Non-Invasive Assessment of Skin Surface Proteins of Psoriasis Vulgaris Patients in Response to Biological Therapy.

Measurements of skin surface biomarkers have enormous value for the detailed assessment of skin conditions, both for clinical application and in skin care. The main goals of the current study were to assess whether expression patterns of skin surface hBD-1, hBD-2, IL-1α, CXCL-1, and CXCL-8, examples of proteins known to be involved in psoriasis pathology, are associated with disease severity and whether expression patterns of these proteins on the skin surface can be used to measure pharmacodynamic effects of biological therapy. In this observational study using transdermal analysis patch (TAP), levels of skin surface IL-1α, hBD-1, hBD-2, CXCL-1/2, and CXCL-8 of psoriasis vulgaris (PV) patients over biological therapy were assessed. The Psoriasis Area Severity Index (PASI) and local score for erythema, induration, and desquamation were determined from the exact same skin area as FibroTx TAP measurements. Thirty-seven adult PV patients were included, of which twenty-three were subjected to anti-TNF-α, seven to anti-IL-17A, and seven to anti-IL12/IL-23 therapy. Significantly higher levels of hBD-1, hBD-2, CXCL-1/2, and CXCL-8 were detected on lesional skin compared to the non-lesional skin of the PV patients. In contrast, lower levels of IL-1α were found in lesional skin compared to non-lesional skin. In addition, we observed that the biomarker expression levels correlate with disease severity. Further, we confirmed that changes in the expression levels of skin surface biomarkers during biological therapy correlate with treatment response. Biomarker expression patterns in response to treatment differed somewhat between treatment subtypes. We observed that, in the case of anti-TNF-α therapy, an increase after a steady decrease in the expression levels of CXCL-1/2 and CXCL-8 occurred before the change in clinical scores. Moreover, response kinetics of skin surface proteins differs between the applied therapies-hBD2 expression responds quickly to anti-IL-17A therapy, CXCL-1/2 to anti-IL-12/23, and levels of CXCL-8 are rapidly down-regulated by IL-17A and IL-12/23 therapy. Our findings confirm that the skin surface hBD-2, IL-1α, CXCL-1/2, and CXCL-8 are markers for the psoriasis severity. Further, data obtained during this study give the basis for the conclusion that skin surface proteins CXCL-1/2 and CXCL-8 may have value as therapeutic biomarkers, thus confirming that measuring the 'molecular root' of inflammation appears to have value in scoring disease severity on its own.

Assessment of soluble skin surface protein levels for monitoring psoriasis vulgaris in adult psoriasis patients using non-invasive transdermal analysis patch: A pilot study.

To improve the care of patients with chronic inflammatory skin conditions, such as psoriasis, there is a need for diagnostic methods that can facilitate personalized medicine. This exploratory pilot study aimed to determine whether non-invasive measurements of inflammation-related proteins from psoriatic skin can be sampled using the FibroTx Transdermal Analysis Patch (TAP) to assess disease severity and monitor pharmacodynamic changes. Ten healthy volunteers and 44 psoriasis vulgaris patients were enrolled in the exploratory pilot study. Skin surface protein measurements for healthy and lesional skin were performed using TAP. Patients' scores of psoriasis activity and severity (PASI) were documented, and differences in the thickness of skin layers were determined using sonography. The study assessed the skin surface protein levels of psoriasis patients undergoing whole-body treatment with narrow-band UVB to evaluate whether the levels of the skin surface proteins IL-1α, IL-1RA CXCL-1/2, and hBD-1 were associated with the disease activity and severity measurements. Using TAP technology, it was observed that there were clear differences in levels of IL-1α, IL-1RA, CXCL-1/2, and hBD-1 between psoriasis lesional and non-lesional skin. In addition, a positive correlation between CXCL-1/2 and desquamation, and between CXCL-1/2 and SLEB thickness was observed. During UVB treatment, the TAP measurements revealed a clear reduction of IL-1RA, CXCL 1/2, and hBD-1 on lesional skin. Further, skin surface measurements of IL-1RA and CXCL-1/2 displayed a different profile than those achieved by visual scoring of local inflammation, thus indicating that measuring the 'molecular root' of inflammation appears to have value as an objective, non-invasive biomarker measurement for scoring disease severity.

