In vitro analysis of osteogenic inhibition in non-union spinal fusion caused by nucleus pulposus cells.

Discectomy and spinal fusion is the gold standard for spinal surgery to relieve pain. However, fusion can be hindered for yet unknown reasons that lead to non-fusions with pseudo-arthrosis. Clinical observations indicate that presence of residual intervertebral disc (IVD) tissue might hinder the ossification. We hypothesize that BMP-antagonists are constantly secreted by IVD cells and potentially prevent the ossification process. Furthermore, L51P, the engineered BMP2 variant, stimulates osseo-induction of bone marrow-derived mesenchymal stem cells (MSC) by antagonizing BMP-inhibitors. Human MSCs, primary nucleus pulposus (NPC) and annulus pulposus cells (AFC) were isolated and expanded in monolayer cultures up to passage 3. IVD cells were seeded in 1.2% alginate beads (4Mio/mL) and separated by culture inserts from MSCs. MSCs were kept in 1:control medium, 2:osteogenic medium±alginate beads, 3:osteogenic medium+NPC (±L51P) and 4:osteogenic medium+AFC (±L51P) for 21 days. Relative gene expression of bone-related genes, alkaline phosphatase assay and histological staining were performed. Osteogenesis of MSCs was hindered as shown by reduced alizarin red staining in the presence of NPC. No such inhibition was observed if co-cultured with alginate only or in the presence of AFC. The results were confirmed on the RNA and protein level. Addition of L51Pto the co- cultures, however, induced mineralization of MSCs in presence of NPC. We demonstrated that NPC secrete BMP-antagonists that prevent osteogenesis of MSCs and L51P can antagonize BMP-antagonists and induce bone formation.

Investigation of bone inhibition in non-union spinal fusion caused by intervertebral disc cells in vitro

Introduction: Discectomy and spinal fusion is the gold standard for spinal surgery to relieve pain. However, fusion can be hindered for yet unknown reasons that lead to non-fusions with pseudo-arthrose. It is hence appealing to develop biomaterials that can enhance bone formation. Clinical observations indicate that presence of residual intervertebral disc (IVD) tissue might hinder the ossification. We hypothesize that BMP-antagonists are constantly secreted by IVD cells and potentially prevent the ossification process. Furthermore, L51P, the engineered BMP2 variant, stimulates osteoinduction of bone marrow-derived mesenchymal stem cells (MSC) by antagonizing BMP-inhibitors. Methods: Human MSCs, primary nucleus pulposus (NPC) and annulus pulposus cells (AFC) were isolated and expanded in monolayer cultures up to passage 3. IVD cells were seeded in 1.2% alginate beads (4Mio/mL) and separated by culture inserts from MSCs in a co-culture set-up. MSCs were kept in 1:control medium, 2:osteogenic medium+alginate control, 3:osteogenic medium+NPC (±L51P) and 4:osteogenic medium+AFC (±L51P) for 21 days. Relative gene expression of bone-related genes, Alkaline Phosphatase (ALP) assay and histological staining were performed. Results: Osteogenesis of MSCs was hindered as shown by reduced alizarin red staining in the presence of NPC. No such inhibition was observed if co-cultured with alginate only or in the presence of AFC. The results were confirmed on the RNA and protein level. Addition of L51P to the co-cultures induced mineralization of MSCs, however a reduced ALP was observed. Conclusion: We demonstrated that NPC secrete BMP-antagonists that prevent osteogenesis of MSCs and L51P can antagonize BMP-antagonists and induce bone formation.

Osteogenic differentiation of bone marrow stromal cells is hindered by the presence of intervertebral disc cells.

