Interactions between Polyunsaturated Fatty Acids and Probiotics

The prevalence of inflammatory based diseases has increased in industrialized countries over the last decades. For allergic diseases, two primary hypotheses have been proposed to explain this phenomenon, namely the hygiene and dietary evolution based hypothesis. Particularly, the reduced early exposure to microbes and an increase in the amount of polyunsaturated fatty acids (especially n-6 PUFA) in the diet have been discussed. Often, these two factors have been studied independently, even though both factors have been shown to possess potential health benefits and their mode of action to share similar mechanisms. The hypothesis of the present study was that demonstrate that PUFA and probiotics are not separate entities as such but do interact with each other. In the present study, we investigated whether maternal diet and atopic status influence the PUFA composition of breast milk and serum fatty acids of infants, and whether the fatty acid absorption and utilization of infant formula fatty acids is affected by supplementation of infant formula with probiotic bacteria (Lactobacillus GG and Bifidobacterium lactis Bb-12). Moreover, we investigated the mechanisms by which different PUFA influence the physicochemical and functional properties of probiotics as well as functionality of epithelial cells in vitro. We demonstrated a carry-over effect of dietary fatty acids from maternal diet via breast milk into infants’ serum lipid fatty acids. Our data confirmed the previously shown allergy –related PUFA level imbalances, though it did not fully support the impaired desaturation and elongation capacity hypothesis. We also showed that PUFA incorporation into phospholipids of infants was influenced by probiotics in infant formula in a strain dependent manner. Especially,Bifidobacterium lactis Bb-12 in infant formula promoted the utilization of n-3 PUFA. Mechanistically, we demonstrated that probiotics (Lactobacillus GG, Lactobacillus casei Shirota and Lactobacillus bulgaricus) did incorporate and interconvert exogenous free PUFA in the growth medium into bacterial fatty acids strain and PUFA dependently. In general, high concentrations of free PUFA inhibited the growth and mucus adhesion of probiotics, whereas low concentrations of specific long chain PUFA were found to promote the growth and mucus adhesion of Lactobacillus casei Shirota. These effects were paralleled with only minor alterations in hydrophobicity and electron donor – electron acceptor properties of lactobacilli. Furthermore, free PUFA were also demonstrated to alter the adhesion capacity of the intestinal epithelial cells; n-6 PUFA tended to inhibit the Caco-2 adhesion of probiotics, whereas n-3 PUFA had either no or minor effects or even promote the bacterial adhesion (especially Lactobacillus casei Shirota) to PUFA treated Caco-2 cells. The results of this study demonstrate the close and bilateral interactions between dietary PUFA and probiotics. Probiotics were shown to influence the absorption and utilization of dietary PUFA, whereas PUFA were shown to alter the functional properties of both probiotics and mucosal epithelia. These findings suggest that a more thorough understanding of interactions between PUFA and intestinal microbiota is a prerequisite, when the beneficial effects of new functional foods containing probiotics are designed and planned for human intervention studies.Siirretty Doriast

Levodopa-Induced Changes in Electromyographic Patterns in Patients with Advanced Parkinson's Disease

Levodopa medication is the most efficient treatment for motor symptoms of Parkinson's disease (PD). Levodopa significantly alleviates rigidity, rest tremor, and bradykinesia in PD. The severity of motor symptoms can be graded with UPDRS-III scale. Levodopa challenge test is routinely used to assess patients' eligibility to deep-brain stimulation (DBS) in PD. Feasible and objective measurements to assess motor symptoms of PD during levodopa challenge test would be helpful in unifying the treatment. Twelve patients with advanced PD who were candidates for DBS treatment were recruited to the study. Measurements were done in four phases before and after levodopa challenge test. Rest tremor and rigidity were evaluated using UPDRS-III score. Electromyographic (EMG) signals from biceps brachii and kinematic signals from forearm were recorded with wireless measurement setup. The patients performed two different tasks: arm isometric tension and arm passive flexion-extension. The electromyographic and the kinematic signals were analyzed with parametric, principal component, and spectrum-based approaches. The principal component approach for isometric tension EMG signals showed significant decline in characteristics related to PD during levodopa challenge test. The spectral approach on passive flexion-extension EMG signals showed a significant decrease on involuntary muscle activity during the levodopa challenge test. Both effects were stronger during the levodopa challenge test compared to that of patients' personal medication. There were no significant changes in the parametric approach for EMG and kinematic signals during the measurement. The results show that a wireless and wearable measurement and analysis can be used to study the effect of levodopa medication in advanced Parkinson's disease.Peer reviewe

