scholarly journals FK506 Induces the TGF-β1/Smad 3 Pathway Independently of Calcineurin Inhibition to Prevent Intervertebral Disk Degeneration

2020 ◽  
Vol 8 ◽  
Author(s):  
Jun Ge ◽  
Yingjie Wang ◽  
Qi Yan ◽  
Cenhao Wu ◽  
Hao Yu ◽  
...  
Keyword(s):  
Cell Function ◽  
Underlying Mechanism ◽  
Optical Microscope ◽  
Immunosuppressive Agent ◽  
Intervertebral Disk ◽  
Mrna Levels ◽  
Matrix Components ◽  
Ivd Degeneration ◽  
Calcineurin Inhibition

BackgroundIntervertebral disk (IVD) degeneration is the most common cause of lower back pain. Inhibiting inflammation is a key strategy for delaying IVD degeneration. Tacrolimus (FK506) is a potent immunosuppressive agent that is also beneficial to chondrocytes via alleviating inflammation. However, the potential function of FK506 in IVD and the underlying mechanisms remain unknown. The current study is aim at exploring the underlying mechanism of FK506 in preventing IVD degeneration.MethodsCell morphology was imaged using an optical microscope. mRNA levels of nucleus pulposus (NP) matrix components were determined by qRT-PCR, and protein expression NP matrix components was assessed by western blotting. A rat caudal IVD degeneration model was established to test for FK506 in vivo.ResultsFK506 improved the morphology of NP cells and the cell function at both the mRNA and protein level. FK506 could attenuate NP degeneration induced by IL-1β. Furthermore, FK506 exerted its function via TGFβ/Smad3 activation instead of through calcineurin inhibition. Inhibition of the TGF-β pathway prevented the protective effect of FK506 on IVD degeneration. In an in vivo study, FK506 injection reversed the development of rat caudal IVD degeneration influenced by Smad3.ConclusionOur current study demonstrates the positive effect of FK506 on delaying the degeneration of IVD via the TGFβ/Smad3 pathway.

scholarly journals Glucolipotoxicity and GLP-1 secretion

2021 ◽  
Vol 9 (1) ◽  
pp. e001905
Author(s):  
Jung-Hee Hong ◽  
Dae-Hee Kim ◽  
Moon-Kyu Lee
Keyword(s):  
Glucose Transporter ◽  
Cell Function ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Gene Expressions ◽  
Simultaneous Treatment ◽  
L Cells ◽  
Triglyceride Accumulation

IntroductionThe concept of glucolipotoxicity refers to the combined, deleterious effects of elevated glucose and/or fatty acid levels.Research design and methodsTo investigate the effects of chronic glucolipotoxicity on glucagon-like peptide-1-(7-36) amide (GLP-1) secretion, we generated glucolipotoxic conditions in human NCI-H716 enteroendocrine cells using either 5 or 25 mM glucose with or without 500 µM palmitate for 72 hours. For in vivo study, we have established a chronic nutrient infusion model in the rat. Serial blood samples were collected for 2 hours after the consumption of a mixed meal to evaluate insulin sensitivity and β-cell function.ResultsChronic glucolipotoxic conditions decreased GLP-1 secretion and the expressions of pCREB, pGSK3β, β-catenin, and TCF7L2 in NCI-H716 cells. Glucolipotoxicity conditions reduced glucose transporter expression, glucose uptake, and nicotinamide-adenine dinucleotide phosphate (NADPH) levels in L-cells, and increased triglyceride accumulation. In contrast, PPARα and ATP levels were reduced, which correlated well with decreased levels of SUR1 and Kir6.2, cAMP contents and expressions of pCAMK2, EPAC and PKA. We also observed an increase in reactive oxygen species production, UCP2 expression and Complex I activity. Simultaneous treatment with insulin restored the GLP-1 secretion. Glucolipotoxic conditions decreased insulin secretion in a time-dependent manner in INS-1 cells, which was recovered with exendin-4 cotreatment. Glucose and SMOFlipid infusion for 6 hours decreased GLP-1 secretion and proglucagon mRNA levels as well as impaired the glucose tolerance, insulin and C-peptide secretion in rats.ConclusionThese results provide evidence for the first time that glucolipotoxicity could affect GLP-1 secretion through changes in glucose and lipid metabolism, gene expressions, and proglucagon biosynthesis and suggest the interrelationship between glucolipotoxicities of L-cells and β-cells which develops earlier than that of L-cells.

