Risk of cancer and serious disease in Danish patients with urgent referral for serious non-specific symptoms and signs of cancer in Funen 2014-2021.

BACKGROUND: In 2011, as the first European country, Denmark introduced the non-organ-specific cancer patient pathway (CPP) for patients presenting with non-specific symptoms and signs of cancer (NSSC). The proportion of patients with cancer over time is unknown. METHODS: A retrospective cohort study of all patients with a NSSC-CPP investigational course in the province of Funen to the Diagnostic Centre in Svendborg from 2014 to 2021 was performed to evaluate the proportion of patients with cancer and serious disease over time. RESULTS: A total of 6698 patients were referred to the NSSC-CPP of which 20.2% had cancer. While the crude referral rate increased from 114 per 100,000 people in 2014 and stabilised to around 214 in 2017-2021, the cancer detection rate of the total yearly new cancers in Funen diagnosed through the NSSC-CPP in DC Svendborg increased from 3 to 6%. CONCLUSIONS: With now high and stable conversion and crude referral rates, the NSSC-CPP is one of the largest CPPs in Denmark as measured by the number of new cancer cases found. Similar urgent referral programmes in other countries might fill an unmet medical need for patients presenting with serious non-specific symptoms and signs of cancer in general practice.

Structural characterization of solvent-based food preparation of jellyfish.

Jellyfish as a potential sustainable food material has recently gained increasing interest. However, with their soft gel-like texture and easy spoilage, it remains challenging to achieve desirable edible structures from jellyfish. The culinary preparation of jellyfish is a complex process and extends beyond conventional cooking methods. In this study, we investigate the transformation of jellyfish into crispy-like structures by manipulating their microstructural and mechanical properties through a solvent-based preparation. The study focuses on the use of "poor solvents", namely ethanol and acetone, and employs rheology measurements and quantitative microscopy techniques to analyze the effects of these solvents on the mechanical properties and microstructure of jellyfish. Our findings reveal that both ethanol and acetone lead to a significant increase in jellyfish hardness and deswelling. Notably, a micro-scale network is formed within the jellyfish matrix, and this network is then mechanically reinforced before a crispy-like texture can be obtained. Our study points to solvent polarity as also being a crucial factor for creating these effects and determines an upper polarity limit in the range of 12.2-12.9 MPa1/2 for added solvents, corresponding to approximately 60% of added ethanol or 70% of added acetone. Our study highlights that solvent-based preparation serves as a "reverse cooking" technique, where mechanical modification rather than traditional softening mechanisms are employed to stabilize and strengthen the microstructures and fibers of jellyfish. By elucidating the underlying mechanisms of solvent-induced stabilization, our findings may facilitate the development of innovative and sustainable culinary practices, paving the way for broader applications of jellyfish and other soft edible materials in the gastronomic landscape.

Cytotoxicity of Prymnesium parvum extracts and prymnesin analogs on epithelial fish gill cells RTgill-W1 and the human colon cell line HCEC-1CT.

Harmful algal blooms kill fish populations worldwide, as exemplified by the haptophyte microalga Prymnesium parvum. The suspected causative agents are prymnesins, categorized as A-, B-, and C-types based on backbone carbon atoms. Impacts of P. parvum extracts and purified prymnesins were tested on the epithelial rainbow trout fish gill cell line RTgill-W1 and on the human colon epithelial cells HCEC-1CT. Cytotoxic potencies ranked A > C > B-type with concentrations spanning from low (A- and C-type) to middle (B-type) nM ranges. Although RTgill-W1 cells were about twofold more sensitive than HCEC-1CT, the cytotoxicity of prymnesins is not limited to fish gills. Both cell lines responded rapidly to prymnesins; with EC50 values for B-types in RTgill-W1 cells of 110 ± 11 nM and 41.5 ± 0.6 nM after incubations times of 3 and 24 h. Results of fluorescence imaging and measured lytic effects suggest plasma membrane interactions. Postulating an osmotic imbalance as mechanisms of toxicity, incubations with prymnesins in media lacking either Cl-, Na+, or Ca2+ were performed. Cl- removal reduced morphometric rearrangements observed in RTgill-W1 and cytotoxicity in HCEC-1CT cells. Ca2+-free medium in RTgill-W1 cells exacerbated effects on the cell nuclei. Prymnesin composition of different P. parvum strains showed that analog composition within one type scarcely influenced the cytotoxic potential, while analog type potentially dictate potency. Overall, A-type prymnesins were the most potent ones in both cell lines followed by the C-types, and lastly B-types. Disturbance of Ca2+ and Cl- ionoregulation may be integral to prymnesin toxicity.