Autoantibody screening of plasma and peritoneal fluid of patients with endometriosis.

STUDY QUESTION: Are there specific autoantibody profiles in patients with endometriosis that are different from those in controls? SUMMARY ANSWER: This study did not reveal a significantly higher prevalence of autoantibodies in the studied groups of patients. WHAT IS KNOWN ALREADY: Various inflammatory factors are postulated to be involved in the pathomechanisms of endometriosis, and a potential link exists with autoimmune diseases, which may also play an important role. As the diagnosis of endometriosis remains invasive, it can only be confirmed using laparoscopy with histopathological examination of tissues. Numerous studies have focused on identifying useful biomarkers to confirm the disease, but without unequivocal effects. Autoantibodies are promising molecules that serve as potential prognostic factors. STUDY DESIGN, SIZE, DURATION: A multicentre, cross-sectional study was conducted over 18 months (between 2018 and 2019), at eight Departments of Obstetrics and Gynaecology in several cities across Poland on 137 patients undergoing laparoscopic examination for the diagnosis of endometriosis. PARTICIPANTS/MATERIALS, SETTINGS, METHODS: During laparoscopy, we obtained plasma samples from 137 patients and peritoneal fluid (PF) samples from 98 patients. Patients with autoimmune diseases were excluded from the study. Autoantibody profiling was performed using HuProt v3.1 human proteome microarrays. MAIN RESULTS AND THE ROLE OF CHANCE: We observed no significant differences in the expression of autoantibodies in the plasma or PF between the endometriosis and control groups. The study revealed that in the PF of women with Stage II endometriosis, compared with other stages, there were significantly higher reactivity signals for ANAPC15 and GABPB1 (adj. P < 0.016 and adj. P < 0.026, respectively; logFC > 1 in both cases). Comparison of the luteal and follicular phases in endometriosis patients revealed that levels of NEIL1 (adj. P < 0.029), MAGEB4 (adj. P < 0.029), and TNIP2 (adj. P < 0.042) autoantibody signals were significantly higher in the luteal phase than in the follicular phase in PF samples of patients with endometriosis. No differences were observed between the two phases of the cycle in plasma or between women with endometriosis and controls. Clustering of PF and plasma samples did not reveal unique autoantibody profiles for endometriosis; however, comparison of PF and plasma in the same patient showed a high degree of concordance. LIMITATIONS, REASONS FOR CAUTION: Although this study was performed using the highest-throughput protein array available, it does not cover the entire human proteome and cannot be used to study potentially promising post-translational modifications. Autoantibody levels depend on numerous factors, such as infections; therefore the autoantibody tests should be repeated for more objective results. WIDER IMPLICATIONS OF THE FINDINGS: Although endometriosis has been linked to different autoimmune diseases, it is unlikely that autoimmune responses mediated by specific autoantibodies play a pivotal role in the pathogenesis of this inflammatory disease. Our study shows that in searching for biomarkers of endometriosis, it may be more efficient to use higher-throughput proteomic microarrays, which may allow the detection of potentially new biomarkers. Only research on such a scale, and possibly with different technologies, can help discover biomarkers that will change the method of endometriosis diagnosis. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by a grant from the Polish Ministry of Health (grant no. 6/6/4/1/NPZ/2017/1210/1352). It was also funded by the Estonian Research Council (grant PRG1076) and the Horizon 2020 Innovation Grant (ERIN; grant no. EU952516), Enterprise Estonia (grant no. EU48695), and MSCA-RISE-2020 project TRENDO (grant no. 101008193). The authors declare that there is no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.

Neuralized family member NEURL1 is a ubiquitin ligase for the cGMP-specific phosphodiesterase 9A.

Neuralized functions as a positive regulator of the Notch pathway by promoting ubiquitination of Notch ligands via its E3 ligase activity, resulting in their efficient endocytosis and signaling. Using a yeast two-hybrid screen, we have identified a cGMP-hydrolysing phosphodiesterase, PDE9A, as a novel interactor and substrate of Neuralized E3 ubiquitin protein ligase 1 (NEURL1). We confirmed this interaction with co-immunoprecipitation experiments and show that both Neuralized Homology Repeat domains of NEURL1 can interact with PDE9A. We also demonstrate that NEURL1 can promote polyubiquitination of PDE9A that leads to its proteasome-mediated degradation mainly via lysine residue K27 of ubiquitin. Our results suggest that NEURL1 acts as a novel regulator of protein levels of PDE9A.

