scholarly journals NK3R Mediates the EGF-Induced SLα Secretion and mRNA Expression in Grass Carp Pituitary

2018 ◽  
Vol 20 (1) ◽  
pp. 91 ◽  
Author(s):  
Xiangfeng Qin ◽  
Cheng Ye ◽  
Xiaoyun Zhou ◽  
Jingyi Jia ◽  
Shaohua Xu ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Grass Carp ◽  
Cell Function ◽  
Cell Types ◽  
Pituitary Cells ◽  
Dependent Manner ◽  
Lower Vertebrates ◽  
Mrna And Protein Expression

Epidermal growth factor (EGF) is a potent regulator of cell function in many cell types. In mammals, the EGF/EGFR system played an important role in both pituitary physiology and pathology. However, it is not clear about the pituitary action of EGF in lower vertebrates. In this study, using grass carp as a model, we found that EGF could stimulate NK3R mRNA and protein expression through pituitary ErbB1 and ErbB2 coupled to MEK/ERK and PI3K/Akt/mTOR pathways. In addition, EGF could also induce pituitary somatolactin α (SLα) secretion and mRNA expression in a dose- and time-dependent manner in vivo and in vitro. The stimulatory actions of EGF on SLα mRNA expression were also mediated by PI3K/Akt/mTOR and MEK/ERK pathways coupled to ErbB1 and ErbB2 activation. Our previous study has reported that neurokinin B (NKB) could also induce SLα secretion and mRNA expression in carp pituitary cells. In the present study, interestingly, we found that EGF could significantly enhance NKB-induced SLα mRNA expression. Further studies found that NK3R antagonist SB222200 could block EGF-induced SLα mRNA expression, indicating an NK3R requirement. Furthermore, cAMP/PKA inhibitors and PLC/PKC inhibitors could both abolish EGF- and EGF+NKB-induced SLα mRNA expression, which further supported that EGF-induced SLα mRNA expression is NK3R dependent.

scholarly journals Mn Inhibits GSH Synthesis via Downregulation of Neuronal EAAC1 and Astrocytic xCT to Cause Oxidative Damage in the Striatum of Mice

2018 ◽  
Vol 2018 ◽  
pp. 1-15 ◽  
Author(s):  
Xinxin Yang ◽  
Haibo Yang ◽  
Fengdi Wu ◽  
Zhipeng Qi ◽  
Jiashuo Li ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Oxidative Damage ◽  
Dependent Manner ◽  
Protein Levels ◽  
Key Factor ◽  
Protein Sulfhydryl ◽  
Mrna And Protein Expression ◽  
The Brain

Excessive manganese (Mn) can accumulate in the striatum of the brain following overexposure. Oxidative stress is a well-recognized mechanism in Mn-induced neurotoxicity. It has been proven that glutathione (GSH) depletion is a key factor in oxidative damage during Mn exposure. However, no study has focused on the dysfunction of GSH synthesis-induced oxidative stress in the brain during Mn exposure. The objective of the present study was to explore the mechanism of Mn disruption of GSH synthesis via EAAC1 and xCT in vitro and in vivo. Primary neurons and astrocytes were cultured and treated with different doses of Mn to observe the state of cells and levels of GSH and reactive oxygen species (ROS) and measure mRNA and protein expression of EAAC1 and xCT. Mice were randomly divided into seven groups, which received saline, 12.5, 25, and 50 mg/kg MnCl2, 500 mg/kg AAH (EAAC1 inhibitor) + 50 mg/kg MnCl2, 75 mg/kg SSZ (xCT inhibitor) + 50 mg/kg MnCl2, and 100 mg/kg NAC (GSH rescuer) + 50 mg/kg MnCl2 once daily for two weeks. Then, levels of EAAC1, xCT, ROS, GSH, malondialdehyde (MDA), protein sulfhydryl, carbonyl, 8-hydroxy-2-deoxyguanosine (8-OHdG), and morphological and ultrastructural features in the striatum of mice were measured. Mn reduced protein levels, mRNA expression, and immunofluorescence intensity of EAAC1 and xCT. Mn also decreased the level of GSH, sulfhydryl, and increased ROS, MDA, 8-OHdG, and carbonyl in a dose-dependent manner. Injury-related pathological and ultrastructure changes in the striatum of mice were significantly present. In conclusion, excessive exposure to Mn disrupts GSH synthesis through inhibition of EAAC1 and xCT to trigger oxidative damage in the striatum.

