Unilateral zebrafish corneal injury induces bilateral cell plasticity supporting wound closure

The cornea, transparent and outermost structure of camera-type eyes, is prone to environmental challenges, but has remarkable wound healing capabilities which enables to preserve vision. The manner in which cell plasticity impacts wound healing remains to be determined. In this study, we report rapid wound closure after zebrafish corneal epithelium abrasion. Furthermore, by investigating the cellular and molecular events taking place during corneal epithelial closure, we show the induction of a bilateral response to a unilateral wound. Our transcriptomic results, together with our TGF-beta receptor inhibition experiments, demonstrate conclusively the crucial role of TGF-beta signaling in corneal wound healing. Finally, our results on Pax6 expression and bilateral wound healing, demonstrate the decisive impact of epithelial cell plasticity on the pace of healing. Altogether, our study describes terminally differentiated cell competencies in the healing of an injured cornea. These findings will enhance the translation of research on cell plasticity to organ regeneration.Peer reviewe

Zebrafish cornea formation and homeostasis reveal a slow maturation process, similarly to terrestrial vertebrates' corneas

Corneal blindness is the fourth leading cause of blindness worldwide. The superficial position of cornea on the eye makes this tissue prone to environmental aggressions, which can have a strong impact on sight. While most corneal pathology studies utilize terrestrial models, the knowledge on zebrafish cornea is too scarce to comprehend its strategy for the maintenance of a clear sight in aquatic environment. In this study, we deciphered the cellular and molecular events during corneal formation and maturation in zebrafish. After describing the morphological changes taking place from 3 days post fertilization (dpf) to adulthood, we analyzed cell proliferation. We showed that label retaining cells appear around 14 to 21dpf. Our cell proliferation study, combined to the study of Pax6a and krtt1c19e expression, demonstrate a long maturation process, ending after 45dpf. This maturation ends with a solid patterning of corneal innervation. Finally, we demonstrated that corneal wounding leads to an intense dedifferentiation, leading to the recapitulation of corneal formation and maturation, via a plasticity period. Altogether, our study deciphers the maturation steps of an aquatic cornea. These findings demonstrate the conservation of corneal formation, maturation and wound healing process in aquatic and terrestrial organisms, and they will enhance the use of zebrafish as model for corneal physiology studies.Peer reviewe

PIM-related kinases selectively regulate olfactory sensations in C. elegans

The mammalian PIM family of serine/threonine kinases regulate several cellular functions, such as cell survival and motility. Since PIM expression is observed in sensory organs, such as olfactory epithelium, we now wanted to explore the physiological roles of PIM kinascs there. As our model organism, we used the Caenorhabditis elegans nematodes, which express two PIM-related kinases, PRK-1 and PRK-2. We demonstrated PRKs to be true PIM orthologs with similar substrate specificity as well as sensitivity to PIM-inhibitory compounds. When we analysed the effects of pan-PIM inhibitors on C. elegans sensory functions, we observed that PRK activity is selectively required to support olfactory sensations to volatile repellents and attractants sensed by AWB and AWC(ON) neurons, respectively, but is dispensable for gustatory sensations. Analyses of prk-deficient mutant strains confirmed these findings and suggested that PRK-1, but not PRK-2 is responsible for the observed effects on olfaction. This regulatory role of PRK-1 is further supported by its observed expression in the head and tail neurons, including AWB and AWC neurons. Based on the evolutionary conservation of NM-related kinases, our data may have implications in regulation of also mammalian olfaction.Peer reviewe

Cold temperature represses daily rhythms in the liver transcriptome of a stenothermal teleost under decreasing day length

The climate-change-driven increase in temperature is occurring rapidly and decreasing the predictability of seasonal rhythms at high latitudes. It is therefore urgent to understand how a change in the relationship between photoperiod and temperature can affect ectotherms in these environments. We tested whether temperature affects daily rhythms of transcription in a cold-adapted salmonid using high-throughput RNA sequencing. Arctic char (Salvelinus alpinus) from a subarctic population were reared at a high and a low temperature (15 and 8 degrees C) for 1 month under natural, decreasing day length during late summer. Liver transcriptomes were compared between samples collected in the middle and towards the end of the light period and in the middle of the dark period. Daily variation in transcription was lower in fish from the low temperature compared with strong daily variation in warm-acclimated fish, suggesting that cold temperatures dampen the cycling of transcriptional rhythms under a simultaneously decreasing day length. Different circadian clock genes had divergent expression patterns, responding either by decreased expression or by increased rhythmicity at 15 degrees C compared with 8 degrees C. The results point out mechanisms that can affect the ability of fish to adapt to increasing temperatures caused by climate change