Graphene-Augmented Nanofiber Scaffolds Trigger Gene Expression Switching of Four Cancer Cell Types.

Three-dimensional (3D) customized scaffolds are anticipated to provide new frontiers in cell manipulation and advanced therapy methods. Here, we demonstrate the application of hybrid 3D porous scaffolds, representing networks of highly aligned self-assembled ceramic nanofibers, for culturing four types of cancer cells. Ultrahigh aspect ratio (∼107) of graphene augmented fibers of tailored nanotopology is shown as an alternative tool to substantially affect cancerous gene expression, eventually due to differences in local biomechanical features of the cell-matrix interactions. Here, we report a clear selective up- and down-regulation of groups of markers for breast cancer (MDA-MB231), colorectal cancer (CaCO2), melanoma (WM239A), and neuroblastoma (Kelly) depending on only fiber orientation and morphology without application of any other stimulus. Changes in gene expression are also revealed for Mitomycin C treatment of MDA-MB231, making the scaffold a suitable platform for testing of anticancer agents. This allows an opportunity for selective "clean" guidance to a deep understanding of mechanisms of cancer cells progressive growth and tumor formation without possible side effects by manipulation with the specific markers.

Hybrid graphene-ceramic nanofibre network for spontaneous neural differentiation of stem cells.

A challenge in regenerative medicine is governed by the need to have control over the fate of stem cells that is regulated by the physical and chemical microenvironment in vitro and in vivo. The differentiation of the stem cells into specific lineages is commonly guided by use of specific culture media. For the first time, we demonstrate that human mesenchymal stem cells are capable of turning spontaneously towards neurogenic lineage when seeded on graphene-augmented, highly anisotropic ceramic nanofibres without special differentiation media, contrary to commonly thought requirement of 'soft' substrates for the same purpose. Furthermore, pro-inflammatory gene expression is simultaneously suppressed, and expression of factors promoting focal adhesion and monocytes taxis is upregulated. This opens new possibilities of using local topo-mechanical cues of the 'graphenized' scaffold surfaces to guide stem cell proliferation and differentiation, which can be used in studies of neurological diseases and cell therapy.

Prostaglandin D2 Receptor DP1 Antibodies Predict Vaccine-induced and Spontaneous Narcolepsy Type 1: Large-scale Study of Antibody Profiling.

BACKGROUND: Neuropathological findings support an autoimmune etiology as an underlying factor for loss of orexin-producing neurons in spontaneous narcolepsy type 1 (narcolepsy with cataplexy; sNT1) as well as in Pandemrix influenza vaccine-induced narcolepsy type 1 (Pdmx-NT1). The precise molecular target or antigens for the immune response have, however, remained elusive. METHODS: Here we have performed a comprehensive antigenic repertoire analysis of sera using the next-generation phage display method - mimotope variation analysis (MVA). Samples from 64 children and adolescents were analyzed: 10 with Pdmx-NT1, 6 with sNT1, 16 Pandemrix-vaccinated, 16 H1N1 infected, and 16 unvaccinated healthy individuals. The diagnosis of NT1 was defined by the American Academy of Sleep Medicine international criteria of sleep disorders v3. FINDINGS: Our data showed that although the immunoprofiles toward vaccination were generally similar in study groups, there were also striking differences in immunoprofiles between sNT1 and Pdmx-NT1 groups as compared with controls. Prominent immune response was observed to a peptide epitope derived from prostaglandin D2 receptor (DP1), as well as peptides homologous to B cell lymphoma 6 protein. Further validation confirmed that these can act as true antigenic targets in discriminating NT1 diseased along with a novel epitope of hemagglutinin of H1N1 to delineate exposure to H1N1. INTERPRETATION: We propose that DP1 is a novel molecular target of autoimmune response and presents a potential diagnostic biomarker for NT1. DP1 is involved in the regulation of non-rapid eye movement (NREM) sleep and thus alterations in its functions could contribute to the disturbed sleep regulation in NT1 that warrants further studies. Together our results also show that MVA is a helpful method for finding novel peptide antigens to classify human autoimmune diseases, possibly facilitating the design of better therapies.

