Comparison of Fucoidans from Saccharina latissima Regarding Age-Related Macular Degeneration Relevant Pathomechanisms in Retinal Pigment Epithelium.

Fucoidans from brown algae are described as anti-inflammatory, antioxidative, and antiangiogenic. We tested two Saccharina latissima fucoidans (SL-FRO and SL-NOR) regarding their potential biological effects against age-related macular degeneration (AMD). Primary porcine retinal pigment epithelium (RPE), human RPE cell line ARPE-19, and human uveal melanoma cell line OMM-1 were used. Cell survival was assessed in tetrazolium assay (MTT). Oxidative stress assays were induced with erastin or H2O2. Supernatants were harvested to assess secreted vascular endothelial growth factor A (VEGF-A) in ELISA. Barrier function was assessed by measurement of trans-epithelial electrical resistance (TEER). Protectin (CD59) and retinal pigment epithelium-specific 65 kDa protein (RPE65) were evaluated in western blot. Polymorphonuclear elastase and complement inhibition assays were performed. Phagocytosis of photoreceptor outer segments was tested in a fluorescence assay. Secretion and expression of proinflammatory cytokines were assessed with ELISA and real-time PCR. Fucoidans were chemically analyzed. Neither toxic nor antioxidative effects were detected in ARPE-19 or OMM-1. Interleukin 8 gene expression was slightly reduced by SL-NOR but induced by SL-FRO in RPE. VEGF secretion was reduced in ARPE-19 by SL-FRO and in RPE by both fucoidans. Polyinosinic:polycytidylic acid induced interleukin 6 and interleukin 8 secretion was reduced by both fucoidans in RPE. CD59 expression was positively influenced by fucoidans, and they exhibited a complement and elastase inhibitory effect in cell-free assay. RPE65 expression was reduced by SL-NOR in RPE. Barrier function of RPE was transiently reduced. Phagocytosis ability was slightly reduced by both fucoidans in primary RPE but not in ARPE-19. Fucoidans from Saccharina latissima, especially SL-FRO, are promising agents against AMD, as they reduce angiogenic cytokines and show anti-inflammatory and complement inhibiting properties; however, potential effects on gene expression and RPE functions need to be considered for further research.

Factors affecting utilization of sexual and reproductive health services among women with disabilities- a mixed-method cross-sectional study from Ilam district, Nepal.

BACKGROUND: Persons with disabilities can have physical, mental, intellectual, or sensory impairments which can hinder their social participation. Despite Sustainable Development Goals call for "universal access to sexual and reproductive health (SRH)", women with disabilities (WwDs) continue to experience barriers to access SRH services in Nepal. This study evaluated factors affecting the utilization of SRH services among WwDs in Ilam district, Nepal. METHODS: A mixed-method study with 384 WwDs of reproductive age was conducted in Ilam district, eastern Nepal. Quantitative data were collected using a structured questionnaire. Relationships between utilization of SRH services and associated factors were explored using multivariate logistic regression analysis. Qualitative data were collected from focus groups with female community health volunteers and interviews with WwDs, health workers and local political leaders. They were audio-recorded, translated and transcribed into English and were thematically analyzed. RESULTS: Among 384 respondents (31% physical; 7% vision,16% hearing, 7% voice&speech,12% mental/psychosocial, 9% intellectual, 18% multiple disabilities), only 15% of them had ever utilized any SRH services. No requirement (57%) and unaware of SRH services (24%) were the major reasons for not utilizing SRH services. A majority (81%) of them reported that the nearest health facility was not disability-inclusive (73%), specifically referring to the inaccessible road (48%). Multivariate analysis showed that being married (AOR = 121.7, 95% CI: 12.206-1214.338), having perceived need for SRH services (AOR = 5.5; 95% CI: 1.419-21.357) and perceived susceptibility to SRH related disease/condition (AOR = 6.0; 95% CI:1.978-18.370) were positively associated with the utilization of SRH services. Qualitative findings revealed that illiteracy, poor socioeconomic status, and lack of information hindered the utilization of SRH services. WwDs faced socioeconomic (lack of empowerment, lack of family support), structural (distant health facility, inaccessible-infrastructure), and attitudinal (stigmatization, bad behaviour of health care providers, perception that SRH is needed only for married) barriers to access SRH services. CONCLUSIONS: Utilization of SRH services among WwDs was very low in Ilam district, Nepal. The findings of this study warrant a need to promote awareness-raising programs to WwDs and their family members, sensitization programs to health service providers, and ensure the provision of disability-inclusive SRH services in all health facilities.

