scholarly journals In Vivo Analysis of the Contribution of Proprotein Convertases to the Processing of FGF23

2021 ◽  
Vol 12 ◽  
Author(s):  
Omar Al Rifai ◽  
Delia Susan-Resiga ◽  
Rachid Essalmani ◽  
John W. M. Creemers ◽  
Nabil G. Seidah ◽  
...  
Keyword(s):  
Fibroblast Growth Factor 23 ◽  
Basic Amino Acid ◽  
Serum Phosphate ◽  
Intact Protein ◽  
Proprotein Convertases ◽  
Endoproteolytic Cleavage ◽  
In Vivo Analysis ◽  
Deficient Mice

Fibroblast growth factor 23 (FGF23) is a hormone secreted from fully differentiated osteoblasts and osteocytes that inhibits phosphate reabsorption by kidney proximal tubules. The full-length (i.e., intact) protein mediates FGF23 endocrine functions, while endoproteolytic cleavage at a consensus cleavage sequence for the proprotein convertases (PCs) inactivates FGF23. Two PCs, furin and PC5, were shown to cleave FGF23 in vitro at RHTR179↓, but whether they are fulfilling this function in vivo is currently unknown. To address this question, we used here mice lacking either or both furin and PC5 in cell-specific manners and mice lacking the paired basic amino acid-cleaving enzyme 4 (PACE4) in all cells. Our analysis shows that furin inactivation in osteoblasts and osteocytes results in a 25% increase in circulating intact FGF23, without any significant impact on serum phosphate levels, whether mice are maintained on a normal or a low phosphate diet. Under conditions of iron deficiency, FGF23 is normally processed in control mice, but its processing is impaired in mice lacking furin in osteoblasts and osteocytes. In contrast, FGF23 is normally cleaved following erythropoietin or IL-1β injections in mice lacking furin or both furin and PC5, and in PACE4-deficient mice. Altogether, these studies suggest that furin is only partially responsible for FGF23 cleavage under certain conditions in vivo. The processing of FGF23 may therefore involve the redundant action of multiple PCs or of other peptidases in osteoblasts, osteocytes and hematopoietic cells.

scholarly journals In vivo analysis of the contribution of proprotein convertases to the processing of FGF23

2021 ◽  
Author(s):  
Omar Al Rifai ◽  
Delia Susan-Resiga ◽  
Rachid Essalmani ◽  
John W.M. Creemers ◽  
Nabil G. Seidah ◽  
...  
Keyword(s):  
Fibroblast Growth Factor 23 ◽  
Basic Amino Acid ◽  
Serum Phosphate ◽  
Intact Protein ◽  
Proprotein Convertases ◽  
Endoproteolytic Cleavage ◽  
In Vivo Analysis ◽  
Deficient Mice

ABSTRACTFibroblast growth factor 23 (FGF23) is a hormone secreted from fully differentiated osteoblasts and osteocytes that inhibits phosphate reabsorption by kidney proximal tubules. The full-length (i.e., intact) protein mediates FGF23 endocrine functions, while endoproteolytic cleavage at a consensus cleavage sequence for the proprotein convertases (PCs) inactivates FGF23. Two PCs, furin and PC5, were shown to cleave FGF23 in vitro at RHTR179↓, but whether they are fulfilling this function in vivo is currently unknown. To address this question we used here mice lacking either or both furin and PC5 in cell-specific manners and mice lacking the paired basic amino acid-cleaving enzyme 4 (PACE4) in all cells. Our analysis shows that furin inactivation in osteoblasts and osteocytes results in a 25% increase in circulating intact FGF23, without any significant impact on serum phosphate levels, whether mice are maintained on a normal or a low phosphate diet. Under conditions of iron deficiency, FGF23 is normally processed in control mice, but its processing is impaired in mice lacking furin in osteoblasts and osteocytes. In contrast, FGF23 is normally cleaved following erythropoietin or IL-1β injections in mice lacking furin or both furin and PC5, and in PACE4-deficient mice. Altogether, these studies suggest that furin is only partially responsible for FGF23 cleavage under certain conditions in vivo. The processing of FGF23 may therefore involve the redundant action of multiple PCs or of other peptidases in osteoblasts, osteocytes and hematopoietic cells.

Multinucleated Osteoclast Formation In Vivo and In Vitro by P2X7 Receptor-Deficient Mice

2003 ◽  
Vol 13 (2-4) ◽  
pp. 12 ◽  
Author(s):  
Alison Gartland ◽  
Katherine A. Buckley ◽  
Robert A. Hipskind ◽  
M. J. Perry ◽  
J. H. Tobias ◽  
...  
Keyword(s):  
P2x7 Receptor ◽  
Osteoclast Formation ◽  
Deficient Mice

scholarly journals Drosophila Hormone Receptor 38 Functions in Metamorphosis: A Role in Adult Cuticle Formation