BACKGROUND Clinical observations indicate that the presence of nucleus pulposus (NP) tissue during spinal fusion hinders the rate of disc ossification. While the underlying mechanism remains unknown, this observation could be due to incomplete removal of NP cells (NPCs) that secrete factors preventing disc calcification, such as bone morphogenetic protein (BMP) antagonists including noggin and members of the DAN (differential screening selected gene aberrative in neuroblastoma) family. METHODS Monolayer human bone marrow-derived mesenchymal stem cells (MSCs) were cocultured withNPCs and annulus fibrosus cells (AFCs) embedded in alginate for 21 days. At the end of coculture, MSCs were stained for mineral deposition by alizarin red, and relative expression of bone-related genes [Runt-related transcription factor 2, (RUNX2), Osteopontin (OPN), and Alkaline phosphatase (ALP)] and ALP activity were analyzed. Relative expression of three BMP antagonists, chordin (CHRD), gremlin (GREM1), and noggin (NOG), was determined in primary human NPCs and AFCs. These cells were also stained for Gremlin and Noggin by immunocytochemistry. RESULTS Alizarin red staining showed that MSC osteogenesis in monolayer cultures was inhibited by coculture with NPCs or AFCs. ALP activity and RT-PCR analyses confirmed these results and demonstrated inhibition of osteogenesis of MSC in the presence of disc cells. NOG was significantly up-regulated in MSCs after coculture. Relative gene expression of intervertebral disc (IVD) cells showed higher expression of GREM1 in NPCs than in AFCs. CONCLUSIONS We show that primary IVD cells inhibit osteogenesis of MSCs. BMP inhibitors NOG, GREM1 and CHRD were expressed in IVD cells. GREM1 appears to be differentially expressed in NPCs and AFCs. Our results have implications for the design and development of treatments for non-union in spinal fusion.

Characterization of a murine type II sodium-phosphate cotransporter expressed in mammalian small intestine

An isoform of the mammalian renal type II Na/Pi-cotransporter is described. Homology of this isoform to described mammalian and nonmammalian type II cotransporters is between 57 and 75%. Based on major diversities at the C terminus, the new isoform is designed as type IIb Na/Pi-cotransporter. Na/Pi-cotransport mediated by the type IIb cotransporter was studied in oocytes of Xenopus laevis. The results indicate that type IIb Na/Pi-cotransport is electrogenic and in contrast to the renal type II isoform of opposite pH dependence. Expression of type IIb mRNA was detected in various tissues, including small intestine. The type IIb protein was detected as a 108-kDa protein by Western blots using isolated small intestinal brush border membranes and by immunohistochemistry was localized at the luminal membrane of mouse enterocytes. Expression of the type IIb protein in the brush borders of enterocytes and transport characteristics suggest that the described type IIb Na/Pi-cotransporter represents a candidate for small intestinal apical Na/Pi-cotransport.

Comprehensive copy number and gene expression profiling of the 17q23 amplicon in human breast cancer

The biological significance of DNA amplification in cancer is thought to be due to the selection of increased expression of a single or few important genes. However, systematic surveys of the copy number and expression of all genes within an amplified region of the genome have not been performed. Here we have used a combination of molecular, genomic, and microarray technologies to identify target genes for 17q23, a common region of amplification in breast cancers with poor prognosis. Construction of a 4-Mb genomic contig made it possible to define two common regions of amplification in breast cancer cell lines. Analysis of 184 primary breast tumors by fluorescence in situ hybridization on tissue microarrays validated these results with the highest amplification frequency (12.5%) observed for the distal region. Based on GeneMap'99 information, 17 known genes and 26 expressed sequence tags were localized to the contig. Analysis of genomic sequence identified 77 additional transcripts. A comprehensive analysis of expression levels of these transcripts in six breast cancer cell lines was carried out by using complementary DNA microarrays. The expression patterns varied from one cell line to another, and several overexpressed genes were identified. Of these, RPS6KB1, MUL, APPBP2, and TRAP240 as well as one uncharacterized expressed sequence tag were located in the two common amplified regions. In summary, comprehensive analysis of the 17q23 amplicon revealed a limited number of highly expressed genes that may contribute to the more aggressive clinical course observed in breast cancer patients with 17q23-amplified tumors.

