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Abstract In advanced drug delivery, versatile liposomal formulations are commonly employed for safer and more accurate therapies. Here we report a method that allows a straightforward production of synthetic monodisperse (~ 100 μm) giant unilamellar vesicles (GUVs) using a microfluidic system. The stability analysis based on the microscopy imaging showed that at ambient conditions the produced GUVs had a half-life of 61 ± 2 h. However, it was observed that ~ 90% of the calcein dye that was loaded into GUVs was transported into a surrounding medium in 24 h, thus indicating that the GUVs may release these small dye molecules without distinguishable membrane disruption. We further demonstrated the feasibility of our method by loading GUVs with larger and very different cargo objects; small soluble fluorescent proteins and larger magnetic microparticles in a suspension. Compared to previously reported microfluidics-based production techniques, the obtained results indicate that our simplified method could be equally harnessed in creating GUVs with less cost, effort and time, which could further benefit studying closed membrane systems.

Chaga mushroom (Inonotus obliquus) is a pathogenic fungus that grows mostly on birch species (Betula pendula Roth and B. pubescens Ehrh.) and has traditionally been used as an anticancer medicine. This study aimed to compare the chemical composition and cytotoxic activity of chagas growing on both Betula spp. on various cancer cell lines. The freeze-dried extracts contained triterpenes inotodiol, lanosterol betulin, and betulinic acid typical to conks growing on Betula species. The cytotoxic activity of chaga growing on Betula pendula and B. pubescens 80% ethanolic extracts against 31 human cancer cell lines was evaluated by a sulforhodamine B assay. Chaga extract showed moderate activity against all cancer cell lines examined; it did not result in high cytotoxicity (IC50 ≤ 20 µg/mL). The strongest inhibitions were observed with chaga (growing on B. pendula) extract on the HepG2 and CAL-62 cell line and with chaga (from B. pubescens) extract on the HepG2 cell line, with IC50 values of 37.71, 43.30, and 49.99 μg/mL, respectively. The chaga extracts from B. pendula exert somewhat stronger effects on most cancer cell lines studied than B. pubescens extracts, which can be attributed to a higher content of inotodiol in B. pendula extracts. This study highlights the potential of chaga as a source of bioactive compounds with selective anticancer properties. To the best of our knowledge, this study is the first investigation of the chemical composition of I. obliquus parasitizing on B. pubescens.

Thrombospondin-4 (THBS4) is a non-structural extracellular matrix molecule associated with tissue regeneration and a variety of pathological processes characterized by increased cell proliferation and migration. However, the mechanisms of how THBS4 regulates cell behavior as well as the pathways contributing to its effects have remained largely unexplored. In the present study we investigated the role of THBS4 in skin regeneration both in vitro and in vivo. We found that THBS4 expression was upregulated in the dermal compartment of healing skin wounds in humans as well as in mice. Application of recombinant THBS4 protein promoted cutaneous wound healing in mice and selectively stimulated migration of primary fibroblasts as well as proliferation of keratinocytes in vitro. By using a combined proteotranscriptomic pathway analysis approach we discovered that β-catenin acted as a hub for THBS4-dependent cell signaling and likely plays a key role in promoting its downstream effects. Our results suggest that THBS4 is an important contributor to wound healing and its incorporation into novel wound healing therapies may be a promising strategy for treatment of cutaneous wounds.

Inonotus obliquus grows in the Northern Hemisphere on some living broadleaved tree species as a pathogen, causing stem rot. In Estonia, the fungus is well known in the Betula species but can also be found on Alnus. Sterile conks of I. obliquus contain different bioactive compounds, but the quantitative and comparative research of these compounds in conks on different host species is limited. In the current work, I. obliquus was isolated and, evidently, determined from Alnus incana (L.) Moench., Alnus glutinosa (L.) Gaertn., and Betula pendula Roth, and the content of bioactive compounds in conks on these hosts were analysed. All the analysed conks sampled from A. incana and B. pendula contained betulin that varied from 111 to 159 µg/g. A significantly (p < 0.05) higher betulinic acid content was found in conks sampled from A. incana when compared with B. pendula: 474–635 and 20–132 µg/g, respectively. However, the conks from Betula were richer in total polyphenols, flavonols, and glucans. The content of inotodiol was quite similar in the conks from A. incana (7455–8961 µg/g) and B. pendula (7881–9057 µg/g). Also, no significant differences in the lanosterol content were found between the samples from these two tree species. To the best of our knowledge, this study is the first investigation of the chemical composition of I. obliquus parasitizing on Alnus. The results demonstrate that the bioactive compounds are promising in conks of I. obliquus growing not only on Betula but also on the Alnus species. It supports the opportunity to cultivate I. obliquus, also on the Alnus species, thus increasing the economic value of growing this tree species in forestry.

