Papyrus Oxyrhynchus 840 : Käännös ja vertailu kanonisiin evankeliumeihin

Tämä pro gradu -tutkielma tarjoaa suomenkielisen käännöksen apokryfiselle Papyrus Oxyrhynchus 840 -evankeliumifragmentille (P.Oxy. 840). Käännös on tehty tätä tutkielmaa varten laaditusta käsikirjoitustekstin editiosta, joka on koottu aiempia P.Oxy. 840:n editioita hyödyntäen. Käännös on varustettu kommenteilla. P.Oxy. 840 sisältää kaksi osittain säilynyttä perikooppia, joista kumpaakaan ei tunneta muista lähteistä. Niistä ensimmäinen on varoituspuhe pahantekijöille koituvasta rangaistuksesta. Toisessa perikoopissa Pelastajaksi kutsuttu Jeesus-hahmo saapuu opetuslapsineen temppeliin, jossa fariseus-ylipappi tulee huomauttamaan, etteivät he ole puhdistautuneet oikein ennen temppeliin tuloa. Tästä muodostuu kiistakeskustelu, jossa ylipappi kuvaa vaaditun puhtausrituaalin. Pelastaja vastaa voi-huudoilla, joilla toteaa ylipapin rituaalin tuovan vain ulkoisen puhtauden eikä sisäistä. Käännöksen jälkeen tutkielmassa verrataan P.Oxy. 840:tä kanonisiin evankeliumeihin ja tehdään arvio siitä, mitkä niistä ovat toimineet lähteinä P.Oxy. 840:lle. Tekstissä on viitteitä Matteuksen, Luukkaan ja Johanneksen evankeliumien hyödyntämiseen. Näistä on saatu temppelikertomukselle kertomusyhteys ja rakenne, joita mukaillen on laadittu uusi evankeliumikertomus. Alun varoituspuheessa on viitteitä Matteuksen ja Luukkaan evankeliumien hyödyntämisestä. Evankeliumeita on todennäköisimmin hyödynnetty kirjoitusta laadittaessa muistinvaraisesti tarkoituksena saada aikaan niitä läheisesti muistuttava teksti. Tutkielmassa rajoitutaan kanonisten vaikutteiden tarkasteluun, mutta on ilmeistä, että P.Oxy. 840:n laatimisessa on hyödynnetty muutakin kuin kanonisista evankeliumeista peräisin olevaa materiaalia. P.Oxy. 840:n sisällöstä nousee esiin neljä piirrettä, jotka antavat viitteitä sen kirjoittajan teologisesta ajattelusta ja tekstin sosiaalisesta kontekstista: σωτήρ-sanan yksinomainen käyttö, synoptisista evankeliumeista saatu käsitys kuoleman jälkeen olevasta rangaistuksesta. Johanneksen evankeliumin hyödyntäminen sekä Raamatun kuvausta laajempi tuntemus juutalaisista puhtauskäytännöistä

A novel fed-batch based cultivation method provides high cell-density and improves yield of soluble recombinant proteins in shaken cultures

<p>Abstract</p> <p>Background</p> <p>Cultivations for recombinant protein production in shake flasks should provide high cell densities, high protein productivity per cell and good protein quality. The methods described in laboratory handbooks often fail to reach these goals due to oxygen depletion, lack of pH control and the necessity to use low induction cell densities. In this article we describe the impact of a novel enzymatically controlled fed-batch cultivation technology on recombinant protein production in <it>Escherichia coli </it>in simple shaken cultures.</p> <p>Results</p> <p>The enzymatic glucose release system together with a well-balanced combination of mineral salts and complex medium additives provided high cell densities, high protein yields and a considerably improved proportion of soluble proteins in harvested cells. The cultivation method consists of three steps: 1) controlled growth by glucose-limited fed-batch to OD₆₀₀~10, 2) addition of growth boosters together with an inducer providing efficient protein synthesis within a 3 to 6 hours period, and 3) a slow growth period (16 to 21 hours) during which the recombinant protein is slowly synthesized and folded. Cell densities corresponding to 10 to 15 g l^-1cell dry weight could be achieved with the developed technique. In comparison to standard cultures in LB, Terrific Broth and mineral salt medium, we typically achieved over 10-fold higher volumetric yields of soluble recombinant proteins.</p> <p>Conclusions</p> <p>We have demonstrated that by applying the novel EnBase^®Flo cultivation system in shaken cultures high cell densities can be obtained without impairing the productivity per cell. Especially the yield of soluble (correctly folded) proteins was significantly improved in comparison to commonly used LB, Terrific Broth or mineral salt media. This improvement is thought to result from a well controlled physiological state during the whole process. The higher volumetric yields enable the use of lower culture volumes and can thus significantly reduce the amount of time and effort needed for downstream processing or process optimization. We claim that the new cultivation system is widely applicable and, as it is very simple to apply, could widely replace standard shake flask approaches.</p

