The Microalgae Phaeodactylum tricornutum Is Well Suited as a Food with Positive Effects on the Intestinal Microbiota and the Generation of SCFA: Results from a Pre-Clinical Study.

Microalgae such as Phaeodactylum tricornutum (PT) are a sustainable source of nutrients, especially eicosapentaenoic acid (EPA), fucoxanthin (Fx), and chrysolaminarin (Chrl), the concentrations of which can vary depending on the culture conditions. We generated three types of diets containing either an EPA- and Fx-rich (EPA/Fx) or Chrl-rich microalgae (with 5, 15, or 25% added to the diet) or an isocaloric control diet (CD). These diets were evaluated over 14 days in young C57BL/6J mice for safety and bioavailability, short-chain fatty acid (SCFA) production, and microbiome analysis. Both microalgae diets increased body weight gain dose-dependently compared to the CD. Microalgae-derived EPA was well absorbed, resulting in increased liver and fat tissue levels and a decrease in the n-6:n-3 ratio in liver tissue. Both microalgae diets increased the production of selected SCFA and decreased the Firmicutes/Bacteriodota ratio, whereas the Chrl-rich diet led to an increase in Akkermansia. Doses of up to 4621 mg Chrl, 920 mg EPA, and 231 mg Fx per kg body weight daily were tolerated without adverse effects. This pre-clinical study shows that PT is suitable for mouse feed, with positive effects on microbiota composition and SCFA production, suggesting beneficial effects on gut health.

Transstadial Transmission and Replication Kinetics of West Nile Virus Lineage 1 in Laboratory Reared Ixodes ricinus Ticks.

West Nile virus (WNV) is a mosquito-borne agent that has also been isolated from several tick species. Vector competence of Ixodes ricinus, one of the most common tick species in Europe, has been poorly investigated for WNV to date. As such, to evaluate the vector competence, laboratory reared Ixodes ricinus nymphs were in vitro fed with WNV lineage 1 infectious blood, allowed to molt, and the resulting females artificially fed to study the virus transmission. Furthermore, we studied the kinetics of WNV replication in ticks after infecting nymphs using an automatic injector. Active replication of WNV was detected in injected nymphs from day 7 post-infection until 28 dpi. In the nymphs infected by artificial feeding, the transstadial transmission of WNV was confirmed molecularly in 46.7% of males, while virus transmission during in vitro feeding of I. ricinus females originating from infected nymphs was not registered. The long persistence of WNV in I. ricinus ticks did not correlate with the transmission of the virus and it is unlikely that I. ricinus represents a competent vector. However, there is a potential reservoir role that this tick species can play, with hosts potentially acquiring the viral agent after ingesting the infected ticks.

Fucoxanthin, A Carotenoid Derived from Phaeodactylum tricornutum Exerts Antiproliferative and Antioxidant Activities In Vitro.

Microalgae contain a multitude of nutrients and can be grown sustainably. Fucoxanthin, a carotenoid from Phaeodactylum tricornutum, could have beneficial health effects. Therefore, we investigated the anti-inflammatory, antioxidative and antiproliferative effects of fucoxanthin derived from this diatom in vitro. The effects of purified fucoxanthin on metabolic activity were assessed in blood mononuclear cells and different cell lines. In cell lines, caspase 3/7 activity was also analyzed. Nitrogen monoxide release and mRNA-expression of proinflammatory cytokines were measured. For antioxidant assays, cell free assays were conducted. Additionally, the antioxidant effect in neutrophils was quantified and glutathione was determined in HeLa cells. The results show that neither did fucoxanthin have anti-inflammatory properties nor did it exert cytotoxic effects on mononuclear cells. However, the metabolic activity of cell lines was decreased up to 58% and fucoxanthin increased the caspase 3/7 activity up to 4.6-fold. Additionally, dose-dependent antioxidant effects were detected, resulting in a 63% decrease in chemiluminescence in blood neutrophils and a 3.3-fold increase in the ratio of reduced to oxidized glutathione. Our studies show that fucoxanthin possesses antiproliferative and antioxidant activities in vitro. Hence, this carotenoid or the whole microalgae P. tricornutum could be considered as a food or nutraceutical in human nutrition, showcasing beneficial health effects.

Lack of liver steatosis in germ-free mice following hypercaloric diets.