Background: While growing in natural environments yeasts can be affected by osmotic stress provoked by high glucose concentrations. The response to this adverse condition requires the HOG pathway and involves transcriptional and posttranscriptional mechanisms initiated by the phosphorylation of this protein, its translocation to the nucleus and activation of transcription factors. One of the genes induced to respond to this injury is YHR087W. It encodes for a protein structurally similar to the N-terminal region of human SBDS whose expression is also induced under other forms of stress and whose deletion determines growth defects at high glucose concentrations. Results: In this work we show that YHR087W expression is regulated by several transcription factors depending on the particular stress condition, and Hot1p is particularly relevant for the induction at high glucose concentrations. In this situation, Hot1p, together to Sko1p, binds to YHR087W promoter in a Hog1p-dependent manner. Several evidences obtained indicate Yhr087wp's role in translation. Firstly, and according to TAP purification experiments, it interacts with proteins involved in translation initiation. Besides, its deletion mutant shows growth defects in the presence of translation inhibitors and displays a slightly slower translation recovery after applying high glucose stress than the wild type strain. Analyses of the association of mRNAs to polysome fractions reveals a lower translation in the mutant strain of the mRNAs corresponding to genes GPD1, HSP78 and HSP104. Conclusions: The data demonstrates that expression of Yhr087wp under high glucose concentration is controlled by Hot1p and Sko1p transcription factors, which bind to its promoter. Yhr087wp has a role in translation, maybe in the control of the synthesis of several stress response proteins, which could explain the lower levels of some of these proteins found in previous proteomic analyses and the growth defects of the deletion strain. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented which discusses the factors causing thrombosis in the carotid artery of women.

Anthropogenic nitrogen deposition is currently causing a more than twofold increase of reactive nitrogen input over large areas in the tropics. Elevated 15N abundance (δ15N) in the growth rings of some tropical trees has been hypothesized to reflect an increased leaching of 15N-depleted nitrate from the soil following anthropogenic nitrogen deposition over the last decades. To find further evidence for altered nitrogen cycling in tropical forests we measured long-term δ15N values in trees from Bolivia, Cameroon and Thailand. We used two different sampling methods. In the first, wood samples were taken in a conventional way: from the pit to the bark across the stem of 28 large trees (the ‘radial’ method). In the second, δ15N values were compared across a fixed diameter (the ‘fixed-diameter’ method). We sampled 400 trees that differed widely in size, but measured δ15N in the stem around the same diameter (20 cm dbh) in all trees. As a result, the growth rings formed around this diameter differed in age and allowed a comparison of δ15N values over time with an explicit control for the potential size-effects on δ15N values. We found a significant increase of tree-ring δ15N across the stem radius of large trees from Bolivia and Cameroon, but no change in tree-ring δ15N values over time was found in any of the study sites when controlling for tree size. This suggests that radial trends of δ15N values within trees reflect tree ontogeny (size development). However, for the trees from Cameroon and Thailand, a low statistical power in the fixed-diameter method prevents to conclude this with high certainty. For the trees from Bolivia, statistical power in the fixed-diameter method was high, showing that the temporal trend in tree-ring δ15N values in the radial method is primarily caused by tree ontogeny and unlikely by a change in nitrogen cycling. We therefore stress to account for tree size before tree-ring δ15N values can be properly interpreted.

