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Mark E. Lauer Metin Aytekin Suzy A. Comhair Jacqueline Loftis Liping Tian Carol F. Farver Vincent C. Hascall Raed A. Dweik 《The Journal of biological chemistry》2014,289(10):6791-6798
We previously reported an altered hyaluronan (HA) metabolism in idiopathic pulmonary arterial hypertension (IPAH) lung tissue and cultured smooth muscle cells. Hyaluronan was present in the smooth muscle cell layer surrounding the pulmonary vasculature and in plexigenic lesions. Additionally, cultured pulmonary artery smooth muscle cells produced spontaneous HA “cable” structures, without additional stimuli, that were leukocyte-adhesive. We now present evidence that the HA that accumulates in IPAH plexigenic lesions is a pathological form of HA in which heavy chains (HCs) from the serum-derived proteoglycan inter-α-inhibitor are covalently attached to the HA backbone to form a pathological HC-HA complex. CD45-positive leukocytes were identified within these HC-HA matrices. Elevated mRNA levels of the enzyme that transfers HCs to HA, known as tumor necrosis factor-stimulated gene 6, were detected in IPAH lung tissue. 相似文献
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Hua He Wei Li David Y. Tseng Shan Zhang Szu-Yu Chen Anthony J. Day Scheffer C. G. Tseng 《The Journal of biological chemistry》2009,284(30):20136-20146
Clinically, amniotic membrane (AM) suppresses inflammation, scarring, and angiogenesis. AM contains abundant hyaluronan (HA) but its function in exerting these therapeutic actions remains unclear. Herein, AM was extracted sequentially with buffers A, B, and C, or separately by phosphate-buffered saline (PBS) alone. Agarose gel electrophoresis showed that high molecular weight (HMW) HA (an average of ∼3000 kDa) was predominantly extracted in isotonic Extract A (70.1 ± 6.0%) and PBS (37.7 ± 3.2%). Western blot analysis of these extracts with hyaluronidase digestion or NaOH treatment revealed that HMW HA was covalently linked with the heavy chains (HCs) of inter-α-inhibitor (IαI) via a NaOH-sensitive bond, likely transferred by the tumor necrosis factor-α stimulated gene-6 protein (TSG-6). This HC·HA complex (nHC·HA) could be purified from Extract PBS by two rounds of CsCl/guanidine HCl ultracentrifugation as well as in vitro reconstituted (rcHC·HA) by mixing HMW HA, serum IαI, and recombinant TSG-6. Consistent with previous reports, Extract PBS suppressed transforming growth factor-β1 promoter activation in corneal fibroblasts and induced mac ro phage apo pto sis. However, these effects were abolished by hyaluronidase digestion or heat treatment. More importantly, the effects were retained in the nHC·HA or rcHC·HA. These data collectively suggest that the HC·HA complex is the active component in AM responsible in part for clinically observed anti-inflammatory and anti-scarring actions.Hyaluronan (HA)4 is widely distributed in extracellular matrices, tissues, body fluids, and even in intracellular compartments (reviewed in Refs. 1 and 2). The molecular weight of HA ranges from 200 to 10,000 kDa depending on the source (3), but can also exist as smaller fragments and oligosaccharides under certain physiological or pathological conditions (1). Investigations over the last 15 years have suggested that low Mr HA can induce the gene expression of proinflammatory mediators and proangiogenesis, whereas high molecular weight (HMW) HA inhibits these processes (4–7).Several proteins have been shown to bind to HA (8) such as aggrecan (9), cartilage link protein (10), versican (11), CD44 (12, 13), inter-α-inhibitor (IαI) (14, 15), and tumor necrosis factor-α stimulated gene-6 protein (TSG-6) (16, 17). IαI consists of two heavy chains (HCs) (HC1 and HC2), both of which are linked through ester bonds to a chondroitin sulfate chain that is attached to the light chain, i.e. bikunin. Among all HA-binding proteins, only the HCs of IαI have been clearly demonstrated to be covalently coupled to HA (14, 18). However, TSG-6 has also been reported to form stable, possibly covalent, complexes with HA, either alone (19, 20) or when associated with HC (21).The formation of covalent bonds between HCs and HA is mediated by TSG-6 (22–24) where its expression is often induced by inflammatory mediators such as tumor necrosis factor-α and interleukin-1 (25, 26). TSG-6 is also expressed in inflammatory-like processes, such as ovulation (21, 27, 28) and cervical ripening (29). TSG-6 interacts with both HA (17) and IαI (21, 24, 30–33), and is essential for covalently transferring HCs on to HA (22–24). The TSG-6-mediated formation of the HC·HA complex has been demonstrated to play a crucial role in female fertility in mice. The HC·HA complex is an integral part of an expanded extracellular “cumulus” matrix around the oocyte, which plays a critical role in successful ovulation and fertilization in vivo (22, 34). HC·HA complexes have also been found at sites of inflammation (35–38) where its pro- or anti-inflammatory role remain arguable (39, 40).Immunostaining reveals abundant HA in the avascular stromal matrix of the AM (41, 42).5 In ophthalmology, cryopreserved AM has been widely used as a surgical graft for ocular surface reconstruction and exerts clinically observable actions to promote epithelial wound healing and to suppress inflammation, scarring, and angiogenesis (for reviews see Refs. 43–45). However, it is not clear whether HA in AM forms HC·HA complex, and if so whether such an HC·HA complex exerts any of the above therapeutic actions. To address these questions, we extracted AM with buffers of increasing salt concentration. Because HMW HA was found to form the HC·HA complex and was mainly extractable by isotonic solutions, we further purified it from the isotonic AM extract and reconstituted it in vitro from three defined components, i.e. HMW HA, serum IαI, and recombinant TSG-6. Our results showed that the HC·HA complex is an active component in AM responsible for the suppression of TGF-β1 promoter activity, linkable to the scarring process noted before by AM (46–48) and by the AM soluble extract (49), as well as for the promotion of macrophage death, linkable to the inflammatory process noted by AM (50) and the AM soluble extract (51). 相似文献
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Recently, we reported HC-HA, a covalent complex formed between heavy chains (HCs) of inter-α-inhibitor (IαI) and hyaluronan (HA) by the catalytic action of tumor necrosis factor (TNF)-stimulated gene-6 (TSG-6), is responsible for human amniotic membrane (AM) anti-inflammatory, anti-scarring, and anti-angiogenic actions. At the present time, the only well characterized source of IαI is serum being produced by the liver. This study showed that AM epithelial and stromal cells and stromal matrix all stained positively for HA, HC 1, 2, and 3, bikunin, and TSG-6. TSG-6 mRNA and protein were constitutively expressed by cultured AM epithelial and stromal cells without being up-regulated by TNF. In serum-free conditions, these cells expressed IαI, leading to the formation of HC-HA complex that contained both HC1 and HC2. In contrast, only HC1 was found in the HC-HA complex purified from AM. Local production of IαI, the HC-TSG-6 intermediate complex, and HC-HA were abolished when cells were treated with siRNA to HC1, HC2, bikunin (all of which impair the biosynthesis of IαI), or TSG-6 but not to HC3. Collectively, these results indicate that AM is another tissue in addition to the liver to constitutively produce IαI and that the HC-HA complex made by this tissue is different from that found at inflammatory sites (e.g. in asthma and arthritis) and in the matrix of the cumulus oocyte complex. 相似文献
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Scavenius C Sanggaard KW Nikolajsen CL Bak S Valnickova Z Thøgersen IB Jensen ON Højrup P Enghild JJ 《Biochimica et biophysica acta》2011,1814(12):1624-1630
