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丙酮酸钠 BioXtra, ≥99%; CAS Number: 113-24-6; EC Number: 204-024-4; Synonyms: 2-氧代丙酸 钠盐,乙酰甲酸 钠盐,α-酮基丙酸 钠盐; Linear Formula: CH3COCOONa; find Sigma-Aldrich-P8574 MSDS, related peer-reviewed papers, technical documents, similar products & more at Sigma-Aldrich

丙酮酸钠 BioXtra, ≥99% | Sigma-Aldrich

丙酮酸钠

Biomarkers and clinical rating scales for sodium pyruvate therapy in patients with mitochondrial disease

Transcriptome and proteome analysis of soleus muscle of hormone-sensitive lipase-null mice

Addition of sodium pyruvate to stored red blood cells attenuates liver injury in a murine transfusion model

Embryonic development after follicle culture is influenced by follicle-stimulating hormone isoelectric point range

Imaging calcium concentration dynamics in small neuronal compartments

Identifying master regulators of biological processes and mapping their downstream gene networks are key challenges in systems biology. We developed a computational method, called iRegulon, to reverse-engineer the transcriptional regulatory network underlying a co-expressed gene set using cis-regulatory sequence analysis. iRegulon implements a genome-wide ranking-and-recovery approach to detect enriched transcription factor motifs and their optimal sets of direct targets. We increase the accuracy of network inference by using very large motif collections of up to ten thousand position weight matrices collected from various species, and linking these to candidate human TFs via a motif2TF procedure. We validate iRegulon on gene sets derived from ENCODE ChIP-seq data with increasing levels of noise, and we compare iRegulon with existing motif discovery methods. Next, we use iRegulon on more challenging types of gene lists, including microRNA target sets, protein-protein interaction networks, and genetic perturbation data. In particular, we over-activate p53 in breast cancer cells, followed by RNA-seq and ChIP-seq, and could identify an extensive up-regulated network controlled directly by p53. Similarly we map a repressive network with no indication of direct p53 regulation but rather an indirect effect via E2F and NFY. Finally, we generalize our computational framework to include regulatory tracks such as ChIP-seq data and show how motif and track discovery can be combined to map functional regulatory interactions among co-expressed genes. iRegulon is available as a Cytoscape plugin from http://iregulon.aertslab.org.

iRegulon: from a gene list to a gene regulatory network using large motif and track collections.

Polycomb Repressive Complex 1 and histone H2A ubiquitination (ubH2A) contribute to embryonic stem cell (ESC) pluripotency by repressing lineage-specific gene expression. However, whether active deubiquitination co-regulates ubH2A levels in ESCs and during differentiation is not known. Here we report that Usp16, a histone H2A deubiquitinase, regulates H2A deubiquitination and gene expression in ESCs, and importantly, is required for ESC differentiation. Usp16 knockout is embryonic lethal in mice, but does not affect ESC viability or identity. Usp16 binds to the promoter regions of a large number of genes in ESCs, and Usp16 binding is inversely correlated with ubH2A levels, and positively correlates with gene expression levels. Intriguingly, Usp16(-/-) ESCs fail to differentiate due to ubH2A-mediated repression of lineage-specific genes. Finally, Usp16, but not a catalytically inactive mutant, rescues the differentiation defects of Usp16(-/-) ESCs. Therefore, this study identifies Usp16 and H2A deubiquitination as critical regulators of ESC gene expression and differentiation.

The histone H2A deubiquitinase Usp16 regulates embryonic stem cell gene expression and lineage commitment.

The human nasal epithelium is an important physical barrier, and a part of the innate immune defense that protect against pathogens. The epithelial cells recognize microbial components by pattern-recognition receptors (PRRs), and thereby trigger an immune response. Even though TLR3, TLR7, TLR9, RIG-I and MDA-5 are all known to respond to viral stimulation, their potential role in chronic airway inflammation triggered by local cytokine release remains to be established. mRNA and corresponding protein expression of TLR3, TLR7, TLR9, RIG-I and MDA-5 were analyzed in nasal biopsies and various upper airway epithelial cell lines using real-time reverse transcription PCR, immunohistochemistry and flow cytometry. Ligand induced, cytokine release, was evaluated with ELISA. Nasal biopsies were found to express TLR3, TLR7, TLR9, RIG-I and MDA-5, with the most abundant expression in the surface epithelium. These receptors were verified in primary human nasal epithelial cell (HNEC) as well as in the airway epithelial cell lines Detroit-562 and FaDu. Poly(I:C) (TLR3) and R-837 (TLR7) stimulation increased secretion of IL-6 and GM-CSF from the nasal mucosa and the epithelial cell lines. CpG (TLR9) stimulation caused release of IL-8 in the nasal mucosa and in FaDu. Poly(I:C)/LyoVec (RIG-I/MDA-5) stimulation activated the secretion of IFN-β in the nasal mucosa. A corresponding release was also detected from HNEC and Detroit-562. The nasal epithelium has the ability to recognize viral intrusion through TLR and RLR receptors, and the subsequent response might have a role in exacerbation of inflammatory diseases like allergic rhinitis and chronic rhinosinusitis.