The elaborate eukaryotic DNA replication machinery evolved from the archaeal ancestors that themselves show considerable complexity. Here we discuss the comparative genomic and phylogenetic analysis of the core replication enzymes, the DNA polymerases, in archaea and their relationships with the eukaryotic polymerases. In archaea, there are three groups of family B DNA polymerases, historically known as PolB1, PolB2 and PolB3. All three groups appear to descend from the last common ancestors of the extant archaea but their subsequent evolutionary trajectories seem to have been widely different. Although PolB3 is present in all archaea, with the exception of Thaumarchaeota, and appears to be directly involved in lagging strand replication, the evolution of this gene does not follow the archaeal phylogeny, conceivably due to multiple horizontal transfers and/or dramatic differences in evolutionary rates. In contrast, PolB1 is missing in Euryarchaeota but otherwise seems to have evolved vertically. The third archaeal group of family B polymerases, PolB2, includes primarily proteins in which the catalytic centers of the polymerase and exonuclease domains are disrupted and accordingly the enzymes appear to be inactivated. The members of the PolB2 group are scattered across archaea and might be involved in repair or regulation of replication along with inactivated members of the RadA family ATPases and an additional, uncharacterized protein that are encoded within the same predicted operon. In addition to the family B polymerases, all archaea, with the exception of the Crenarchaeota, encode enzymes of a distinct family D the origin of which is unclear. We examine multiple considerations that appear compatible with the possibility that family D polymerases are highly derived homologs of family B. The eukaryotic DNA polymerases show a highly complex relationship with their archaeal ancestors including contributions of proteins and domains from both the family B and the fam

This paper aims at studying the problems of aviation noise and air, soil and water pollution in Tallinn Airport and the implementation of measures to make the airport more environmentally friendly. At present Tallinn Airport has two stationary noise level monitoring terminals and one mobile terminal for metering and modelling the noise level caused by aircraft taking-off and landing. Research shows that Tallinn Airport has not yet exceeded the noise limits stipulated by regulations. The area surrounding Tallinn Airport has been divided into four noise zones. Zone 1, where the noise level does not exceed 55 dB, is suitable for a majority of types of buildings, whereas Zone 4, where the noise level exceeds 70 dB and the highest level measured was 105 dB, is absolutely unacceptable as a building area. In recent years the number of flights flying over the residential areas of Tallinn has been reduced significantly, i.e. the number of flights taking off from runway 26 and landing on runway 08. In suitable weather conditions, noisy aircraft are directed to land on RWY 26 and take off from RWY08. Thanks to the measures mentioned above, air pollution from aircraft exhaust gases has been reduced considerably in Tallinn. After the completion of the military airfield at Ämari in the nearest future, it is going to be possible to transfer the cargo flights, usually made by large noisy jets, from Tallinn Airport to Ämari. Various measures have also been taken to reduce the pollution of soil and water within the area of Tallinn Airport.

We examined sensitivity of audiovisual temporal order in adolescents with Autism Spectrum Disorder (ASD) using an audiovisual Temporal Order Judgment (TOJ) task. In order to assess domain-specific impairments, the stimuli varied in social complexity from simple flash/beeps to videos of a handclap or a speaking face. Compared to typically-developing controls, individuals with ASD were generally less sensitive in judgments of audiovisual temporal order (larger Just Noticeable Differences, JNDs), but there was no specific impairment with social stimuli. This suggests that people with ASD suffer from a more general impairment in audiovisual temporal processing.