Genetics ◽  
1998 ◽  
Vol 149 (3) ◽  
pp. 1465-1475 ◽  
Author(s):  
T Kozlova ◽  
G V Pokholkova ◽  
G Tzertzinis ◽  
J D Sutherland ◽  
I F Zhimulev ◽  
...  
Keyword(s):  
Pupal Stage ◽  
Transcription Unit ◽  
P Element ◽  
Specific Response ◽  
Orphan Receptors ◽  
Mrna Isoforms ◽  
A Cell ◽  
In Vivo Analysis

Abstract DHR38 is a member of the steroid receptor superfamily in Drosophila homologous to the vertebrate NGFI-B-type orphan receptors. In addition to binding to specific response elements as a monomer, DHR38 interacts with the USP component of the ecdysone receptor complex in vitro, in yeast and in a cell line, suggesting that DHR38 might modulate ecdysone-triggered signals in the fly. We characterized the molecular structure and expression of the Dhr38 gene and initiated an in vivo analysis of its function(s) in development. The Dhr38 transcription unit spans more than 40 kb in length, includes four introns, and produces at least four mRNA isoforms differentially expressed in development; two of these are greatly enriched in the pupal stage and encode nested polypeptides. We characterized four alleles of Dhr38: a P-element enchancer trap line, l(2)02306, which shows exclusively epidermal staining in the late larval, pre-pupal and pupal stages, and three EMS-induced alleles. Dhr38 alleles cause localized fragility and rupturing of the adult cuticle, demonstrating that Dhr38 plays an important role in late stages of epidermal metamorphosis.

FcγRIII (CD16)-Deficient Mice Show IgG Isotype-Dependent Protection to Experimental Autoimmune Hemolytic Anemia

Blood ◽  
1998 ◽  
Vol 92 (11) ◽  
pp. 3997-4002 ◽  
Author(s):  
Dirk Meyer ◽  
Carsten Schiller ◽  
Jürgen Westermann ◽  
Shozo Izui ◽  
Wouter L. W. Hazenbos ◽  
...  
Keyword(s):  
Hemolytic Anemia ◽  
Autoimmune Hemolytic Anemia ◽  
Knock Out ◽  
Effector Cells ◽  
Igg Isotypes ◽  
Knock Out Mice ◽  
Deficient Mice ◽  
Experimental Autoimmune

Abstract In autoimmune hemolytic anemia (AIHA), there is accumulating evidence for an involvement of FcγR expressed by phagocytic effector cells, but demonstration of a causal relationship between individual FcγRs and IgG isotypes for disease development is lacking. Although the relevance of IgG isotypes to human AIHA is limited, we could show a clear IgG isotype dependency in murine AIHA using pathogenic IgG1 (105-2H) and IgG2a (34-3C) autoreactive anti–red blood cell antibodies in mice defective for FcγRIII, and comparing the clinical outcome to those in wild-type mice. FcγRIII-deficient mice were completely resistent to the pathogenic effects of 105-2H monoclonal antibody, as shown by a lack of IgG1-mediated erythrophagocytosis in vitro and in vivo. In addition, the IgG2a response by 34-3C induced a less severe but persistent AIHA in FcγRIII knock-out mice, as documented by a decrease in hematocrit. Blocking studies indicated that the residual anemic phenotype induced by 34-3C in the absence of FcγRIII reflects an activation of FcγRI that is normally coexpressed with FcγRIII on macrophages. Together these results show that the pathogenesis of AIHA through IgG1-dependent erythrophagocytosis is exclusively mediated by FcγRIII and further suggest that FcγRI, in addition to FcγRIII, contributes to this autoimmune disease when other IgG isotypes such as IgG2a are involved.

scholarly journals Transcriptional Regulation of PEN-2, a Key Component of the γ-Secretase Complex, by CREB

2006 ◽  
Vol 26 (4) ◽  
pp. 1347-1354 ◽  
Author(s):  
Ruishan Wang ◽  
Yun-wu Zhang ◽  
Ping Sun ◽  
Runzhong Liu ◽  
Xian Zhang ◽  
...  
Keyword(s):  
Transcriptional Regulation ◽  
Transcriptional Activity ◽  
Start Codon ◽  
Gamma Secretase ◽  
Mobility Shift ◽  
Endoproteolytic Cleavage ◽  
Notch Cleavage ◽  
Translational Start