scholarly journals Tenovin-6 impairs autophagy by inhibiting autophagic flux

2017 ◽  
Vol 8 (2) ◽  
pp. e2608-e2608 ◽  
Author(s):  
Hongfeng Yuan ◽  
Brandon Tan ◽  
Shou-Jiang Gao
Keyword(s):  
Cell Types ◽  
Inhibitory Effect ◽  
Lymphocytic Leukemia ◽  
Autophagic Flux ◽  
Dependent Manner ◽  
Autophagy Pathway ◽  
Sarcoma Cells ◽  
Regulatory Functions

Abstract Tenovin-6 has attracted significant interest because it activates p53 and inhibits sirtuins. It has anti-neoplastic effects on multiple hematopoietic malignancies and solid tumors in both in vitro and in vivo studies. Tenovin-6 was recently shown to impair the autophagy pathway in chronic lymphocytic leukemia cells and pediatric soft tissue sarcoma cells. However, whether tenovin-6 has a general inhibitory effect on autophagy and whether there is any involvement with SIRT1 and p53, both of which are regulators of the autophagy pathway, remain unclear. In this study, we have demonstrated that tenovin-6 increases microtubule-associated protein 1 light chain 3 (LC3-II) level in diverse cell types in a time- and dose-dependent manner. Mechanistically, the increase of LC3-II by tenovin-6 is caused by inhibition of the classical autophagy pathway via impairing lysosomal function without affecting the fusion between autophagosomes and lysosomes. Furthermore, we have revealed that tenovin-6 activation of p53 is cell type dependent, and tenovin-6 inhibition of autophagy is not dependent on its regulatory functions on p53 and SIRT1. Our results have shown that tenovin-6 is a potent autophagy inhibitor, and raised the precaution in interpreting results where tenovin-6 is used as an inhibitor of SIRT1.

scholarly journals Mitochondrial Calcium Uptake Capacity Modulates Neocortical Excitability

2013 ◽  
Vol 33 (7) ◽  
pp. 1115-1126 ◽  
Author(s):  
Basavaraju G Sanganahalli ◽  
Peter Herman ◽  
Fahmeed Hyder ◽  
Sridhar S Kannurpatti
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Cell Types ◽  
Dependent Manner ◽  
Evoked Responses ◽  
Blood Oxygen Level ◽  
Calcium Uniporter ◽  
Sensory Evoked

Local calcium (Ca2 +) changes regulate central nervous system metabolism and communication integrated by subcellular processes including mitochondrial Ca2 + uptake. Mitochondria take up Ca2 + through the calcium uniporter (mCU) aided by cytoplasmic microdomains of high Ca2 +. Known only in vitro, the in vivo impact of mCU activity may reveal Ca2 + -mediated roles of mitochondria in brain signaling and metabolism. From in vitro studies of mitochondrial Ca2 + sequestration and cycling in various cell types of the central nervous system, we evaluated ranges of spontaneous and activity-induced Ca2 + distributions in multiple subcellular compartments in vivo. We hypothesized that inhibiting (or enhancing) mCU activity would attenuate (or augment) cortical neuronal activity as well as activity-induced hemodynamic responses in an overall cytoplasmic and mitochondrial Ca2 + -dependent manner. Spontaneous and sensory-evoked cortical activities were measured by extracellular electrophysiology complemented with dynamic mapping of blood oxygen level dependence and cerebral blood flow. Calcium uniporter activity was inhibited and enhanced pharmacologically, and its impact on the multimodal measures were analyzed in an integrated manner. Ru360, an mCU inhibitor, reduced all stimulus-evoked responses, whereas Kaempferol, an mCU enhancer, augmented all evoked responses. Collectively, the results confirm aforementioned hypotheses and support the Ca2 + uptake-mediated integrative role of in vivo mitochondria on neocortical activity.

scholarly journals Administration of cardiac mesenchymal cells modulates innate immunity in the acute phase of myocardial infarction in mice

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Yi Kang ◽  
Marjan Nasr ◽  
Yiru Guo ◽  
Shizuka Uchida ◽  
Tyler Weirick ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Innate Immunity ◽  
Mechanism Of Action ◽  
Cell Activation ◽  
Cell Function ◽  
Immune Cell ◽  
Mesenchymal Cell ◽  
Cell Types