scholarly journals Fully reduced HMGB1 accelerates the regeneration of multiple tissues by transitioning stem cells to GAlert

2018 ◽  
Vol 115 (19) ◽  
pp. E4463-E4472 ◽  
Author(s):  
Geoffrey Lee ◽  
Ana Isabel Espirito Santo ◽  
Stefan Zwingenberger ◽  
Lawrence Cai ◽  
Thomas Vogl ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Tissue Regeneration ◽  
Cell Function ◽  
Elective Surgery ◽  
High Mobility ◽  
Underlying Mechanism ◽  
Clinical Scenarios ◽  
Stem And Progenitor Cells

A major discovery of recent decades has been the existence of stem cells and their potential to repair many, if not most, tissues. With the aging population, many attempts have been made to use exogenous stem cells to promote tissue repair, so far with limited success. An alternative approach, which may be more effective and far less costly, is to promote tissue regeneration by targeting endogenous stem cells. However, ways of enhancing endogenous stem cell function remain poorly defined. Injury leads to the release of danger signals which are known to modulate the immune response, but their role in stem cell-mediated repair in vivo remains to be clarified. Here we show that high mobility group box 1 (HMGB1) is released following fracture in both humans and mice, forms a heterocomplex with CXCL12, and acts via CXCR4 to accelerate skeletal, hematopoietic, and muscle regeneration in vivo. Pretreatment with HMGB1 2 wk before injury also accelerated tissue regeneration, indicating an acquired proregenerative signature. HMGB1 led to sustained increase in cell cycling in vivo, and using Hmgb1−/− mice we identified the underlying mechanism as the transition of multiple quiescent stem cells from G0 to GAlert. HMGB1 also transitions human stem and progenitor cells to GAlert. Therefore, exogenous HMGB1 may benefit patients in many clinical scenarios, including trauma, chemotherapy, and elective surgery.

scholarly journals The Circ_0001367/miR-545-3p/LUZP1 Axis Regulates Cell Proliferation, Migration and Invasion in Glioma Cells

2021 ◽  
Vol 11 ◽  
Author(s):  
Xuchen Dong ◽  
Peng Zhang ◽  
Liang Liu ◽  
Haoran Li ◽  
Shan Cheng ◽  
...  
Keyword(s):  
Leucine Zipper ◽  
Cell Function ◽  
Glioma Cells ◽  
Malignant Tumour ◽  
Underlying Mechanism ◽  
Circular Rna ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Loss Of Function

Glioma is the most common primary intracranial malignant tumour in adults. It has a high incidence and poses a serious threat to human health. Circular RNA is a hotspot of cancer research. In this study, we aimed to explore the role of circ_0001367 in gliomagenesis and the underlying mechanism. First, qRT-PCR was conducted, which showed that circ_0001367 level was downregulated in glioma tissues and cells. Next, gain-of-function and loss-of-function assays were performed, which indicated that circ_0001367 inhibited the proliferation, migration and invasion of glioma cells. Subsequent bioinformatics analysis, dual-luciferase reporter assays, RNA immunoprecipitation assays and cell function assays demonstrated that circ_0001367 inhibited the proliferation, migration and invasion of glioma cells by absorbing miR-545-3p and thereby regulating the expression of leucine zipper protein (LUZP1). Finally, an in vivo experiment was conducted, which demonstrated that circ_0001367 inhibited glioma growth in vivo by modulating miR-545-3p and LUZP1. Taken together, the results of this study demonstrate that the circ_0001367/miR-545-3p/LUZP1 axis may be a novel target for glioma therapy.

scholarly journals Integrin α6 upregulation in nucleus pulposus cell under high oxygen tension attenuates intervertebral disc degeneration