Analyses of regulatory CD4(+)CD25(+)FOXP3(+) T cells and observations from peripheral T cell subpopulation markers during the development of type 1 diabetes in children

Our aim was to study whether the aberrant amount or function of regulatory T cells is related to the development of type 1 diabetes (T1D) in children. We also set out to investigate the balance of different T cell subtype markers during the T1D autoimmune process. Treg cells were quantified with flow cytometric assay, and the suppression capacity was analysed with a carboxyfluorescein succinimidyl ester (CFSE)-based T cell suppression assay in children in various phases of T1D disease process and in healthy autoantibody-negative control children. The mRNA expression of different T cell subpopulation markers was analysed with real-time qPCR method. The proportion and suppression capacity of regulatory T cells were similar in seroconverted children at an early stage of beta cell autoimmunity and also in children with T1D when compared to healthy and autoantibody-negative children. Significant differences were observed in the mRNA expression of different T cell subpopulation markers in prediabetic children with multiple (2) autoantibodies and in children with newly diagnosed T1D when compared to the control children. In conclusion, there were no quantitative or functional differences in regulatory T cells between the case and control groups in any phase of the autoimmune process. Decreased mRNA expression levels of T cell subtype markers were observed in children with multiple islet autoantibodies and in those with newly diagnosed T1D, probably reflecting an exhaustion of the immune system after the strong immune activation during the autoimmune process or a generally aberrant immune response related to the progression of the disease.Peer reviewe

Tonsillar granuloma associated with hypogammaglobulinemia

Background Rare tonsillar granulomas may be caused for example by infections, malignancies or sarcoidosis. Granulomas also occur in inborn errors of immunity (IEI) such as common variable immunodeficiency (CVID) with B cell maturation defects and hypogammaglobulinemia. CVID shares various features with sarcoidosis and drug-induced secondary hypogammaglobulinemia; careful consideration of differential diagnosis between these conditions is warranted. Case presentation A 29-year-old female with epilepsy developed dysphagia, dyspnea and impaired exercise tolerance. Obstruction caused by swollen lingual tonsil and edema in the epiglottis and arytenoid mucosa were found. Lingual tonsil and epiglottis biopsies displayed non-necrotizing granulomas. There was no evidence of viral, bacterial, mycobacterial or fungal infections. Chest X-ray, computerized tomography of chest and ultrasound of neck and abdomen remained unremarkable. Positron emission tomography/computed tomography (PET/CT) showed laryngeal enhancement. Empiric antimicrobials combined with prednisolone were insufficient to control her disease. In immunological evaluation, the patient had normal counts of B and T cells. Proportions of CD27(+)memory B cells (30.3%) and IgD(-)IgM(-)CD27(+)switched memory B cells (7.2%; normal range 6.5-29.2%) were normal. Percentage of activated CD21(low)B cells was high (6.6%; normal range 0.6-3.5%). IgG (3.5 g/L; normal range 6.77-15.0 g/l) and all IgG subclass concentrations were low. Anti-polysaccharide responses were impaired, with 3/10 serotypes reaching a level of 0.35 mu g/ml after immunization with Pneumovax(R). The findings were consistent with hypogammaglobulinemia resembling CVID, possibly secondary to antiepileptic medication. Her dyspnea and dysphagia responded favorably to subcutaneous IgG and rituximab. Conclusions Tonsillar granulomas can be the presenting and only clinical feature of B cell deficiency, highlighting the diversity of symptoms and findings in primary or secondary immunodeficiencies.Peer reviewe

