expression of the Rous sarcoma virus (RSV) promoter-driven expression of enhanced green fluorescent protein (EGFP) in fibroblasts of transgenic quails. We analyzed the direct influence of CpG methylation of the RSV promoter on the transcriptional activity of delivered transgenes. Embryonic fibroblasts collected from homozygous transgenic quail (TQ2) were treated with 50 µM of DNA methyltransferase inhibitor followed by 5-aza-2'-deoxycytidine (5-azadC) for 48 h, and changes in expression were then analyzed by flow cytometry. The results show a significant increase of EGFP expression in TQ2 embryonic fibroblasts (QEFs) (2.64% to 79.84%). Subsequent methylation-specific amplification revealed that 5-azadC significantly reduced the CpG methylation status in the RSV promoters of the QEFs (86.42 to 48.41%); even after 5-azadC was withdrawn, CpG methylation remained decreased in expanded culture (16.28%). Further analysis showed that potential transcription factor binding sites existed in the CpG methylation site of the RSV promoter. These results may provide the basis for understanding the epigenetic mechanism responsible for transgenic animal production and genetic preservation.

The expression profile of apoptosis-related genes in the chicken as a human epithelial ovarian cancer model.

The purpose of our study was to examine the expression pattern of apoptosis-related genes in normal and cancerous ovaries of the hen. Localization of apoptosis-related gene mRNA was investigated in cancerous ovaries using in situ hybridization. The expression patterns of apoptosis-related genes were confirmed with RT-PCR in normal and cancerous ovaries. Differences of expression level between normal ovaries and ovarian cancers were analyzed using quantitative RT-PCR. In both normal and cancerous chicken ovaries, the expression of CASP1, CASP2, CASP3, CASP6, CASP8 and CASP9 were detected through RT-PCR analysis. The expression of BCL2, BCL2L1 and BID were confirmed in normal and cancerous ovaries of the hen. Quantitative RT-PCR showed that CASP1 expression was significantly increased in cancerous ovaries compared with normal ovaries, whereas BID expression was decreased. Our results showed a resistance to removal of abnormal cells via apoptosis in cancerous ovaries of the hen. Collectively, this phenomenon is closely associated with the dysregulation of CASP1 and BID expression in chicken ovarian cancer.

Matrix metalloproteinase 3 is a stromal marker for chicken ovarian cancer.

Matrix metalloproteinases (MMPs) are involved in the degradation of the extracellular matrix and basement membranes. Due to this, MMPs have been thought to promote invasion and metastasis of cancer cells and angiogenesis in tumors. Even though the chicken is a useful animal model for studying human ovarian cancer, no reports exist of the MMP expression pattern in chicken ovarian cancer. Therefore, we investigated the expression pattern of MMPs in chicken ovarian cancer. Results of RT-PCR and quantitative RT-PCR analyses showed MMP3 to be over-expressed in cancerous hen ovaries. In situ hybridization analysis of cancerous chicken ovaries showed that MMP3 mRNA was predominantly localized in the stroma, which is similar to MMP3 expression in human cancers. The results suggest that the expression pattern of MMP3 mRNA in chicken ovarian cancer is similar to that in various types of human cancer. Moreover, MMP3 potentially plays a significant role in developing ovarian cancer in chickens. The cell type-specific expression of MMP3 makes this gene a unique marker for ovarian cancer in chickens.

Molecular and biological aspects of early germ cell development in interspecies hybrids between chickens and pheasants.

Interspecific hybrids provide insights into fundamental genetic principles, and may prove useful for biotechnological applications and as tools for the conservation of endangered species. In the present study, interspecies hybrids were generated between the Korean ring-necked pheasant (Phasianus colchicus) and the White Leghorn chicken (Gallus gallus domesticus). We determined whether these hybrids were good recipients for the production of germline chimeric birds. PCR-based species-specific amplification and karyotype analyses showed that the hybrids inherited genetic material from both parents. Evaluation of biological function indicated that the growth rates of hybrids during the exponential phase (body weight/week) were similar to those of the pheasant but not the chicken, and that the incubation period for hatching was significantly different from that of both parents. Primordial germ cells (PGCs) of hybrids reacted with a pheasant PGC-specific antibody and circulated normally in blood vessels. The peak time of hybrid PGC migration was equivalent to that of the pheasant. In late embryonic stages, germ cells were detected by the QCR1 antibody on 15 d male gonads and were normally localized in the seminiferous cords. We examined the migration ability and developmental localization of exogenous PGCs transferred into the blood vessels of 63 h hybrid embryos. Donor-derived PGCs reacted with a donor-specific antibody were detected on 7 d gonads and the seminiferous tubules of hatchlings. Therefore, germ cell transfer into developing embryos of an interspecies hybrid can be efficiently used for the conservation of threatened animals and endangered species, and many biotechnological applications.

Basic fibroblast growth factor activates MEK/ERK cell signaling pathway and stimulates the proliferation of chicken primordial germ cells.