2021 ◽  
Author(s):  
Zeng Xu ◽  
Jiancheng Zheng ◽  
Ying Zhang ◽  
Huiqiao Wu ◽  
Bin Sun ◽  
...  
Keyword(s):  
Oxygen Tension ◽  
Protective Factor ◽  
Cell Function ◽  
Hypoxia Inducible Factor ◽  
Nucleus Pulposus Cell ◽  
Intervertebral Disk ◽  
High Oxygen ◽  
Low Oxygen ◽  
Induced Apoptosis

AbstractThe destruction of low oxygen microenvironment played critical roles in the pathogenesis of intervertebral disk degeneration (IVDD). In this study, high oxygen tension (HOT) treatment upregulated integrin α6(ITG α6) expression, which could be alleviated by blocking PI3K/AKT signaling pathway. And the levels of ITG α6 expression were increased in the NP tissue from IVDD patients and IVDD rat model with mild degeneration, which were reduced as degeneration degree increases. Further studies found that ITG α6 could protect NP cells against HOT-induced apoptosis and oxidative stress, and protect NP cells from HOT-inhibited ECM proteins synthesis. ITG α6 upregulation by HOT contributed to maintain a NP tissue homeostasis through the interaction with hypoxia inducible factor-1α (HIF-1α). Furthermore, silencing of ITG α6 in vivo could obviously accelerate puncture-induced IVDD. Taken together, ITG α6 upregulation by HOT in NP cells might be a protective factor in IVDD as well as restore NP cell function.

Inosine Monophosphate Dehydrogenase II Mutant (Thr-333-Ile + Ser-351-Tyr) Does Not Confer Resistance to Mycophenolic Acid In Vivo.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5226-5226
Author(s):  
Mark A. Schroeder ◽  
Michael P. Rettig ◽  
Julie K. Ritchey ◽  
John F. DiPersio
Keyword(s):  
T Cells ◽  
Cell Function ◽  
Immunosuppressive Agents ◽  
Retroviral Vectors ◽  
Immunosuppressive Agent ◽  
Inosine Monophosphate Dehydrogenase ◽  
Over Expression ◽  
Donor T Cells

Abstract Several immunosuppressive agents are currently being evaluated for their ability to inhibit host immune responses that prevent donor engraftment after reduced intensity allogeneic BMT. Unfortunately, these immunosuppressive agents have the potential to inhibit the beneficial graft-versus-infection and graft-versus-leukemia effects mediated by mature donor T cells that are present in the graft. One potential approach to retain donor T cell function in immunosuppressed recipients after allogeneic BMT is to genetically modify the donor T cells with a drug resistance gene that confers protection from the effects of the immunosuppressive agent. Mycophenolate mofetil (MMF), the morpholino-ethyl ester precursor of the active compound mycophenolic acid (MPA), is an immunosuppressive agent that prevents cell proliferation by inhibiting the enzyme inosine monophosphate dehydrogenase (IMPDH) II and has been used in conjunction with other immunosuppressants to promote donor engraftment and limit graft versus host disease after nonmyeloablative BMT. IMPDH II is the rate-limiting enzyme in de novo guanosine synthesis and is preferentially expressed in activated T and B cells. A number of investigators have identified IMPDH II mutants that have altered MPA binding capacity and normal guanosine synthesis in vitro (BBA2002;1594:27, JBC.1997;272:961, BBA.1994;1217:156). One prototypic mutant (IMPDH*; Thr-333-Ile, Ser-351-Tyr) has been used for in vivo selection of donor T-cells in a canine model (Blood2003;102:3696a) We hypothesized that over expression of IMPDH* in donor T cells would provide resistance to MPA in vitro and MMF in vivo, thus allowing MMF to be used in vivo to inhibit recipient immune responses while maintaining normal donor T cell function. In our initial studies, we transduced murine A20 cells (a B-lymphoblastic leukemia) with retroviral vectors encoding IMPDH or IMPDH* fused to the C-terminus of our previously described CD34/HSV-TK chimeric suicide gene. This strategy enabled us to rapidly and efficiently select genetically modified cells using a well-established CD34 immunoselection technique. Unfortunately, the CD34/TK/IMPDH* fusion protein failed to confer MPA resistance to A20 cells in vitro. To determine whether this failure to confer MPA resistance was caused by fusing IMPDH* to CD34/TK, we generated bicistronic retroviral vectors that expressed either IMPDH or IMPDH* and EGFP using an internal ribosome entry site (IRES). Human Jurkat T cells were efficiently transduced with the bicistronic vectors and GFP+ cells were selected using a MoFlo cell sorter. As before, forced expression of IMPDH* failed to confer MPA resistance to the transduced and selected cells in vitro (IC50 = 0.5uM). Interestingly, chronic selection of transduced Jurkat cells in 1 uM MPA for 2 weeks led to a 4 fold increase in the IC50 (IC50 = 2uM MPA). These studies suggest that the resistance to MPA observed in vivo by others for IMPDH* may not be derived from the aforementioned point mutations in IMPDH but rather to over expression or up-regulation of IMPDH II, which may act as a sink for MPA. Alternatively, differences between species and cell lines, introduction of novel mutations during selection, or alternative salvage pathways may be activated resulting in MPA resistance. We subsequently have generated six additional mutants of the MPA binding site based on in vitro data and x-ray crystallography of the MPA binding site and we have began expressing them in our in vivo model. Analysis of these mutants is ongoing.

