scholarly journals Glycosylation Profile of the Transferrin Receptor in Gestational Iron Deficiency and Early-Onset Severe Preeclampsia

2019 ◽  
Vol 2019 ◽  
pp. 1-9 ◽  
Author(s):  
Alejandra María Gómez-Gutiérrez ◽  
Beatriz Elena Parra-Sosa ◽  
Julio Cesar Bueno-Sánchez
Keyword(s):  
Iron Deficiency ◽  
Western Blot ◽  
Blot Analysis ◽  
Transferrin Receptor ◽  
Lectin Binding ◽  
Hypoxia Inducible Factor ◽  
Severe Preeclampsia ◽  
Hypoxia Inducible Factor 1 ◽  
Lectin Blot ◽  
Low Expression

Objective. To examine the expression of hypoxia-inducible factor-1α(HIF-1α), TfR1, and TfR1-attached terminal monosaccharides in placentas of women with IDAP and severe preeclampsia.Methods. TfR1 and HIF-1αwere detected by western blot. Immunoadsorption of TfR1 was performed to characterize the terminal monosaccharides by specific lectin binding.Results. There was no difference in the expression of TfR1 and HIF-1αbetween groups. Lectin blot analysis pointed out an overexpression of galactoseβ1-4N-acetylglucosamine (Gal-GlcNAc) and mannose in severe preeclampsia.Conclusion. The increase in Gal-GlcNAc may be due to the increased presence of antennary structures and the mannose glycans of TfR1 may indicate the presence of misfolded or incomplete proteins. These findings may be associated with the low expression of placental TfR1 in women with preeclampsia.

scholarly journals Synthesis and glycosylation of CD52, the major 'maturation-associated' antigen of rat spermatozoa, in the cauda epididymidis

Reproduction ◽  
2001 ◽  
pp. 435-446 ◽  
Author(s):  
P Derr ◽  
CH Yeung ◽  
TG Cooper ◽  
C Kirchhoff
Keyword(s):  
Blot Analysis ◽  
Cdna Sequence ◽  
Northern Blot Analysis ◽  
Lectin Binding ◽  
Transcriptional Level ◽  
Lectin Blot ◽  
Molecular Masses ◽  
Rat Epididymis ◽  
Sperm Membrane ◽  
Cauda Epididymidis

A western and lectin blot analysis was performed of the major 'maturation-associated' antigen of rat spermatozoa, which is the rat counterpart of human CD52. In the absence of a suitable antibody, direct study of this approximately 26 kDa antigen, named previously SMemG, had been difficult. In the present study, these problems were overcome by raising a polyclonal antibody against a chemosynthetic peptide predicted from the cDNA sequence of the antigen. The antibody bound to a glycoprotein of rat cauda epididymidal tissue and spermatozoa, this glycoprotein was cleaved by phosphatidylinositol-specific phospholipase C and, after deglycosylation, was reduced to approximately 6 kDa. Northern blot analysis confirmed that the CD52 mRNA was transcribed only post-testicularly, and antibody binding to testicular and sperm proteins of different molecular masses was shown to be nonspecific. Flow cytometry also indicated that the antigen was inserted into the sperm membrane during epididymal transit. Moreover, despite the presence of CD52 mRNA in all parts of the rat epididymis, only the 'long' mRNA molecules of the cauda region were efficiently translated and the antigen glycosylated, indicating that expression of rat CD52 is regulated on a post-transcriptional level. Lectin binding and deglycosylation studies supported the contention that there is extensive mucin-type O-glycosylation of rat CD52. In rats, there was no indication of complex N-linked carbohydrates similar to those described for human CD52.

Association of GRP78, HIF-1α and BAG3 Expression with the Severity of Chronic Lymphocytic Leukemia

2020 ◽  
Vol 20 (4) ◽  
pp. 429-436
Author(s):  
Roghayeh Ijabi ◽  
Parisa Roozehdar ◽  
Reza Afrisham ◽  
Hemen Moradi-Sardareh ◽  
Saeed Kaviani ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Western Blot ◽  
Disease Progression ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Strong Association ◽  
Hypoxia Inducible Factor ◽  
Lymphocytic Leukemia ◽  
Rt Pcr ◽  
Metabolic Factors

Introduction: Parallel with the progression of Chronic Lymphocytic Leukemia (CLL), the levels of 78KDa Glucose-Regulated Protein (GRP78) and Hypoxia-Inducible Factor 1 alpha (HIF-1α) are increased as they may activate the induction of anti-apoptotic proteins such as BCL2 Associated Athanogene 3 (BAG3). Previous studies have indicated that there is a positive correlation among GRP78, HIF-1α and BAG3. Objective: This study aimed to evaluate the effect of metabolic factors involved in invasive CLL on apoptotic factors. Methods: A case-control study was conducted on 77 patients diagnosed with CLL along with 100 healthy individuals. Cell blood count was performed for all participants. According to Binet's classification, CLL patients were divided into different groups. B cells were isolated from the peripheral blood of CLL patients by binding to anti-CD19 beads. The expression of BAG3, GRP78 and HIF-1α genes was analyzed using the RT-PCR method. To confirm the results of RT-PCR, western blot analysis was carried out. Results: The results showed that there was a strong association among the expression of BAG3, GRP78 and HIF-1α. The stage of CLL in patients was highly correlated with the expression rate of each gene (p<0.001). Accordingly, the western blot analysis indicated that the concentrations of GRP78 and HIF-1α were significantly higher than the expression of BAG3, considering the stage of CLL. Conclusion: It was shown that increased expression of GRP78 and HIF-1α could result in the elevation of BAG3, as well as the disease progression. Therefore, the role of these metabolic factors might be more pronounced compared with the anti-apoptotic agents to monitor disease progression in CLL patients.

