Fine-tuning of the signalling network controlling morphogenesis and stem cell development in teeth

Tooth development is regulated by sequential and reciprocal interactions between epithelium and mesenchyme. The molecular mechanisms underlying this regulation are conserved and most of the participating molecules belong to several signalling families. Research focusing on mouse teeth has uncovered many aspects of tooth development, including molecular and evolutionary specifi cs, and in addition offered a valuable system to analyse the regulation of epithelial stem cells. In mice the spatial and temporal regulation of cell differentiation and the mechanisms of patterning during development can be analysed both in vivo and in vitro. Follistatin (Fst), a negative regulator of TGFβ superfamily signalling, is an important inhibitor during embryonic development. We showed the necessity of modulation of TGFβ signalling by Fst in three different regulatory steps during tooth development. First we showed that tinkering with the level of TGFβ signalling by Fst may cause variation in the molar cusp patterning and crown morphogenesis. Second, our results indicated that in the continuously growing mouse incisors asymmetric expression of Fst is responsible for the labial-lingual patterning of ameloblast differentiation and enamel formation. Two TGFβ superfamily signals, BMP and Activin, are required for proper ameloblast differentiation and Fst modulates their effects. Third, we identifi ed a complex signalling network regulating the maintenance and proliferation of epithelial stem cells in the incisor, and showed that Fst is an essential modulator of this regulation. FGF3 in cooperation with FGF10 stimulates proliferation of epithelial stem cells and transit amplifying cells in the labial cervical loop. BMP4 represses Fgf3 expression whereas Activin inhibits the repressive effect of BMP4 on the labial side. Thus, Fst inhibits Activin rather than BMP4 in the cervical loop area and limits the proliferation of lingual epithelium, thereby causing the asymmetric maintenance and proliferation of epithelial stem cells. In addition, we detected Lgr5, a Wnt target gene and an epithelial stem cell marker in the intestine, in the putative epithelial stem cells of the incisor, suggesting that Lgr5 is a marker of incisor stem cells but is not regulated by Wnt/β-catenin signalling in the incisor. Thus the epithelial stem cells in the incisor may not be directly regulated by Wnt/β-catenin signalling. In conclusion, we showed in the mouse incisors that modulating the balance between inductive and inhibitory signals constitutes a key mechanism regulating the epithelial stem cells and ameloblast differentiation. Furthermore, we found additional support for the location of the putative epithelial stem cells and for the stemness of these cells. In the mouse molar we showed the necessity of fi ne-tuning the signalling in the regulation of the crown morphogenesis, and that altering the levels of an inhibitor can cause variation in the crown patterning.Hampaiden kehitystä säätelee epiteelin ja mesenkyymin välinen vuorovaikutus. Molekyylien säätelemään vuorovaikutuksen tärkeimmät osallistujat kuuluvat muutamamaan yleiseen signalointimolekyyliperheeseen. Follistatin (Fst) on TGFβ signalointimolekyyliperheen negatiivinen säätelijä ja myös tärkeä inhibiittori kehityksen aikana. Työmme todisti, että Fst on välttämätön TGFβ perheen signaloinnin säätelijänä kolmessa eri vaiheessa hampaan kehitystä. Ensiksi osoitimme, että Fst säätelyn avulla on mahdollista muokata poskihampaiden nystermien rakentumista ja kruunun muodostumista. Toiseksi tuloksemme esittävät, että jatkuvasti kasvavien hiiren etuhampaiden epäsymmetrinen rakenne on Fst säätelemää. Fst, Activinin ja BMP:n vuorovaikutus johtaa ameloblastien erilaistumiseen ja kiilteen muodostumiseen ainoastaan labiaalipuolelle etuhammasta. Kolmanneksi tunnistimme monimutkaisen signalointiverkoston, joka säätelee etuhampaan epiteliaalisen kantasolupopulaation ylläpitoa ja osoitimme, että Fst on tämän verkoston välttämätön säätelijä. BMP4, FGF, Activin ja Fst vuorovaikutuksen seurauksena labiaalisen kervikaalisen silmun epiteliaalista kantasolupopulaatiota ylläpidetään aktiivisesti, kun taas linguaalipuolella kervikaalisen silmun epiteelisolujen proliferaatio on hyvin vähäistä. Näiden lisäksi tutkimme mahdollisen kantasolumarkkerigeenin, Lgr5, ekspressoitumista etuhampaan epiteeliaalisissa soluissa. Geenin ekspressoitumisalue ehdottaa sen olevan kantasolumarkkeri myös etuhampaassa. Tuloksemme osoittavat, että hiiren etuhampaan epäsymmetristä rakennetta säätelee indusoivien ja inhiboivien molekyylien verkosto. Hiiren poskihampaissa hienovarainen säätely taas muokkaa kruunun rakennetta ja jo pienet vaihtelut säätelygeenien tasoissa voivat saada aikaan suuria muutoksia rakenteeseen

An integrated gene regulatory network controls stem cell proliferation in teeth.