Functional effects of Toll-like receptor (TLR)3, 7, 9, RIG-I and MDA-5 stimulation in nasal epithelial cells.

Targeted drug delivery with antibody-drug conjugates such as the HER2-directed ado-trastuzumab emtansine (T-DM1) has emerged as a powerful strategy for cancer therapy. We show that T-DM1 is particularly effective in eliciting antitumor immunity in patients with early breast cancer (WSG-ADAPT trial) and in a HER2-expressing orthotopic tumor model. In the latter, despite primary resistance to immunotherapy, combined treatment with T-DM1 and anti-CTLA-4/PD-1 (cytotoxic T lymphocyte-associated protein-4/programmed cell death protein-1) was curative because it triggered innate and adaptive immunity. Tumor rejection was accompanied by massive T cell infiltration, TH1 (T helper 1) cell polarization, and, notably, a substantial increase in regulatory T cells. Depletion of regulatory T cells resulted in inflammation and tissue damage, implying their essential role in protecting the host during therapy. This study provides insights into the mechanisms of T-DM1's therapeutic activity and a rationale for potential therapeutic combination strategies with immunotherapy.

Trastuzumab emtansine (T-DM1) renders HER2+ breast cancer highly susceptible to CTLA-4/PD-1 blockade.

Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.

A general method to improve fluorophores for live-cell and single-molecule microscopy.

The cytotoxic T lymphocyte antigen-4 (CTLA-4)-blocking antibody ipilimumab induces immune-mediated long-term control of metastatic melanoma in a fraction of patients. Although ipilimumab undoubtedly exerts its therapeutic effects via immunostimulation, thus far clinically useful, immunologically relevant biomarkers that predict treatment efficiency have been elusive. Here, we show that neutralization of IL-2 or blocking the α and β subunits of the IL-2 receptor (CD25 and CD122, respectively) abolished the antitumor effects and the accompanying improvement of the ratio of intratumoral T effector versus regulatory cells (Tregs), which were otherwise induced by CTLA-4 blockade in preclinical mouse models. CTLA-4 blockade led to the reduction of a suppressive CD4(+) T cell subset expressing Lag3, ICOS, IL-10 and Egr2 with a concomitant rise in IL-2-producing effector cells that lost FoxP3 expression and accumulated in regressing tumors. While recombinant IL-2 improved the therapeutic efficacy of CTLA-4 blockade, the decoy IL-2 receptor α (IL-2Rα, sCD25) inhibited the anticancer effects of CTLA-4 blockade. In 262 metastatic melanoma patients receiving ipilimumab, baseline serum concentrations of sCD25 represented an independent indicator of overall survival, with high levels predicting resistance to therapy. Altogether, these results unravel a role for IL-2 and IL-2 receptors in the anticancer activity of CTLA-4 blockade. Importantly, our study provides the first immunologically relevant biomarker, namely elevated serum sCD25, that predicts resistance to CTLA-4 blockade in patients with melanoma.

Anticancer immunotherapy by CTLA-4 blockade: obligatory contribution of IL-2 receptors and negative prognostic impact of soluble CD25.