Changes in elbow flexion EMG morphology during adjustment of deep brain stimulator in advanced Parkinson's disease

Publisher Copyright: © 2022 Ruonala et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Objective Deep brain stimulation (DBS) is an effective treatment for motor symptoms of advanced Parkinson's disease (PD). Currently, DBS programming outcome is based on a clinical assessment. In an optimal situation, an objectively measurable feature would assist the operator to select the appropriate settings for DBS. Surface electromyographic (EMG) measurements have been used to characterise the motor symptoms of PD with good results; with proper methodology, these measurements could be used as an aid to program DBS. Methods Muscle activation measurements were performed for 13 patients who had advanced PD and were treated with DBS. The DBS pulse voltage, frequency, and width were changed during the measurements. The measured EMG signals were analysed with parameters that characterise the EMG signal morphology, and the results were compared to the clinical outcome of the adjustment. Results The EMG signal correlation dimension, recurrence rate, and kurtosis changed significantly when the DBS settings were changed. DBS adjustment affected the signal recurrence rate the most. Relative to the optimal settings, increased recurrence rates (median ± IQR) 1.1 ± 0.5 (-0.3 V), 1.3 ± 1.1 (+0.3 V), 1.7 ± 0.4 (-30 Hz), 1.7 ± 0.8 (+30 Hz), 2.0 ± 1.7 (+30 μs), and 1.5 ± 1.1 (DBS off) were observed. With optimal stimulation settings, the patients' Unified Parkinson's Disease Rating Scale motor part (UPDRS-III) score decreased by 35% on average compared to turning the device off. However, the changes in UPRDS-III arm tremor and rigidity scores did not differ significantly in any settings compared to the optimal stimulation settings. Conclusion Adjustment of DBS treatment alters the muscle activation patterns in PD patients. The changes in the muscle activation patterns can be observed with EMG, and the parameters calculated from the signals differ between optimal and non-optimal settings of DBS. This provides a possibility for using the EMG-based measurement to aid the clinicians to adjust the DBS.Peer reviewe

A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in E. coli

Quantitating intracellular oxidative damage caused by reactive oxygen species (ROS) is of interest in many fields of biological research. The current systems primarily rely on supplemented oxygen-sensitive substrates that penetrate the target cells, and react with ROS to produce signals that can be monitored with spectroscopic or imaging techniques. The objective here was to design a new non-invasive analytical strategy for measuring ROS-induced damage inside living cells by taking advantage of the native redox sensor system of E. coli. The developed plasmid-based sensor relies on an oxygen-sensitive transcriptional repressor IscR that controls the expression of a fluorescent marker in vivo. The system was shown to quantitatively respond to oxidative stress induced by supplemented H2O2 and lowered cultivation temperatures. Comparative analysis with fluorescence microscopy further demonstrated that the specificity of the reporter system was equivalent to the commercial chemical probe (CellROX). The strategy introduced here is not dependent on chemical probes, but instead uses a fluorescent expression system to detect enzyme-level oxidative damage in microbial cells. This provides a cheap and simple means for analysing enzyme-level oxidative damage in a biological context in E. coli

Quantitative bioimage analytics enables measurement of targeted cellular stress response induced by celastrol-loaded nanoparticles