Evaluation of the Effects of Fucoidans from Fucus Species and Laminaria hyperborea against Oxidative Stress and Iron-Dependent Cell Death.

Fucoidans are algal polysaccharides that exhibit protective properties against oxidative stress. The aim of this study was to investigate different fucoidans from brown seaweeds for their ability to protect against iron-dependent oxidative stress (ferroptosis), a main hallmark of retinal and brain diseases, including hemorrhage. We investigated five new high-molecular weight fucoidan extracts from Fucus vesiculosus, F. serratus, and F. distichus subsp. evanescens, a previously published Laminaria hyperborean extract, and commercially available extracts from F. vesiculosus and Undaria pinnatifida. We induced oxidative stress by glutathione depletion (erastin) and H2O2 in four retinal and neuronal cell lines as well as primary cortical neurons. Only extracts from F. serratus, F. distichus subsp. evanescens, and Laminaria hyperborea were partially protective against erastin-induced cell death in ARPE-19 and OMM-1 cells, while none of the extracts showed beneficial effects in neuronal cells. Protective fucoidans also attenuated the decrease in protein levels of the antioxidant enzyme GPX4, a key regulator of ferroptosis. This comprehensive analysis demonstrates that the antioxidant abilities of fucoidans may be cell type-specific, besides depending on the algal species and extraction method. Future studies are needed to further characterize the health-benefiting effects of fucoidans and to determine the exact mechanism underlying their antioxidative abilities.

Influence of Fucoidan Extracts from Different Fucus Species on Adult Stem Cells and Molecular Mediators in In Vitro Models for Bone Formation and Vascularization.

Fucoidans, sulfated polysaccharides extracted from brown algae, are marine products with the potential to modulate bone formation and vascularization processes. The bioactivity and safety of fucoidans are highly associated with their chemical structure, which may vary with algae species and extraction method. Thus, in depth evaluation of fucoidan extracts in terms of endotoxin content, cytotoxicity, and their detailed molecular biological impact on the individual cell types in bone is needed. In this study, we characterized fucoidan extracts from three different Fucus species including Fucus vesiculosus (Fv), Fucus serratus (Fs), and Fucus distichus subsp. evanescens (Fe) for their chemical features, endotoxin content, cytotoxicity, and bioactive effects on human outgrowth endothelial cells (OEC) and human mesenchymal stem cells (MSC) as in vitro models for bone function and vascularization. Extracts contained mainly high molecular weight (HMW) fucoidans and were free of endotoxins that may cause inflammation or influence vascularization. OEC tolerated fucoidan concentrations up to 200 µg/mL, and no indication of cytotoxicity was observed. The inflammatory response, however, investigated by real-time PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) and endothelial barrier assessed by impedance measurement differed for the individual extracts. MSC in comparison with endothelial cells were more sensitive to fucoidans and showed partly reduced metabolic activity and proliferation at higher doses of fucoidans. Further results for MSC indicated impaired osteogenic functions in alkaline phosphatase and calcification assays. All tested extracts consistently lowered important molecular mediators involved in angiogenesis, such a VEGF (vascular endothelial growth factor), ANG-1 (angiopoietin 1), and ANG-2 (angiopoietin 2), as indicated by RT-PCR and ELISA. This was associated with antiangiogenic effects at the functional level using selected extracts in co-culture models to mimic bone vascularization processes during bone regeneration or osteosarcoma.