Abstract Although cardiac mesenchymal cell (CMC) therapy mitigates post-infarct cardiac dysfunction, the underlying mechanisms remain unidentified. It is acknowledged that donor cells are neither appreciably retained nor meaningfully contribute to tissue regeneration—suggesting a paracrine-mediated mechanism of action. As the immune system is inextricably linked to wound healing/remodeling in the ischemically injured heart, the reparative actions of CMCs may be attributed to their immunoregulatory properties. The current study evaluated the consequences of CMC administration on post myocardial infarction (MI) immune responses in vivo and paracrine-mediated immune cell function in vitro. CMC administration preferentially elicited the recruitment of cell types associated with innate immunity (e.g., monocytes/macrophages and neutrophils). CMC paracrine signaling assays revealed enhancement in innate immune cell chemoattraction, survival, and phagocytosis, and diminished pro-inflammatory immune cell activation; data that identifies and catalogues fundamental immunomodulatory properties of CMCs, which have broad implications regarding the mechanism of action of CMCs in cardiac repair.

scholarly journals Extracellular SPARC cooperates with TGF-β signalling to induce pro-fibrotic activation of systemic sclerosis patient dermal fibroblasts

Rheumatology ◽  
2019 ◽  
Vol 59 (9) ◽  
pp. 2258-2263 ◽  
Author(s):  
Tiago Carvalheiro ◽  
Beatriz Malvar Fernández ◽  
Andrea Ottria ◽  
Barbara Giovannone ◽  
Wioleta Marut ◽  
...  
Keyword(s):  
Protein Expression ◽  
Therapeutic Target ◽  
Dermal Fibroblasts ◽  
Dependent Manner ◽  
Healthy Controls ◽  
Multiple Organs ◽  
Extracellular Matrix Components ◽  
Mrna And Protein Expression

Abstract Objectives SSc is an autoimmune disease characterized by inflammation, vascular injury and excessive fibrosis in multiple organs. Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein that regulates processes involved in SSc pathology, such as inflammation and fibrosis. In vivo and in vitro studies have implicated SPARC in SSc, but it is unclear if the pro-fibrotic effects of SPARC on fibroblasts are a result of intracellular signalling or fibroblast interactions with extracellular SPARC hampering further development of SPARC as a potential therapeutic target. This study aimed to analyse the potential role of exogenous SPARC as a regulator of fibrosis in SSc. Methods Dermal fibroblasts from both healthy controls and SSc patients were stimulated with SPARC alone or in combination with TGF-β1, in the absence or presence of a TGF receptor 1 inhibitor. mRNA and protein expression of extracellular matrix components and other fibrosis-related mediators were measured by quantitative PCR and western blot. Results Exogenous SPARC induced mRNA and protein expression of collagen I, collagen IV, fibronectin 1, TGF-β and SPARC by dermal fibroblasts from SSc patients, but not from healthy controls. Importantly, exogenous SPARC induced the activation of the tyrosine kinase SMAD2 and pro-fibrotic gene expression induced by SPARC in SSc fibroblasts was abrogated by inhibition of TGF-β signalling. Conclusion These results indicate that exogenous SPARC is an important pro-fibrotic mediator contributing to the pathology driving SSc but in a TGF-β dependent manner. Therefore, SPARC could be a promising therapeutic target for reducing fibrosis in SSc patients, even in late states of the disease.

scholarly journals Direct Stimulatory Effects of Oxytocin in Female Rat Gonadotrophs and Somatotrophs In Vitro: Comparison With Lactotrophs

Endocrinology ◽  
2014 ◽  
Vol 156 (2) ◽  
pp. 600-612 ◽  
Author(s):  
Arturo E. Gonzalez-Iglesias ◽  
Patrick A. Fletcher ◽  
José A. Arias-Cristancho ◽  
Ruth Cristancho-Gordo ◽  
Cleyde V. Helena ◽  
...  
Keyword(s):  
Receptor Antagonist ◽  
Cell Types ◽  
Pituitary Hormones ◽  
Prolactin Secretion ◽  
Female Rats ◽  
Pituitary Cells ◽  
Female Rat ◽  
Dependent Manner ◽  
Rat Pituitary Cells

The peptide oxytocin (OT) is secreted by hypothalamic neurons and exerts numerous actions related to reproduction. OT stimulation of prolactin secretion in female rats is important during the estrous cycle, pregnancy, and lactation. Here we report that OT also stimulates transients of intracellular Ca2+ concentration in somatotrophs and gonadotrophs as well as the release of GH and LH in a dose-dependent manner with EC50 values that closely correspond to the ligand affinity of the OT receptor (OTR). Remarkably, the hormone-releasing effect of OT in these two cell types is 2 orders of magnitude more sensitive than that in lactotrophs. The specific OTR agonist [Thr4,Gly7]-oxytocin acutely stimulated the release of LH, GH, and prolactin from female rat pituitary cells in primary culture and increased intracellular Ca2+ concentration in gonadotrophs, somatotrophs, and lactotrophs. In these three cell types, the effects on hormone release and intracellular Ca2+ of both OT and [Thr4,Gly7]oxytocin were abolished by the specific OT receptor antagonist desGly-NH2-d(CH2)5[D-Tyr2,Thr4]OVT but not by the highly selective vasopressin V1a receptor antagonist, d(CH2)5[Tyr(Me)2,Dab5]AVP. Furthermore, 10 nM arginine vasopressin stimulated LH and GH release comparably with a dose of OT that was at least 10 times lower. Finally, the presence of the OTR-like immunoreactivity could be observed in all three cell types. Taken together, these results show that OT directly stimulates gonadotrophs, somatotrophs, and lactotrophs through OT receptors and suggest that OT signaling may serve to coordinate the release of different pituitary hormones during specific physiological conditions.

