scholarly journals Regulatory effects of TLR2 on megakaryocytic cell function

Blood ◽  
2011 ◽  
Vol 117 (22) ◽  
pp. 5963-5974 ◽  
Author(s):  
Lea M. Beaulieu ◽  
Elaine Lin ◽  
Kristine M. Morin ◽  
Kahraman Tanriverdi ◽  
Jane E. Freedman
Keyword(s):  
Cell Function ◽  
Mammalian Target Of Rapamycin ◽  
Matrix Proteins ◽  
Protein Levels ◽  
Erk Mapk ◽  
Normal Platelet ◽  
Immune Mediated ◽  
Regulatory Effects ◽  
Specific Ligand

Abstract TLR2, a functional, inflammatory-related receptor, is known to be expressed on megakaryocytes and platelets and to lead to infection and immune-mediated activation of platelets; however, the role of this receptor in megakaryocytes is not understood. Using Meg-01 cells and mouse megakaryocytes, we found that NFκB, ERK-MAPK, and PI3K/Akt pathways, known downstream pathways of TLRs, are activated by Pam3CSK4, a TLR2-specific ligand. In addition, transcription factors associated with megakaryocyte maturation, GATA-1, NF-E2, and mammalian target of rapamycin (mTOR), are all increased in the presence of Pam3CSK4. The effect of Pam3CSK4 on megakaryocyte maturation was verified by the increase in DNA content and adhesion to extracellular matrix proteins by TLR2-dependent stimulation. In addition, TLR2 stimulation resulted in an increase in reactive oxygen species (ROS) production. Gene expression and protein levels of GP1b, CD41, MCP-1, COX2, NFκB1, and TLR2 were up-regulated in megakaryocytes after TLR2 stimulation through NFκB, PI3K/Akt, and ERK-MAPK pathways. Treatment of wild-type mice with Pam3CSK4 resulted in a return to normal platelet levels and an increase in megakaryocyte maturation, which did not occur in the TLR2−/− mice. Therefore, inflammation, through TLR2, can increase maturation and modulate the phenotype of megakaryocytes, contributing to the interrelationship between inflammation and hemostasis.

scholarly journals Nox4 Mediates RANKL-induced ER-Phagy and Osteoclastogenesis via Activating ROS/PERK/eIF-2α/ATF4 Pathway

2020 ◽  
Author(s):  
Jing Sun ◽  
wugui chen ◽  
Songtao Li ◽  
Sizhen Yang ◽  
Ying Zhang ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Bone Resorption ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Nuclear Factor Κb ◽  
Protein Levels ◽  
Unfolded Protein ◽  
Regulatory Effects ◽  
Protein Response

Abstract Background: Receptor activator of nuclear factor-κB ligand (RANKL) has been found to induce osteoclastogenesis and bone resorption. However, the underlying molecular mechanisms remain unclear. Methods: Osteoclastogenesis was evaluated by number of TRAP-positive multinuclear (≥3) osteoclasts, bone resorption pits and expression levels of related genes. Autophagy activity were evaluated by LC3-II/LC3-I ratio, number of autophagic vacuoles and adenovirus-mRFP-GFP-tagged LC3 reporting system; Inhibitor chloroquine (CQ) was used to verified the role of autophagy in RANKL-induced osteoclastogenesis; Via downregulating Nox4 with inhibitor (DPI) and retrovirus-conveyed shRNA, we further explored the importance of Nox4 in RANKL-induced autophagy and osteoclastogenesis, as well as the regulatory effects of Nox4 on nonmitochondrial reactive oxygen species (ROS) and PERK/eIF-2α/ATF4 pathway. Intracellular ROS scavenger (NAC), mitochondrial-targeted antioxidant (MitoTEMPO) and inhibitor of PERK (GSK2606414) were also employed to investigate the role of ROS and PERK/eIF-2α/ATF4 pathway in RANKL-induced autophagy and osteoclastogenesis. Results: RANKL markedly increased autophagy, while CQ treatment caused reduction of RANKL-induced autophagy and osteoclastogenesis. Consistent with the increased autophagy, the protein levels of Nox4 were significantly increased, and Nox4 was selectively localized within the endoplasmic reticulum (ER) after RANKL stimulation. DPI and shRNA efficiently decreased the protein level and (or) activity of Nox4 in the ER and inhibited RANKL-induced autophagy and osteoclastogenesis. Mechanistically, we found that Nox4 regulates RANKL-induced autophagy activation and osteoclastogenesis by stimulating the production of nonmitochondrial ROS. Additionally, Nox4-derived nonmitochondrial ROS dramatically activate PERK/eIF-2α/ATF4, which is a critical unfolded protein response (UPR)-related signaling pathway during ER stress. Blocking the activation of the PERK/eIF-2α/ATF4 signaling pathway either by Nox4 shRNA, ROS antioxidant or PERK inhibitor (GSK2606414) treatment significantly inhibited endoplasmic reticulum autophagy (ER-phagy) during RANKL-induced osteoclastogenesis. Conclusions: Our findings provide new insights into the processes of RANKL-induced osteoclastogenesis and will help the development of new therapeutic strategies for osteoclastogenesis-related diseases.