Evaluation of a Brown Seaweed Extract from Dictyosiphon foeniculaceus as a Potential Therapeutic Agent for the Treatment of Glioblastoma and Uveal Melanoma.

Ingredients of brown seaweed like fucoidans are often described for their beneficial biological effects, that might be interesting for a medical application. In this study, we tested an extract from Dictyosiphon foeniculaceus (DF) to evaluate the effects in glioblastoma and uveal melanoma, looking for a possible anti-cancer treatment. We investigated toxicity, VEGF (vascular endothelial growth factor) secretion and gene expression of tumor and non-tumor cells. SVGA (human fetal astrocytes), the human RPE (retinal pigment epithelium) cell line ARPE-19, the tumor cell line OMM-1 (human uveal melanoma), and two different human primary glioblastoma cultures (116-14 and 118-14) were used. Tests for cell viability were conducted with MTS-Assay (3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium), and the proliferation rate was determined with cell counting. VEGF secretion was assessed with ELISA (enzyme-linked immunosorbent assay). The gene expression of VEGF receptor 1 (VEGFR1), VEGF receptor 2 (VEGFR2) and VEGF-A was determined with real-time qPCR (quantitative polymerase chain reaction). DF lowered the cell viability of OMM-1. Proliferation rates of ARPE-19 and OMM-1 were decreased. The VEGF secretion was inhibited in ARPE-19 and OMM-1, whereas it was increased in SVGA and 116-14. The expression of VEGFR1 was absent and not influenced in OMM-1 and ARPE-19. VEGFR2 expression was lowered in 116-14 after 24 h, whereas VEGF-A was increased in 118-14 after 72 h. The extract lowered cell viability slightly and was anti-proliferative depending on the cell type investigated. VEGF was heterogeneously affected. The results in glioblastoma were not promising, but the anti-tumor properties in OMM-1 could make them interesting for further research concerning cancer diseases in the human eye.

Effects of Fucoidans from Five Different Brown Algae on Oxidative Stress and VEGF Interference in Ocular Cells.

BACKGROUND: Fucoidans are interesting for potential usage in ophthalmology, and especially age-related macular degeneration. However, fucoidans from different species may vary in their effects. Here, we compare fucoidans from five algal species in terms of oxidative stress protection and vascular endothelial growth factor (VEGF) interference in ocular cells. METHODS: Brown algae (Fucus vesiculosus, Fucus distichus subsp. evanescens, Fucus serratus, Laminaria digitata, Saccharina latissima) were harvested and fucoidans isolated by hot-water extraction. Fucoidans were tested in several concentrations (1, 10, 50, and 100 µg/mL). Effects were measured on a uveal melanoma cell line (OMM-1) (oxidative stress), retinal pigment epithelium (RPE) cell line ARPE19 (oxidative stress and VEGF), and primary RPE cells (VEGF). Oxidative stress was induced by H2O2 or tert-Butyl hydroperoxide (TBHP). Cell viability was investigated with methyl thiazolyl tetrazolium (MTT or MTS) assay, and VEGF secretion with ELISA. Affinity to VEGF was determined by a competitive binding assay. RESULTS: All fucoidans protected OMM-1 from oxidative stress. However, in ARPE19, only fucoidan from Saccharina latissima was protective. The affinity to VEGF of all fucoidans was stronger than that of heparin, and all reduced VEGF secretion in ARPE19. In primary RPE, only the fucoidan from Saccharina latissima was effective. CONCLUSION: Among the fucoidans from five different species, Saccharina latissima displayed the most promising results concerning oxidative stress protection and reduction of VEGF secretion.

Effects of Crude Fucus distichus Subspecies evanescens Fucoidan Extract on Retinal Pigment Epithelium Cells-Implications for Use in Age-Related Macular Degeneration.