Increased fibrinogen levels at diagnosis are associated with adverse outcome in patients with acute myeloid leukemia

Increased plasma fibrinogen levels are associated with shortened overall survival (OS) in some solid tumor types. In contrast, the prognostic significance of varying fibrinogen levels in acute myeloid leukemia (AML) at diagnosis is unknown. In this study, we assessed the prognostic significance of fibrinogen levels in AML patients. In a comprehensive retrospective single-center study, we determined the survival rates of 375 consecutive AML patients undergoing at least one cycle of intensive chemotherapy induction treatment. Patients were dichotomized between low (<4.1 g/L) and high fibrinogen levels (≥4.1 g/L) at diagnosis of AML before initiation of treatment. Subsequently, quartile ranges were applied to analyze the association of varying fibrinogen levels on survival. We observed that the rates of complete remission, early death, and admission to intensive care unit were equal in the low versus high fibrinogen group. However, OS was significantly better in the low fibrinogen group (27.3 vs 13.5 months; p = 0.0009) as well as progression-free survival (12.3 vs 7.8 months; p = 0.0076). This survival difference remained significant in the multivariate analysis (p = 0.003). Assessing quartiles of fibrinogen values, we further confirmed this observation. Our data suggest that high fibrinogen levels at diagnosis of AML are associated with unfavorable OS and progression-free survival but not with increased mortality during induction treatment.

Maan mahtavimmat hybridisankarit. Supersankarit ja sankaruus The Avengers –sarjakuvissa 1970-luvun Yhdysvalloissa.

Tutkielma tarkastelee The Avengers –sarjakuvien supersankareita ja sankaruutta osana amerikkalaisten sankaritarinoiden sekä –myyttien jatkumoa, kiinnittäen erityistä huomiota myös siihen, miten yhteiskunnassa tapahtuvat sosiaaliset ja kulttuuriset murrokset näkyvät aikansa sarjakuvissa. Supersankarit ovat vain yksi, varsin nuori sankaruuden muoto amerikkalaisen populaarikulttuurin historiassa, ja tutkimuksen tarkoitus onkin selvittää, mistä moderni supersankari on saanut inspiraationsa. Supersankari ei kuitenkaan ole yksiselitteinen, staattinen sankaruuden ilmentymä, vaan se elää ja muuttuu aikansa mukana siinä missä muukin kulttuuri. Tutkimuksen lähdeaineistona ovat kaikki Marvel Comicsin 1970-luvulla julkaisemat The Avengers –sarjakuvat, joita vuosikymmenen aikana ehdittiin julkaista 118 kappaletta. Huomion keskiössä ovat nimenomaan Marvelin supersankarit, joiden uniikki inhimillisyys ja samaistuttava epätäydellisyys aikanaan mullistivat käsityksiä supersankareista ja sankaruudesta ylipäänsä. Tutkielma sijoittuu Yhdysvaltoihin, jossa sarjakuvat alun perin julkaistiin, ja kiinnostuksen kohteena ovat nimenomaan amerikkalaista yhteiskuntaa ravistelleet mullistukset kyseisenä vuosikymmenenä. Pääpainotus tutkielmassa on sarjakuvien kytkeminen amerikkalaisen sankaruuden ja kylmän sodan konteksteihin. Mikään populaarikulttuurin ilmiö ei synny tyhjiössä, eivätkä näin ollen syntyneet Marvelin supersankaritkaan, vaan heidän juurensa voidaan nähdä klassisissa amerikkalaisissa sankaritarinoissa, uuden ajan menestystarinoissa sekä sosiaalisiin ja kulttuurisiin muutoksiin perustuvissa mahdollisuuksien tarinoissa. Myös kylmä sota sekä sen luoma epävarmuuden ja pelon ilmapiiri vaikuttivat siihen, millainen supersankari 1970-luvun sarjakuvissa seikkaili, ja millaiseksi hän vuosikymmenen kuluessa muuttui. The Avengers –sarjakuvien supersankarit voidaan nähdä niin kulttuurisen historiansa kuin aikansakin muovaamina hybrideinä. Ei ole vain yhtä sankarimyyttiä, josta Marvelin supersankarit ovat saaneet inspiraationsa, vaan heissä yhdistyy niin klassisten kuin modernienkin sankaritarinoiden piirteitä. Supersankarit heijastelevat myös vahvasti yhteiskunnan poliittisia, sosiaalisia ja kulttuurisia muutoksia, ottaen niistä oppia ja kasvaen niiden mukana.