Epithelial stem cells reside in specific niches that regulate their self-renewal and differentiation, and are responsible for the continuous regeneration of tissues such as hair, skin, and gut. Although the regenerative potential of mammalian teeth is limited, mouse incisors grow continuously throughout life and contain stem cells at their proximal ends in the cervical loops. In the labial cervical loop, the epithelial stem cells proliferate and migrate along the labial surface, differentiating into enamel-forming ameloblasts. In contrast, the lingual cervical loop contains fewer proliferating stem cells, and the lingual incisor surface lacks ameloblasts and enamel. Here we have used a combination of mouse mutant analyses, organ culture experiments, and expression studies to identify the key signaling molecules that regulate stem cell proliferation in the rodent incisor stem cell niche, and to elucidate their role in the generation of the intrinsic asymmetry of the incisors. We show that epithelial stem cell proliferation in the cervical loops is controlled by an integrated gene regulatory network consisting of Activin, bone morphogenetic protein (BMP), fibroblast growth factor (FGF), and Follistatin within the incisor stem cell niche. Mesenchymal FGF3 stimulates epithelial stem cell proliferation, and BMP4 represses Fgf3 expression. In turn, Activin, which is strongly expressed in labial mesenchyme, inhibits the repressive effect of BMP4 and restricts Fgf3 expression to labial dental mesenchyme, resulting in increased stem cell proliferation and a large, labial stem cell niche. Follistatin limits the number of lingual stem cells, further contributing to the characteristic asymmetry of mouse incisors, and on the basis of our findings, we suggest a model in which Follistatin antagonizes the activity of Activin. These results show how the spatially restricted and balanced effects of specific components of a signaling network can regulate stem cell proliferation in the niche and account for asymmetric organogenesis. Subtle variations in this or related regulatory networks may explain the different regenerative capacities of various organs and animal species

An Integrated Gene Regulatory Network Controls Stem Cell Proliferation in Teeth

Epithelial stem cells reside in specific niches that regulate their self-renewal and differentiation, and are responsible for the continuous regeneration of tissues such as hair, skin, and gut. Although the regenerative potential of mammalian teeth is limited, mouse incisors grow continuously throughout life and contain stem cells at their proximal ends in the cervical loops. In the labial cervical loop, the epithelial stem cells proliferate and migrate along the labial surface, differentiating into enamel-forming ameloblasts. In contrast, the lingual cervical loop contains fewer proliferating stem cells, and the lingual incisor surface lacks ameloblasts and enamel. Here we have used a combination of mouse mutant analyses, organ culture experiments, and expression studies to identify the key signaling molecules that regulate stem cell proliferation in the rodent incisor stem cell niche, and to elucidate their role in the generation of the intrinsic asymmetry of the incisors. We show that epithelial stem cell proliferation in the cervical loops is controlled by an integrated gene regulatory network consisting of Activin, bone morphogenetic protein (BMP), fibroblast growth factor (FGF), and Follistatin within the incisor stem cell niche. Mesenchymal FGF3 stimulates epithelial stem cell proliferation, and BMP4 represses Fgf3 expression. In turn, Activin, which is strongly expressed in labial mesenchyme, inhibits the repressive effect of BMP4 and restricts Fgf3 expression to labial dental mesenchyme, resulting in increased stem cell proliferation and a large, labial stem cell niche. Follistatin limits the number of lingual stem cells, further contributing to the characteristic asymmetry of mouse incisors, and on the basis of our findings, we suggest a model in which Follistatin antagonizes the activity of Activin. These results show how the spatially restricted and balanced effects of specific components of a signaling network can regulate stem cell proliferation in the niche and account for asymmetric organogenesis. Subtle variations in this or related regulatory networks may explain the different regenerative capacities of various organs and animal species