Targeted alternative splicing of TAF4: a new strategy for cell reprogramming.

Reprogramming of somatic cells has become a versatile tool for biomedical research and for regenerative medicine. In the current study, we show that manipulating alternative splicing (AS) is a highly potent strategy to produce cells for therapeutic applications. We demonstrate that silencing of hTAF4-TAFH activity of TAF4 converts human facial dermal fibroblasts to melanocyte-like (iMel) cells. iMel cells produce melanin and express microphthalmia-associated transcription factor (MITF) and its target genes at levels comparable to normal melanocytes. Reprogramming of melanoma cells by manipulation with hTAF4-TAFH activity upon TAFH RNAi enforces cell differentiation towards chondrogenic pathway, whereas ectoptic expression of TAF4 results in enhanced multipotency and neural crest-like features in melanoma cells. In both cell states, iMels and cancer cells, hTAF4-TAFH activity controls migration by supporting E- to N-cadherin switches. From our data, we conclude that targeted splicing of hTAF4-TAFH coordinates AS of other TFIID subunits, underscoring the role of TAF4 in synchronised changes of Pol II complex composition essential for efficient cellular reprogramming. Taken together, targeted AS of TAF4 provides a unique strategy for generation of iMels and recapitulating stages of melanoma progression.

Graphene-augmented nanofiber scaffolds demonstrate new features in cells behaviour.

Three-dimensional (3D) customized scaffolds capable to mimic a native extracellular matrix open new frontiers in cells manipulation and advanced therapy. The major challenge is in a proper substrate for in vitro models on engineered scaffolds, capable to modulate cells differentiation. Here for the first time we demonstrate novel design and functionality of the 3D porous scaffolds of aligned, self-assembled ceramic nanofibers of ultra-high anisotropy ratio (~10(7)), augmented into graphene shells. This unique hybrid nano-network allows an exceptional combination of selective guidance stimuli of stem cells differentiation, immune reactions variations, and local immobilization of cancer cells, which was not available before. The scaffolds were shown to be able to direct human mesenchymal stem cells (important for stimulation of neuronal and muscle cells) preferential orientation, to suppress major inflammatory factors, and to localize cancer cells; all without additions of specific culture media. The selective downregulation of specific cytokines is anticipated as a new tool for understanding of human immune system and ways of treatment of associated diseases. The effects observed are self-regulated by cells only, without side effects, usually arising from use of external factors. New scaffolds may open new horizons for stem cells fate control such as towards axons and neurites regeneration (Alzheimer's disease) as well as cancer therapy development.

GLI2 cell-specific activity is controlled at the level of transcription and RNA processing: Consequences to cancer metastasis.

High activity of GLI family zinc finger protein 2 (GLI2) promotes tumor progression. Removal of the repressor domain at the N terminus (GLI2∆N) by recombinant methods converts GLI2 into a powerful transcriptional activator. However, molecular mechanisms leading to the formation of GLI2∆N activator proteins have not been established. Herein we report for the first time that the functional activities of GLI2 are parted into different protein isoforms by alternative promoter usage, selection of alternative splicing, transcription initiation and termination sites. Functional studies using melanoma cells revealed that transcriptional regulation of GLI2 is TGFbeta-dependent and supports the predominant production of GLI2∆N and C-terminally truncated GLI2 (GLI2∆C) isoforms in cells with high migratory and invasive phenotype. Taken together, these results highlight the role of transcription and RNA processing as major processes in the regulation of GLI2 activity with severe impacts in cancer development.

TAF4 controls differentiation of human neural progenitor cells through hTAF4-TAFH activity.