The G1/S transition is a crucial decision point in the cell cycle. At G1/S, there is an abrupt switch from a state of high CDK inhibitor (CKI) levels and low S-phase CDK activity to a state of high S-phase CDK activity and degraded CKI. In budding yeast, this transition is triggered by phosphorylation of the Cdk1 inhibitor Sic1 at multiple sites by G1-phase CDK (Cln1,2-Cdk1) and S-phase CDK (Clb5,6-Cdk1) complexes. Using mathematical modeling we demonstrate that the mechanistic basis for the abruptness and irreversibility of the G1/S transition is the highly specific phosphorylation of Sic1 by S-phase CDK complex. This switch is generated by a double negative feedback loop in which S-CDK1 phosphorylates Sic1, thus targeting it for destruction, and thereby liberating further S-CDK1 from the inhibitory Sic1-S-CDK1 complex. Our model predicts that the abruptness of the switch depends upon a strong binding affinity within the Sic1-S-CDK inhibitory complex. In vitro phosphorylation analysis using purified yeast proteins revealed that free Clb5-Cdk1 can create positive feedback by phosphorylating Sic1 that is bound in the inhibitory complex, and that Sic1 inhibits Clb5-Cdk1 with a sub-nanomolar inhibition constant. Our model also predicts that if the G1-phase CDK complex is too efficient at targeting Sic1 for destruction, then G1/S becomes a smooth and readily reversible transition. We propose that the optimal role for the G1-phase CDK in the switch would not be to act as a kinase activity directly responsible for abrupt degradation of CKI, but rather to act as a priming signal that initiates a positive feedback loop driven by emerging free S-phase CDK.

In recent years, concern over airport noise and its impact on surrounding communities has been spurred by the continual growth in air traffic together with urban development in close proximity to many airports around the world. This paper aims at studying the cases of Tallinn Airport and Tartu Airport, both in Estonia, with a focus on evaluating the airport noise and its impact on the neighbouring residential districts. Tallinn Airport is located within the city limits (only 4 kilometres from the city centre), and therefore the inhabited areas are also exposed to aviation noise. The main water catchment area for the city – Lake Ülemiste – and the nearest residential district are situated about 0.5 km from the airport. In 2003 stationary noise monitoring equipment was installed at Tallinn Airport to measure the noise level caused by starting and landing aircraft so that the necessary measures and sanctions can be taken. At Tartu Airport, the noise was recurrently measured in 2008, by using a TES 1352A sound level meter. The monitoring system at Tallinn Airport and the measurements at Tartu Airport enable operating noise and aircraft noise to be analyzed and predicted as a way to differentiate the landing fees for aircraft depending on noise and flight trajectory. According to the noise monitoring and measurements results, noise maps can be drawn. Santrauka Pastaraisiais metais nerimo faktorius, atsirades del triukšmo, sukeliamo oro uostuose ir jo poveikio aplinkiniams gyvenamiesiems rajonams, tapo rimtesnis, nes gauseja orlaiviu, taip pat oro uostai isikuria per arti gyvenamuju rajonu. Šiu tyrimu tikslas buvo nustatyti Talino ir Tartu oro uostu keliamo triukšmo lygi bei jo itaka aplinkiniams gyvenamiesiems rajonams. Tarptautinis Talino oro uotas yra isikūres miesto ribose (tik 4 km atstumu nuo miesto centro). Tai reiškia, kad nemažai gyventoju kenčia nuo orlaiviu keliamo triukšmo. Pati artimiausia gyvenviete yra nutolusi nuo oro uosto tik 0,5 km atstumu. Julemistes ežeras, kuris yra miesto geriamojo vandens šaltinis, taip pat yra oro uosto apylinkese. 2003 m. Talino oro uoste buvo pastatyta triukšmo lygio matavimo iranga, kuri turejo ne tik nustatyti kylančiu ir besileidžiančiu orlaiviu sukeliamo triukšmo lygi, bet ir numatyti būtinas priemones jam sumažinti bei taikyti sankcijas. 2008 m. Tartu oro uoste triukšmas matuotas kelis kartus naudojant triukšmo matuokli TES 1352A. Ši triukšmo matavimo sistema leidžia pirmiausia išanalizuoti ir nustatyti orlaiviu bei kitos irangos keliamo triukšmo lygi ir tuomet diferencijuoti rinkliavas už nusileidima, kurios priklauso nuo sukeliamo triukšmo lygio ir skrydžio trajektorijos. Priklausomai nuo triukšmo kontroles ir jos rezultatu, galima nupiešti triukšmo žemelapius. First Publish Online: 14 Oct 2010 Reikšminiai žodžiai: oro uosto triukšmas, oro uosto aplinkos įtaka, triukšmo matavimas, triukšmo lygis, triukšmingiausi orlaiviai, triukšmo žemėlapiai, sumažinimo schemos