Fucoidan extracts may have beneficial effects in age-related macular degeneration(AMD). Over-the-counter fucoidan preparations are generally undefined, crude extracts. In thisstudy, we investigated the effect of a crude fucoidan extract from Fucus distichus subspeciesevanescens (Fe) on the retinal pigment epithelium (RPE). Fe extract was investigated for chemicalcomposition and molar mass. It was tested in primary RPE and RPE cell line ARPE19. Oxidativestress was induced with tert-butyl hydroperoxide, cell viability evaluated with MTT assay, VEGFsecretion assessed in ELISA. Phagocytosis was evaluated in a fluorescence microscopic assay.Wound healing ability was tested in a scratch assay. Additionally, the inhibition of elastase andcomplement system by Fe extract was studied. The Fe extract contained about 61.9% fucose andhigh amounts of uronic acids (26.2%). The sulfate content was not as high as expected (6.9%). It wasnot toxic and not protective against oxidative stress. However, Fe extract was able to reduce VEGFsecretion in ARPE19. Phagocytosis was also reduced. Concerning wound healing, a delay could beobserved in higher concentrations. While some beneficial effects could be found, it seems tointerfere with RPE function, which may reduce its beneficial effects in AMD treatment.

In vivo and in vitro identification of Z-BOX C - a new bilirubin oxidation end product.

A new bilirubin oxidation end product (BOX) was isolated and characterized. The formation of the so-called Z-BOX C proceeds from bilirubin via propentdyopents as intermediates. This BOX was detected in pathological human bile samples using liquid chromatography/mass spectrometry and has potential relevance for liver dysfunction and cerebral vasospasms.

Binding modes of potential anti-prion phytochemicals to PrPC structures in silico.

BACKGROUND: Prion diseases involve the conversion of a normal, cell-surface glycoprotein (PrPC) into a misfolded pathogenic form (PrPSc). One possible strategy to inhibit PrPSc formation is to stabilize the native conformation of PrPC and interfere with the conversion of PrPC to PrPSc. Many compounds have been shown to inhibit the conversion process, however, no promising drugs have been identified to cure prion diseases. OBJECTIVE: This study aims to identify potential anti-prion compounds from plant phytochemicals by integrating traditional ethnobotanical knowledge with modern in silico drug design approaches. MATERIALS AND METHODS: In the current study medicinal phytochemicals were docked with swapped and non-swapped crystal structures of PrPCin silico to identify potential anti-prions to determine their binding modes and interactions. RESULTS: Eleven new phytochemicals were identified based on their binding energies and pharmacokinetic properties. The binding sites and interactions of the known and new anti-prion compounds are similar, and differences in binding modes occur in structures with very subtle differences in side chain conformations. Binding of these compounds poses steric hindrance to neighbouring molecules. Residues shown to be associated with the inhibition of PrPC to PrPSc conversion form interactions with most of the compounds. CONCLUSION: Identified compounds might act as potent inhibitors of PrPC to PrPSc conversion. These might be attractive candidates for the development of novel anti-prion therapy although further tests in vitro cell cultures and in vivo mouse models are needed to confirm these findings.

The Hidden Cell-to-Cell Trail of α-Synuclein Aggregates.

The progressive accumulation of insoluble aggregates of the presynaptic protein alpha-synuclein (α-Syn) is a hallmark of neurodegenerative disorders including Parkinson's disease (PD), Multiple System Atrophy, and Dementia with Lewy Bodies, commonly referred to as synucleinopathies. Despite considerable progress on the structural biology of these aggregates, the molecular mechanisms mediating their cell-to-cell transmission, propagation, and neurotoxicity remain only partially understood. Numerous studies have highlighted the stereotypical spatiotemporal spreading of pathological α-Syn aggregates across different tissues and anatomically connected brain regions over time. Experimental evidence from various cellular and animal models indicate that α-Syn transfer occurs in two defined steps: the release of pathogenic α-Syn species from infected cells, and their uptake via passive or active endocytic pathways. Once α-Syn aggregates have been internalized, little is known about what drives their toxicity or how they interact with the endogenous protein to promote its misfolding and subsequent aggregation. Similarly, unknown genetic factors modulate different cellular responses to the aggregation and accumulation of pathogenic α-Syn species. Here we discuss the current understanding of the molecular phenomena associated with the intercellular spreading of pathogenic α-Syn seeds and summarize the evidence supporting the transmission hypothesis. Understanding the molecular mechanisms involved in α-Syn aggregates transmission is essential to develop novel targeted therapeutics against PD and related synucleinopathies.