scholarly journals PKM2 Is Required to Activate Myeloid Dendritic Cells from Patients with Severe Aplastic Anemia

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Chunyan Liu ◽  
Mengying Zheng ◽  
Ting Wang ◽  
Huijuan Jiang ◽  
Rong Fu ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Aplastic Anemia ◽  
Immune Status ◽  
Bone Marrow Failure ◽  
Severe Aplastic Anemia ◽  
Myeloid Dendritic Cells ◽  
Protein Levels ◽  
Immune Mediated ◽  
Normal Controls

Severe aplastic anemia (SAA) is an autoimmune disease in which bone marrow failure is mediated by activated myeloid dendritic cells (mDCs) and T lymphocytes. Recent research has identified a strong immunomodulatory effect of pyruvate kinase M2 (PKM2) on dendritic cells in immune-mediated diseases. In this study, we aimed to explore the role of PKM2 in the activation of mDCs in SAA. We observed conspicuously higher levels of PKM2 in mDCs from SAA patients compared to normal controls at both the gene and protein levels. Concurrently, we unexpectedly discovered that after the mDC-specific downregulation of PKM2, mDCs from patients with SAA exhibited weakened phagocytic activity and significantly decreased and shortened dendrites relative to their counterparts from normal controls. The expression levels of the costimulatory molecules CD86 and CD80 were also reduced on mDCs. Our results also suggested that PKM2 knockdown in mDCs reduced the abilities of these cells to promote the activation of CD8+ T cells (CTLs), leading to the decreased secretion of cytotoxic factors by the latter cell type. These findings demonstrate that mDC activation requires an elevated intrinsic PKM2 level and that PKM2 improves the immune status of patients with SAA by enhancing the functions of mDCs and, consequently, CTLs.

scholarly journals Multi-Omics Integration Highlights the Role of Ubiquitination in CCl4-Induced Liver Fibrosis

2020 ◽  
Vol 21 (23) ◽  
pp. 9043
Author(s):  
Maria Mercado-Gómez ◽  
Fernando Lopitz-Otsoa ◽  
Mikel Azkargorta ◽  
Marina Serrano-Maciá ◽  
Sofia Lachiondo-Ortega ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Cell Function ◽  
Gene Array ◽  
Matrix Proteins ◽  
Nuclear Antigen ◽  
Post Translational Modification ◽  
Physiological Processes ◽  
Ubiquitin System ◽  
Proliferating Cell