Targeting Hepcidin with Antisense Oligonucletides Improves Anemia Endpoints in Mice.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 269-269 ◽  
Author(s):  
Jeffrey R. Crosby ◽  
William A. Gaarde ◽  
Jarrett Egerston ◽  
Robert McKay ◽  
Yingqing Sun ◽  
...  
Keyword(s):  
Serum Iron ◽  
Cell Function ◽  
Iron Absorption ◽  
Mrna Levels ◽  
Erythroid Progenitor ◽  
Primary Mouse ◽  
Progenitor Cell Proliferation ◽  
Hepcidin Mrna

Abstract Anemia is one of the more common blood disorders and is associated with a number of diseases, including chronic kidney disease, chronic inflammation, and certain types of cancer. Under these conditions iron is essential as it is required for erythroid progenitor cell proliferation and red cell function. Hepcidin is a liver-derived growth factor that regulates iron absorption in the GI tract and iron absorption and release in tissues. Furthermore, hepcidin overexpression has been strongly linked mechanistically as a mediator of decreased iron availability and anemia. We have utilized an antisense approach to investigate the role of hepcidin in animal models of anemia and as a potential therapeutic approach for the treatment of this disorder in humans. Second-generation 2′-O-methoxyethyl chimeric antisense oligonucleotides (ASOs) were screened in isolated primary mouse hepatocytes, followed by in vivo screening in mice, for the ability to reduce hepcidin mRNA levels. ASO treatment resulted in a reduction of hepcidin mRNA in liver which was associated with a significant increase in serum iron levels. The best hepcidin ASO was then tested in a mouse model of turpentine induced hypoferremia and anemia to determine the role of hepcidin in regulating serum iron and anemia endpoints. Mice were treated (I.P. twice/weekly) with hepcidin or control ASO for two weeks at varying doses prior to a single subcutaneous injection of turpentine. Turpentine treatment 16 hours post-injection produced a significant reduction in serum iron levels and at two weeks resulted in reduced RBC numbers, hematocrit and hemoglobin levels. Treatment with the hepcidin ASO resulted in a dose dependent improvement in all of these endpoints while the control oligonucleotide had no effect. Studies are in progress to further characterize the pharmacological activity of hepcidin ASO in additional models of anemia and results from these on-going studies will also be presented.

scholarly journals Antiobesity Effects of UnripeRubus coreanusMiquel and Its Constituents: AnIn VitroandIn VivoCharacterization of the Underlying Mechanism

2016 ◽  
Vol 2016 ◽  
pp. 1-15 ◽  
Author(s):  
Dool-Ri Oh ◽  
Yujin Kim ◽  
Eun-jin Choi ◽  
Hunmi-Lee ◽  
Myung-A Jung ◽  
...  
Keyword(s):  
Body Weight ◽  
Adipose Tissue ◽  
Bioactive Compounds ◽  
Lipid Accumulation ◽  
Weight Control ◽  
Underlying Mechanism ◽  
Serum Glucose ◽  
Mrna Levels ◽  
Dependent Manner