Focusing the GWAS Lens on days to flower using latent variable phenotypes derived from global multienvironment trials.

Adaptation constraints within crop species have resulted in limited genetic diversity in some breeding programs and areas where new crops have been introduced, for example, for lentil (Lens culinaris Medik.) in North America. An improved understanding of the underlying genetics involved in phenology-related traits is valuable knowledge to aid breeders in overcoming limitations associated with unadapted germplasm and expanding their genetic diversity by introducing new, exotic material. We used a large, 18 site-year, multienvironment dataset phenotyped for phenology-related traits across nine locations and over 3 yr along with accompanying latent variable phenotypes derived from a photothermal model and principal component analysis (PCA) of days from sowing to flower (DTF) data for a lentil diversity panel (324 accessions), which has also been genotyped with an exome capture array. Genome-wide association studies (GWAS) on DTF across multiple environments helped confirm associations with known flowering-time genes and identify new quantitative trait loci (QTL), which may contain previously unknown flowering time genes. Additionally, the use of latent variable phenotypes, which can incorporate environmental data such as temperature and photoperiod as both GWAS traits and as covariates, strengthened associations, revealed additional hidden associations, and alluded to potential roles of the associated QTL. Our approach can be replicated with other crop species, and the results from our GWAS serve as a resource for further exploration into the complex nature of phenology-related traits across the major growing environments for cultivated lentil.

Pseudomonas aeruginosa as a culprit of cervical necrotizing fasciitis: A case report.

INTRODUCTION AND IMPORTANCE: Necrotizing fasciitis is usually a polymicrobial infection and odontogenic source is usually the foci for infection in the neck region. Cervical necrotizing fasciitis due to Pseudomonas is a rare and potentially fatal complication in diabetic patients. The study highlights the importance of early intervention to improve the outcome of the patient. CASE PRESENTATION: We report a case of a 48-year female who presented with neck pain for 10 days. On further investigations, she had diabetic ketoacidosis, and a culture of the wound showed Pseudomonas. With appropriate antibiotics and surgical intervention, her condition gradually improved. CLINICAL DISCUSSION: Necrotizing fasciitis in the neck region with Pseudomonas without odontogenic infections is a rare occurrence. Early medical and surgical intervention leads to a better outcome. The location of the infection and its extensions can affect the prognosis. CONCLUSION: Physicians should be aware of cervical necrotizing fasciitis as a complication in diabetic ketoacidosis and install early treatment to improve survivability and the outcome.

Why do people purchase antibiotics over-the-counter? A qualitative study with patients, clinicians and dispensers in central, eastern and western Nepal.