Tonsillar granuloma associated with hypogammaglobulinemia

Background Rare tonsillar granulomas may be caused for example by infections, malignancies or sarcoidosis. Granulomas also occur in inborn errors of immunity (IEI) such as common variable immunodeficiency (CVID) with B cell maturation defects and hypogammaglobulinemia. CVID shares various features with sarcoidosis and drug-induced secondary hypogammaglobulinemia; careful consideration of differential diagnosis between these conditions is warranted. Case presentation A 29-year-old female with epilepsy developed dysphagia, dyspnea and impaired exercise tolerance. Obstruction caused by swollen lingual tonsil and edema in the epiglottis and arytenoid mucosa were found. Lingual tonsil and epiglottis biopsies displayed non-necrotizing granulomas. There was no evidence of viral, bacterial, mycobacterial or fungal infections. Chest X-ray, computerized tomography of chest and ultrasound of neck and abdomen remained unremarkable. Positron emission tomography/computed tomography (PET/CT) showed laryngeal enhancement. Empiric antimicrobials combined with prednisolone were insufficient to control her disease. In immunological evaluation, the patient had normal counts of B and T cells. Proportions of CD27(+)memory B cells (30.3%) and IgD(-)IgM(-)CD27(+)switched memory B cells (7.2%; normal range 6.5-29.2%) were normal. Percentage of activated CD21(low)B cells was high (6.6%; normal range 0.6-3.5%). IgG (3.5 g/L; normal range 6.77-15.0 g/l) and all IgG subclass concentrations were low. Anti-polysaccharide responses were impaired, with 3/10 serotypes reaching a level of 0.35 mu g/ml after immunization with Pneumovax(R). The findings were consistent with hypogammaglobulinemia resembling CVID, possibly secondary to antiepileptic medication. Her dyspnea and dysphagia responded favorably to subcutaneous IgG and rituximab. Conclusions Tonsillar granulomas can be the presenting and only clinical feature of B cell deficiency, highlighting the diversity of symptoms and findings in primary or secondary immunodeficiencies.Peer reviewe

Analyses of regulatory CD4+CD25+FOXP3+ T cells and observations from peripheral T cell subpopulation markers during the development of type 1 diabetes in children

Our aim was to study whether the aberrant amount or function of regulatory T cells is related to the development of type 1 diabetes (T1D) in children. We also set out to investigate the balance of different T cell subtype markers during the T1D autoimmune process. Treg cells were quantified with flow cytometric assay, and the suppression capacity was analysed with a carboxyfluorescein succinimidyl ester (CFSE)-based T cell suppression assay in children in various phases of T1D disease process and in healthy autoantibody-negative control children. The mRNA expression of different T cell subpopulation markers was analysed with real-time qPCR method. The proportion and suppression capacity of regulatory T cells were similar in seroconverted children at an early stage of beta cell autoimmunity and also in children with T1D when compared to healthy and autoantibody-negative children. Significant differences were observed in the mRNA expression of different T cell subpopulation markers in prediabetic children with multiple (≥2) autoantibodies and in children with newly diagnosed T1D when compared to the control children. In conclusion, there were no quantitative or functional differences in regulatory T cells between the case and control groups in any phase of the autoimmune process. Decreased mRNA expression levels of T cell subtype markers were observed in children with multiple islet autoantibodies and in those with newly diagnosed T1D, probably reflecting an exhaustion of the immune system after the strong immune activation during the autoimmune process or a generally aberrant immune response related to the progression of the disease.</p

Heterozygous premature termination in zinc-finger domain of Kruppel-like factor 2 gene associates with dysregulated immunity