The Mixoplankton Database (MDB): Diversity of photo-phago-trophic plankton in form, function, and distribution across the global ocean.

Protist plankton are major members of open-water marine food webs. Traditionally divided between phototrophic phytoplankton and phagotrophic zooplankton, recent research shows many actually combine phototrophy and phagotrophy in the one cell; these protists are the "mixoplankton." Under the mixoplankton paradigm, "phytoplankton" are incapable of phagotrophy (diatoms being exemplars), while "zooplankton" are incapable of phototrophy. This revision restructures marine food webs, from regional to global levels. Here, we present the first comprehensive database of marine mixoplankton, bringing together extant knowledge of the identity, allometry, physiology, and trophic interactivity of these organisms. This mixoplankton database (MDB) will aid researchers that confront difficulties in characterizing life traits of protist plankton, and it will benefit modelers needing to better appreciate ecology of these organisms with their complex functional and allometric predator-prey interactions. The MDB also identifies knowledge gaps, including the need to better understand, for different mixoplankton functional types, sources of nutrition (use of nitrate, prey types, and nutritional states), and to obtain vital rates (e.g. growth, photosynthesis, ingestion, factors affecting photo' vs. phago' -trophy). It is now possible to revisit and re-classify protistan "phytoplankton" and "zooplankton" in extant databases of plankton life forms so as to clarify their roles in marine ecosystems.

Impacts of inorganic nutrients on the physiology of a mixoplanktonic ciliate and its cryptophyte prey.

Many marine planktonic ciliates retain functional chloroplasts from their photosynthetic prey and use them to incorporate inorganic carbon via photosynthesis. While this strategy provides the ciliates with carbon, little is known about their ability to incorporate major dissolved inorganic nutrients, such as nitrogen and phosphorus. Here, we studied how ciliates respond to different concentrations of dissolved inorganic nitrogen and phosphorus. Specifically, we tested the direct and indirect effects of nutrient availability on the ciliate Strombidium cf. basimorphum fed the cryptophyte prey Teleaulax amphioxeia. We assessed responses in the rates of growth, ingestion, photosynthesis, inorganic nutrient uptake, and excretion. Our results show that the prey changed its carbon content depending on the nutrient concentrations. Low inorganic nutrient concentrations increased S. cf. basimorphum growth and prey ingestion. The higher carbon content of the prey under these low nutrient conditions likely supported the growth of the ciliate, while the higher carbon:nutrient stoichiometry of the prey led to the higher ingestion rates. The low carbon content of the prey at high nutrient concentrations resulted in reduced growth of S. cf. basimorphum, which indicates that carbon acquired via photosynthesis in the ciliate cannot compensate for the ingestion of prey with low carbon content. In conclusion, our findings show S. cf. basimorphum is not able to utilize dissolved inorganic nitrogen and phosphorus for growth, and this species seems to be well adapted to exploit its prey when grown at low nutrient conditions.

Mannan-BAM, TLR ligands, and anti-CD40 immunotherapy in established murine pancreatic adenocarcinoma: understanding therapeutic potentials and limitations.