Expression of general transcription factor and co-activator TAF4 varies during development and in the processes of cell differentiation with suggested connection to neurodegenerative diseases. Here, we show that expression of TAF4 alternative splice variants is different in various regions of the human brain, substantiating the role of alternative splicing of TAF4 in the regulation of neural development and brain function. Most of the described splicing events affect the TAFH homology domain of TAF4 (hTAF4-TAFH). Besides, differentiated towards neural lineages, normal human neural progenitors (NHNPs) lose canonical full-length TAF4 isoform. To study the effects of hTAF4-TAFH splicing on neuronal differentiation, we used RNAi approach to target hTAF4-TAFH-encoding domain in NHNPs. Results show that inactivation of hTAF4-TAFH domain accelerates differentiation of human neural progenitor cells. Conversely, enhanced expression of TAF4 suppresses differentiation and keeps neural progenitor cells in a stem cell-like state. Finally, we provide data on the involvement of TP53 and noncanonical WNT signaling pathways in mediating effects of TAF4 on neuronal differentiation. Overall, our data suggest that specific isoforms of TAF4 may selectively and efficiently control neurogenesis.

LXXLL peptide converts transportan 10 to a potent inducer of apoptosis in breast cancer cells.

Degenerate expression of transcription coregulator proteins is observed in most human cancers. Therefore, in targeted anti-cancer therapy development, intervention at the level of cancer-specific transcription is of high interest. The steroid receptor coactivator-1 (SRC-1) is highly expressed in breast, endometrial, and prostate cancer. It is present in various transcription complexes, including those containing nuclear hormone receptors. We examined the effects of a peptide that contains the LXXLL-motif of the human SRC-1 nuclear receptor box 1 linked to the cell-penetrating transportan 10 (TP10), hereafter referred to as TP10-SRC1LXXLL, on proliferation and estrogen-mediated transcription of breast cancer cells in vitro. Our data show that TP10-SRC1LXXLL induced dose-dependent cell death of breast cancer cells, and that this effect was not affected by estrogen receptor (ER) status. Surprisingly TP10-SRC1LXXLL severely reduced the viability and proliferation of hormone-unresponsive breast cancer MDA-MB-231 cells. In addition, the regulation of the endogenous ERα direct target gene pS2 was not affected by TP10-SRC1LXXLL in estrogen-stimulated MCF-7 cells. Dermal fibroblasts were similarly affected by treatment with higher concentrations of TP10-SRC1LXXLL and this effect was significantly delayed. These results suggest that the TP10-SRC1LXXLL peptide may be an effective drug candidate in the treatment of cancers with minimal therapeutic options, for example ER-negative tumors.

Protein kinase inhibitor SU6668 attenuates positive regulation of Gli proteins in cancer and multipotent progenitor cells.

Observations that Glioma-associated transcription factors Gli1 and Gli2 (Gli1/2), executers of the Sonic Hedgehog (Shh) signaling pathway and targets of the Transforming Growth Factor ß (TGF-ß) signaling axis, are involved in numerous developmental and pathological processes unveil them as attractive pharmaceutical targets. Unc-51-like serine/threonine kinase Ulk3 has been suggested to play kinase activity dependent and independent roles in the control of Gli proteins in the context of the Shh signaling pathway. This study aimed at investigating whether the mechanism of generation of Gli1/2 transcriptional activators has similarities regardless of the signaling cascade evoking their activation. We also elucidate further the role of Ulk3 kinase in regulation of Gli1/2 proteins and examine SU6668 as an inhibitor of Ulk3 catalytic activity and a compound targeting Gli1/2 proteins in different cell-based experimental models. Here we demonstrate that Ulk3 is required not only for maintenance of basal levels of Gli1/2 proteins but also for TGF-ß or Shh dependent activation of endogenous Gli1/2 proteins in human adipose tissue derived multipotent stromal cells (ASCs) and mouse immortalized progenitor cells, respectively. We show that cultured ASCs possess the functional Shh signaling axis and differentiate towards osteoblasts in response to Shh. Also, we demonstrate that similarly to Ulk3 RNAi, SU6668 prevents de novo expression of Gli1/2 proteins and antagonizes the Gli-dependent activation of the gene expression programs induced by either Shh or TGF-ß. Our data suggest SU6668 as an efficient inhibitor of Ulk3 kinase allowing manipulation of the Gli-dependent transcriptional outcome.

Variable expression of lineage regulators in differentiated stromal cells indicates distinct mechanisms of differentiation towards common cell fate.