In this study, we show that inter-α-inhibitor is a substrate for both factor XIIIa and tissue transglutaminase. These enzymes catalyze the incorporation of dansylcadaverine and biotin-pentylamine, revealing that inter-α-inhibitor contains reactive Gln residues within all three subunits. These findings suggest that transglutaminases catalyze the covalent conjugation of inter-α-inhibitor to other proteins. This was demonstrated by the cross-linking between inter-α-inhibitor and fibrinogen by either factor XIIIa or tissue transglutaminase. Finally, using quantitative mass spectrometry, we show that inter-α-inhibitor is cross-linked to the fibrin clot in a 1:20 ratio relative to the known factor XIIIa substrate α2-antiplasmin. This interaction may protect fibrin or other Lys-donating proteins from adventitious proteolysis by increasing the local concentration of bikunin. In addition, the reaction may influence the TSG-6/heavy Chain 2-mediated transfer of heavy chains observed during inflammation. 相似文献
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I. I. Galkin O. Yu. Pletjushkina R. A. Zinovkin V. V. Zakharova I. S. Birjukov B. V. Chernyak E. N. Popova 《Biochemistry. Biokhimii?a》2014,79(2):124-130
Increased serum level of tumor necrosis factor α (TNFα) causes endothelial dysfunction and leads to serious vascular pathologies. TNFα signaling is known to involve reactive oxygen species (ROS). Using mitochondria-targeted antioxidant SkQR1, we studied the role of mitochondrial ROS in TNFα-induced apoptosis of human endothelial cell line EAhy926. We found that 0.2 nM SkQR1 prevents TNFα-induced apoptosis. SkQR1 has no influence on TNFα-dependent proteolytic activation of caspase-8 and Bid, but it inhibits cytochrome c release from mitochondria and cleavage of caspase-3 and its substrate PARP. SkQ analogs lacking the antioxidant moieties do not prevent TNFα-induced apoptosis. The antiapoptotic action of SkQR1 may be related to other observations made in these experiments, namely SkQR1-induced increase in Bcl-2 and corresponding decrease in Bax as well as p53. These results indicate that mitochondrial ROS production is involved in TNFα-initiated endothelial cell death, and they suggest the potential of mitochondria-targeted antioxidants as vasoprotectors. 相似文献
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Yang Wang Zhi-Hao Huang Yang-Jia Li Gui-Wei He Ru-Yuan Yu Jie Yang Wan-Ting Liu Bin Li Qing-Yu He 《Apoptosis : an international journal on programmed cell death》2016,21(12):1438-1446
Emerging evidence suggested that necroptosis has essential functions in many human inflammatory diseases, but the molecular mechanisms of necroptosis remain unclear. Here, we employed SILAC quantitatively dynamic proteomics to compare the protein changes during TNF-α-induced necroptosis at different time points in murine fibrosarcoma L929 cells with caspase-8 deficiency, and then performed the systematical analysis on the signaling networks involved in the progress using bioinformatics methods. Our results showed that a total of 329, 421 and 378 differentially expressed proteins were detected at three stages of necroptosis, respectively. Gene ontology and ingenuity pathway analysis (IPA) revealed that the proteins regulated at early stages of necroptosis (2, 6 h) were mainly involved in mitochondria dysfunction, oxidative phosphorylation and Nrf-2 signaling, while the expression levels of the proteins related to ubiquitin, Nrf-2, and NF-κB pathways were found to have changes at last stages of necroptosis (6, 18 h). Taken together, we demonstrated for the first time that dysfunction of mitochondria and ubiquitin–proteasome signaling contributed to the initiation and execution of necroptosis. These findings may provide clues for the identification of important regulators in necroptosis and the development of novel therapeutic strategies for the related diseases. 相似文献
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Fenglian Xu Juliane Proft Sarah Gibbs Bob Winkfein Jadah N. Johnson Naweed Syed Janice E. A. Braun 《PloS one》2010,5(6)
Background
Cysteine string protein (CSPα) is a synaptic vesicle protein that displays unique anti-neurodegenerative properties. CSPα is a member of the conserved J protein family, also called the Hsp40 (heat shock protein of 40 kDa) protein family, whose importance in protein folding has been recognized for many years. Deletion of the CSPα in mice results in knockout mice that are normal for the first 2–3 weeks of life followed by an unexplained presynaptic neurodegeneration and premature death. How CSPα prevents neurodegeneration is currently not known. As a neuroprotective synaptic vesicle protein, CSPα represents a promising therapeutic target for the prevention of neurodegenerative disorders.Methodology/Principal Findings