Kruppel-like factor 2 (KLF2) is a transcription factor with significant roles in development, maturation, differentiation, and proliferation of several cell types. In immune cells, KLF2 regulates maturation and trafficking of lymphocytes and monocytes. KLF2 participates in regulation of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kappa B) pathway. Although pulmonary arterial hypertension (PAH) related to KLF2 genetic variant has been suggested, genetic role of KLF2 associated with immune dysregulation has not been described. We identified a family whose members suffered from lymphopenia, autoimmunity, and malignancy. Whole exome sequencing revealed a KLF2 p.(Glu318Argfs*87) mutation disrupting the highly conserved zinc finger domain. We show a reduced amount of KLF2 protein, defective nuclear localization and altered protein-protein interactome. The phenotypically variable positive cases presented with B and T cell lymphopenia and abnormalities in B and T cell maturation including low naive T cell counts and low CD27(+)IgD(-)IgM(-) switched memory B cells. KLF2 target gene (CD62L) expression was affected. Although the percentage of (CD25(+)FOXP3(+), CD25(+)CD127(-)) regulatory T cells (Treg) was high, the naive Treg cells (CD45RA(+)) were absent. Serum IgG1 levels were low and findings in one case were consistent with common variable immunodeficiency (CVID). Transcription of NF-kappa beta pathway genes and p65/RelA phosphorylation were not significantly affected. Inflammasome activity, transcription of genes related with JAK/STAT pathway and interferon signature were also comparable to controls. Evidence of PAH was not found. In conclusion, KLF2 variant may be associated with familial immune dysregulation. Although the KLF2 deficient family members in our study suffered from lymphopenia, autoimmunity or malignancy, additional study cohorts are required to confirm our observations.Peer reviewe

Haploinsufficiency of A20 impairs protein–protein interactome and leads into caspase-8-dependent enhancement of NLRP3 inflammasome activation

Objectives TNFAIP3 encodes A20 that negatively regulates nuclear factor kappa light chain enhancer of activated B cells (NF-κB), the major transcription factor coordinating inflammatory gene expression. TNFAIP3 polymorphisms have been linked with a spectrum of inflammatory and autoimmune diseases and, recently, loss-of-function mutations in A20 were found to cause a novel inflammatory disease ‘haploinsufficiency of A20’ (HA20). Here we describe a family with HA20 caused by a novel TNFAIP3 loss-of-function mutation and elucidate the upstream molecular mechanisms linking HA20 to dysregulation of NF-κB and the related inflammasome pathway.Methods NF-κB activation was studied in a mutation-expressing cell line using luciferase reporter assay. Physical and close-proximity protein–protein interactions of wild-type and TNFAIP3 p.(Lys91*) mutant A20 were analysed using mass spectrometry. NF-κB -dependent transcription, cytokine secretion and inflammasome activation were compared in immune cells of the HA20 patients and control subjects.Results The protein–protein interactome of p.(Lys91*) mutant A20 was severely impaired, including interactions with proteins regulating NF-κB activation, DNA repair responses and the NLR family pyrin domain containing 3 (NLRP3) inflammasome. The p.(Lys91*) mutant A20 failed to suppress NF-κB signalling, which led to increased NF-κB -dependent proinflammatory cytokine transcription. Functional experiments in the HA20 patients’ immune cells uncovered a novel caspase-8-dependent mechanism of NLRP3 inflammasome hyperresponsiveness that mediated the excessive secretion of interleukin-1β and interleukin-18.Conclusions The current findings significantly deepen our understanding of the molecular mechanisms underlying HA20 and other diseases associated with reduced A20 expression or function, paving the way for future therapeutic targeting of the pathway.Peer reviewe

Herpes simplex virus 2 encephalitis in a patient heterozygous for a TLR3 mutation

Susceptibility to herpes simplex virus type 1 (HSV-1) encephalitis (HSE-1) in otherwise healthy individuals, in the course of primary infection, can be caused by single-gene inborn errors of Toll-like receptor 3 (TLR3) dependent, interferon (IFN)-alpha/beta-mediated immunity,(1,2) or by single-gene inborn errors of snoRNA31.(3) These variations underlie infections of the forebrain, whereas mutations of DBR1 underlie infections of the brainstem.(3) HSV-2 encephalitis (HSE-2) is typically observed in neonates, albeit also rarely in older children and adults.(4) Its manifestations include altered level of consciousness, cranial neuropathies or more extensive brainstem encephalitis, hemiparesis, hemisensory loss, and permanent neurologic deficit.(4) MRI in HSE-2 may show normal findings, nonspecific white matter, orbitofrontal, mesial temporal lobe, or brainstem lesions. Inborn errors of immunity underlying HSE-2 have not been described.Non peer reviewe