Mesenchymal stem cells (MSCs) possess a multi-lineage differentiation capacity that makes them important players in the field of regenerative medicine. MSC populations derived from different tissues or donors have been shown to exhibit variable gene expression patterns. Further, it is widely acknowledged that MSC isolates are heterogeneous mixtures of cells at different developmental stages. However, the heterogeneity of expression of lineage regulators has not been linked to differentiation potential of different MSC populations towards mesenchymal lineages. Here, we analyzed variation of expression of differentiation markers across whole population and between single differentiating cells of multipotent stromal cell populations derived from adipose tissue (AdMSCs) and skin (FBs) of seven donors. The results of the analyses show that all cell populations exhibit similar differentiation potential towards adipocyte, osteoblast and chondrocyte lineages despite tissue type- and donor-specific variations of expression of differentiation-associated genes. Further, we detected variable expression of lineage regulators in individual differentiating cells. Together, our data indicate that single cells of stromal cell populations could use distinct molecular mechanisms to reach a common cell fate.

Development of TAP, a non-invasive test for qualitative and quantitative measurements of biomarkers from the skin surface.

BACKGROUND: The skin proteome contains valuable information on skin condition, but also on how skin may evolve in time and may respond to treatments. Despite the potential of measuring regulatory-, effector- and structural proteins in the skin for biomarker applications in clinical dermatology and skin care, convenient diagnostic tools are lacking. The aim of the present study was to develop a highly versatile and non-invasive diagnostic tool for multiplex measurements of protein biomarkers from the surface of skin. RESULTS: The Transdermal Analyses Patch (TAP) is a novel molecular diagnostic tool that has been developed to capture biomarkers directly from skin, which are quantitatively analyzed in spot-ELISA assays. Optimisation of protocols for TAP production and biomarker analyses makes TAP measurements highly specific and reproducible. In measurements of interleukin-1α (IL-1α), IL-1 receptor antagonist (IL-1RA) and human ß-defensin (hBD-1) from healthy skin, TAP appears far more sensitive than skin lavage-based methods using ELISA. No side-effects were observed using TAP on human skin. CONCLUSION: TAP is a practical and valuable new skin diagnostic tool for measuring protein-based biomarkers from skin, which is convenient to use for operators, with minimal burden for patients.

Alternative splicing targeting the hTAF4-TAFH domain of TAF4 represses proliferation and accelerates chondrogenic differentiation of human mesenchymal stem cells.

Transcription factor IID (TFIID) activity can be regulated by cellular signals to specifically alter transcription of particular subsets of genes. Alternative splicing of TFIID subunits is often the result of external stimulation of upstream signaling pathways. We studied tissue distribution and cellular expression of different splice variants of TFIID subunit TAF4 mRNA and biochemical properties of its isoforms in human mesenchymal stem cells (hMSCs) to reveal the role of different isoforms of TAF4 in the regulation of proliferation and differentiation. Expression of TAF4 transcripts with exons VI or VII deleted, which results in a structurally modified hTAF4-TAFH domain, increases during early differentiation of hMSCs into osteoblasts, adipocytes and chondrocytes. Functional analysis data reveals that TAF4 isoforms with the deleted hTAF4-TAFH domain repress proliferation of hMSCs and preferentially promote chondrogenic differentiation at the expense of other developmental pathways. This study also provides initial data showing possible cross-talks between TAF4 and TP53 activity and switching between canonical and non-canonical WNT signaling in the processes of proliferation and differentiation of hMSCs. We propose that TAF4 isoforms generated by the alternative splicing participate in the conversion of the cellular transcriptional programs from the maintenance of stem cell state to differentiation, particularly differentiation along the chondrogenic pathway.

CCL5/CCR1 axis regulates multipotency of human adipose tissue derived stromal cells.

Several potential clinical applications of stem cells rely on their capacity to migrate into sites of inflammation where they contribute to tissue regeneration processes. Inflammatory signals are partially mediated by chemokines acting via their receptors expressed on the target cells. Data concerning the repertoire and biological activities of chemokine receptors in human adipose tissue derived stromal cells (ADSCs) are limited. Here we show that CCR1 is one of the few chemokine receptors expressed in ADSCs at a high level. CCR1 expression varies in ADSCs derived from different donors. It sharply decreases in the early phase of ADSCs in vitro propagation, but further demonstrates relative stability. Expression of CCR1 positively correlates with expression of SOX2, OCT4 and NANOG, transcription factors responsible for maintenance of the stemness properties of the cells. We demonstrate that signaling via CCL5/CCR1 axis triggers migration of ADSCs, activates ERK and AKT kinases, stimulates NFκB transcriptional activity and culminates in increased proliferation of CCR1(+) cells accompanied with up-regulation of SOX2, OCT4 and NANOG expression. Our data suggest that chemokine signaling via CCR1 may be involved in regulation of stemness of ADSCs.

