Frederiksendalsgaard8877

The obligate intracellular bacterium, Chlamydia pneumoniae, has been identified as a risk factor for several chronic inflammatory diseases in addition to respiratory tract infections. The dissemination of C. pneumoniae from respiratory tract to secondary sites of infection occurs via infected monocyte / macrophage line cells, in which C. pneumoniae can persist as an antibiotic-refractory phenotype. To allow more detailed studies on the epithelium-monocyte/macrophage transition of the infection, new in vitro bioassays are needed. To this end, a coculture system with human continuous cell lines was established. Respiratory epithelial HL cells were infected with C. pneumoniae and THP-1 monocytes were added into the cultures at 67 h post infection. After a 5 h coculture, THP-1 cells were collected with a biotinylated HLA antibody and streptavidin-coated magnetic beads and C. pneumoniae genome copy numbers in THP-1 determined by quantitative PCR. The assay was optimized for cell densities, incubation time, THP-1 separation technique and buffer composition, and its robustness was demonstrated by a Z' value of 0.6. The mitogen-activated protein kinase (MAPK) inhibitors SP600125 (JNK inhibitor), SB203580 (p38 inhibitor) and FR180204 (ERK inhibitor) suppressed the transfer of C. pneumoniae from HL to THP-1 cells, making them suitable positive controls for the assay. Based on analysis of separate steps of the process, the MAPK inhibitors suppress the bacterial entry to THP-1 cells. The transfer of C. pneumoniae from epithelium to phagocytes represents a crucial step in the establishment of persistent infections by this pathogen, and the presented methods enables future studies to block this process by therapeutic means. AIMS Steroid receptor coactivator-1 (SRC-1) is a key coactivator for the efficient transcriptional activity of steroids in the regulation of hippocampal functions. However, the effect of SRC-1 on hippocampal memory processes remains unknown. Our aim was to investigate the roles of hippocampal SRC-1 in the consolidation and reconsolidation of contextual fear memory in mice. MAIN METHODS Contextual fear conditioning paradigm was constructed in adult male C57BL/6 mice to examine the fear learning and memory processes. Adeno-associated virus (AAV) vector-mediated RNA interference (RNAi) was infused into hippocampus to block hippocampal SRC-1 level. Immunofluorescent staining was used to detect the efficiency of transfection. MEDICA16 inhibitor High plus maze and open field test were used to determine anxiety and locomotor activity. Western blot analyses were used to detect the expression of SRC-1 and synaptic proteins in the hippocampus. KEY FINDINGS We first showed that the expression of SRC-1 was regulated by fear conditioning training in a time-dependent manner, and knockdown of SRC-1 impaired contextual fear memory consolidation without affecting innate anxiety or locomotor activity. In addition, hippocampal SRC-1 was also regulated by the retrieval of contextual fear memory, and downregulation of SRC-1 disrupted fear memory reconsolidation. Moreover, knockdown of SRC-1 reversed the increased GluR1 and PSD-95 levels induced by contextual fear memory retrieval. SIGNIFICANCE Our data indicate that hippocampal SRC-1 is required for the consolidation and reconsolidation of contextual fear memory, and SRC-1 may be a potential therapeutic target for mental disorders that are involved in hippocampal memory dysfunction. AIMS The depot-specific differences in lipidome of visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) reflect heterogeneity of white adipose tissue (WAT), which plays a central role in its distinct response to outside stimuli. However, the detailed lipidome of depot-specific WAT is largely unknown, especially the minor constitutes including phospholipid and sphingolipid. MATERIALS AND METHODS To investigate this field, we applied a high-coverage targeted lipidomics approach of VAT and SAT in male C57BL/6J mice to compare the basal level of their lipid profiles. Applying microarray and quantitative real-time polymerase chain reaction, we analyzed the transcriptome of twodepot-specific WAT and verified the differences in individual genes. KEY FINDINGS In total, 342 lipid species from 19 lipid classes were identified. Our results showed the composition of TAG and FFA were different in length of chain and saturation. Interestingly, low abundance phospholipid, sphingolipid and cardiolipin were significantly higher in SAT. Lipid correlation network analysis vindicated that TAG and phospholipid formed distinct subnet and had more connections with other lipid species. Enriched ontology analysis of gene screened from LIPID MAPS and microarray suggested the differences were mainly involved in lipid metabolism, insulin resistance and inflammatory response. SIGNIFICANCE Our comprehensive lipidomics and transcriptomics analyses revealed differences in lipid composition and lipid metabolism of two depot-specific WAT, which would offer new insights into the investigation of heterogeneity of visceral and subcutaneous white adipose tissue. T-cell based immunity is mediated through specific T cell receptor (TCR) recognition of a small antigenic peptide in complex with a host immune molecule, major histocompatibility complex (pMHC). The interaction of a TCR and its pMHC ligand is generally quite weak, degenerate and biophysically unfavorable. Yet, the resulting immune response is extremely effective, being both sensitive and specific. Recent observations indicate that the TCR is an anisotropic mechanosensor. The force sensed by TCR's recognition module is transmitted to the non-covalently associated signal transduction module. Multiple biophysical methods reveal that the molecular mechanism for TCR-pMHC interaction under force required to induce T cell signaling is linked to "catch bond" formation between a TCR and its cognate ligand pMHC. This kind of dynamic non-covalent bond actually increases the bond lifetime by deforming the molecule to make the interaction lock tighter. The key observation is that the more stimulatory the antigenic peptide, the more pronounced the catch bond and immune response.