Pancreatic adenocarcinoma is one of the leading causes of cancer-related deaths, and its therapy remains a challenge. Our proposed therapeutic approach is based on the intratumoral injections of mannan-BAM, toll-like receptor ligands, and anti-CD40 antibody (thus termed MBTA therapy), and has shown promising results in the elimination of subcutaneous murine melanoma, pheochromocytoma, colon carcinoma, and smaller pancreatic adenocarcinoma (Panc02). Here, we tested the short- and long-term effects of MBTA therapy in established subcutaneous Panc02 tumors two times larger than in previous study and bilateral Panc02 models as well as the roles of CD4+ and CD8+ T lymphocytes in this therapy. The MBTA therapy resulted in eradication of 67% of Panc02 tumors with the development of long-term memory as evidenced by the rejection of Panc02 cells after subcutaneous and intracranial transplantations. The initial Panc02 tumor elimination is not dependent on the presence of CD4+ T lymphocytes, although these cells seem to be important in long-term survival and resistance against tumor retransplantation. The resistance was revealed to be antigen-specific due to its inability to reject B16-F10 melanoma cells. In the bilateral Panc02 model, MBTA therapy manifested a lower therapeutic response. Despite numerous combinations of MBTA therapy with other therapeutic approaches, our results show that only simultaneous application of MBTA therapy into both tumors has potential for the treatment of the bilateral Panc02 model.

Physiological Responses of Mesodinium major to Irradiance, Prey Concentration and Prey Starvation.

Ciliates within the Mesodinium rubrum/Mesodinium major species complex harbor chloroplasts and other cell organelles from specific cryptophyte species. Mesodinium major was recently described, and new studies indicate that blooms of M. major are just as common as blooms of M. rubrum. Despite this, the physiology of M. major has never been studied and compared to M. rubrum. In this study, growth, food uptake, chlorophyll a and photosynthesis were measured at six different irradiances, when fed the cryptophyte, Teleaulax amphioxeia. The results show that the light compensation point for growth of M. major was significantly higher than for M. rubrum. Inorganic carbon uptake via photosynthesis contributed by far most of total carbon uptake at most irradiances, similar to M. rubrum. Mesodinium major cells contain ~four times as many chloroplast as M. rubrum leading to up to ~four times higher rates of photosynthesis. The responses of M. major to prey starvation and refeeding were also studied. Mesodinium major was well adapted to prey starvation, and 51 d without prey did not lead to mortality. Mesodinium major quickly recovered from prey starvation when refed, due to high ingestion rates of > 150 prey/predator/d.

Retraction Note to: Bismuth adjuvant ameliorates adverse effects of high-dose chemotherapy in patients with multiple myeloma and malignant lymphoma undergoing autologous stem cell transplantation: a randomised, double-blind, prospective pilot study.

This article has been retracted. Please see the retraction notice for more detail: https://doi.org/10.1007/s00520-020-05770-w.

Karmitoxin: An Amine-Containing Polyhydroxy-Polyene Toxin from the Marine Dinoflagellate Karlodinium armiger.

Marine algae from the genus Karlodinium are known to be involved in fish-killing events worldwide. Here we report for the first time the chemistry and bioactivity of a natural product from the newly described mixotrophic dinoflagellate Karlodinium armiger. Our work describes the isolation and structural characterization of a new polyhydroxy-polyene named karmitoxin. The structure elucidation work was facilitated by use of 13C enrichment and high-field 2D NMR spectroscopy, where 1H-13C long-range correlations turned out to be very informative. Karmitoxin is structurally related to amphidinols and karlotoxins; however it differs by containing the longest carbon-carbon backbone discovered for this class of compounds, as well as a primary amino group. Karmitoxin showed potent nanomolar cytotoxic activity in an RTgill-W1 cell assay as well as rapid immobilization and eventual mortality of the copepod Acartia tonsa, a natural grazer of K. armiger.

Bismuth adjuvant ameliorates adverse effects of high-dose chemotherapy in patients with multiple myeloma and malignant lymphoma undergoing autologous stem cell transplantation: a randomised, double-blind, prospective pilot study.

