Коррекция состояния иммунной системы крыс с адъювантным артритом введением липидной фракции плаценты

Введення ліпідної фракції плаценти, отриманої методом кріогенного молекулярного фракціонування, на фоні розвитку ад’ювантного артриту оказує корегуючий вплив у відношенні як вмісту, так і функціональної активності регуляторних Т-клітин регіональних лімфовузлів, що призводить до зниження інтенсивності клінічних ознак захворювання.Injection of placental lipid fraction obtained by cryogenic molecular fractionation method on the background of adjuvant arthritis development has a correcting influence in relation to both content and functional activity of regulatory T-cells of regional lymph nodes which leads to a decrease in intensity of clinical signs of the disease

MKS1 regulates ciliary INPP5E levels in Joubert syndrome

Background Joubert syndrome (JS) is a recessive ciliopathy characterised by a distinctive brain malformation \u27the molar tooth sign\u27. Mutations in \u3e 27 genes cause JS, and mutations in 12 of these genes also cause Meckel-Gruber syndrome (MKS). The goals of this work are to describe the clinical features of MKS1- related JS and determine whether disease causing MKS1 mutations affect cellular phenotypes such as cilium number, length and protein content as potential mechanisms underlying JS. Methods We measured cilium number, length and protein content (ARL13B and INPP5E) by immunofluorescence in fibroblasts from individuals with MKS1-related JS and in a three-dimensional (3D) spheroid rescue assay to test the effects of diseaserelated MKS1 mutations. Results We report MKS1 mutations (eight of them previously unreported) in nine individuals with JS. A minority of the individuals with MKS1-related JS have MKS features. In contrast to the truncating mutations associated with MKS, all of the individuals with MKS1- related JS carry \u3e1 non-truncating mutation. Fibroblasts from individuals with MKS1-related JS make normal or fewer cilia than control fibroblasts, their cilia are more variable in length than controls, and show decreased ciliary ARL13B and INPP5E. Additionally, MKS1 mutant alleles have similar effects in 3D spheroids. Conclusions MKS1 functions in the transition zone at the base of the cilium to regulate ciliary INPP5E content, through an ARL13B-dependent mechanism. Mutations in INPP5E also cause JS, so our findings in patient fibroblasts support the notion that loss of INPP5E function, due to either mutation or mislocalisation, is a key mechanism underlying JS, downstream of MKS1 and ARL13B

Impaired SARS-CoV-2 specific T-cell response in patients with severe COVID-19

Cellular immune responses are of pivotal importance to understand SARS-CoV-2 pathogenicity. Using an enzyme-linked immunosorbent spot (ELISpot) interferon-γ release assay with wild-type spike, membrane and nucleocapsid peptide pools, we longitudinally characterized functional SARS-CoV-2 specific T-cell responses in a cohort of patients with mild, moderate and severe COVID-19. All patients were included before emergence of the Omicron (B.1.1.529) variant. Our most important finding was an impaired development of early IFN-γ-secreting virus-specific T-cells in severe patients compared to patients with moderate disease, indicating that absence of virus-specific cellular responses in the acute phase may act as a prognostic factor for severe disease. Remarkably, in addition to reactivity against the spike protein, a substantial proportion of the SARS-CoV-2 specific T-cell response was directed against the conserved membrane protein. This may be relevant for diagnostics and vaccine design, especially considering new variants with heavily mutated spike proteins. Our data further strengthen the hypothesis that dysregulated adaptive immunity plays a central role in COVID-19 immunopathogenesis

TCF21 hypermethylation regulates renal tumor cell clonogenic proliferation and migration

We recently identified hypermethylation at the gene promoter of transcription factor 21 (TCF21) in clear cell sarcoma of the kidney (CCSK), a rare pediatric renal tumor. TCF21 is a transcription factor involved in tubular epithelial development of the kidney and is a candidate tumor suppressor. As there are no in vitro models of CCSK, we employed a well-established clear cell renal cell carcinoma (ccRCC) cell line, 786-O, which also manifests high methylation at the TCF21 promoter, with consequent low TCF21 expression. The tumor suppressor function of TCF21 has not been functionally addressed in ccRCC cells; we aimed to explore the functional potential of TCF21 expression in ccRCC cells in vitro. 786-O clones stably transfected with either pBABE-TCF21-HA construct or pBABE vector alone were functionally analyzed. We found that ectopic expression of TCF21 in 786-O cells results in a trend toward decreased cell proliferation (not significant) and significantly decreased migration compared with mock-transfected 786-O cells. Although the number of colonies established in colony formation assays was not different between 786-O clones, colony size was significantly reduced in 786-O cells expressing TCF21. To investigate whether the changes in migration were due to epithelial-to-mesenchymal transition changes, we interrogated the expression of selected epithelial and mesenchymal markers. Although we observed upregulation of mRNA and protein levels of epithelial marker E-cadherin in clones overexpressing TCF21, this did not result in surface expression of E-cadherin as measured by fluorescence-activated cell sorting and immunofluorescence. Furthermore, mRNA expression of the mesenchymal markers vimentin (VIM) and SNAI1 was not significantly decreased in TCF21-expressing 786-O cells, while protein levels of VIM were markedly decreased. We conclude that re-expression of TCF21 in renal cancer cells that have silenced their endogenous TCF21 locus through hypermethylation results in reduced clonogenic proliferation, reduced migration, and reduced mesenchymal-like characteristics, suggesting a tumor suppressor function for transcription factor 21