The cellular stress response, which provides protection against proteotoxic stresses, is characterized by the activation of heat shock factor 1 and the formation of nuclear stress bodies (nSBs). In this study, we developed a computerized method to quantify the formation and size distribution of nSBs, as stress response induction is of interest in cancer research, neurodegenerative diseases, and in other pathophysiological processes. We employed an advanced bioimaging and analytics workflow to enable quantitative detailed subcellular analysis of cell populations even down to single-cell level. This type of detailed analysis requires automated single cell analysis to allow for detection of both size and distribution of nSBs. For specific induction of nSB we used mesoporous silica nanoparticles (MSNs) loaded with celastrol, a plant-derived triterpene with the ability to activate the stress response. To enable specific targeting, we employed folic acid functionalized nanoparticles, which yields targeting to folate receptor expressing cancer cells. In this way, we could assess the ability to quantitatively detect directed and spatio-temporal nSB induction using 2D and 3D confocal imaging. Our results demonstrate successful implementation of an imaging and analytics workflow based on a freely available, general-purpose software platform, BioImageXD, also compatible with other imaging modalities due to full 3D/4D and high-throughput batch processing support. The developed quantitative imaging analytics workflow opens possibilities for detailed stress response examination in cell populations, with significant potential in the analysis of targeted drug delivery systems related to cell stress and other cytoprotective cellular processes.</p

Internalization of novel non-viral vector TAT-streptavidin into human cells

BACKGROUND: The cell-penetrating peptide derived from the Human immunodeficiency virus-1 transactivator protein Tat possesses the capacity to promote the effective uptake of various cargo molecules across the plasma membrane in vitro and in vivo. The objective of this study was to characterize the uptake and delivery mechanisms of a novel streptavidin fusion construct, TAT(47–57)-streptavidin (TAT-SA, 60 kD). SA represents a potentially useful TAT-fusion partner due to its ability to perform as a versatile intracellular delivery vector for a wide array of biotinylated molecules or cargoes. RESULTS: By confocal and immunoelectron microscopy the majority of internalized TAT-SA was shown to accumulate in perinuclear vesicles in both cancer and non-cancer cell lines. The uptake studies in living cells with various fluorescent endocytic markers and inhibiting agents suggested that TAT-SA is internalized into cells efficiently, using both clathrin-mediated endocytosis and lipid-raft-mediated macropinocytosis. When endosomal release of TAT-SA was enhanced through the incorporation of a biotinylated, pH-responsive polymer poly(propylacrylic acid) (PPAA), nuclear localization of TAT-SA and TAT-SA bound to biotin was markedly improved. Additionally, no significant cytotoxicity was detected in the TAT-SA constructs. CONCLUSION: This study demonstrates that TAT-SA-PPAA is a potential non-viral vector to be utilized in protein therapeutics to deliver biotinylated molecules both into cytoplasm and nucleus of human cells