PIM-Related Kinases Selectively Regulate Olfactory Sensations in Caenorhabditis elegans

The mammalian PIM family of serine/threonine kinases regulate several cellular functions, such as cell survival and motility. Because PIM expression is observed in sensory organs, such as olfactory epithelium, we now wanted to explore the physiological roles of PIM kinases there. As our model organism, we used the Caenorhabditis elegans nematodes, which express two PIM-related kinases, PRK-1 and PRK-2. We demonstrated PRKs to be true PIM orthologs with similar substrate specificity as well as sensitivity to PIM-inhibitory compounds. When we analyzed the effects of pan-PIM inhibitors on C. elegans sensory functions, we observed that PRK activity is selectively required to support olfactory sensations to volatile repellents and attractants sensed by AWB and AWCON neurons, respectively, but is dispensable for gustatory sensations. Analyses of prk-deficient mutant strains confirmed these findings and suggested that PRK-1, but not PRK-2 is responsible for the observed effects on olfaction. This regulatory role of PRK-1 is further supported by its observed expression in the head and tail neurons, including AWB and AWC neurons. Based on the evolutionary conservation of PIM-related kinases, our data may have implications in regulation of also mammalian olfaction.</p

Zebrafish as a model for studying the maturation and pathophysiology of the corneal epithelium

The cornea is the transparent tissue on the eye surface, where light enters the ocular machinery. It has two main roles: light refraction, and protection of the inner eye components. Proper light transmittance requires corneal transparency, which can be compromised by injuries and corneal diseases. Understanding the physiology of the cornea in health and disease is thus important for proper treatment of corneal malfunction, and prevention of corneal blindness. Up to date, the majority of research relies on rodent models and cell-based systems. In this thesis, I studied the corneal formation and injury response in the zebrafish, which has many beneficial traits as a model organism, is known for its regenerative capacity, but has been somewhat overlooked in corneal studies. This study focused on the epithelium, the outermost cellular layer of the cornea. First, I delineated the epithelial maturation timeframe, by studying the cell appearance, proliferation, gene expression, and the innervation pattern in young fish. The analysis revealed two main phases: a morphogenetic period of circa three weeks, leading to epithelial stratification, and a maturation period that lasted up to later juvenile stages. These observations have implications for future corneal studies in young fish. In this thesis, I also applied a mechanical surface abrasion method to study the injury response on the zebrafish corneal epithelium. The results showed that a central corneal wound close within three hours in a process that does not necessitate proliferation. The data also revealed consistent downregulation of the gene pax6a on the migrating epithelium, and showed the importance of TGFβ signaling in the healing process. Due to the small amount of previous research on the zebrafish cornea, there is a lack of markers that would enable the identification of epithelial cell populations, and refined experimentation with genetic tools. In this work, I identified a type I keratin, krtt1c19e, as a potential marker for limbal stem/progenitor cells in the mature cornea. Overall, these results demonstrate the utility of the zebrafish in corneal studies, and open up new avenues for corneal research in this species.Sarveiskalvo on silmän etuosan läpinäkyvä kudos, jonka kautta valo kulkeutuu silmään. Sarveiskalvolla on kaksi tehtävää: se taittaa silmään tulevaa valoa, ja suojaa silmän sisempiä osia. Silmän pinnan vauriot ja sairaudet voivat johtaa sarveiskalvon samentumiseen ja näkökyvyn heikkenemiseen. Uusien hoitomuotojen kehitys edellyttää sekä terveen että vioittuneen kudoksen toiminnan tuntemista. Sarveiskalvon tutkimuksessa käytetään tällä hetkellä paljolti jyrsijöitä ja solumalleja. Tässä väitöskirjassa tutkin sarveiskalvon muotoutumista ja vauriovastetta seeprakalassa. Tällä lajilla on monia malliorganismille hyödyllisiä ominaisuuksia, ja huomattava potentiaali korjata kudosvaurioita, mutta toistaiseksi sitä on käytetty sarveiskalvon tutkimuksessa vähän. Tämän projektin kohteena oli sarveiskalvon uloin kerros, epiteeli. Aluksi tutkimme, millaisella aikajänteellä epiteeli muotoutuu alkionkehityksen jälkeen nuorissa kaloissa. Tuloksemme viittaavat kaksivaiheiseen kehitykseen, jossa ensimmäisen noin kolmen viikon kuluessa epiteeli saavuttaa tyypillisen muotonsa. Toinen vaihe, jonka aikana tapahtuu lopullinen kudoksen kypsyminen, kestää kuitenkin vielä useita viikkoja. Nämä tulokset ovat hyödyllisiä tuleville, nuorten kalojen sarveiskalvoa käsitteleville tutkimuksille. Sovelsimme tässä projektissa myös pintahaavametodia, jonka avulla tutkimme epiteelin vauriovastetta. Tulostemme mukaan sarveiskalvon keskialueelle tehty haava sulkeutui kolmessa tunnissa, ilman solujakautumista. Tuloksemme osoittivat myös, että geenin pax6 ilmeneminen alentui vaurioituneessa kudoksessa, ja että TGFβ-välitteinen signalointi on edellytys haavan nopealle sulkeutumiselle. Seeprakalan sarveiskalvon geeniekspressiota on toistaiseksi tutkittu vähän. Tässä projektissa havaitsimme, että tyypin I keratiinia koodaava geeni krtt1c19e ilmenee aikuisissa kaloissa kantasolut sisältävällä alueella, limbuksessa. Täten kyseinen geeni merkinnee kanta- tai esiastesoluja seeprakalan sarveiskalvon epiteelissä. Tulevaisuudessa tätä löydöstä voidaan hyödyntää tutkittaessa seeprakalan sarveiskalvon uusiutumista yksityiskohtaisemmin. Kokonaisuudessaan tuloksemme avaavat uusia mahdollisuuksia lisätutkimukselle, ja osoittavat, että seeprakala soveltuu sarveiskalvon tutkimusmalliksi