scholarly journals Inhibition of Orbivirus Replication by Aurintricarboxylic Acid

2020 ◽  
Vol 21 (19) ◽  
pp. 7294
Author(s):  
Celia Alonso ◽  
Sergio Utrilla-Trigo ◽  
Eva Calvo-Pinilla ◽  
Luis Jiménez-Cabello ◽  
Javier Ortego ◽  
...  
Keyword(s):  
Cell Types ◽  
Dependent Manner ◽  
Biting Midges ◽  
Lethal Challenge ◽  
Concentration Dependent Manner ◽  
Antiviral Compound ◽  
Aurintricarboxylic Acid ◽  
And Control

Bluetongue virus (BTV) and African horse sickness virus (AHSV) are vector-borne viruses belonging to the Orbivirus genus, which are transmitted between hosts primarily by biting midges of the genus Culicoides. With recent BTV and AHSV outbreaks causing epidemics and important economy losses, there is a pressing need for efficacious drugs to treat and control the spread of these infections. The polyanionic aromatic compound aurintricarboxylic acid (ATA) has been shown to have a broad-spectrum antiviral activity. Here, we evaluated ATA as a potential antiviral compound against Orbivirus infections in both mammalian and insect cells. Notably, ATA was able to prevent the replication of BTV and AHSV in both cell types in a time- and concentration-dependent manner. In addition, we evaluated the effect of ATA in vivo using a mouse model of infection. ATA did not protect mice against a lethal challenge with BTV or AHSV, most probably due to the in vivo effect of ATA on immune system regulation. Overall, these results demonstrate that ATA has inhibitory activity against Orbivirus replication in vitro, but further in vivo analysis will be required before considering it as a potential therapy for future clinical evaluation.

scholarly journals Effect of GHRH and GHRP-2 treatment in vitro on GH secretion and levels of GH, pituitary transcription factor-1, GHRH-receptor, GH-secretagogue-receptor and somatostatin receptor mRNAs in ovine pituitary cells

2004 ◽  
pp. 235-242 ◽  
Author(s):  
M Yan ◽  
M Hernandez ◽  
R Xu ◽  
C Chen
Keyword(s):  
Transcription Factor ◽  
Somatostatin Receptor ◽  
Receptor Subtypes ◽  
Pituitary Cells ◽  
Dependent Manner ◽  
Gh Secretion ◽  
Somatostatin Receptor Subtypes ◽  
Transcription Factor 1

OBJECTIVE: Growth hormone (GH)-releasing hormone (GHRH) and GH-releasing peptides (GHRPs) stimulate the release of GH through their specific receptors on somatotropes. Combined GHRH and GHRP administration causes a synergistic GH release in vivo by an unknown mechanism. The current study focuses on the direct action of GHRH and GHRP on several molecular targets in somatotropes. DESIGN AND METHODS: To clarify the mechanism of action, ovine somatotropes were used to measure the expression of mRNAs encoding for GH, pituitary transcription factor-1 (Pit-1), GH-secretagogue receptor (GHS-R), GHRH-R, somatostatin receptor subtypes (sst-1 and sst-2) and GH release after GHRH and GHRP-2 treatment for 0.5, 1, 1.5 and 2 h. RESULTS: GHRH (10 nM), GHRP-2 (100 nM) and combined GHRH-GHRP-2 increased the levels of GH mRNA and GH release from 0.5 to 2 h in a time-dependent manner. The levels of Pit-1, GHRH-R and GHS-R mRNA were increased after 0.5 h treatment of cells with GHRH and GHRP-2. The levels of sst-1 but not sst-2 mRNA were significantly increased after 0.5 and 1 h of GHRH treatment. In contrast, both sst-1 and sst-2 mRNA expression was inhibited after 0.5-2 h of GHRP treatment. CONCLUSIONS: These data demonstrate a direct in vitro modification of ovine somatotropes by GHRH and GHRP-2 resulting in altered GHRH-R, GHS-R, Pit-1, sst-1, sst-2 and GH gene expression; this may underlie the regulatory action of GHRH and GHRP-2 on GH secretion.