Proteomic Signature of Mesenchymal Stromal Cell-Derived Small Extracellular Vesicles

Small extracellular vesicles (EVs) are 50–200 nm vesicles secreted by most cells. They are considered as mediators of intercellular communication, and EVs from specific cell types, in particular mesenchymal stem/stromal cells (MSCs), offer powerful therapeutic potential, and can provide a novel therapeutic strategy. They appear promising and safe (as EVs are non-self-replicating), and eventually MSC-derived EVs (MSC-EVs) may be developed to standardized, off-the-shelf allogeneic regenerative and immunomodulatory therapeutics. Promising pre-clinical data have been achieved using MSCs from different sources as EV-producing cells. Similarly, a variety EV isolation and characterization methods have been applied. Interestingly, MSC-EVs obtained from different sources and prepared with different methods show in vitro and in vivo therapeutic effects, indicating that isolated EVs share a common potential. Here, well-characterized and controlled, publicly available proteome profiles of MSC-EVs are compared to identify a common MSC-EV protein signature that might be coupled to the MSC-EVs’ common therapeutic potential. This protein signature may be helpful in developing MSC-EV quality control platforms required to confirm the identity and test for the purity of potential therapeutic MSC-EVs

Proteomic Signature of Mesenchymal Stromal Cell-Derived Small Extracellular Vesicles

Small extracellular vesicles (EVs) are 50–200 nm vesicles secreted by most cells. They are considered as mediators of intercellular communication, and EVs from specific cell types, in particular mesenchymal stem/stromal cells (MSCs), offer powerful therapeutic potential, and can provide a novel therapeutic strategy. They appear promising and safe (as EVs are non-self-replicating), and eventually MSC-derived EVs (MSC-EVs) may be developed to standardized, off-the-shelf allogeneic regenerative and immunomodulatory therapeutics. Promising pre-clinical data have been achieved using MSCs from different sources as EV-producing cells. Similarly, a variety EV isolation and characterization methods have been applied. Interestingly, MSC-EVs obtained from different sources and prepared with different methods show in vitro and in vivo therapeutic effects, indicating that isolated EVs share a common potential. Here, well-characterized and controlled, publicly available proteome profiles of MSC-EVs are compared to identify a common MSC-EV protein signature that might be coupled to the MSC-EVs’ common therapeutic potential. This protein signature may be helpful in developing MSC-EV quality control platforms required to confirm the identity and test for the purity of potential therapeutic MSC-EVs

Exosomes from hypoxic endothelial cells have increased collagen crosslinking activity through up-regulation of lysyl oxidase-like 2

Exosomes are important mediators of intercellular communication. Additionally, they contain a variety of components capable of interacting with the extracellular matrix (ECM), including integrins, matrix metalloproteinases and members of the immunoglobin superfamily. Despite these observations, research on exosome-ECM interactions is limited. Here, we investigate whether the exosome-associated lysyl oxidase family member lysyl oxidase-like 2 (LOXL2) is involved in ECM remodelling. We found that LOXL2 is present on the exterior of endothelial cell (EC)-derived exosomes, placing it in direct vicinity of the ECM. It is up-regulated twofold in EC-derived exosomes cultured under hypoxic conditions. Intact exosomes from hypoxic EC and LOXL2 overexpressing EC show increased activity in a fluorometric lysyl oxidase enzymatic activity assay as well as in a collagen gel contraction assay. Concordantly, knockdown of LOXL2 in exosome-producing EC in both normal and hypoxic conditions reduces activity of exosomes in both assays. Our findings show for the first time that ECM crosslinking by EC-derived exosomes is mediated by LOXL2 under the regulation of hypoxia, and implicate a role for exosomes in hypoxia-regulated focal ECM remodelling, a key process in both fibrosis and wound healing