Here, we demonstrate that the flavonoid quercetin promotes formation of stable CSPα-CSPα dimers and that quercetin-induced dimerization is dependent on the unique cysteine string region. Furthermore, in primary cultures of Lymnaea neurons, quercetin induction of CSPα dimers correlates with an inhibition of synapse formation and synaptic transmission suggesting that quercetin interfers with CSPα function. Quercetin''s action on CSPα is concentration dependent and does not promote dimerization of other synaptic proteins or other J protein family members and reduces the assembly of CSPα:Hsc70 units (70kDa heat shock cognate protein).Conclusions/Significance
Quercetin is a plant derived flavonoid and popular nutritional supplement proposed to prevent memory loss and altitude sickness among other ailments, although its precise mechanism(s) of action has been unclear. In view of the therapeutic promise of upregulation of CSPα and the undesired consequences of CSPα dysfunction, our data establish an essential proof of principle that pharmaceutical agents can selectively target the neuroprotective J protein CSPα. 相似文献11.
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Constantin R. Popescu Megan R. Sutherland Anik Cloutier Geneviève Beno?t Mariane Bertagnolli Catherine Yzydorczyk Nathalie Germain Véronique Phan Martine Lelièvre-Pegorier Hervé Sartelet Anne Monique Nuyt 《PloS one》2013,8(12)
Preterm neonates are exposed at birth to high oxygen concentrations relative to the intrauterine environment. We have previously shown in a rat model that a hyperoxic insult results in a reduced nephron number in adulthood. Therefore, the aim of this study was to determine the effects of transient neonatal hyperoxia exposure on nephrogenesis. Sprague-Dawley rat pups were raised in 80% O2 or room air from P3 to P10. Pups (n = 12/group, 6 males and 6 females) were sacrificed at P5 (during active nephrogenesis) and at P10 (after the completion of nephrogenesis). Hyperoxia exposure resulted in a significant reduction in both nephrogenic zone width and glomerular diameter at P5, and a significantly increased apoptotic cell count; however, nephron number at P10 was not affected. HIF-1α expression in the developing kidney was significantly reduced following hyperoxia exposure. Systemic administration of the HIF-1α stabilizer dimethyloxalylglycine (DMOG) resulted in enhanced expression of HIF-1α and improved nephrogenesis: kidneys from hyperoxia-exposed pups treated with DMOG exhibited a nephrogenic zone width and glomerular diameter similar to room-air controls. These findings demonstrate that neonatal hyperoxia exposure results in impaired nephrogenesis, which may be at least in part HIF-1α-mediated. Although nephron number was not significantly reduced at the completion of nephrogenesis, early indicators of maldevelopment suggest the potential for accelerated nephron loss in adulthood. Overall, this study supports the premise that prematurely born neonates exposed to high oxygen levels after birth are vulnerable to impaired renal development. 相似文献
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TNFα is a pleotropic cytokine that initiates many downstream signaling pathways, including NF-κB activation, MAP kinase activation
and the induction of both apoptosis and necrosis. TNFα has shown to lead to reactive oxygen species generation through activation
of NADPH oxidase, through mitochondrial pathways, or other enzymes. As discussed, ROS play a role in potentiation or inhibition
of many of these signaling pathways. We particularly discuss the role of sustained JNK activation potentiated by ROS, which
generally is supportive of apoptosis and “necrotic cell death” through various mechanisms, while ROS could have inhibitory
or stimulatory roles in NF-κB signaling. 相似文献
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Xiaoxuan Wang Hualing Sun Huan Liu Li Ma Chenxi Jiang Haiqing Liao Shihan Xu Junbo Xiang Zhengguo Cao 《Journal of cellular physiology》2019,234(12):22719-22730