Liver fibrosis is the excessive accumulation of extracellular matrix proteins that occurs in chronic liver disease. Ubiquitination is a post-translational modification that is crucial for a plethora of physiological processes. Even though the ubiquitin system has been implicated in several human diseases, the role of ubiquitination in liver fibrosis remains poorly understood. Here, multi-omics approaches were used to address this. Untargeted metabolomics showed that carbon tetrachloride (CCl4)-induced liver fibrosis promotes changes in the hepatic metabolome, specifically in glycerophospholipids and sphingolipids. Gene ontology analysis of public deposited gene array-based data and validation in our mouse model showed that the biological process “protein polyubiquitination” is enriched after CCl4-induced liver fibrosis. Finally, by using transgenic mice expressing biotinylated ubiquitin (bioUb mice), the ubiquitinated proteome was isolated and characterized by mass spectrometry in order to unravel the hepatic ubiquitinated proteome fingerprint in CCl4-induced liver fibrosis. Under these conditions, ubiquitination appears to be involved in the regulation of cell death and survival, cell function, lipid metabolism, and DNA repair. Finally, ubiquitination of proliferating cell nuclear antigen (PCNA) is induced during CCl4-induced liver fibrosis and associated with the DNA damage response (DDR). Overall, hepatic ubiquitome profiling can highlight new therapeutic targets for the clinical management of liver fibrosis.

scholarly journals Hypoxia-inducible factor (HIF1alpha) inhibition modulates cumulus cell function and affects bovine oocyte maturation in vitro†

2020 ◽  
Author(s):  
Aslihan Turhan ◽  
Miguel Tavares Pereira ◽  
Gerhard Schuler ◽  
Ulrich Bleul ◽  
Mariusz P Kowalewski
Keyword(s):  
Oocyte Maturation ◽  
Cell Function ◽  
Hypoxia Inducible Factor ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Negative Effects ◽  
Protein Levels ◽  
Maturation In Vitro

Abstract Various metabolic and hormonal factors expressed in cumulus cells are positively correlated with the in vitro maturation (IVM) of oocytes. However, the role of hypoxia sensing both during maturation of cumulus–oocyte complexes (COCs) as well as during the resumption of meiosis remains uncertain. HIF1alpha plays major roles in cellular responses to hypoxia, and here we investigated its role during bovine COC maturation by assessing the expression of related genes in cumulus cells. COCs were divided into the following groups: immature (control), in vitro matured (IVM/control), or matured in the presence of a blocker of HIF1alpha activity (echinomycin, IVM/E). We found an inhibition of cumulus cell expansion in IVM/E, compared with the IVM/control. Transcript levels of several factors (n = 13) were assessed in cumulus cells. Decreased expression of HAS2, TNFAIP6, TMSB4, TMSB10, GATM, GLUT1, CX43, COX2, PTGES, and STAR was found in IVM/E (P < 0.05). Additionally, decreased protein levels were detected for STAR, HAS2, and PCNA (P < 0.05), while activated-Caspase 3 remained unaffected in IVM/E. Progesterone output decreased in IVM/E. The application of PX-478, another blocker of HIF1alpha expression, yielded identical results. Negative effects of HIF1alpha suppression were further observed in the significantly decreased oocyte maturation and blastocyst rates from COCs matured with echinomycin (P < 0.05) or PX-478 (P < 0.05). These results support the importance of HIF1alpha for COC maturation and subsequent embryo development. HIF1alpha is a multidirectional factor controlling intercellular communication within COCs, steroidogenic activity, and oocyte development rates, and exerting effects on blastocyst rates.

scholarly journals A Versatile Role of Mammalian Target of Rapamycin in Human Dendritic Cell Function and Differentiation

2010 ◽  
Vol 185 (7) ◽  
pp. 3919-3931 ◽  
Author(s):  
Michael Haidinger ◽  
Marko Poglitsch ◽  
Rene Geyeregger ◽  
Sudhir Kasturi ◽  
Maximilian Zeyda ◽  
...  
Keyword(s):  
Dendritic Cell ◽  
Cell Function ◽  
Mammalian Target Of Rapamycin ◽  
Target Of Rapamycin ◽  
Human Dendritic Cell ◽  
Dendritic Cell Function

scholarly journals Role of Epigallocatechin Gallate in Glucose, Lipid, and Protein Metabolism and L-Theanine in the Metabolism-Regulatory Effects of Epigallocatechin Gallate