PURPOSE: High-dose chemotherapy prior to autologous stem cell transplantation (ASCT) leads to adverse effects including mucositis, neutropenia and bacteremia. To reduce the toxicity, we treated myeloma and lymphoma patients with peroral bismuth as an adjuvant to chemotherapy to convey cytoprotection in non-malignant cells. METHODS: This trial was a prospective, randomised, double-blind, placebo-controlled pilot study of hematological inpatients (n = 50) receiving bismuth or placebo tablets, in order to identify any potential superiority of bismuth on toxicity from chemotherapy. RESULTS: We show for the first time that bismuth significantly reduces grade 2 stomatitis, febrile neutropenia and infections caused by melphalan in multiple myeloma, where adverse effects also were significantly linked to gender. In lymphoma patients, bismuth significantly reduces diarrhoea relative to placebo. Also, lymphoma patients' adverse effects were linked to gender. For the first time, bismuth is demonstrated as a safe strategy against chemotherapy's toxicity without interfering with intentional anti-cancer efficiency. Also, we show how gender significantly influences various adverse effects and response to treatment in both multiple myeloma and malignant lymphomas. CONCLUSION: These results may impact clinical prevention of chemotherapy's cytotoxicity in certain patient groups, and also, this study may direct further attention towards the impact of gender during the course and treatment outcome of malignant disorders.

HPLC-HRMS Quantification of the Ichthyotoxin Karmitoxin from Karlodinium armiger.

Being able to quantify ichthyotoxic metabolites from microalgae allows for the determination of ecologically-relevant concentrations that can be simulated in laboratory experiments, as well as to investigate bioaccumulation and degradation. Here, the ichthyotoxin karmitoxin, produced by Karlodinium armiger, was quantified in laboratory-grown cultures using high-performance liquid chromatography (HPLC) coupled to electrospray ionisation high-resolution time-of-flight mass spectrometry (HRMS). Prior to the quantification of karmitoxin, a standard of karmitoxin was purified from K. armiger cultures (80 L). The standard was quantified by fluorescent derivatisation using Waters AccQ-Fluor reagent and derivatised fumonisin B1 and fumonisin B2 as standards, as each contain a primary amine. Various sample preparation methods for whole culture samples were assessed, including six different solid phase extraction substrates. During analysis of culture samples, MS source conditions were monitored with chloramphenicol and valinomycin as external standards over prolonged injection sequences (>12 h) and karmitoxin concentrations were determined using the response factor of a closely eluting iturin A2 internal standard. Using this method the limit of quantification was 0.11 µg·mL-1, and the limit of detection was found to be 0.03 µg·mL-1. Matrix effects were determined with the use of K. armiger cultures grown with 13C-labelled bicarbonate as the primary carbon source.

Preparing giant unilamellar vesicles (GUVs) of complex lipid mixtures on demand: Mixing small unilamellar vesicles of compositionally heterogeneous mixtures.

Giant unilamellar vesicles (GUVs) are simple model membrane systems of cell-size, which are instrumental to study the function of more complex biological membranes involving heterogeneities in lipid composition, shape, mechanical properties, and chemical properties. We have devised a method that makes it possible to prepare a uniform sample of ternary GUVs of a prescribed composition and heterogeneity by mixing different populations of small unilamellar vesicles (SUVs). The validity of the protocol has been demonstrated by applying it to ternary lipid mixture of DOPC, DPPC, and cholesterol by mixing small unilamellar vesicles (SUVs) of two different populations and with different lipid compositions. The compositional homogeneity among GUVs resulting from SUV mixing is quantified by measuring the area fraction of the liquid ordered-liquid disordered phases in giant vesicles and is found to be comparable to that in GUVs of the prescribed composition produced from hydration of dried lipids mixed in organic solvent. Our method opens up the possibility to quickly increase and manipulate the complexity of GUV membranes in a controlled manner at physiological buffer and temperature conditions. The new protocol will permit quantitative biophysical studies of a whole new class of well-defined model membrane systems of a complexity that resembles biological membranes with rafts.

Dynamic Cluster Analysis: An Unbiased Method for Identifying A + 2 Element Containing Compounds in Liquid Chromatographic High-Resolution Time-of-Flight Mass Spectrometric Data.