scholarly journals The role of C1GALT1C1 in lipopolysaccharide-induced IgA1 aberrant O-glycosylation in IgA nephropathy

2010 ◽  
Vol 33 (1) ◽  
pp. 5 ◽  
Author(s):  
Lin-Shen Xie ◽  
Wei Qin ◽  
Jun-Ming Fan ◽  
Jun Huang ◽  
Xi-Sheng Xie ◽  
...  
Keyword(s):  
Correlation Analysis ◽  
Western Blot ◽  
Iga Nephropathy ◽  
Blot Analysis ◽  
Real Time Pcr ◽  
Binding Assay ◽  
Lectin Binding ◽  
Vicia Villosa ◽  
Normal Controls

Purpose: IgA1 aberrant O-glycosylation is one of the main pathogenetic features of IgA nephropathy (IgAN). This study attempted to determine the role of C1GALT1C1 in aberrant IgA1 O-glycosylation induced by lipopolysaccharide (LPS) and identify potential therapeutic targets in IgAN. Methods: Lymphocytes isolated from 22 patients with IgAN and 17 normal controls were cultured for 3 to 7 days with or without LPS and 5-azacytidine (5-AZA). Expression levels of C1GALT1C1 mRNA and protein were measured by real-time PCR and Western blot analysis, respectively. Concentration of IgA1 and level of O-glycosylation were determined by ELISA and Vicia villosa (VV) lectin-binding assay. Correlation analysis was performed between the expression of C1GALT1C1 protein and IgA1 O-glycosylation. Results: Lymphocytes from patients with IgAN secreted more IgA1 than that from normal controls after LPS stimulation (P=0.26, 0.002 and 0.005 on the 3rd, 5th and 7th day, respectively) which could be inhibited by 5-AZA (P=0.001, 0.025 and 0.001 on the 3rd, 5th and 7th day, respectively). Moreover, LPS stimulation could obviously inhibit C1GALT1C1 expression in patients with IgAN (decreased by 71%, 82% and 92% on the 3rd, 5th and 7th day, respectively; P < 0.001), and cause a significant decrease of IgA1 O-glycosylation compared with normal controls (P=0.004, 0.003 and 0.03 on the 3rd, 5th and 7th day, respectively). When 5-AZA was added, the level of C1GALT1C1 expression increased dramatically (1.98, 5.53 and 8.97 times on the 3rd, 5th and 7th day, respectively; P < 0.001) along with an increase of IgA1 O-glycosylation (P=0.295, 0.09 and 0.003 on the 3rd, 5th and 7th day, respectively). However, normal controls showed no significant change in C1GALT1C1 expression and IgA1 O-glycosylation after LPS stimulation (P > 0.05). Conclusion: LPS induced IgA1 aberrant O-glycosylation and suppressed C1GALT1C1 expression in patients with IgAN. Upregulation of C1GALT1C1 expression by 5-AZA could reverse the IgA1 aberrant O-glycosylation. These results suggest that C1GALT1C1 may play a key role in the regulation of IgA1 O-glycosylation.

A Novel DNM2 Mutation Displaying Embryonic Lethality and Impaired Transferrin Uptake Identified in a Mouse ENU Mutagenesis Screen for Genes Perturbing Erythropoiesis

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 608-608 ◽  
Author(s):  
Fiona C Brown ◽  
Michael Collett ◽  
Phillip J Robinson ◽  
James C Whisstock ◽  
Douglas J. Hilton ◽  
...  
Keyword(s):  
Iron Deficiency ◽  
Western Blot ◽  
Blot Analysis ◽  
Human Disease ◽  
Western Blot Analysis ◽  
Deficiency Anemia ◽  
Wild Type ◽  
Enu Mutagenesis ◽  
Mutagenesis Screen ◽  
Transferrin Uptake