Tiimiorganisaation kyvykkyyden kehittäminen

Opinnäytetyön tarkoituksena oli tutkia kohdeyrityksen ohjelmisto-osaston organisaation toimintaa. Lähtökohtana tutkimukselle oli ohjelmisto-osaston ja tiimien nykytilan selvittäminen ja sidosryhmien odotusten syvempi ymmärtäminen. Tämän päivän järjestelmätoimituksissa ohjelmistot ovat isossa roolissa. Ohjelmistoilla pystytään luomaan sellaista lisäarvoa asiakkaille, jota ei pelkällä laitteistolla pystytä luomaan. Tämän vuoksi on hyvin tärkeää, että yrityksen ohjelmisto-osaaminen kehittyy, toimintatavat ovat määriteltyjä ja uusia asioita kyetään ottamaan käyttöön mahdollisimman tehokkaasti. Opinnäytetyössä pyrittiin selvittämään, mitkä ovat organisaation kipukohdat ja miten toimintaa voitaisiin kehittää, jotta se olisi mahdollisimman tarkoituksenmukaista ja organisoituminen tukisi toiminnan kehittymistä parhaalla mahdollisella tavalla. Opinnäytetyössä haastateltiin eri sidosryhmiä teemahaastattelun keinoin. Teemahaastattelujen tulokset koottiin miellekarttaan, jossa asiat ryhmiteltiin viiteen eri luokkaan. Teemahaastattelujen, luokittelun ja havaintojen pohjalta muodostettiin näkemys ohjelmisto-osaston haasteista. Tutkimus toteutettiin konstruktiivisella tutkimusotteella, jossa on tarkoituksena luoda uusi konstruktio ja näin ratkaista reaalimaailman ongelma. Kaikki ihmisen kehittämät mallit, prosessit ja organisaatiorakenteet ovat konstruktioita. Tässä tutkimuksessa luotiin uusi konstruktio kohdeyrityksen ohjelmisto-osaston organisaatiorakenteeksi.The purpose of this thesis was to study the organizational activities of the target company's software department. The starting point for the research was to find out the current state of the software department and teams and to gain a deeper understanding of stakeholders’ expectations. Software plays a big role in today's automation system deliveries. Software could create added value for customers that hardware alone cannot bring. For this reason, it is very important that the company's software capability develops, operating methods are defined, and new things can be introduced as efficiently as possible. With this thesis, the aim was to better understand the pain points of the organization and how the operations could be developed so that the operations would be as appropriate as possible, and the organization would support the development of the activities in the best possible way. In this thesis, different stakeholders were interviewed by means of a thematic interview. The results of the thematic interviews were collected into a mind map, where the issues are grouped into five different categories. Based on the thematic interviews, classification and observations, a vision of the challenges of the software department was formed. This research was carried out with a constructive research approach aimed at creating some new construction and thus solving some real-world problem. All man-made models, processes, and organization structures are constructions. In this research, a new construction was created for the organization structure of the target company's software department

Developing Bioimage Informatics – from Microscopy to Software Solutions – with alpha2beta1 Integrin as a Case Study