Basware FPM-järjestelmän käyttöönotto : Case: Isku-Yhtymä Oy

Kvalitatiivisen tutkielman tavoitteena oli selvittää, kuinka toteuttaa Isku-Yhtymä Oy:lle Basware Financial Performance Management -järjestelmän luotettava käyttöönotto. FPM-järjestelmä on laaja ohjelmistokokonaisuus, joten prosessia tarkasteltiin tasekirjan ja rahoituslaskelman tuottamisen näkökulmasta. Työn teoreettisen viitekehyksen ensimmäisessä kappaleessa perehdytään ulkoiseen laskentatoimeen konsernitasolla sekä tasekirjan ja rahoituslaskelman tuottamiseen. Toisessa kappaleessa tutustutaan sähköiseen taloushallintoon, tuodaan esiin tunnetuimpia talousohjausjärjestelmiä sekä käsitellään järjestelmän käyttöönottoprojektin toteutusta. Työn empiirinen osio alkaa toimeksiantoyrityksen ja Basware FPM-järjestelmän esittelyllä, jonka jälkeen pureudutaan tutkimustuloksiin ja tehtyihin johtopäätöksiin. Tutkimuksen avulla saatiin selville käyttöönottoprojektin kannalta tärkeät työvaiheet sekä käsitys siitä, mitä toimenpiteitä luotettava FPMjärjestelmän käyttöönotto edellytti. Projektin onnistumisen kannalta tärkeimmiksi nousivat toimivan tilikartan rakentaminen sekä sisäisten erien täsmäytysraportin teko. Käyttöönottoprojektin aikana havaittiin myös tarve kirjallisille käyttöohjeille. Erityisesti ulkomaisten tytäryhtiöiden osalta, koska tallennettavien tietojen tulee olla samassa muodossa kuin emoyhtiöllä.The purpose of this thesis was to define how to execute reliable implementation of Basware’s Financial Performance Management (FPM) software. This case study was executed at Isku-Yhtymä Oy. FPM is extensive software, so its implementation at Isku was examined from the perspective of financial statements. The theoretical framework of this thesis explores the financial accounting of a consolidated company. More specifically, this section discusses how to produce financial statements and cash flow statements. The second part of the theoretical section explores digital financial management and financial management software. The second part also explores software implementation in general. The empirical part of the thesis begins by introducing Isku-Yhtymä Oy and the FPM software. The empirical part also includes interview results and conclusions. Based on the results of this thesis, all the important stages on software implementation were defined. In addition, the thesis describes the reliable implementation of Basware’s FPM. The software needs to have a functional account scheme and a separate reconciliation report. During the project, it became evident that written instructions are needed. This is particularly important for subsidiaries abroad because all saved data needs to be in the same format as the parent company’s data