Merosin-deficient congenital muscular dystrophy type-1A (MDC1A) is characterized by progressive muscular dystrophy and dysmyelinating neuropathy caused by mutations of the α2 chain of laminin-211, the predominant laminin isoform of muscles and nerves. MDC1A has no available treatment so far, although preclinical studies showed amelioration of the disease by the overexpression of miniagrin (MAG). MAG reconnects orphan laminin-211 receptors to other laminin isoforms available in the extracellular matrix of MDC1A mice. Mesoangioblasts (MABs) are vessel-associated progenitors that can form the skeletal muscle and have been shown to restore defective protein levels and motor skills in animal models of muscular dystrophies. As gene therapy in humans still presents challenging technical issues and limitations, we engineered MABs to overexpress MAG to treat MDC1A mouse models, thus combining cell to gene therapy. MABs synthesize and secrete only negligible amount of laminin-211 either in vitro or in vivo. MABs engineered to deliver MAG and injected in muscles of MDC1A mice showed amelioration of muscle histology, increased expression of laminin receptors in muscle, and attenuated deterioration of motor performances. MABs did not enter the peripheral nerves, thus did not affect the associated peripheral neuropathy. Our study demonstrates the potential efficacy of combining cell with gene therapy to treat MDC1A.

Mesoangioblast delivery of miniagrin ameliorates murine model of merosin-deficient congenital muscular dystrophy type 1A.

Tenomodulin (Tnmd) is a well-known gene marker for the tendon and ligament lineage, but its exact functions in these tissues still remain elusive. In this study, we investigated Tnmd loss of function in mouse tendon stem/progenitor cells (mTSPC) by implicating a previously established Tnmd knockout (KO) mouse model. mTSPC were isolated from control and Tnmd KO tail tendons and their stemness features, such as gene marker profile, multipotential, and self-renewal, were compared. Immunofluorescence and reverse transcriptase-polymerase chain reaction analyses for stem cell-, tenogenic-, osteogenic-, and chondrogenic-related genes confirmed their stemness and lineage specificity and demonstrated no profound differences between the two genotypes. Multipotential was not significantly affected since both cell types differentiated successfully into adipogenic, osteogenic, and chondrogenic lineages. In contrast, self-renewal assays validated that Tnmd KO TSPC exhibit significantly reduced proliferative potential, which was also reflected in lower Cyclin D1 levels. When analyzing possible cellular mechanisms behind the observed decreased self-renewability of Tnmd KO TSPC, we found that cellular senescence plays a major role, starting earlier and cumulating more in Tnmd KO compared with control TSPC. This was accompanied with augmented expression of the cell cycle inhibitor p53. Finally, the proliferative effect of Tnmd in TSPC was confirmed with transient transfection of Tnmd cDNA into Tnmd KO TSPC, which rescued their proliferative deficit. Taken together, we can report that loss of Tnmd affects significantly the self-renewal and senescence properties, but not the multipotential of TSPC.

Loss of tenomodulin results in reduced self-renewal and augmented senescence of tendon stem/progenitor cells.

The molecular mechanisms of mechanotransduction in regulating mesenchymal stem cell (MSC) chondrogenesis are not fully understood and represent an area of growing investigation. In this study, human MSC was subjected to chondrogenic differentiation in chitosan-coated poly L-lactide-co-ɛ-caprolactone scaffolds under free swelling or deferral dynamic compression conditions. The effect of deferral dynamic compression to MSC chondrogenesis and late stage hypertrophy development was investigated, and the involvement of TGF-β/SMAD pathway and integrin β1 signaling was analyzed. Deferral dynamic compression enhanced cartilage formation and suppressed chondrocyte hypertrophy. Differential cell morphology and cytoskeletal organization were induced under dynamic compression, together with the activation of TGF-β/Activin/Nodal and suppression of the BMP/GDP signaling. This was accompanied by the repression of integrin/FAK/ERK signaling in the non-hypertrophic cells when compared to the free swelling samples. Inhibition studies blocking TGF-β/Activin/Nodal signaling heightened hypertrophy, activate BMP/SMAD1/5/8 and integrin signaling, while inhibition of integrin-ECM interaction suppressed hypertrophy and activate TGF-β/SMAD2/3 in the free-swelling samples. This study demonstrates the roles of TGF-β/SMAD and integrin signaling, and suggests cross-talk between these two signaling pathways, in regulating the compression-driven hypertrophy development.

Cross-talk between TGF-beta/SMAD and integrin signaling pathways in regulating hypertrophy of mesenchymal stem cell chondrogenesis under deferral dynamic compression.