Hoitopolun kehittäminen ennaltaehkäisevään päihdehoitotyöhön

Opinnäytetyön tarkoituksena oli luoda ennaltaehkäisevän päihdehoitotyön hoitopolku Haminan kaupungille. Hoitopolun tavoitteena oli parantaa päihdeasiakkaan selviytymistä omassa elämässään ja päihdehoitotyön piirissä, selkiyttää päihdehuollon toimintamallia ja eri ammattiryhmien työtä sekä turvata asiakkaan toimiva, asiakaslähtöinen hoito jo ongelmien synnyn varhaisvaiheessa. Työ on luonteeltaan kehittämistehtävä, jossa päihdetyön hoitopolku on laadittu teoreettisen viitekehyksen, Haminan A-klinikan palveluesimiehen teemahaastattelun sekä työelämän edustajien toiveiden pohjalta. Hoitopolun rakenne noudattaa hoitotyön prosessimallia. Teemahaastatteluaineisto analysoitiin käyttäen deduktiivista strukturoitua sisällön analyysia. Teemahaastattelun keskeisimpiä tulosten mukaan Haminan päihdepalveluiden kehittämisen tulisi kohdistua olemassa olevien rakenteiden vahvistamiseen ja jatkuvuuden turvaamiseen sekä yhteistyön edistämiseen eri toimijoiden välillä, sillä päihdepalvelut sellaisenaan on koettu toimivina ja kysyntään pystytään vastaamaan yksilöllisesti. A-klinikalle hakeutuvien asiakkaiden ongelmien vaikeusaste on hyvin vaihteleva ja päihdeongelman lisäksi myös muita ongelmia esiintyy laaja-alaisesti. Haminassa on suhteellisen paljon päihteidenkäyttäjiä, ja erityisesti mietojen huumausai-neiden käyttö on lisääntynyt. Uusimman haasteen muodostavat muuntohuumeet. A-klinikan rooli ennaltaehkäisevässä päihdehoitotyössä sijoittuukin lähinnä sekundaari- ja tertiaaritasoille. Suurin asiakasryhmä A-klinikalla ovat työikäiset. Teemahaastattelu on reliaabeli, sillä kaksi tutkijaa on päätynyt samoihin tulkintoihin aineistosta. Sitä voidaan pitää myös validina, koska haastattelu vastasi sille asetettuihin tavoitteisiin. Myös haastateltava luki teema-alueittain auki kirjoitetun haastatteluaineiston ja esitti siihen täsmennys- ja korjausehdotuksia, mikä lisää tulkinnan luotettavuutta. Kehittämistyön tuotos, valmis hoitopolku, on Haminan kaupungin edustajan mukaan tarkoitustaan vastaava. Se on kaksiosainen kaavio, jonka ensimmäisessä osassa kuvataan hoitotyön prosessi vaiheittain tarpeen määrittelystä arviointiin. Toisessa osassa Haminan päihdepalvelut yhteystietoineen on järjestetty asiakkaan ikäryhmän mukaisesti organisaatiokartaksi. Näin jokainen haminalainen saadaan ohjatuksi hakemaan apua tarkoituksenmukaisesta paikasta, eikä työntekijän aikaa kulu yhteystietojen etsimiseenThe purpose of this thesis was to produce a care pathway of preventive substance abuse nursing for the citizens of Hamina. The objective of the care pathway was to improve the coping abilities of substance abuse clients in their own lives and substance abuse nursing, clarify the approach of substance abuse nursing and the work of different occupational groups, as well as to ensure the client’s purpose-built and customer-oriented care at an early stage of problems. The approach of the thesis is a development task where building care pathway is based on theoretical frame of reference, the theme interview of Hamina´s A-clinic’s superior and the wishes of the commissioner. The structure of the care pathway follows nursing’s process model. The theme interview material was analyzed with deductive and structured content analysis. According to the results of the interview, the development of Hamina’s substance abuse care should focus on strengthening existing structures and ensuring continuity. Together with improving cooperation between different factors since substance abuse services are perceived workable and the demand can be met individually. The severity of the substance abuse problems of the clients who seek admittance to the A-clinic varies a lot and additional problems occur within a wide range. In Hamina they have a relatively high amount of substance abusers; especially mild drug usage has increased. Designer drugs are the current challenge. A-clinic’s role in preventing substance abuse is a secondary or tertiary stage of prevention. The widest client group in A-clinic is people of a working age. The theme interview was reliable as the two researchers ended up with similar interpretations. It can be deemed as valid when the interview corresponds to the objectives. Also, the interviewee read the themed interview material and made some suggestions which clarified and increased the reliability.According to the commissioner, the goals of the development task; finished care pathway, have achieved its purpose. It is a two-piece chart where the nursing process is described step by step at the first section of the chart. At the second part Hamina’s substance abuse services and their contact details is organized by age groups creating a organization map. This is how every citizen of Hamina will be advised to seek help from the appropriate places, eliminating the need for health care workers to have to spend time looking for contact detail