Zebrafish Corneal Wound Healing : From Abrasion to Wound Closure Imaging Analysis

As the transparent surface of the eye, the cornea is instrumental for clear sight. Due to its location, this tissue is prone to environmental insults. Indeed, the eye injuries most frequently encountered clinically are those to the cornea. While corneal wound healing has been extensively studied in small mammals (i.e., mice, rats, and rabbits), corneal physiology studies have neglected other species, including zebrafish, despite zebrafish being a classic research model. This report describes a method of performing a corneal abrasion on zebrafish. The wound is performed in vivo on anesthetized fish using an ocular burr. This method allows for a reproducible epithelial wound, leaving the rest of the eye intact. After abrasion, wound closure is monitored over the course of 3 h, after which the wound is reepithelialized. By using scanning electron microscopy, followed by image processing, the epithelial cell shape, and apical protrusions can be investigated to study the various steps during corneal epithelial wound closure. The characteristics of the zebrafish model permit study of the epithelial tissue physiology and the collective behavior of the epithelial cells when the tissue is challenged. Furthermore, the use of a model deprived of the influence of the tear film can produce new answers regarding corneal response to stress. Finally, this model also allows the delineation of the cellular and molecular events involved in any epithelial tissue subjected to a physical wound. This method can be applied to the evaluation of drug effectiveness in preclinical testing.Peer reviewe

Toimintatuokioiden suunnittelu muistinvirkistysmateriaalipakettiin

Opinnäytetyön toiminnallinen osuus tehtiin toimeksiantona Jyväskylän seudun Dementiayhdistys ry:n hallinnoimaan Kotihoitoa tukevaa kuntoutusta Keski-Suomen kuntiin -projektiin. Projektin yhtenä osa-alueena on muistinvirkistysmateriaalipaketin valmistus, joka on suunnattu etenevää muistisairautta sairastavien ryhmien ohjaajille ja sairastuneiden omaisille. Muistisairaiden lisäksi sitä voi käyttää myös muiden ikääntyneiden kanssa. Opinnäytetyön toiminnallisena osuutena suunniteltiin 29 toimintatuokiota muistinvirkistysmateriaalipaketin 22 teemaan. Toimintatuokioiden kohderyhmänä ovat lievää ja keskivaikeaa muistisairautta sairastavien ryhmät. Toimintatuokiot suunniteltiin käytännön työvälineeksi auttamaan ja helpottamaan muistisairaiden ryhmien ohjaajien työtä. Toiminnat analysoitiin ja muokattiin toimintaterapiassa käytettävän kognitiivis-hahmotuksellisen viitekehyksen mukaisesti tukien jäljellä olevien kykyjen käyttöä. Opinnäytetyön tietoperusta perustuu tietoon etenevistä muistisairauksista, niiden vaikutuksesta ihmisen toimintakykyyn sekä toimintaterapian teorioihin toiminnan merkityksestä ihmiselle. Toimintatuokiot pyrittiin suunnittelemaan mahdollisimman monipuolisiksi käyttäen erilaisia toiminnallisia menetelmiä, kuten kädentaitoja, luovia menetelmiä, muistelua ja aivovoimistelua. Toimintatuokioiden suunnittelussa huomioitiin muistihäiriöiden tuomat vaikeudet toimintakykyyn sekä pyrittiin tukemaan ja vireyttämään muistitoimintoja.The thesis was made as an assignment for project called The Home Care Supporting Rehabilitation to Central Finland's Communes which is managed by Jyväskylä region’s Dementia Association. One part of the project is to create a material package for memory refreshning. The material package is planned for the use of instructors who lead groups for persons with dementia and for the use of relatives of persons with dementia. It can also be used with other elderly people. The aim of the thesis was to plan 29 sessions to the 22 themes of the material package. The target group of the sessions is persons with mild or moderate dementia. Sessions were designed as a practical tool to assist and facilitate the work of instructors who lead dementia groups. Activities were analyzed and modified by cognitive-perceptual frame of reference which supports the use of remaining capabilities. The frame of reference is used in occupational therapy. The knowledge base of the thesis is based on the knowledge of dementia and its impact on persons’ functional capacity and also on the meaning of activities for people (based on the theories of occupational therapy). Sessions were planned versatile by using functional methods as hand crafts, creative methods and reminiscence. While planning the sessions, attention was paid to the difficulties that dementia causes and to the refreshning of memory functions