Kuntamuutoksen tekijät : Espoon vanhusten palvelujen kotihoidon esimiesten muutostuki -kehittämishankkeen loppuraportti

Vanhusten palveluihin kohdistuu nyt ja tulevaisuudessa isoja haasteita, kun väestö ikääntyy ja kotihoidon tarve kasvaa. Miten tehdä palveluja tehokkaammin ja hyödyntää uutta teknologiaa palvelutapahtumien raportoinnissa? Miten esimiehet voivat tukea työntekijöitä arjen haasteissa? Miten kotihoito luo uudenlaista kumppanuutta asiakkaiden ja heidän omaistensa kanssa? Näitä muutoshaasteita tutkittiin yhdessä Espoon kotihoidon esimiesten ja työntekijöiden kanssa.1

Single-cell characterization of leukemic and non-leukemic immune repertoires in CD8(+) T-cell large granular lymphocytic leukemia

T cell large granular lymphocytic leukemia (T-LGLL) is a rare lymphoproliferative disorder of mature, clonally expanded T cells, where somatic-activating STAT3 mutations are common. Although T-LGLL has been described as a chronic T cell response to an antigen, the function of the non-leukemic immune system in this response is largely uncharacterized. Here, by utilizing single-cell RNA and T cell receptor profiling (scRNA+TCR alpha beta-seq), we show that irrespective of STAT3 mutation status, T-LGLL clonotypes are more cytotoxic and exhausted than healthy reactive clonotypes. In addition, T-LGLL clonotypes show more active cell communication than reactive clones with non-leukemic immune cells via costimulatory cell-cell interactions, monocyte-secreted proinflammatory cytokines, and T-LGLL-clone-secreted IFN gamma. Besides the leukemic repertoire, the non-leukemic T cell repertoire in T-LGLL is also more mature, cytotoxic, and clonally restricted than in other cancers and autoimmune disorders. Finally, 72% of the leukemic T-LGLL clonotypes share T cell receptor similarities with their non-leukemic repertoire, linking the leukemic and non-leukemic repertoires together via possible common target antigens. Our results provide a rationale to prioritize therapies that target the entire immune repertoire and not only the T-LGLL clonotype. T cell large granular lymphocytic leukemia (T-LGLL) is a lymphoproliferative disorder involving clonally expanded T cell clones and is not fully understood. Here the authors show that the rest of the immune repertoire is interconnected with the T-LGLL clonotype(s) and is more mature, cytotoxic and clonally restricted than in other cancers and autoimmune disorders.Peer reviewe

Anti-COX-2 autoantibody is a novel biomarker of immune aplastic anemia

In immune aplastic anemia (IAA), severe pancytopenia results from the immune-mediated destruction of hematopoietic stem cells. Several autoantibodies have been reported, but no clinically applicable autoantibody tests are available for IAA. We screened autoantibodies using a microarray containing >9000 proteins and validated the findings in a large international cohort of IAA patients (n = 405) and controls (n = 815). We identified a novel autoantibody that binds to the C-terminal end of cyclooxygenase 2 (COX-2, aCOX-2 Ab). In total, 37% of all adult IAA patients tested positive for aCOX-2 Ab, while only 1.7% of the controls were aCOX-2 Ab positive. Sporadic non-IAA aCOX-2 Ab positive cases were observed among patients with related bone marrow failure diseases, multiple sclerosis, and type I diabetes, whereas no aCOX-2 Ab seropositivity was detected in the healthy controls, in patients with non-autoinflammatory diseases or rheumatoid arthritis. In IAA, anti-COX-2 Ab positivity correlated with age and the HLA-DRB1*15:01 genotype. 83% of the >40 years old IAA patients with HLA-DRB1*15:01 were anti-COX-2 Ab positive, indicating an excellent sensitivity in this group. aCOX-2 Ab positive IAA patients also presented lower platelet counts. Our results suggest that aCOX-2 Ab defines a distinct subgroup of IAA and may serve as a valuable disease biomarker.Peer reviewe

Single-cell characterization of leukemic and non-leukemic immune repertoires in CD8+ T-cell large granular lymphocytic leukemia

T cell large granular lymphocytic leukemia (T-LGLL) is a rare lymphoproliferative disorder of mature, clonally expanded T cells, where somatic-activating STAT3 mutations are common. Although T-LGLL has been described as a chronic T cell response to an antigen, the function of the non-leukemic immune system in this response is largely uncharacterized. Here, by utilizing single-cell RNA and T cell receptor profiling (scRNA+TCRαβ-seq), we show that irrespective of STAT3 mutation status, T-LGLL clonotypes are more cytotoxic and exhausted than healthy reactive clonotypes. In addition, T-LGLL clonotypes show more active cell communication than reactive clones with non-leukemic immune cells via costimulatory cell-cell interactions, monocyte-secreted proinflammatory cytokines, and T-LGLL-clone-secreted IFN gamma. Besides the leukemic repertoire, the non-leukemic T cell repertoire in T-LGLL is also more mature, cytotoxic, and clonally restricted than in other cancers and autoimmune disorders. Finally, 72% of the leukemic T-LGLL clonotypes share T cell receptor similarities with their non-leukemic repertoire, linking the leukemic and non-leukemic repertoires together via possible common target antigens. Our results provide a rationale to prioritize therapies that target the entire immune repertoire and not only the T-LGLL clonotype.</p

IFN-α with dasatinib broadens the immune repertoire in patients with chronic-phase chronic myeloid leukemia