Dynamic cluster analysis (DCA) is an automated, unbiased technique which can identify Cl, Br, S, and other A + 2 element containing metabolites in liquid chromatographic high-resolution mass spectrometric data. DCA is based on three features, primarily the previously unutilized A + 1 to A + 2 isotope cluster spacing which is a strong classifier in itself but improved with the addition of the monoisotopic mass, and the well-known A:A+2 intensity ratio. Utilizing only the A + 1 to A + 2 isotope cluster spacing and the monoisotopic mass it was possible to filter a chromatogram for metabolites which contain Cl, Br, and S. Screening simulated isotope patterns of the Antibase Natural Products Database it was determined that the A + 1 to A + 2 isotope cluster spacing can be used to correctly classify 97.4% of molecular formulas containing these elements, only misclassifying a few metabolites which were either over 2800 u or metabolites which contained other A + 2 elements, such as Cu, Ni, Mg, and Zn. It was determined that with an interisotopic mass accuracy of 1 ppm, in a fully automated process, using all three parameters, it is possible to specifically filter a chromatogram for S containing metabolites with monoisotopic masses less than 825 u. Furthermore, it was possible to specifically filter a chromatogram for Cl and Br containing metabolites with monoisotopic masses less than 1613 u. Here DCA is applied on (i) simulated isotope patterns of the Antibase natural products databases, (ii) LC-QTOF data of reference standards, and (iii) LC-QTOF data of crude extracts of 10 strains of laboratory grown cultures of the microalga Prymnesium parvum where it identified known metabolites of the prymnesin series as well as over 20 previously undescribed prymnesin-like molecular features.

Nordic MCL3 study: 90Y-ibritumomab-tiuxetan added to BEAM/C in non-CR patients before transplant in mantle cell lymphoma.

The main objective of the MCL3 study was to improve outcome for patients not in complete remission (CR) before transplant by adding (90)Y-ibritumomab-tiuxetan (Zevalin) to the high-dose regimen. One hundred sixty untreated, stage II-IV mantle cell lymphoma patients <66 years received rituximab (R)-maxi-CHOP (cyclophosphamide, hydroxydaunorubicin, vincristine, and prednisone) alternating with R-high-dose cytarabine (6 cycles total), followed by high-dose BEAM/C (bis-chloroethylnitrosourea, etoposide, cytarabine, and melphalan or cyclophosphamide) and autologous stem cell transplantation from 2005 to 2009. Zevalin (0.4 mCi/kg) was given to responders not in CR before transplant. Overall response rate pretransplant was 97%. The outcome did not differ from that of the historic control: the MCL2 trial with similar treatment except for Zevalin. Overall survival (OS), event-free survival (EFS), and progression-free survival (PFS) at 4 years were 78%, 62%, and 71%, respectively. For responding non-CR patients who received Zevalin, duration of response was shorter than for the CR group. Inferior PFS, EFS, and OS were predicted by positron emission tomography (PET) positivity pretransplant and detectable minimal residual disease (MRD) after transplant. In conclusion, positive PET and MRD were strong predictors of outcome. Intensification with Zevalin may be too late to improve the outcome of patients not in CR before transplant. This trial was registered at www.clinicaltrials.gov as #NCT00514475.

Chemical Diversity, Origin, and Analysis of Phycotoxins.

Microalgae, particularly those from the lineage Dinoflagellata, are very well-known for their ability to produce phycotoxins that may accumulate in the marine food chain and eventually cause poisoning in humans. This includes toxins accumulating in shellfish, such as saxitoxin, okadaic acid, yessotoxins, azaspiracids, brevetoxins, and pinnatoxins. Other toxins, such as ciguatoxins and maitotoxins, accumulate in fish, where, as is the case for the latter compounds, they can be metabolized to even more toxic metabolites. On the other hand, much less is known about the chemical nature of compounds that are toxic to fish, the so-called ichthyotoxins. Despite numerous reports of algal blooms causing massive fish kills worldwide, only a few types of compounds, such as the karlotoxins, have been proven to be true ichthyotoxins. This review will highlight marine microalgae as the source of some of the most complex natural compounds known to mankind, with chemical structures that show no resemblance to what has been characterized from plants, fungi, or bacteria. In addition, it will summarize algal species known to be related to fish-killing blooms, but from which ichthyotoxins are yet to be characterized.

Chemodiversity of Ladder-Frame Prymnesin Polyethers in Prymnesium parvum.