Nutrients ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 4120
Author(s):  
Ling Lin ◽  
Li Zeng ◽  
An Liu ◽  
Dongyin Yuan ◽  
Yingqi Peng ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Insulin Receptor ◽  
Protein Metabolism ◽  
Glycogen Synthase ◽  
Mammalian Target Of Rapamycin ◽  
Epigallocatechin Gallate ◽  
Control Group ◽  
Nutrient Metabolism ◽  
Regulatory Effects

Epigallocatechin gallate (EGCG) and L-theanine (LTA) are important bioactive components in tea that have shown promising effects on nutrient metabolism. However, whether EGCG alone or combined with LTA can regulate the glucose, lipid, and protein metabolism of healthy rats remains unclear. Therefore, we treated healthy rats with EGCG or the combination of EGCG and LTA (EGCG+LTA) to investigate the effects of EGCG on nutrient metabolism and the role of LTA in the metabolism-regulatory effects of EGCG. The results showed that compared with the control group, EGCG activated insulin and AMP-activated protein kinase (AMPK) signals, thus regulating glucose, lipid, and protein metabolism. Compared with EGCG, EGCG+LTA enhanced hepatic and muscle glycogen levels and suppressed phosphorylation of AMPK, glycogen synthase 2, mammalian target of rapamycin, and ribosomal protein S6 kinase. In addition, EGCG+LTA inhibited the expression of liver kinase B1, insulin receptor and insulin receptor substrate, and promoted the phosphorylation level of acetyl-CoA carboxylase. Furthermore, both EGCG and EGCG+LTA were harmless for young rats. In conclusion, EGCG activated AMPK and insulin pathways, thereby promoting glycolysis, glycogen, and protein synthesis and inhibiting fatty acid (FA) and cholesterol synthesis. However, LTA cooperated with EGCG to promote glycogen metabolism and suppressed the effect EGCG on FA and protein synthesis via AMPK signals.

scholarly journals KIF15 supports spermatogenesis via its effects on Sertoli cell microtubule, actin, vimentin, and septin cytoskeletons

Endocrinology ◽  
2021 ◽  
Vol 162 (4) ◽  
Author(s):  
Siwen Wu ◽  
Lixiu Lv ◽  
Linxi Li ◽  
Lingling Wang ◽  
Baiping Mao ◽  
...  
Keyword(s):  
Basement Membrane ◽  
Sertoli Cell ◽  
Cell Function ◽  
Seminiferous Epithelium ◽  
Loss Of Function ◽  
Specific Expression ◽  
Specific Distribution ◽  
Regulatory Effects

Abstract Throughout spermatogenesis, cellular cargoes including haploid spermatids are required to be transported across the seminiferous epithelium, either toward the microtubule (MT) plus (+) end near the basement membrane at stage V, or to the MT minus (−) end near the tubule lumen at stages VI to VIII of the epithelial cycle. Furthermore, preleptotene spermatocytes, differentiated from type B spermatogonia, are transported across the Sertoli cell blood-testis barrier (BTB) to enter the adluminal compartment. Few studies, however, have been conducted to explore the function of MT-dependent motor proteins to support spermatid transport during spermiogenesis. Herein, we examined the role of MT-dependent and microtubule plus (+) end–directed motor protein kinesin 15 (KIF15) in the testis. KIF15 displayed a stage-specific expression across the seminiferous epithelium, associated with MTs, and appeared as aggregates on the MT tracks that aligned perpendicular to the basement membrane and laid across the entire epithelium. KIF15 also tightly associated with apical ectoplasmic specialization, displaying strict stage-specific distribution, apparently to support spermatid transport across the epithelium. We used a loss-of-function approach by RNAi to examine the role of KIF15 in Sertoli cell epithelium in vitro to examine its role in cytoskeletal-dependent Sertoli cell function. It was noted that KIF15 knockdown by RNAi that reduced KIF15 expression by ~70% in Sertoli cells with an established functional tight junction barrier impeded the barrier function. This effect was mediated through remarkable changes in the cytoskeletal organization of MTs, but also actin-, vimentin-, and septin-based cytoskeletons, illustrating that KIF15 exerts its regulatory effects well beyond microtubules.