BACKGROUND: Long-term maintenance of avian primordial germ cells (PGCs) in vitro has tremendous potential because it can be used to deepen our understanding of the biology of PGCs. A transgenic bioreactor based on the unique migration of PGCs toward the recipients' sex cord via the bloodstream and thereby creating a germline chimeric bird has many potential applications. However, the growth factors and the signaling pathway essential for inducing proliferation of chicken PGCs are unknown. METHODOLOGY/PRINCIPAL FINDINGS: Therefore, we conducted this study to investigate the effects of various combinations of growth factors on the survival and proliferation of PGCs under feeder-free conditions. We observed proliferation of PGCs in media containing bFGF. Subsequent characterization confirmed that the cultured PGCs maintained expression of PGC-specific markers, telomerase activity, normal migrational activity, and germline transmission. We also found that bFGF activates the mitogen-activated protein kinase kinase/extracellular-signal regulated kinase (MEK/ERK) signaling. Also, the expression of 133 transcripts was reversibly altered by bFGF withdrawal. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that chicken PGCs can be maintained in vitro without any differentiation or dedifferentiation in feeder free culture conditions, and subsequent analysis revealed that bFGF is one of the key factors that enable proliferation of chicken PGCs via MEK/ERK signaling regulating downstream genes that may be important for PGC proliferation and survival.

Increased expression of cysteine cathepsins in ovarian tissue from chickens with ovarian cancer.

BACKGROUND: Cysteine cathepsins (CTSs) are involved in the degradation and remodeling of the extracellular matrix and are associated with cell transformation, differentiation, motility, and adhesion. These functions are also related to cancer cell invasion and metastasis. Chickens spontaneously develop epithelial ovarian cancer and are therefore a good animal model for human ovarian cancer. However, no studies have investigated the expression of CTSs in chickens with ovarian cancer. METHODS: Cancerous (n = 5) and normal (n = 3) ovaries were collected from 2-to 3-year-old hens, and ovarian tissue samples were collected for study. Ovarian cancers were evaluated with hematoxylin and eosin staining. Reverse transcriptase and quantitative PCR analyses, in situ hybridization analysis were performed to examine the mRNA expression pattern of three CTSs in detail, and protein expression of CTSB was evaluated. RESULTS: The CTSB, CTSC, and CTSS genes were highly expressed in cancerous chicken ovaries. Messenger RNAs for the three CTSs were localized to a nodule area, a major characteristic of cancerous ovaries, but the three CTSs showed no specific localization in normal ovaries. Immunoreactive CTSB protein was present in the nodule area of cancerous ovaries. CONCLUSION: Our results suggest that CTSB, CTSC, and CTSS have important functions in the development of epithelial ovarian cancer.

Molecular cloning and comparative analysis of immunoglobulin heavy chain genes from Phasianus colchicus, Meleagris gallopavo, and Coturnix japonica.

To date, immunoglobulin (Ig) genes have only been fully characterized in a small number of aves, which pose a major obstacle to understanding Ig evolution. Thus, we cloned the cDNAs of three immunoglobulin classes, IgA, IgM, and IgY, from Phasianus colchicus, Coturnix japonica, and Meleagris gallopavo. Multiple sequence alignments revealed that the highest degree of sequence homology in all Ig classes was observed between pheasant and turkey whereas the degree of homology between the galliforms and non-galliforms was relatively low compared to that among the galliforms. When the constant region domains of the four human Ig classes were compared with the corresponding regions in aves, the average percent homology between human CH2 and avian CH3, and between human CH3 and avian CH4, was greater than between identical domains in IgA and IgY, which are in partial agreement with the hypothesis that the avian CH2 domain evolved to form the mammalian hinge via domain condensation. Phylogenetic analysis showed that the galliform Ig heavy chain constant regions were divided into quail and the common ancestor of chicken, turkey, and pheasant, and that chicken was separated from turkey and pheasant, which were grouped together. These results add to our knowledge of galliform Igs and the diversification of these genes.

Production of biofunctional recombinant human interleukin 1 receptor antagonist (rhIL1RN) from transgenic quail egg white.

Oviduct-specific expression of heterologous recombinant proteins in transgenic birds is a promising technology for the large-scale production of therapeutic proteins in eggs. We describe the production of recombinant human interleukin 1 receptor antagonist (rhIL1RN) in the eggs of transgenic quails. To drive tissue-specific expression of rhIL1RN, a 1.35-kb fragment of the chicken ovalbumin promoter, which contains both the steroid-dependent regulatory element and the negative regulatory element, was used. A transgenic quail was generated by microinjection of a concentrated stock of lentivirus into stage X blastodermal cells. A single copy of the transgene was integrated into the seventh intron of the gene for conserved oligomeric golgi complex protein 5 (COG5) on chromosome 1. As expected, rhIL1RN expression was restricted to oviductal tissue, and the amount of protein deposited in the eggs of homozygous transgenic quails ranged from 88.7 to 233.8 ng/ml. Transgene expression was conserved from the G(1) generation to the G(4) generation, and there was no evidence of transgene silencing. In a bioassay using the EL4.NOB-1/CTLL-2 coculture system, no significant difference was observed between the egg-produced rhIL1RN and a commercially available rhIL1RN (anakinra).