Blooms of the microalga Prymnesium parvum cause devastating fish kills worldwide, which are suspected to be caused by the supersized ladder-frame polyether toxins prymnesin-1 and -2. These toxins have, however, only been detected from P. parvum in rare cases since they were originally described two decades ago. Here, we report the isolation and characterization of a novel B-type prymnesin, based on extensive analysis of 2D- and 3D-NMR data of natural as well as 90% (13)C enriched material. B-type prymnesins lack a complete 1,6-dioxadecalin core unit, which is replaced by a short acyclic C2 linkage compared to the structure of the original prymnesins. Comparison of the bioactivity of prymnesin-2 with prymnesin-B1 in an RTgill-W1 cell line assay identified both compounds as toxic in the low nanomolar range. Chemical investigations by liquid chromatography high-resolution mass spectrometry (LC-HRMS) of 10 strains of P. parvum collected worldwide showed that only one strain produced the original prymnesin-1 and -2, whereas four strains produced the novel B-type prymnesin. In total 13 further prymnesin analogues differing in their core backbone and chlorination and glycosylation patterns could be tentatively detected by LC-MS/HRMS, including a likely C-type prymnesin in five strains. Altogether, our work indicates that evolution of prymnesins has yielded a diverse family of fish-killing toxins that occurs around the globe and has significant ecological and economic impact.

Noise robustness of a combined phase retrieval and reconstruction method for phase-contrast tomography.

Classical reconstruction methods for phase-contrast tomography consist of two stages: phase retrieval and tomographic reconstruction. A novel algebraic method combining the two was suggested by Kostenko et al. [Opt. Express21, 12185 (2013)OPEXFF1094-408710.1364/OE.21.012185], and preliminary results demonstrated improved reconstruction compared with a given two-stage method. Using simulated free-space propagation experiments with a single sample-detector distance, we thoroughly compare the novel method with the two-stage method to address limitations of the preliminary results. We demonstrate that the novel method is substantially more robust toward noise; our simulations point to a possible reduction in counting times by an order of magnitude.

Sublethal concentrations of ichthyotoxic alga Prymnesium parvum affect rainbow trout susceptibility to viral haemorrhagic septicaemia virus.

Ichthyotoxic algal blooms are normally considered a threat to maricultured fish only when blooms reach lethal cell concentrations. The degree to which sublethal algal concentrations challenge the health of the fish during blooms is practically unknown. In this study, we analysed whether sublethal concentrations of the ichthyotoxic alga Prymnesium parvum affect the susceptibility of rainbow trout Oncorhynchus mykiss to viral haemorrhagic septicaemia virus (VHSV). During exposure to sublethal algal concentrations, the fish increased production of mucus on their gills. When fish were exposed to the algae for 12 h prior to the addition of virus, a marginal decrease in the susceptibility to VHSV was observed compared to fish exposed to VHSV without algae. If virus and algae were added simultaneously, inclusion of the algae increased mortality by 50% compared to fish exposed to virus only, depending on the experimental setup. We concluded that depending on the local exposure conditions, sublethal concentrations of P. parvum could affect susceptibility of fish to infectious agents such as VHSV.

Ocean acidification with (de)eutrophication will alter future phytoplankton growth and succession.

Human activity causes ocean acidification (OA) though the dissolution of anthropogenically generated CO2 into seawater, and eutrophication through the addition of inorganic nutrients. Eutrophication increases the phytoplankton biomass that can be supported during a bloom, and the resultant uptake of dissolved inorganic carbon during photosynthesis increases water-column pH (bloom-induced basification). This increased pH can adversely affect plankton growth. With OA, basification commences at a lower pH. Using experimental analyses of the growth of three contrasting phytoplankton under different pH scenarios, coupled with mathematical models describing growth and death as functions of pH and nutrient status, we show how different conditions of pH modify the scope for competitive interactions between phytoplankton species. We then use the models previously configured against experimental data to explore how the commencement of bloom-induced basification at lower pH with OA, and operating against a background of changing patterns in nutrient loads, may modify phytoplankton growth and competition. We conclude that OA and changed nutrient supply into shelf seas with eutrophication or de-eutrophication (the latter owing to pollution control) has clear scope to alter phytoplankton succession, thus affecting future trophic dynamics and impacting both biogeochemical cycling and fisheries.