Pathological fibrosis is driven by a feedback loop in which the fibrotic extracellular matrix is both a cause and consequence of fibroblast activation. However, the molecular mechanisms underlying this process remain poorly understood. Here we identify yes-associated protein (YAP) (homolog of drosophila Yki) and transcriptional coactivator with PDZ-binding motif (TAZ) (also known as Wwtr1), transcriptional effectors of the Hippo pathway, as key matrix stiffness-regulated coordinators of fibroblast activation and matrix synthesis. YAP and TAZ are prominently expressed in fibrotic but not healthy lung tissue, with particularly pronounced nuclear expression of TAZ in spindle-shaped fibroblastic cells. In culture, both YAP and TAZ accumulate in the nuclei of fibroblasts grown on pathologically stiff matrices but not physiologically compliant matrices. Knockdown of YAP and TAZ together in vitro attenuates key fibroblast functions, including matrix synthesis, contraction, and proliferation, and does so exclusively on pathologically stiff matrices. Profibrotic effects of YAP and TAZ operate, in part, through their transcriptional target plasminogen activator inhibitor-1, which is regulated by matrix stiffness independent of transforming growth factor-β signaling. Immortalized fibroblasts conditionally expressing active YAP or TAZ mutant proteins overcome soft matrix limitations on growth and promote fibrosis when adoptively transferred to the murine lung, demonstrating the ability of fibroblast YAP/TAZ activation to drive a profibrotic response in vivo. Together, these results identify YAP and TAZ as mechanoactivated coordinators of the matrix-driven feedback loop that amplifies and sustains fibrosis.

Mechanosignaling through YAP and TAZ drives fibroblast activation and fibrosis.

The phagocytosis of apoptotic cells and associated vesicles (efferocytosis) by DCs is an important mechanism for both self tolerance and host defense. Although some of the engulfment ligands involved in efferocytosis have been identified and studied in vitro, the contributions of these ligands in vivo remain ill defined. Here, we determined that during Mycobacterium tuberculosis (Mtb) infection, the engulfment ligand annexin1 is an important mediator in DC cross-presentation that increases efferocytosis in DCs and intrinsically enhances the capacity of the DC antigen-presenting machinery. Annexin1-deficient mice were highly susceptible to Mtb infection and showed an impaired Mtb antigen-specific CD8+ T cell response. Importantly, annexin1 expression was greatly downregulated in Mtb-infected human blood monocyte-derived DCs, indicating that reduction of annexin1 is a critical mechanism for immune evasion by Mtb. Collectively, these data indicate that annexin1 is essential in immunity to Mtb infection and mediates the power of DC efferocytosis and cross-presentation.

Annexin1 regulates DC efferocytosis and cross-presentation during Mycobacterium tuberculosis infection.

Mesenchymal stem cells (MSCs) are a promising cell type for the repair of damaged cartilage in osteoarthritis (OA). However, OA synovial fluid and factors secreted by synovium impede chondrogenic differentiation of MSCs, and the mechanism responsible for this effect remains unclear. In this study, we sought to investigate whether M1 and M2 synovial macrophages can contribute to the inhibition of MSC chondrogenesis. The constitution of synovial macrophage subsets was analysed by immunohistochemical staining of human OA synovium sections for CD86 (M1 marker) and CD206 (M2 marker). To assess the effect of synovial macrophages on chondrogenesis, collagen type II (COL2) and aggrecan (ACAN) gene expression were compared between MSCs undergoing chondrogenic differentiation in medium conditioned (CM) by human OA synovial explants, human synovial macrophages and fibroblasts, or peripheral blood derived primary human monocytes differentiated towards an M1 or M2 phenotype. OA synovium contained both M1 and M2 macrophages. Medium conditioned by synovial macrophages (CD45 + plastic adherent cells) down-regulated chondrogenic gene expression by MSCs. Additionally, CM of M1 polarised monocytes significantly decreased COL2 and ACAN gene expression by MSCs; this effect was not observed for treatment with CM of M2 polarised monocytes. MSC chondrogenesis is inhibited by OA synovium CM through factors secreted by synovial macrophages and our findings suggest that M1 polarised subsets are potential mediators of this anti-chondrogenic effect. Modulation of macrophage phenotype may serve as a beneficial strategy to maximise the potential of MSCs for efficient cartilage repair.

Human osteoarthritic synovium impacts chondrogenic differentiation of mesenchymal stem cells via macrophage polarisation state.

