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Sofia Johansson

Publications and source records attributed to Sofia Johansson.

8 recordsLinked to original sources
PubMed2006

Validation of endogenous controls for quantitative gene expression analysis: application on brain cortices of human chronic alcoholics.

Real-time PCR is frequently used for gene expression quantification due to its methodological sensitivity and reproducibility. The gene expression is quantified by normalization to one or more reference genes, usually beta-actin (ACTB), glyceraldehyde-3-phosphate dehydrogenase (GAPD) or to ribosomal RNA (18S). However, different environmental or pathological conditions might also influence the expression of normalizing genes, which could severely skew the interpretation of quantitative results. This study evaluates whether 16 genes frequently used as endogenous controls in expression studies, can serve as such for comparison of human brain tissues of chronic alcoholics and control subjects. The prefrontal and motor cortices that are affected differently by chronic alcohol consumption were analyzed. The reference genes that have no or small differences in expression in alcoholics and control subjects, were found to be specific for each region: beta-actin (ACTB) and ribosomal large P0 (RPLP0) for the prefrontal cortex while importin 8 (IPO8) and RNA polymerase II (POLR2A) for the motor cortex. Four out of sixteen analyzed genes demonstrated significant differences in expression between alcoholics and controls: phosphoglycerate kinase (PGK1), hypoxanthine phosphoribosyl transferase (HPRT1) and peptidylprolyl isomerase A (PPIA) in the motor cortex and beta-2-microglobulin (B2M) in the prefrontal cortex. Our study demonstrates the importance of validation of endogenous control genes prior to real-time PCR analysis of human brain tissues. Prescribed and non-prescribed drugs, pathological or environmental conditions along with alcohol abuse may differentially influence expression of reference genes.

Adult↗
PubMed2006

LUMA (LUminometric Methylation Assay)--a high throughput method to the analysis of genomic DNA methylation.

Changes in genomic DNA methylation are recognized as important events in normal and pathological cellular processes, contributing both to normal development and differentiation as well as cancer and other diseases. Here, we report a novel method to estimate genome-wide DNA methylation, referred to as LUminometric Methylation Assay (LUMA). The method is based on combined DNA cleavage by methylation-sensitive restriction enzymes and polymerase extension assay by Pyrosequencing. The method is quantitative, highly reproducible and easy to scale up. Since no primary modification of genomic DNA, such as bisulfite treatment, is needed, the total assay time is only 6 h. In addition, the assay requires only 200-500 ng of genomic DNA and incorporates an internal control to eliminate the problem of varying amounts of starting DNA. The accuracy and linearity of LUMA were verified by in vitro methylated lambda DNA. In addition, DNA methylation levels were assessed by LUMA in DNA methyltransferase knock-out cell lines and after treatment with the DNA methyltransferase inhibitor (5-AzaCytidine). The LUMA assay may provide a useful method to analyze genome-wide DNA methylation for a variety of physiological and pathological conditions including etiologic, diagnostic and prognostic aspects of cancer.

Animals↗
PubMed2006

Natural killer cells and malaria.

Malaria, caused by the infection with parasites of the germs Plasmodium, is one of the three most important infectious diseases worldwide, along with tuberculosis and infection with human immunodeficiency virus. Natural killer (NK) cells are lymphocytes classically involved in the early defense against viral infections and intracytoplasmic bacterial infections and are also implicated during the course of tumor development and allogeneic transplantation. These cells display important cytotoxic activity and produce high levels of proinflammatory cytokines. In both mouse and human models of malaria, NK cells appear to be a major source of interferon-gamma during the early phase of infection. In humans, indirect signaling through monocytes/macrophages required to optimally stimulate NK cell activity. However, the in vivo functions of NK cells during malaria are still enigmatic, and many issues remain to be dissected, such as the molecular basis of the direct recognition of iRBCs by NK cells.

Animals↗
PubMed2005

NK cells: elusive players in autoimmunity.