Effects of crude and highly purified bovine inhibin (Mr 32 000 form) on gonadotrophin production by ovine pituitary cells in vitro: inhibin enhances gonadotrophin-releasing hormone-induced release of LH

1990 ◽  
Vol 127 (1) ◽  
pp. 149-159 ◽  
Author(s):  
S. Muttukrishna ◽  
P. G. Knight
Keyword(s):  
Primary Cultures ◽  
Suppressive Effect ◽  
Pituitary Cells ◽  
Dependent Manner ◽  
Α Subunit ◽  
Releasing Hormone ◽  
Lh Release ◽  
Gonadotrophin Releasing Hormone

ABSTRACT Primary cultures of ovine pituitary cells (from adult ewes) were used to investigate the actions of steroid-free bovine follicular fluid (bFF) and highly-purified Mr 32 000 bovine inhibin on basal and gonadotrophin-releasing hormone (GnRH)-induced release of FSH and LH. Residual cellular contents of each hormone were also determined allowing total gonadotrophin content/well to be calculated. As in rats, both crude and highly purified inhibin preparations promoted a dose (P < 0·001)- and time (P < 0·001)-dependent suppression of basal and GnRH-induced release of FSH as well as an inhibition of FSH synthesis, reflected by a fall in total FSH content/well. However, while neither inhibin preparation affected basal release of LH or total LH content/well, GnRH-induced LH release was significantly (P< 0·001) increased by the presence of either bFF (+ 75%) or highly-purified inhibin (+ 64%) in a dose- and time-dependent manner. This unexpected action of bFF on GnRH-induced LH release was abolished in the presence of 5 μl specific anti-inhibin serum, confirming that the response was indeed mediated by inhibin. Furthermore, neither oestradiol-17β (1 pmol/l–10 nmol/l) nor monomeric α-subunit of bovine inhibin (2·5–40 ng/ml) significantly affected basal or GnRH-induced release of LH. These in-vitro findings for the ewe lend support to a number of recent in-vivo observations and indicate that, in addition to its well-documented suppressive effect on the synthesis and secretion of FSH, inhibin may actually facilitate LH release in this species, in marked contrast to its action in the rat. Journal of Endocrinology (1990) 127, 149–159

A Device to Study the Effects of Stretch Gradients on Cell Behavior

2011 ◽  
Vol 133 (10) ◽  
Author(s):  
William J. Richardson ◽  
Richard P. Metz ◽  
Michael R. Moreno ◽  
Emily Wilson ◽  
James E. Moore
Keyword(s):  
Mrna Expression ◽  
Cell Function ◽  
Cyclic Stretch ◽  
Radial Deformation ◽  
Rigid Fixation ◽  
Cell Stretching ◽  
3T3 Fibroblasts ◽  
Collagen Iii

Mechanical forces are key regulators of cell function with varying loads capable of modulating behaviors such as alignment, migration, phenotype modulation, and others. Historically, cell-stretching experiments have employed mechanically simple environments (e.g., uniform uniaxial or equibiaxial stretches). However, stretch distributions in vivo can be highly non-uniform, particularly in cases of disease or subsequent to interventional treatments. Herein, we present a cell-stretching device capable of subjecting cells to controllable gradients in biaxial stretch via radial deformation of circular elastomeric membranes. By including either a defect or a rigid fixation at the center of the membrane, various gradients are generated. Capabilities of the device were quantified by tracking marked positions of the membrane while applying various loads, and experimental feasibility was assessed by conducting preliminary experiments with 3T3 fibroblasts and 10T1/2 cells subjected to 24 h of cyclic stretch. Quantitative real-time PCR was used to measure changes in mRNA expression of a profile of genes representing the major smooth muscle phenotypes. Genes associated with the contractile state were both upregulated (e.g., calponin) and downregulated (e.g., α-2-actin), and genes associated with the synthetic state were likewise both upregulated (e.g., SKI-like oncogene) and downregulated (e.g., collagen III). In addition, cells aligned with an orientation perpendicular to the maximal stretch direction. We have developed an in vitro cell culture device that can produce non-uniform stretch environments similar to in vivo mechanics. Cells stretched with this device showed alignment and altered mRNA expression indicative of phenotype modulation. Understanding these processes as they relate to in vivo pathologies could enable a more accurately targeted treatment to heal or inhibit disease, either through implantable device design or pharmaceutical approaches.

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