ABSTRACT Gamma-secretase, which is responsible for the intramembranous cleavage of Alzheimer's β-amyloid precursor protein (APP), the signaling receptor Notch, and many other substrates, is a multiprotein complex consisting of at least four components: presenilin (PS), nicastrin, APH-1, and PEN-2. Despite the fact that PEN-2 is known to mediate endoproteolytic cleavage of full-length PS and APH-1 and nicastrin are required for maintaining the stability of the complex, the detailed physiological function of each component remain elusive. Unlike that of PS, the transcriptional regulation of PEN-2, APH-1, and nicastrin has not been investigated. Here, we characterized the upstream regions of the human PEN-2 gene and identified a 238-bp fragment located 353 bp upstream of the translational start codon as the key region necessary for the promoter activity. Further analysis revealed a CREB binding site located in the 238-bp region that is essential for the transcriptional activity of the PEN-2 promoter. Mutation of the CREB site abolished the transcriptional activity of the PEN-2 promoter. Electrophoretic mobility shift assays and chromatin immunoprecipitation analysis showed the binding of CREB to the PEN-2 promoter region both in vitro and in vivo. Activation of the CREB transcriptional factor by forskolin dramatically promoted the expression of PEN-2 mRNA and protein, whereas the other components of the γ-secretase complex remained unaffected. Forskolin treatment slightly increases the secretion of soluble APPα and Aβ without affecting Notch cleavage. These results demonstrate that expression of PEN-2 is regulated by CREB and suggest that the specific control of PEN-2 expression may imply additional physiological functions uniquely assigned to PEN-2.

scholarly journals Mice Lacking the Inhibitory Collagen Receptor LAIR-1 Exhibit a Mild Thrombocytosis and Hyperactive Platelets

2017 ◽  
Vol 37 (5) ◽  
pp. 823-835 ◽  
Author(s):  
Christopher W. Smith ◽  
Steven G. Thomas ◽  
Zaher Raslan ◽  
Pushpa Patel ◽  
Maxwell Byrne ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Kinase Activity ◽  
Thrombus Formation ◽  
Physiological Role ◽  
Src Family Kinase ◽  
Glycoprotein Vi ◽  
Collagen Receptor ◽  
Deficient Mice

Objective— Leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) is a collagen receptor that belongs to the inhibitory immunoreceptor tyrosine-based inhibition motif–containing receptor family. It is an inhibitor of signaling via the immunoreceptor tyrosine-based activation motif–containing collagen receptor complex, glycoprotein VI-FcRγ-chain. It is expressed on hematopoietic cells, including immature megakaryocytes, but is not detectable on platelets. Although the inhibitory function of LAIR-1 has been described in leukocytes, its physiological role in megakaryocytes and in particular in platelet formation has not been explored. In this study, we investigate the role of LAIR-1 in megakaryocyte development and platelet production by generating LAIR-1–deficient mice. Approach and Results— Mice lacking LAIR-1 exhibit a significant increase in platelet counts, a prolonged platelet half-life in vivo, and increased proplatelet formation in vitro. Interestingly, platelets from LAIR-1–deficient mice exhibit an enhanced reactivity to collagen and the glycoprotein VI–specific agonist collagen-related peptide despite not expressing LAIR-1, and mice showed enhanced thrombus formation in the carotid artery after ferric chloride injury. Targeted deletion of LAIR-1 in mice results in an increase in signaling downstream of the glycoprotein VI–FcRγ-chain and integrin αIIbβ3 in megakaryocytes because of enhanced Src family kinase activity. Conclusions— Findings from this study demonstrate that ablation of LAIR-1 in megakaryocytes leads to increased Src family kinase activity and downstream signaling in response to collagen that is transmitted to platelets, rendering them hyper-reactive specifically to agonists that signal through Syk tyrosine kinases, but not to G-protein–coupled receptors.

scholarly journals In vitro and in vivo evaluation of diamond-coated strips

2016 ◽  
Vol 87 (3) ◽  
pp. 455-459 ◽  
Author(s):  
Roberta Lione ◽  
Francesca Gazzani ◽  
Chiara Pavoni ◽  
Stefano Guarino ◽  
Vincenzo Tagliaferri ◽  
...  
Keyword(s):  
Contact Point ◽  
Sem Analysis ◽  
Tribological Test ◽  
In Vivo Evaluation ◽  
Wear Performance ◽  
Abrasive Grains ◽  
Abrasive Power ◽  
In Vivo Analysis

ABSTRACT Objective: To test in vitro and in vivo the wear performance of diamond-coated strips by means of tribological testing and scanning electronic microscope (SEM). Materials and Methods: To evaluate the in vitro wear performance, a tribological test was performed by a standard tribometer. The abrasive strips slid against stationary, freshly extracted premolars fixed in resin blocks, at a 2-newton load. At the end of the tribological test, the residual surface of the strip was observed by means of SEM analysis, which was performed every 50 meters until reaching 300 meters. For the in vivo analysis, the strip was used for 300 seconds, corresponding to 250 meters. Results: The strips presented a fenestrated structure characterized by diamond granules alternating with voids. After the first 50 meters, it was possible to observe tooth material deposited on the surface of the strips and a certain number of abrasive grains detached. The surface of the strip after 250 meters appeared smoother and therefore less effective in its abrasive power. After 300 seconds of in vivo utilization of the strip, it was possible to observe the detachment of diamond abrasive grains, the near absence of the grains and, therefore, loss of abrasive power. Conclusions: Under ideal conditions, after 5 minutes (300 meters) of use, the strip loses its abrasive capacity by about 60%. In vivo, a more rapid loss of abrasive power was observed due to the greater load applied by the clinician in forcing the strip into the contact point.

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