Natural killer (NK) cells were once regarded as relatively simple cells of the innate immune system. However, they are now revealing themselves as multifunctional regulatory cells that are present throughout the body. The role of NK cells in autoimmunity is attracting increased attention, although the picture is clouded by a conflicting literature that presents disease-promoting as well as disease-protective roles. In this review, we discuss how NK cells might use these dual roles, and suggest that future studies should focus on the impact of the anatomical localization of NK cells as well as the cytokine environment in which NK cells act in individual autoimmune diseases.

Autoimmunity↗
PubMed2005

Natural killer cell education in mice with single or multiple major histocompatibility complex class I molecules.

The ability of murine NK cells to reject cells lacking self MHC class I expression results from an in vivo education process. To study the impact of individual MHC class I alleles on this process, we generated mice expressing single MHC class I alleles (K(b), D(b), D(d), or L(d)) or combinations of two or more alleles. All single MHC class I mice rejected MHC class I-deficient cells in an NK cell-dependent way. Expression of K(b) or D(d) conveyed strong rejection of MHC class I-deficient cells, whereas the expression of D(b) or L(d) resulted in weaker responses. The educating impact of weak ligands (D(b) and L(d)) was further attenuated by the introduction of additional MHC class I alleles, whereas strong ligands (K(b) and D(d)) maintained their educating impact under such conditions. An analysis of activating and inhibitory receptors in single MHC class I mice suggested that the educating impact of a given MHC class I molecule was controlled both by the number of NK cells affected and by the strength of each MHC class I-Ly49 receptor interaction, indicating that NK cell education may be regulated by a combination of qualitative and quantitative events.

Alleles↗
PubMed2005

Computational modeling of human natural killer cell development suggests a selection process regulating coexpression of KIR with CD94/NKG2A.

Natural killer cells fail to lyse target cells expressing sufficient levels of self MHC class I molecules, providing one mechanism to secure self tolerance. Inhibition of lysis is mediated by inhibitory receptors expressed by NK cells, such as the murine Ly49 receptors, human KIR receptors and CD94/NKG2A, expressed by both species. To ensure that most, if not all, NK cells express at least one inhibitory receptor for self MHC class I, selection processes have been postulated for murine NK cells regulating the number and identity of inhibitory receptors expressed by each cell. The presence of similar selection processes in human NK cells has not been demonstrated. In previous studies using mathematical modeling we have shown that, in the Ly49 system, the sequential model (in which gene expression and selection operate simultaneously) is most likely to explain the observed expression frequencies. We also predicted the parameters (such as receptor-ligand binding affinity levels) under which the models fit with the observed frequencies. This study aims to evaluate whether these models may be valid in the human system. Our data suggest that if selection operates during human NK cell development, it affects the co-expression of CD94/NKG2A and KIR rather than KIR expression alone, and is more likely to be governed by the two-step selection model.

Animals↗
PubMed2004

Loss or mismatch of MHC class I is sufficient to trigger NK cell-mediated rejection of resting lymphocytes in vivo - role of KARAP/DAP12-dependent and -independent pathways.

A prediction from the "missing self" hypothesis is that down-regulation of MHC class I on resting hematopoietic cells should be sufficient to make them susceptible to NK cell killing. Using a method enabling kinetic and quantitative assessments of NK cell-mediated rejection responses in vivo, we here show that resting hematopoietic cells from beta(2)-microglobulin-deficient (beta(2)m(-/-)) mice were rapidly rejected in unmanipulated C57BL/6 (B6) mice. In situations of allelic MHC class I mismatches rejection occurred but required longer time. beta(2)m(-/-) donor cells pre-activated with concanavalin A were more efficiently eliminated compared to resting cells, as were MHC(-) tumor cells. When recipient mice were pretreated with an IFN inducer to activate NK cells, rejection was also enhanced. The signaling adaptor KARAP/DAP12 was dispensable for rejection of beta(2)m(-/-) cells (lacking MHC) but critical for rejection of BALB/c cells (mismatched MHC) in unmanipulated B6 recipients. In contrast, B6 recipients with pre-activated NK cells rejected BALB/c cells in a KARAP/DAP12-independent fashion. Loss or mismatch of MHC class I in resting cells was thus sufficient to convey susceptibility to NK cell rejection. However, activation of the effector or the target enhanced rejection and shifted the balance between different signaling pathways involved.

Adaptor Proteins, Signal Transducing↗