Tooth cementum is a bone-like mineralized tissue and serves as a microbial barrier against invasion and destruction. Cementum is also responsible for tooth stability and defending pulp from outside stimuli, which is formed by cementoblasts. Although it is crucial for periodontal and periapical diseases, the mechanisms underlying the pathophysiological changes of cementoblasts and their inflammatory responses remain unclear. MiR-181b is found to modulate vascular inflammation and endotoxin tolerance. In this study, miR-181b-5p was downregulated in tumor necrosis factor-α (TNF-α)-stimulated cementoblasts, whereas proinflammatory molecules increased. The mouse periapical lesions have similar results, which imitate an inflammatory environment for cementoblasts in vivo. The bioinformatics analysis and dual luciferase reporter assay suggested that miR-181b-5p targeted interleukin-6 (IL-6). Overexpressing miR-181b-5p negatively regulated IL-6 and proinflammatory chemokine. Western blot analysis and luciferase activity reporter assay verified that miR-181b-5p weakened the NF-κB activity. Hence, miR-181b-5p moderated proinflammatory chemokine production by targeting IL-6 in cementoblasts and NF-κB signaling pathway was involved. Furthermore, miR-181b-5p promoted cementoblast apoptosis, which may enhance the resolution of inflammation. Overall, our data revealed that miR-181b-5p was a negative regulator of TNF-α-induced inflammatory responses in cementoblasts. 相似文献
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EVER1 and 2 confer resistance to cutaneous oncogenic human papillomavirus infections by downregulating the activating protein 1 (AP-1) signaling pathway. Defects in their expression are associated with susceptibility to epidermodysplasia verruciformis, which is characterized by persistent β-HPV infection, tumor necrosis factor alpha (TNF-α) overproduction in keratinocytes and the development of skin cancers. TNF-α-induced apoptosis is a key defense strategy, preventing the persistence of the virus within cells, but the role of EVER proteins in this cell death mechanism triggered by extrinsic stimuli is unknown. We show here that EVER2 induces TNF-α- and TRAIL-dependant apoptosis. It interacts with the N-terminal domain of TRADD, impairs the recruitment of TRAF2 and RIPK1 and promotes apoptosis. The skin cancer-associated EVER2 I306 allele results in an impaired TRADD–EVER2 interaction, with lower levels of cell death following treatment with TNF-α. These data highlight a new, critical function of EVER2 in controlling cell survival in response to death stimuli. 相似文献
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Zhang L Xing D Chen M 《Apoptosis : an international journal on programmed cell death》2008,13(7):950-958
Bcl-2 family proteins are implicated as essential regulators in tumor necrosis factor-α (TNFα)-induced apoptosis. BimL, a BH3-only member of Bcl-2 family, can directly or indirectly activate the proapoptotic Bax and the subsequent mitochondrial
apoptotic pathway. However, the molecular mechanism of BimL activating Bax activation during TNFα-induced apoptosis is not fully understood. In this study, the role of BimL in Bax activation during TNFα-induced apoptosis was investigated in differentiated PC12 and MCF7 cells, with real-time single-cell
analysis. The experimental results show that Bax translocated to mitochondria and cytochrome c (Cyt c) released from mitochondria
after TNFα treatment. Furthermore, SP600125 (specific inhibitor of JNK) could inhibit the Cyt c release from mitochondria.
Co-immunoprecipitation results show that, the interaction between Bcl-xL and Bax decreased after TNFα treatment, while that between Bcl-xL and BimL increased. Bax did not co-immunoprecipitate with BimL before or after the TNFα treatment. In addition, the increased interaction between BimL and Bcl-xL was dynamically monitored by using fluorescence resonance energy transfer (FRET) technique. Most importantly, there was no
evidence of BimL redistribution to mitochondria until cell apoptosis. By comprehensively analyzing these data, it is concluded that BimL displaces Bcl-xL in the mitochondria and promotes Bax translocation during TNFα-induced apoptosis. 相似文献
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《Carbohydrate research》1990,203(1):C3-C8