INTRODUCTION: Over-the-counter (OTC) use of antibiotics contributes to the burgeoning rise in antimicrobial resistance (AMR). Drawing on qualitative research methods, this article explores the characteristics of OTC sales of antibiotic in Nepal, its drivers and implications for policy. METHODS: Data were collected in and around three tertiary hospitals in eastern, western and central Nepal. Using pre-defined guides, a mix of semi-structured interviews and focus group discussions were conducted with dispensers at drug stores, patients attending a hospital and clinicians. Interviews were audio-recorded, translated and transcribed into English and coded using a combination of an inductive and deductive approach. RESULTS: Drug shops were the primary location where patients engaged with health services. Interactions were brief and transactional: symptoms were described or explicit requests for specific medicine made, and money was exchanged. There were economic incentives for clients and drug stores: patients were able to save money by bypassing the formal healthcare services. Clinicians described antibiotics as easily available OTC at drug shops. Dispensing included the empirical use of broad-spectrum antibiotics, often combining multiple antibiotics, without laboratory diagnostic and drug susceptibility testing. Inappropriately short regimens (2-3 days) were also offered without follow-up. Respondents viewed OTC antibiotic as a convenient alternative to formal healthcare, the access to which was influenced by distance, time and money. Respondents also described the complexities of navigating various departments in hospitals and little confidence in the quality of formal healthcare. Clinicians and a few dispensers expressed concerns about AMR and referred to evadable policies around antibiotics use and poor enforcement of regulation. CONCLUSIONS: The findings point to the need for clear policy guidance and rigorous implementation of prescription-only antibiotics.

Size distribution and chain conformation of six different fucoidans using size-exclusion chromatography with multiple detection.

Fucoidans represent an intriguing class of fucose-containing sulfated polysaccharides. The biological activities of these polysaccharides are related to their compositional and structural parameters, whereby their degree of sulfation, as well as molecular weight (MW) distribution and chain conformation play important roles. Modern NMR and mass spectrometry techniques allow elucidating details of the glycan structure, but not the structure of the whole molecules in their native state. Accordingly, the knowledge about the latter of the fucoidans is currently still limited. Contrary to traditional MW determination by SEC with column calibration, SEC with triple detection provides not only the absolute Mw, but can also give information on additional molecule characteristics. In the present study, we used this method to compare six fucoidans extracted from Fucus vesiculosus (FV), F. serratus (FS), F. evanescens (FE), Dictyosiphon foeniculaceus (DF), Laminaria digitata (LD), and Saccharina latissima (SL) concerning their molar mass (Mw, Mn, Mp, dispersity) and size (rms radius, Rh) characteristics and distribution as well as their chain conformation in solution. The tested fucoidans displayed considerable structural diversity including large differences in their MW profiles and showed to be heterogeneously composed. Fuc-FV and Fuc-SL showed the broadest MW distributions, those from Fuc-FE and Fuc-DF the narrowest ones. Most of the fucoidan fractions (except for Fuc-DF) turned out to exist as expanded flexible chains in PBS solution. The conformation data suggest branched structures with partly long side chains. The knowledge obtained by this study is useful for further fractionation and structural characterization as well as the interpretation of the bioactivity differences between the various fucoidans.

Genomic selection for lentil breeding: Empirical evidence.

Genomic selection (GS) is a marker-based selection initially suggested for livestock breeding and is being encouraged for crop breeding. Several statistical models are used to implement GS; however, none have been tested for use in lentil (Lens culinaris Medik.) breeding. This study was conducted to compare the accuracy of different GS models and prediction scenarios based on empirical data and to make recommendations for designing genomic selection strategies for lentil breeding. We evaluated nine single-trait (ST) models, two multiple-trait (MT) models, and a model that incorporates genotype × environment interaction (GEI) using populations from a lentil diversity panel and two recombinant inbred lines (RILs). The lines in all populations were phenotyped for five phenological traits and genotyped using a custom exome capture assay. Within-population, across-population, and across-environment genomic predictions were made. Prediction accuracy varied among the evaluated models, populations, prediction scenarios, and traits. Single-trait models showed similar accuracy in the absence of large effect quantitative trait loci (QTL) but BayesB outperformed all models when there were QTL with relatively large effects. Models that accounted for GEI and MT-GS models increased prediction accuracy for a low heritability trait by up to 66 and 14%, respectively. Moderate to high accuracies were obtained for within-population (range of .36-.85) and across-environment (range of .19-.89) predictions but across-population prediction accuracy was very low. Results suggest that GS can be implemented in lentil breeding to make predictions within populations and across environments, but across-population prediction should not be considered when the population size is small.