RNA-seq analysis reveals different dynamics of differentiation of human dermis- and adipose-derived stromal stem cells.

BACKGROUND: Tissue regeneration and recovery in the adult body depends on self-renewal and differentiation of stem and progenitor cells. Mesenchymal stem cells (MSCs) that have the ability to differentiate into various cell types, have been isolated from the stromal fraction of virtually all tissues. However, little is known about the true identity of MSCs. MSC populations exhibit great tissue-, location- and patient-specific variation in gene expression and are heterogeneous in cell composition. METHODOLOGY/PRINCIPAL FINDINGS: Our aim was to analyze the dynamics of differentiation of two closely related stromal cell types, adipose tissue-derived MSCs (AdMSCs) and dermal fibroblasts (FBs) along adipogenic, osteogenic and chondrogenic lineages using multiplex RNA-seq technology. We found that undifferentiated donor-matched AdMSCs and FBs are distinct populations that stay different upon differentiation into adipocytes, osteoblasts and chondrocytes. The changes in lineage-specific gene expression occur early in differentiation and persist over time in both AdMSCs and FBs. Further, AdMSCs and FBs exhibit similar dynamics of adipogenic and osteogenic differentiation but different dynamics of chondrogenic differentiation. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that stromal stem cells including AdMSCs and dermal FBs exploit different molecular mechanisms of differentiation to reach a common cell fate. The early mechanisms of differentiation are lineage-specific and are similar for adipogenic and osteogenic differentiation but are distinct for chondrogenic differentiation between AdMSCs and FBs.

Human dermal fibroblasts exhibit delayed adipogenic differentiation compared with mesenchymal stem cells.

Human dermal fibroblasts (FBs) express mesenchymal stem cell (MSC)-specific cell surface markers and differentiate into several cell types under appropriate conditions. Molecular mechanisms controlling the early stages of differentiation of dermal FBs and MSCs isolated from different sources have not been well studied. Here, we have analyzed the cell type-specific changes of adipose tissue-derived mesenchymal stem cells (AdMSCs) and dermal FBs in the process of differentiation into adipocytes and osteoblasts. Analysis of gene expression in the course of adipogenic differentiation of AdMSCs and FBs isolated from the same individuals revealed a time lag in the induction of adipogenesis-related genes in FBs compared with AdMSCs, a phenomenon not previously described. Further, preliminary evidence suggests that delayed adipogenesis of FBs is related to the delayed induction of preadipocyte transcription factor ZNF423 in FBs. These findings clearly show that AdMSCs and FBs have similar developmental potential but different molecular control mechanisms of initial stages of adipogenic differentiation.

N-terminally truncated BAF57 isoforms contribute to the diversity of SWI/SNF complexes in neurons.

The SWItch/Sucrose NonFermentable, a nucleosome remodeling complex (SWI/SNF) chromatin-remodelling complexes act upon the nucleosomal structure and regulate transcription, replication, repair of chromatin and splicing. In this study, we present evidence that human, mouse and rat genes encoding one of the SWI/SNF complex subunits, BAF57, undergo neuron-specific splicing of exons II, III and IV. Alternative splicing yields in at least three isoforms of BAF57 protein that have truncated N-termini (N-BAF57s). The transcripts encoding N-BAF57 isoforms are predominantly expressed in the nervous system. The biochemical fractionation data supported by the results of the co-immunoprecipitation analysis show that N-BAF57 isoforms associate into protein complexes together with Brg1, Brm, BAF155 and BAF170. Transient over-expression of N-BAF57 isoforms in non-neural cells affects the level of expression of certain neuron-restrictive silencer element-containing genes. Together these data suggest that neuronal isoforms of BAF57 contribute to functional SWI/SNF complexes regulating neurogenesis.