Background. The objective of the present study was to perform a bioguided fractionation of unripeRubus coreanusMiquel (uRC) and evaluate the lipid accumulation system involvement in its antiobesity activity as well as study the uRC mechanism of action.Results. After the fractionation, the BuOH fraction of uRC (uRCB) was the most active fraction, suppressing the differentiation of 3T3-L1 adipocytes in a dose-dependent manner. Moreover, after an oral administration for 8 weeks in HFD-induced obese mice, uRCB (10 and 50 mg/kg/day) produced a significant decrease in body weight, food efficiency ratio, adipose tissue weight and LDL-cholesterol, serum glucose, TC, and TG levels. Similarly, uRCB significantly suppressed the elevated mRNA levels of PPARγin the adipose tissuein vivo. Next, we investigated the antiobesity effects of ellagic acid, erycibelline, 5-hydroxy-2-pyridinemethanol, m-hydroxyphenylglycine, and 4-hydroxycoumarin isolated from uRCB. Without affecting cell viability, five bioactive compounds decreased the lipid accumulation in the 3T3-L1 cells and the mRNA expression levels of key adipogenic genes such as PPARγ, C/EBPα, SREBP-1c, ACC, and FAS.Conclusion. These results suggest that uRC and its five bioactive compounds may be a useful therapeutic agent for body weight control by downregulating adipogenesis and lipogenesis.

Growth hormone-induced STAT5B regulates Star gene expression through a cooperation with cJUN in mouse MA-10 Leydig cells

Endocrinology ◽  
2021 ◽  
Author(s):  
Pierre-Olivier Hébert-Mercier ◽  
Francis Bergeron ◽  
Nicholas M Robert ◽  
Samir Mehanovic ◽  
Kenley Joule Pierre ◽  
...  
Keyword(s):  
Gene Expression ◽  
Growth Hormone ◽  
Leydig Cell ◽  
Leydig Cells ◽  
Cell Function ◽  
Mrna Levels ◽  
Leydig Cell Function ◽  
Star Gene

Abstract Leydig cells produce androgens that are essential for male sex differentiation and reproductive function. Leydig cell function is regulated by several hormones and signaling molecules, including growth hormone (GH). Although GH is known to upregulate Star gene expression in Leydig cells, its molecular mechanism of action remains unknown. The STAT5B transcription factor is a downstream effector of GH signaling in other systems. While STAT5B is present in both primary and Leydig cell lines, its function in these cells has yet to be ascertained. Here we report that treatment of MA-10 Leydig cells with GH or overexpression of STAT5B induces Star mRNA levels and increases steroid hormone output. The mouse Star promoter contains a consensus STAT5B element (TTCnnnGAA) at -756 bp to which STAT5B binds in vitro (EMSA and supershift) and in vivo (ChIP) in a GH-induced manner. In functional promoter assays, STAT5B was found to activate a -980 bp mouse Star reporter. Mutating the -756 bp element prevented STAT5B binding but did not abrogate STAT5B-responsiveness. STAT5B was found to functionally cooperate with DNA-bound cJUN. The STAT5B/cJUN cooperation was only observed in Leydig cells and not in Sertoli or fibroblast cells, indicating that additional Leydig cell-enriched transcription factors are required. The STAT5B/cJUN cooperation was lost only when both STAT5B and cJUN elements were mutated. In addition to identifying the Star gene as a novel target for STAT5B in Leydig cells, our data provide important new insights into the mechanism of GH and STAT5B action in the regulation of Leydig cell function.

scholarly journals Anti-inflammatory and Pro-apoptotic Effects of 18beta-Glycyrrhetinic Acid In Vitro and In Vivo Models of Rheumatoid Arthritis