Abstract Abstract 608 Forward-genetic screens have become a powerful method to study the pathogenesis of human disease and gene function. Chemical mutagenesis in mice using the mutagen, N-ethyl N-nitrosourea (ENU), has shown to be highly successful in elucidating novel genes or alleles in a variety of biological pathways, describing new functions of existing genes, and establishing mouse models that accurately recapitulate human disease. Advances in mapping strategies and deep sequencing technologies has dramatically simplified mutation detection, making ENU mutagenesis screens a feasible tool to study specific organ systems. To identify novel alleles regulating erythropoiesis, our laboratory has undertaken a dominant ENU mutagenesis screen. In this screen, the G1 progeny were screened at seven weeks of age for abnormalities in red cell indices (MCV, MCH, and HCT) using an automated hematological analyser. Here, we describe the identification of mice with a missense mutation of the large GTPase Dynamin 2 (DNM2) leading to an amino acid substitution V235G, predicted to lie within the nucleotide binding pocket for GTP. Western blot analysis for DNM2 protein revealed 50% protein levels in heterozygotes, suggesting that the point mutation leads to loss of protein rather than a dominant negative effect. Inherited DNM2 mutations are associated with autosomal dominant Charcot Tooth Myopathy (CTM) and Centronuclear Myopathy (CNM), but no recognised blood disorders. Heterozygous DNM2V235G displayed hypochromic, microcytic anemia – HGB (15 g/dl compared to 16.5 g/dl in wild type mice), MCV (41.3 fl compared to 45.6 fl in wild type mice) and MCH (12.7 pg compared to 14.5 pg in wild type mice), but no obvious neuropathy or myopathy. Homozygosity was lethal before embryonic (E) day 8.5. DNM2 is an essential component in clathrin-mediated endocytosis, which is required for uptake of transferrin into red cells for incorporation of heme. Accordingly, endocytosis assays for transferrin uptake by FACS and confocal microscopy revealed reduced uptake in heterozygotes, explaining the microcytic hypochromic anemia. Western blot analysis for ferritin demonstrated reduced cellular ferritin, indicating cellular iron deficiency. Thus, this mouse model provides the first in vivo evidence that haplo-insufficiency of DNM2 can lead to iron deficiency anemia. Disclosures: No relevant conflicts of interest to declare.

scholarly journals MiR-186 inhibited aerobic glycolysis in gastric cancer via HIF-1α regulation

Oncogenesis ◽  
2016 ◽  
Vol 5 (5) ◽  
pp. e224-e224 ◽  
Author(s):  
L Liu ◽  
Y Wang ◽  
R Bai ◽  
K Yang ◽  
Z Tian
Keyword(s):  
Gastric Cancer ◽  
Gastric Cancer Cell ◽  
Aerobic Glycolysis ◽  
Hypoxia Inducible Factor ◽  
Xenograft Tumor ◽  
Hypoxia Inducible Factor 1 ◽  
Programmed Death ◽  
Gastric Cancer Cell Lines ◽  
Low Expression

Abstract Deregulation of microRNAs in human malignancies has been well documented, among which microRNA-186 (miR-186) has an antiproliferative role in some carcinomas. Here we demonstrate that low expression of miR-186 facilitates aerobic glycolysis in gastric cancer. MiR-186 suppresses cell proliferation induced by hypoxia inducible factor 1 alpha (HIF-1α) in gastric cancer cell lines MKN45 and SGC7901. Cellular glycolysis, including cellular glucose uptake, lactate, ATP/ADP and NAD+/NADH ratios, are also inhibited by miR-186. The negative regulation of miR-186 on HIF-1α effects its downstream targets, including programmed death ligand 1 and two glycolytic key enzymes, hexokinase 2 and platelet-type phosphofructokinase. The antioncogenic effects of miR-186 are proved by in vivo xenograft tumor experiment. The results demonstrate that the miR-186/HIF-1α axis has an antioncogenic role in gastric cancer.

Vascular adaptations to hypoxia: molecular and cellular mechanisms regulating vascular tone

2007 ◽  
Vol 43 ◽  
pp. 105-120 ◽  
Author(s):  
Michael L. Paffett ◽  
Benjimen R. Walker
Keyword(s):  
Vascular Tone ◽  
Cellular Proliferation ◽  
Hypoxic Pulmonary Vasoconstriction ◽  
Hypoxia Inducible Factor ◽  
Systemic Circulation ◽  
Cellular Mechanisms ◽  
Hypoxia Inducible Factor 1 ◽  
Pulmonary Vasoconstriction ◽  
Adaptive Mechanisms ◽  
Adaptive Processes

Several molecular and cellular adaptive mechanisms to hypoxia exist within the vasculature. Many of these processes involve oxygen sensing which is transduced into mediators of vasoconstriction in the pulmonary circulation and vasodilation in the systemic circulation. A variety of oxygen-responsive pathways, such as HIF (hypoxia-inducible factor)-1 and HOs (haem oxygenases), contribute to the overall adaptive process during hypoxia and are currently an area of intense research. Generation of ROS (reactive oxygen species) may also differentially regulate vascular tone in these circulations. Potential candidates underlying the divergent responses between the systemic and pulmonary circulations may include Nox (NADPH oxidase)-derived ROS and mitochondrial-derived ROS. In addition to alterations in ROS production governing vascular tone in the hypoxic setting, other vascular adaptations are likely to be involved. HPV (hypoxic pulmonary vasoconstriction) and CH (chronic hypoxia)-induced alterations in cellular proliferation, ionic conductances and changes in the contractile apparatus sensitivity to calcium, all occur as adaptive processes within the vasculature.

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