PURPOSE: Experimental liver steatosis induced by overfeeding is associated with enhanced gut permeability and endotoxin translocation to the liver. We examined the role of the gut microbiota for steatosis formation by performing the feeding experiments in mice raised under conventional and germ-free (GF) housing. METHODS: Adult wild-type and GF mice were fed a Western-style diet (WSD) or a control diet (CD), the latter combined with liquid fructose supplementation (F) or not, for 8 weeks. Markers of liver steatosis and gut permeability were measured after intervention. RESULTS: Mice fed a WSD increased body weight compared to those fed a CD (p < 0.01) under conventional, but not under GF conditions. Increased liver weight, liver-to-body-weight ratio and hepatic triglycerides observed in both the WSD and the CD + F groups, when compared with the CD group, were not apparent under GF conditions, whereas elevated plasma triglycerides were visible (p < 0.05). Wild-type mice fed a WSD or a CD + F, respectively, had thinner adherent mucus layer compared to those fed a CD (p < 0.01), whereas GF mice had always a thin mucus layer independently of the diet. GF mice fed a CD showed increased plasma levels of FITC-dextran 4000 (1.9-fold, p < 0.05) and intestinal fatty acid-binding protein-2 (2.4-fold, p < 0.05) compared with wild-type mice. CONCLUSIONS: GF housing results in an impaired weight gain and a lack of steatosis following a WSD. Also the fructose-induced steatosis, which is unrelated to body weight changes, is absent in GF mice. Thus, diet-induced experimental liver steatosis depends in multiple ways on intestinal bacteria.

Bioavailability and Safety of Nutrients from the Microalgae Chlorella vulgaris, Nannochloropsis oceanica and Phaeodactylum tricornutum in C57BL/6 Mice.

Microalgae are rich in macronutrients and therefore, they have been proposed as a potential future food source preserving natural resources. Here, we studied safety and bioavailability of algae nutrients in mice. Three microalgae species, Chlorella vulgaris, Nannochloropsis oceanica and Phaeodactylum tricornutum, were studied after ball mill disruption at different doses (5%, 15% and 25% dry weight) for 14 days. In response to all three algae diets, we observed a weight gain similar or superior to that in response to the control diet. No substantial differences in organ weights nor gut length occurred. Protein bioavailability from the algae diets did not differ from the control diet ranging from 58% to 77% apparent biological value. Fat absorption was lower for microalgae compared to soy oil in control diets, albeit still substantial. High liver eicosapentaenoic acid levels were measured following feeding with N. oceanica, the algae richest in omega-3 fatty acids. Neither histological nor serum analyses revealed any heart, kidney or liver toxicity induced by any of the algae diets. Algae-rich diets were thus well accepted, well tolerated and suitable for the maintenance of body weight and normal organ function. No toxicological effects were observed.

Cholinergic responses of ophthalmic arteries in M3 and M5 muscarinic acetylcholine receptor knockout mice.

PURPOSE: To determine the functional role of M(3) and M(5) muscarinic acetylcholine receptor subtypes in ophthalmic arteries using gene-targeted mice. METHODS: Muscarinic receptor gene expression was quantified in murine ophthalmic arteries using real-time PCR. To test the functional relevance of M(3) and M(5) receptors, ophthalmic arteries from mice deficient in either subtype (M3R(-/-), M5R(-/-), respectively) and wild-type controls were isolated, cannulated with micropipettes, and pressurized. Changes in luminal vessel diameter in response to muscarinic and nonmuscarinic receptor agonists were measured by video microscopy. RESULTS: With the use of real-time PCR, all five muscarinic receptor subtypes were detected in ophthalmic arteries. However, mRNA levels of M(1), M(3), and M(5) receptors were higher than those of M(2) and M(4) receptors. In functional studies, after preconstriction with phenylephrine, acetylcholine and carbachol produced concentration-dependent dilations of ophthalmic arteries that were similar in M5R(-/-) and wild-type mice. Strikingly, cholinergic dilation of ophthalmic arteries was almost completely abolished in M3R(-/-) mice. Deletion of either M(3) or M(5) receptor did not affect responses to nonmuscarinic vasodilators such as bradykinin or nitroprusside. CONCLUSIONS: These findings provide the first evidence that M(3) receptors are critically involved in cholinergic regulation of diameter in murine ophthalmic arteries.