Hemolytic Uremic Syndrome Is an Immune Mediated Disease: Role of Anti-CD36 Antibodies.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3990-3990
Author(s):  
Gail Rock ◽  
William Clark ◽  
Marion Sternbach ◽  
Maria Kolajova ◽  
Peter McLaine
Keyword(s):  
Hemolytic Uremic Syndrome ◽  
Molecular Mimicry ◽  
Platelet Lysate ◽  
Mechanism Of Formation ◽  
Normal Platelet ◽  
Immune Mediated ◽  
Uremic Syndrome ◽  
Membrane Bound ◽  
Immunological Pathogenesis

Abstract Hemolytic uremic syndrome is a disorder in which platelet microthrombi are formed which have a particular propensity to deposit in the kidney microvasculature resulting in impaired renal function and thrombocytopenia. The mechanism of formation of these microthrombi is not known. In this study, we showed that plasma from five adult and six pediatric cases of HUS caused aggregation and release of adenosine triphosphate from normal platelets. The plasma reacted against platelet lysate in a protein blot and all samples showed reactivity against a band at 88kDa corresponding to the membrane antigen CD36. This was confirmed by probing with Mo91, a monoclonal antibody to CD36. CD36 was also identified in the immune complex formed by incubation of patient plasmas with normal platelet lysate. In other studies, bands of 32kDa and 7.7kDa were obtained when purified verotoxin was protein blotted and probed with either patient plasma or with anti-CD36 antibody Mo91 suggesting structural homologies between CD36 and verotoxin. The data support the concept of an immunological pathogenesis for HUS and suggest that molecular mimicry involving one or both of the homologous domains in membrane-bound CD36 and verotoxin lead to the development of antibodies capable of inducing the pathophysiological events characteristic of HUS.

scholarly journals SIRT5 regulates autophagy and apoptosis in gastric cancer cells

2021 ◽  
Vol 49 (2) ◽  
pp. 030006052098635
Author(s):  
Wen Gu ◽  
Qinyi Qian ◽  
Yinkai Xu ◽  
Xiaolan Xu ◽  
Liping Zhang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cancer Cells ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Mammalian Target Of Rapamycin ◽  
Gastric Cancer Cells ◽  
Protein Levels ◽  
Amp Activated Protein Kinase ◽  
Related Proteins

Objective Accumulating evidence illustrates that sirtuins (SIRTs) regulate autophagy and apoptosis in cancer cells; however, the role of SIRT5 in gastric cancer (GC) cells remains unknown. In this study, we examined the role of SIRT5 in GC cells. Methods We detected SIRT5 protein levels in freshly collected samples from patients with GC. Next, we studied the function of SIRT5 in autophagy. Furthermore, the signaling pathway through which SIRT5 enhanced autophagy in GC cells was detected. In addition, we established a GC cell apoptosis model to analyze the role of SIRT5 in apoptosis. Results SIRT5 expression was downregulated in GC tissues. We discovered that SIRT5 promoted autophagy in GC cells. We demonstrated that SIRT5 enhanced autophagy in GC cells via the AMP-activated protein kinase–mammalian target of rapamycin signaling pathway. In addition, SIRT5 was degraded during apoptosis in GC cells. Meanwhile, we observed that calpains and caspase-related proteins were associated with SIRT5-related GC cell apoptosis. Conclusions SIRT5 is a crucial regulator of autophagy and apoptosis in GC cell lines that can maintain the balance of autophagy and apoptosis.

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