A series of trials in the UK as part of the Ofcom TV white spaces pilot

“Copyright © [2014] IEEE. Reprinted from 1st International Workshop on Cognitive Cellular Systems 2014 . ISBN: 978-1-4799-4139-1 .This material is posted here with permission of the IEEE. Internal or personal use of this material is permitted. However, permission to reprint/republish this material for advertising or promotional purposes or for creating new collective works for resale or redistribution must be obtained from the IEEE by writing to pubs-permissions@ieee.org. By choosing to view this document, you agree to all provisions of the copyright laws protecting it.”

Some initial results and observations from a series of trials within the Ofcom TV white spaces pilot

“Copyright © [2015] IEEE. Reprinted from Vehicular Technology Conference (VTC Spring), 2015 IEEE 81st. ISBN: 978-1-4799-8088-8. This material is posted here with permission of the IEEE. Internal or personal use of this material is permitted. However, permission to reprint/republish this material for advertising or promotional purposes or for creating new collective works for resale or redistribution must be obtained from the IEEE by writing to pubs-permissions@ieee.org. By choosing to view this document, you agree to all provisions of the copyright laws protecting it.”

Miten matematiikkaa tulisi opettaa? Oppilaiden ajatuksia matematiikan opetuksesta

Tässä tutkimuksessa selvitettiin oppilaiden näkemyksiä hyvästä ja huonosta matematiikan opetuksesta. Tutkimuksen tarkoituksena oli selvittää, millaisia elementtejä oppilaat liittävät hyvään ja huonoon matematiikan tuntiin. Oppilaiden ajatusten perusteella pohdittiin myös kehitysideoita matematiikan opetukseen. Tutkimusjoukoksi valikoitui erään länsisuomalaisen keskisuuren koulun viides- ja kuudesluokkalaiset. Tutkimukseen osallistui yhteensä 37 oppilasta. Tutkimus toteutettiin empiirisenä tutkimuksena ja aihetta lähestyttiin kvalitatiivisesta näkökulmasta. Aineisto kerättiin käyttäen passiivista eläytymismenetelmää. Tutkittaville annettiin kirjallinen orientaatio eli kehyskertomus, ja eläytyminen tapahtui kirjoittamalla lyhyt kertomus aiheesta. Vastausten luokittelussa pohjana käytettiin didaktisen kolmion opetustapahtuman osa-alueita. Oppilaat liittivät hyvään ja huonoon matematiikan tuntiin vahvasti opettajan toiminnan. Tutkimustulosten mukaan opettajan tulee opettaa asiat selkeästi ja ymmärrettävästi. Pitkä opettajan pitämä teoriaosuus koettiin huonona asiana. Tutkimuksessa työrauha tuli esille sekä hyviä että huonoja tunteja kuvaavissa kertomuksissa. Kaikkien oppilaiden osallistuminen ja hyvä mieli nousivat myös esiin. Kirjoitelmista kävi ilmi, että matematiikan tehtävien tulisi haastaa oppilaat, heidän oman tasonsa mukaan. Matematiikan tunneilla asioita tulisi tehdä yhdessä toiminnallisesti. Tulosten perusteella näyttäisi siltä, että oppilaat arvostavat yhdistelmää, jossa toiminnallisuus ja uudet opetusmenetelmät ovat käytössä perinteisen opetuksen ja tehtävien tekemisen rinnalla. Opetustilanteiden tulisi olla vastavuoroisia, kuten myös opetuksen suunnittelun. Olisi tärkeää, että opettajilla olisi rohkeutta kysyä oppilailtaan opetukseen liittyvistä asioista ja toteuttaa oppilaiden ideoita.