Biokuvainformatiikan kehittäminen – mikroskopiasta ohjelmistoratkaisuihin – sovellusesimerkkinä α2β1-integriini Kun ihmisen genomi saatiin sekvensoitua vuonna 2003, biotieteiden päätehtäväksi tuli selvittää eri geenien tehtävät, ja erilaisista biokuvantamistekniikoista tuli keskeisiä tutkimusmenetelmiä. Teknologiset kehitysaskeleet johtivat erityisesti fluoresenssipohjaisten valomikroskopiatekniikoiden suosion räjähdysmäiseen kasvuun, mutta mikroskopian tuli muuntua kvalitatiivisesta tieteestä kvantitatiiviseksi. Tämä muutos synnytti uuden tieteenalan, biokuvainformatiikan, jonka on sanottu mahdollisesti mullistavan biotieteet. Tämä väitöskirja esittelee laajan, poikkitieteellisen työkokonaisuuden biokuvainformatiikan alalta. Väitöskirjan ensimmäinen tavoite oli kehittää protokollia elävien solujen neliulotteiseen konfokaalimikroskopiaan, joka oli yksi nopeimmin kasvavista biokuvantamismenetelmistä. Ihmisen kollageenireseptori α2β1-integriini, joka on tärkeä molekyyli monissa fysiologisissa ja patologisissa prosesseissa, oli sovellusesimerkkinä. Työssä saavutettiin selkeitä visualisointeja integriinien liikkeistä, yhteenkeräytymisestä ja solun sisään siirtymisestä, mutta työkaluja kuvainformaation kvantitatiiviseen analysointiin ei ollut. Väitöskirjan toiseksi tavoitteeksi tulikin tällaiseen analysointiin soveltuvan tietokoneohjelmiston kehittäminen. Samaan aikaan syntyi biokuvainformatiikka, ja kipeimmin uudella alalla kaivattiin erikoistuneita tietokoneohjelmistoja. Tämän väitöskirjatyön tärkeimmäksi tulokseksi muodostui näin ollen BioImageXD, uudenlainen avoimen lähdekoodin ohjelmisto moniulotteisten biokuvien visualisointiin, prosessointiin ja analysointiin. BioImageXD kasvoi yhdeksi alansa suurimmista ja monipuolisimmista. Se julkaistiin Nature Methods -lehden biokuvainformatiikkaa käsittelevässä erikoisnumerossa, ja siitä tuli tunnettu ja laajalti käytetty. Väitöskirjan kolmas tavoite oli soveltaa kehitettyjä menetelmiä johonkin käytännönläheisempään. Tehtiin keinotekoisia piidioksidinanopartikkeleita, joissa oli "osoitelappuina" α2β1-integriinin tunnistavia vasta-aineita. BioImageXD:n avulla osoitettiin, että nanopartikkeleilla on potentiaalia lääkkeiden täsmäohjaussovelluksissa. Tämän väitöskirjatyön yksi perimmäinen tavoite oli edistää uutta ja tuntematonta biokuvainformatiikan tieteenalaa, ja tämä tavoite saavutettiin erityisesti BioImageXD:n ja sen lukuisten julkaistujen sovellusten kautta. Väitöskirjatyöllä on merkittävää potentiaalia tulevaisuudessa, mutta biokuvainformatiikalla on vakavia haasteita. Ala on liian monimutkainen keskimääräisen biolääketieteen tutkijan hallittavaksi, ja alan keskeisin elementti, avoimen lähdekoodin ohjelmistokehitystyö, on aliarvostettu. Näihin seikkoihin tarvitaan useita parannuksia,After the human genome was sequenced in 2003, the major task of biosciences became to find out the functions of different genes, and various bioimaging techniques became crucial research tools. Technological advances resulted in especially fluorescence-based light microscopy techniques to explode in popularity, but microscopy needed to transform from a qualitative science to a quantitative one. This transition resulted in a new field of science, bioimage informatics, which has been said to have the potential to revolutionize biosciences. This thesis describes an extensive, multidisciplinary body of work in bioimage informatics. The first aim of the thesis was to develop protocols for four-dimensional confocal microscopy of living cells, one of the fastest growing methods of bioimaging. Human collagen receptor α2β1 integrin, an important molecule in several physiological and pathological processes, was used as a case study. Clear visualizations of the integrins moving, clustering and going inside cells were obtained, but there were no tools to quantify the image data. The second aim of the thesis then became to develop computer software capable of such quantification. At the same time, bioimage informatics was born, and specialized software was the most critical requirement of the new field. The main result of this thesis therefore became BioImageXD, a novel open source software package for visualizing, processing and analyzing multidimensional bioimages. BioImageXD grew to become one of the largest and most versatile in its field. It was published in a special bioimage informatics issue of Nature Methods, and became well known and widely used. The third aim of the thesis was to apply the developed methods to something more practical, and artificial silica nanoparticles were created, with antibodies against α2β1 integrin as "address labels". BioImageXD was successfully used to show that the nanoparticles have potential in targeted drug delivery applications. A major underlying aim of this thesis was to promote the new and unknown science of bioimage informatics, and this aim was reached especially through BioImageXD and its numerous published applications. The work done in this thesis has considerable future potential, but bioimage informatics faces serious challenges. The field is too complex for the average biomedical researcher to master, and there is lack of recognition for the most important element, open source software development. Several improvements in these aspects are required, or alternatively researchers should consider obtaining bioimage informatics services from designated units or companies.Siirretty Doriast

On the Complexity of Electrostatic Suspension Stabilization of Functionalized Silica Nanoparticles for Biotargeting and Imaging Applications

Different means of attaching streptavidin to surface functionalized silica particles with a diameter of 240 nm were investigated with special focus on suspension stability for electrostatically stabilized suspensions. The influence of two different fluorescent dyes covalently linked to the streptavidin on suspension stability was also studied. The results clearly show that the stability of the suspensions is crucially dependent on all functional groups present on the surface. The surface functions may either directly affect the effective surface charge if the functions contain charged groups, or indirectly by affecting the relative concentration of charged groups on the particle surface. Poly(ethylene imine)-functionalized silica particles, where the polymer is grown by surface hyperbranching polymerization, are shown to be promising candidates for bioapplications, as the zeta-potential can remain strongly positive even under biologically relevant conditions