Methods for assessment of innovative medical technologies during early stages of development.

Conventional Health Technology Assessment (HTA) is usually conducted at a point in time at which the development of the respective technology may no longer be influenced. By this time developers and/or purchasers may have misinvested resources. Thus the demand for Technology Assessment (TA) which incorporates appropriate methods during early development stages of a technology becomes apparent. Against this health political background, the present report describes methods for a development-accompanying assessment of innovative medical technologies. Furthermore, international research programmes set out to identify or apply such methods will be outlined. A systematic literature search as well as an extensive manual literature search are carried out in order to obtain literature and information. The greatest units of the identified methods consist of assessment concepts, decision support methods, modelling approaches and methods focusing on users and their knowledge. Additionally, several general-purpose concepts have been identified. The identified research programmes INNO-HTA and MATCH (Multidisciplinary-Assessment-of-Technology-Centre-for-Healthcare) are to be seen as pilot projects which so far have not been able to generate final results. MATCH focuses almost entirely on the incorporation of the user-perspective regarding the development of non-pharmaceutical technologies, whereas INNO-HTA is basically concerned with the identification and possible advancement of methods for the early, socially-oriented technology assessment. Most references offer only very vague descriptions of the respective method and the application of greatly differing methods seldom exceeds the character of a pilot implementation. A standardisation much less an institutionalisation of development-accompanying assessment cannot be recognized. It must be noted that there is no singular method with which development-accompanying assessment should be carried out. Instead, a technology and evaluation specific method selection seem to be necessary as medical innovations are diverse and none of the methods are exhaustive. Because of a variety of issues (e. g. ideal time of evaluation, lack of data and uncertainty of data) a development-accompanying assessment should not replace a comprehensive HTA, but rather form a possible preceding step in a multi-staged HTA-process. A final appraisal of the methods for development-accompanying assessment cannot be made based on the available sources. However, the present review may serve as a starting point for further development and application of these methods as well as further examination of the concept of development-accompanying assessment. There is a substantial need for further research concerning the application, validation and comparison of the various methods for development-accompanying assessment.

Procedures and Criteria for the regulation of innovative non-medicinal technologies into the benefit catalogue of solidly financed health care insurances.

Because great interest in an efficient range of effective medicinal innovations and achievements has arisen, many countries have introduced procedures to regulate the adoption of innovative non-medicinal technologies into the benefit catalogue of solidly financed health care insurances. With this as a background, this report will describe procedures for the adoption of innovative non-medicinal technologies by solidly financed health care insurances in Germany, England, Australia and Switzerland. This report was commissioned by the German Agency for Health Technology Assessment at the German Institute for Medical Documentation and Information.In order to find the relevant literature and information, systematic literature research, a hand search and a written survey were carried out. All the selected documents (chosen according to defined criteria for inclusion and exclusion) were qualitatively evaluated, summarized and presented on a chart using a framework developed for this purpose. All the countries in this report require that some innovative non-medicinal technologies undergo evaluation by a central governing body. This evaluation is a prerequisite for adoption into the benefit catalogue. The process of evaluation can differ (e. g. the people and institutions concerned, the division of the synthesis of evidence and overall evaluation, processing the evidence). Similarities do exist, such as the size and composition of the governing bodies or the overreaching criteria according to which institutions must make their recommendations. This is how all the countries examined in this report determine how the benefits and effectiveness of the innovations, as well as their cost-effectiveness, can be chosen as criteria for the evaluation. Furthermore, there are many criteria which differ from country to country (social and ethical aspects, possible effects on the health system, etc.) and which are also relevant to an evaluation. The preferred types of clinical studies for these evaluations are randomized controlled trials. However, all institutions do allow for other types of evidence (e. g. expert opinion) when no other study types of a higher evidence level are available. In addition, all the countries are willing to allow unpublished or confidential information (e. g. from manufacturers) to be included in an evaluation. It is important to remember that the decisions made by the central governing bodies do not necessarily become conditions for the introduction of innovative non-medicinal technologies. There is a host of other requirements which determine how these innovations can be introduced. This means that a large number of non-medicinal technologies make it into the medical care system via these other decision-making processes. Often, these innovations are unevaluated and differ from region to region. Every country has established a system of observation and registration for medicinal products. These systems are meant to document any incidents with the innovations and to confer responsibility on certain organizations. All in all, no country has a central authority which systematically investigates the effects of newly introduced innovative non-medicinal technologies on medical care in general. However, Australia and England both carry out a review of innovations in some areas (e. g. by means of special commissions). In principle, the starting point for improving regulations of innovative non-medicinal technologies lies in the extension of transparency, the shortening of decision-making time (especially the central decision-making processes), the further development of evaluation methods, more flexibility and increased capacity in the governing bodies' decision-making processes and also, if needed, in the creation of a single authority to act as contact for people who are interested in introducing an innovation into the benefit catalogue.More research is required, especially in the area of decentralized decision-makers and how they actually decide whether or not to introduce innovative technologies into the core care system (methods, criteria, etc.). In view of this, it would also be interesting to see how the application of innovations actually happens in practice once their adoption has been approved by the corresponding governing bodies.