2021 ◽  
Vol 12 ◽  
Author(s):  
Yunhui Feng ◽  
Liyan Mei ◽  
Maojie Wang ◽  
Qingchun Huang ◽  
Runyue Huang
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Cycle ◽  
Cell Apoptosis ◽  
Underlying Mechanism ◽  
Mrna Levels ◽  
Pathological Changes ◽  
Collagen Induced Arthritis ◽  
Tnf Α

18β-Glycyrrhetinic acid (18β-GA), an active component from Glycyrrhiza glabra L. root (licorice), has been demonstrated to be able to protect against inflammatory response and reduce methotrexate (MTX)-derived toxicity. This study was therefore designed to test the therapeutic possibility of 18β-GA on rheumatoid arthritis (RA) and to explore the underlying mechanism. LPS or TNF-α-induced inflammatory cell models and collagen-induced arthritis (CIA) animal models were applied in this study. Real-time quantitative PCR (RT-qPCR) was used to measure the mRNA levels of various cytokines and FOXO family members. The protein levels of molecules in the MAPK/NF-κB signaling pathway were analyzed using western blot. The cell proliferation assay and colony-forming assay were used to test the influence of 18β-GA on cell viability. The cell apoptosis assay and cell cycle assay were performed to detect the effect of 18β-GA on cell proliferative capacity by using flow cytometry. Hematoxylin and eosin (H&E) staining was performed to evaluate pathological changes after drug administration. The enzyme-linked immunosorbent assay (ELISA) was carried out for the detection of cytokines in serum. In vitro, we found that 18β-GA decreased the mRNA levels of IL-1β, IL-6, and COX-2 by inhibiting the MAPK/NF-κB signaling pathway in MH7A and RAW264.7 cell lines. Moreover, 18β-GA was able to suppress cell viability, trigger cell apoptosis, and G1 phase cell cycle arrest in our in vitro studies. 18β-GA dramatically enhanced the mRNA level of FOXO3 in both TNF-α- and LPS-induced inflammation models in vitro. Interestingly, after analyzing GEO datasets, we found that the FOXO3 gene was significantly decreased in the RA synovial tissue as compared to healthy donors in multiple microarray studies. In vivo, 18β-GA exhibited a promising therapeutic effect in a collagen-induced arthritis mouse model by alleviating joint pathological changes and declining serum levels of TNF-α, IL-1β, and IL-6. Finally, we observed that 18β-GA administration could mitigate liver damage caused by collagen or MTX. Collectively, the current study demonstrates for the first time that 18β-GA can inhibit inflammation and proliferation of synovial cells, and the underlying mechanism may be associated with its inhibition of MAPK/NF-κB signaling and promotion of FOXO3 signaling. Therefore, 18β-GA is expected to be a new drug candidate for RA therapy.

scholarly journals Oct1 and OCA-B are selectively required for CD4 memory T cell function

2015 ◽  
Vol 212 (12) ◽  
pp. 2115-2131 ◽  
Author(s):  
Arvind Shakya ◽  
Alon Goren ◽  
Alex Shalek ◽  
Cody N. German ◽  
Jeremy Snook ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Function ◽  
Underlying Mechanism ◽  
Potential Method ◽  
Memory Failure ◽  
Lysine Demethylase ◽  
Poor Memory

Epigenetic changes are crucial for the generation of immunological memory. Failure to generate or maintain these changes will result in poor memory responses. Similarly, augmenting or stabilizing the correct epigenetic states offers a potential method of enhancing memory. Yet the transcription factors that regulate these processes are poorly defined. We find that the transcription factor Oct1 and its cofactor OCA-B are selectively required for the in vivo generation of CD4+ memory T cells. More importantly, the memory cells that are formed do not respond properly to antigen reencounter. In vitro, both proteins are required to maintain a poised state at the Il2 target locus in resting but previously stimulated CD4+ T cells. OCA-B is also required for the robust reexpression of multiple other genes including Ifng. ChIPseq identifies ∼50 differentially expressed direct Oct1 and OCA-B targets. We identify an underlying mechanism involving OCA-B recruitment of the histone lysine demethylase Jmjd1a to targets such as Il2, Ifng, and Zbtb32. The findings pinpoint Oct1 and OCA-B as central mediators of CD4+ T cell memory.

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