Importance: Evidence regarding corticosteroid use for severe coronavirus disease 2019 (COVID-19) is limited., Objective: To determine whether hydrocortisone improves outcome for patients with severe COVID-19., Design, Setting, and Participants: An ongoing adaptive platform trial testing multiple interventions within multiple therapeutic domains, for example, antiviral agents, corticosteroids, or immunoglobulin. Between March 9 and June 17, 2020, 614 adult patients with suspected or confirmed COVID-19 were enrolled and randomized within at least 1 domain following admission to an intensive care unit (ICU) for respiratory or cardiovascular organ support at 121 sites in 8 countries. Of these, 403 were randomized to open-label interventions within the corticosteroid domain. The domain was halted after results from another trial were released. Follow-up ended August 12, 2020., Interventions: The corticosteroid domain randomized participants to a fixed 7-day course of intravenous hydrocortisone (50 mg or 100 mg every 6 hours) (n = 143), a shock-dependent course (50 mg every 6 hours when shock was clinically evident) (n = 152), or no hydrocortisone (n = 108)., Main Outcomes and Measures: The primary end point was organ support-free days (days alive and free of ICU-based respiratory or cardiovascular support) within 21 days, where patients who died were assigned -1 day. The primary analysis was a bayesian cumulative logistic model that included all patients enrolled with severe COVID-19, adjusting for age, sex, site, region, time, assignment to interventions within other domains, and domain and intervention eligibility. Superiority was defined as the posterior probability of an odds ratio greater than 1 (threshold for trial conclusion of superiority >99%)., Results: After excluding 19 participants who withdrew consent, there were 384 patients (mean age, 60 years; 29% female) randomized to the fixed-dose (n = 137), shock-dependent (n = 146), and no (n = 101) hydrocortisone groups; 379 (99%) completed the study and were included in the analysis. The mean age for the 3 groups ranged between 59.5 and 60.4 years; most patients were male (range, 70.6%-71.5%); mean body mass index ranged between 29.7 and 30.9; and patients receiving mechanical ventilation ranged between 50.0% and 63.5%. For the fixed-dose, shock-dependent, and no hydrocortisone groups, respectively, the median organ support-free days were 0 (IQR, -1 to 15), 0 (IQR, -1 to 13), and 0 (-1 to 11) days (composed of 30%, 26%, and 33% mortality rates and 11.5, 9.5, and 6 median organ support-free days among survivors). The median adjusted odds ratio and bayesian probability of superiority were 1.43 (95% credible interval, 0.91-2.27) and 93% for fixed-dose hydrocortisone, respectively, and were 1.22 (95% credible interval, 0.76-1.94) and 80% for shock-dependent hydrocortisone compared with no hydrocortisone. Serious adverse events were reported in 4 (3%), 5 (3%), and 1 (1%) patients in the fixed-dose, shock-dependent, and no hydrocortisone groups, respectively., Conclusions and Relevance: Among patients with severe COVID-19, treatment with a 7-day fixed-dose course of hydrocortisone or shock-dependent dosing of hydrocortisone, compared with no hydrocortisone, resulted in 93% and 80% probabilities of superiority with regard to the odds of improvement in organ support-free days within 21 days. However, the trial was stopped early and no treatment strategy met prespecified criteria for statistical superiority, precluding definitive conclusions., Trial Registration: ClinicalTrials.gov Identifier: NCT02735707.

Descending Necrotizing Mediastinitis (DNM) is a polymicrobic, dangerous and often fatal process, arising from head or neck infections and spreading along the deep fascial cervical planes, descending into the mediastinum. It can rapidly progress to sepsis and can frequently lead to death. It has a high mortality rate, up to 40% in the different series, as described in the literature. Surgical and therapeutic management has been discussed for long time especially in an elderly patient population. The literature has been reviewed in order to evaluate different pathogenesis and evolution and to recognise a correct therapeutic management., Competing Interests: Conflict of Interests: The authors declare that they have no conflict of interests.