Regulation of expression of the retinoic acid metabolizing enzyme CYP26A1 in uteri of ovariectomized mice after treatment with ovarian steroid hormones.

The retinoic acid (RA) synthesizing enzymes, retinaldehyde dehydrogenases (RALDH), are expressed in specific spatial and temporal patterns in uterine tissues during estrous cycle and early pregnancy in mice. Expression of RALDH1 and 2 has been shown to be induced by estrogen treatment within the uterus. In this study, we determined the influence of progesterone and 17-ss-estradiol on the uterine expression of the RA-metabolizing enzyme CYP26A1 after specific time intervals (1, 4, 24, and 48 hr after treatment of ovariectomized mice). In a following experiment, we investigated the influence of gestagen (promegestone 0.3 mg/kg body weight), estrogen (estradiol 3 microg/kg), their combination, as well as the antagonizing anti-progesterone hormone (RU 486 10 mg/kg) on the uterine expression of CYP26A1. Expression of CYP26A1 was localized using in situ hybridization and quantified using RT-PCR. CYP26A1 mRNA expression was strongly--although transiently--induced in uterine endometrial epithelial and glandular cells after administration of gestagen or the combination of gestagen + estrogen, but not by estrogen alone. These observations were confirmed by semi-quantitative RT-PCR experiments on whole uteri. Thus, we show that the expression of CYP26A1 in endometrial epithelial cells is regulated by progesterone and not significantly influenced by co-administration of estrogen. These data indicate an additional level of hormonal control of endogenous RA levels in the mouse uterus, where its synthesis would rely on estrogen-dependent expression of RALDH enzymes, whereas its active metabolism would be triggered by progesterone-induced CYP26A1 expression.

Regulation of expression of the retinoic acid-synthesising enzymes retinaldehyde dehydrogenases in the uteri of ovariectomised mice after treatment with oestrogen, gestagen and their combination.

The active metabolite of vitamin A, retinoic acid (RA), plays an important role in the female reproductive system. The synthesis of RA is tightly regulated by the activity of retinaldehyde dehydrogenases (Raldh). Among these, Raldh1 and Raldh2 exhibit specific temporal and spatial expression patterns in the mouse uterus, both during the oestrous cycle and early pregnancy. In the present study, we have assessed whether oestradiol and progesterone directly influence the uterine expression of Raldh1 and Raldh2 in ovariectomised mice. We investigated the effect of gestagen (promegestone 0.3 mg kg(-1) bodyweight), oestrogen (oestradiol 3 microg kg(-1) bodyweight) and their combination on the uterine expression of Raldh2. Expression was analysed using in situ hybridisation and quantified using real-time detection reverse transcription-polymerase chain reaction. The results show that the expression of Raldh2 is rapidly (within 1-4 h) induced in stromal cells by oestrogen, but not by gestagen, treatment, whereas combined oestrogen + gestagen treatment leads to a more prolonged (48 h) response. In contrast, oestrogen, but not progesterone, treatment downregulates (within 4-24 h) Raldh1 expression in the uterine glandular epithelium. We conclude that the uterine RA concentrations are regulated by oestrogens via an effect on the expression of the Raldh synthesising enzymes. Such a regulation is consistent with the natural fluctuations of Raldh expression during the oestrous cycle, early pregnancy and blastocyst implantation.