Following bidirectional communication, the conceptus and the uterine epithelium must establish a proper cell-cell interaction, resulting in the progression of implantation processes. To clarify the mechanism of conceptus attachment to the uterine endometrium, we studied whether vascular cell adhesion molecule (VCAM1) was expressed in bovine conceptuses or endometrium during the peri-attachment period. Uterine VCAM1 expression was minimal in day 17 (day 0=day of estrus) cyclic and pregnant animals, but increased between days 20 and 22 of pregnancy. In the intercaruncular regions, VCAM1 protein was localized to the luminal and glandular epithelia, whereas in the caruncular regions, VCAM1 protein was detected in the stroma and endothelia of the uterine endometrium. In cultured endometrial epithelial cells (EECs), VCAM1 expression was up-regulated when treated with uterine flushings or growth factor and further increased when EECs were cocultured with bovine trophoblast CT1 cells. VCAM1 expression in CT1 cells was also up-regulated with the use of uterine flushings, and further increased when these cells were cocultured with EECs. Expression of VCAM1 receptor, integrin α 4 (ITGA4) mRNA, increased significantly in day 22 conceptuses. In day 22 pregnant uteri, VCAM1 protein was found in both EECs and conceptuses, but ITGA4 was localized only to trophoblasts. These observations indicate that cell-cell interactions between conceptuses and uterine epithelial cells are required for sufficient VCAM1 and ITGA4 expression in the bovine species and suggest that uterine VCAM1 and conceptus ITGA4 play a role in the establishment of conceptus adhesion to the uterine endometrium.

Involvement of VCAM1 in the bovine conceptus adhesion to the uterine endometrium.

Autophagy inhibition has been widely accepted as a promising therapeutic strategy in cancer, while the lack of effective and specific autophagy inhibitors hinders its application. Here we found that liensinine, a major isoquinoline alkaloid, inhibits late-stage autophagy/mitophagy through blocking autophagosome-lysosome fusion. This effect is likely achieved via inhibiting the recruitment of RAB7A to lysosomes but not to autophagosomes. We further investigated the effects of autophagy inhibition by liensinine on the therapeutic efficacy of chemotherapeutic drugs and found that cotreatment of liensinine markedly decreased the viability and increased apoptosis in breast cancer cells treated with various chemotherapeutic agents. Mechanistically, we found that inhibition of autophagy/mitophagy by liensinine enhanced doxorubicin-mediated apoptosis by triggering mitochondrial fission, which resulted from dephosphorylation and mitochondrial translocation of DNM1L. However, blocking autophagosome/mitophagosome formation by pharmacological or genetic approaches markedly attenuated mitochondrial fission and apoptosis in cells with combinatatorial treatment. Moreover, liensinine was synergized with doxorubicin to inhibit tumor growth in MDA-MB-231 xenograft in vivo. Our findings suggest that liensinine could potentially be further developed as a novel autophagy/mitophagy inhibitor, and a combination of liensinine with classical chemotherapeutic drugs could represent a novel therapeutic strategy for treatment of breast cancer.

A novel autophagy/mitophagy inhibitor liensinine sensitizes breast cancer cells to chemotherapy through DNM1L-mediated mitochondrial fission.

Conditions such as muscular dystrophies (MDs) that affect both cardiac and skeletal muscles would benefit from therapeutic strategies that enable regeneration of both of these striated muscle types. Protocols have been developed to promote induced pluripotent stem cells (iPSCs) to differentiate toward cardiac or skeletal muscle; however, there are currently no strategies to simultaneously target both muscle types. Tissues exhibit specific epigenetic alterations; therefore, source-related lineage biases have the potential to improve iPSC-driven multilineage differentiation. Here, we determined that differential myogenic propensity influences the commitment of isogenic iPSCs and a specifically isolated pool of mesodermal iPSC-derived progenitors (MiPs) toward the striated muscle lineages. Differential myogenic propensity did not influence pluripotency, but did selectively enhance chimerism of MiP-derived tissue in both fetal and adult skeletal muscle. When injected into dystrophic mice, MiPs engrafted and repaired both skeletal and cardiac muscle, reducing functional defects. Similarly, engraftment into dystrophic mice of canine MiPs from dystrophic dogs that had undergone TALEN-mediated correction of the MD-associated mutation also resulted in functional striatal muscle regeneration. Moreover, human MiPs exhibited the same capacity for the dual differentiation observed in murine and canine MiPs. The findings of this study suggest that MiPs should be further explored for combined therapy of cardiac and skeletal muscles.

丙酮酸钠

BioXtra, ≥99%

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