In chronic myeloid leukemia (CML), combination therapies with tyrosine kinase inhibitors (TKIs) aim to improve the achievement of deep molecular remission that would allow therapy discontinuation. IFN-alpha is one promising candidate, as it has long-lasting effects on both malignant and immune cells. In connection with a multicenter clinical trial combining dasatinib with IFN-alpha in 40 patients with chronic-phase CML (NordCML007, NCT01725204), we performed immune monitoring with single-cell RNA and T cell receptor (TCR) sequencing (n = 4, 12 samples), bulk TCR beta sequencing (n = 13, 26 samples), flow cytometry (n = 40, 106 samples), cytokine analyses (n = 17, 80 samples), and ex vivo functional studies (n = 39, 80 samples). Dasatinib drove the immune repertoire toward terminally differentiated NK and CD8+ T cells with dampened functional capabilities. Patients with dasatinib-associated pleural effusions had increased numbers of CD8(+) recently activated effector memory T (Temra) cells. In vitro, dasatinib prevented CD3-induced cell death by blocking TCR signaling. The addition of IFN-alpha reversed the terminally differentiated phenotypes and increased the number of costimulatory intercellular interactions and the number of unique putative epitope-specific TCR clusters. In vitro IFN-alpha had costimulatory effects on TCR signaling. Our work supports the combination of IFN-alpha with TKI therapy, as IFN-alpha broadens the immune repertoire and restores immunological function.Peer reviewe

Yersinia pseudotuberculosis serotyyppi O:9 lipopolysakkaridin O-antigeenin biosynteesiä ohjaavan geeniklusterin kloonaaminen ja sekvensointi

Yersinia pseudotuberculosis on gramnegatiivinen bakteeri, jonka soluseinässä oleva ulkomembraani koostuu pääosin lipopolysakkaridista sekä fosfolipideistä ja proteiineista. Lipopolysakkaridi on tärkeä antigeeninen rakenne, joka koostuu lipidi A:sta, ydinoligosakkaridista sekä O-polysakkaridista eli O-antigeenista. O-antigeenisen osan rakenne vaihtelee hyvin paljon eri bakteereilla ja samankin bakteerilajin eri kannoilla, mihin perustuu bakteerikantojen jako eri serotyyppeihin. Tutkimuksen kohteena oli Yersinia pseudotuberculosis serotyypin O:9 lipopolysakkaridin O-antigeenin biosynteesiä ohjaavan geeniklusterin kloonaaminen ja sekvensointi. Kloonausvektorina käytettiin kosmidia, jolloin puhutaan kosmidikloonauksesta. Lyhyesti kuvattuna tämä tehtiin seuraavasti: Yersinia pseudotuberculosis O:9 genominen DNA digestoitiin Sau3AI-restriktioentsyymillä ja kosmidi pHC79 BamHI:llä. Tuotteet ligoitiin, pakattiin in vitro lambda-faagiin ja transduktoitiin E. coli LE 392 -soluihin. Kosmidikirjaston valmistuttua se seulottiin. Geeniklusterin geeneistä wzz-geenin sekvenssi on tiedossa, mikä on mahdollistanut spesifisen PCR- eli polymeraasiketjureaktiomenetelmän kehittämisen geeniklusterin identifioimiseksi. wzz-positiivisten kloonien sisältämät kosmidit eristettiin ja kosmidien inserttien ääripäät sekvensoitiin. Sekvensoinnista saatuja tuloksia verrattiin geenipankin Yersinia pseudotuberculosis -kantojen DNA-sekvensseihin ja varsin ajoissa heräsi epäilys siitä, että insertit sisältävät monta lyhyttä Sau3AI-digestiotuotetta, jotka ovat ligoituneet toisiinsa. Kosmidikirjasto tehtiin uudelleen. Suuri määrä genomista DNA:ta digestoitiin partiaalisesti ja tällä kertaa lyhyet Sau3AI-digestiotuotteet poistettiin sokerigradienttiajolla. Saadut pitkät genomiset DNA-fragmentit ligoitiin kosmidiin, pakattiin ja transduktoitiin E. coliin. Tämän seurauksena saatiin yli 20000 kloonin kosmidikirjasto. Positiivisten kloonien seulonta tulee olemaan seuraava tehtävä tässä projektissa.The aim of this research was to clone the O-antigen gene cluster of Yersinia pseudotuberculosis serotype O:9 and to perform a sequence analysis. The cloning vector was a cosmid, hence the cloning is called cosmid cloning. Briefly, Yersinia pseudotuberculosis O:9 genomic DNA was digested with restriction enzyme Sau3AI and cosmid pHC79 was digested with BamHI. Digestion products were directly ligated, packaged in vitro into a lambda-phage using a packaging extract and transduced into E. coli LE 392 cells. The created cosmid library was then screened for positive clones that carry the O-antigen gene cluster. The sequence of the wzz-gene of the gene cluster is known which made it possible to design a specific PCR (polymerase chain reaction) method for the identification of the presence of the gene cluster. wzz-positive cosmids were isolated and the extreme ends of the inserts in the cosmids were sequenced. The obtained sequences were compared to known genome sequences of Yersinia pseudotuberculosis strains in data banks. The results indicated that the constructed genomic library was of poor quality since inserts contained many short Sau3AI fragments ligated to each other. Therefore a new cosmid library was created. A large amount of genomic DNA was digested partially by Sau3AI and this time the short Sau3AI fragments were removed from the digest using sucrose gradient fractionation. The obtained long genomic DNA fragments were ligated to the cosmid vector, packaged and transduced into E. coli. This resulted in cosmid library with >20000 clones. The screening for positive clones will be the next task in the project