scholarly journals Immunoanalysis indicates that the sodium iodide symporter is not overexpressed in intracellular compartments in thyroid and breast cancers

2009 ◽  
Vol 160 (2) ◽  
pp. 215-225 ◽  
Author(s):  
Isabelle Peyrottes ◽  
Valerie Navarro ◽  
Alejandro Ondo-Mendez ◽  
Didier Marcellin ◽  
Laurent Bellanger ◽  
...  
Keyword(s):  
Western Blot ◽  
Specific Binding ◽  
Basolateral Membrane ◽  
Breast Cancers ◽  
Sodium Iodide Symporter ◽  
Intracellular Staining ◽  
Thyroid Cells ◽  
Breast Tumours ◽  
Intracellular Compartments ◽  
Initial Objective

ObjectiveThe active transport of iodide into thyroid cells is mediated by the Na+/I− symporter (NIS) located in the basolateral membrane. Strong intracellular staining with anti-NIS antibodies has been reported in thyroid and breast cancers. Our initial objective was to screen tumour samples for intracellular NIS staining and then to study the mechanisms underlying the altered subcellular localization of the transporters.MethodsImmunostaining using three different anti-NIS antibodies was performed on paraffin-embedded tissue sections from 93 thyroid or breast cancers. Western blot experiments were carried out to determine the amount of NIS protein in 20 samples.ResultsUsing three different anti-NIS antibodies, we observed intracellular staining in a majority of thyroid tumour samples. Control immunohistochemistry and western blot experiments indicated that this intracellular staining was due to non-specific binding of the antibodies. In breast tumours, very weak intracellular staining was observed in some samples. Western blot experiments suggest that this labelling is also non-specific.ConclusionsOur results strongly indicate that the NIS protein level is low in thyroid and breast cancers and that the intracellular staining obtained with anti-NIS antibodies corresponds to a non-specific signal. Accordingly, to increase the efficiency of radiotherapy for thyroid cancers and to enable the use of radioiodine in the diagnosis and therapy of breast tumours, improving NIS targeting to the plasma membrane will not be sufficient. Instead, increasing the expression level of NIS should remain the major goal of this field.

scholarly journals Identification and Characterization of a Putative Human Iodide Transporter Located at the Apical Membrane of Thyrocytes

2002 ◽  
Vol 87 (7) ◽  
pp. 3500-3503 ◽  
Author(s):  
Anne-Marie Rodriguez ◽  
Barbara Perron ◽  
Ludovic Lacroix ◽  
Bernard Caillou ◽  
Gérard Leblanc ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Human Kidney ◽  
Protein Product ◽  
Sodium Iodide Symporter ◽  
Pcr Cloning ◽  
Thyroid Cells ◽  
Iodide Transport ◽  
Acid Protein

Iodide transport by thyrocytes is a two step process involving transporters located either in the basal or in the apical membranes of the cell. The sodium iodide symporter (NIS) is localized in the basolateral membrane facing the bloodstream and mediates iodide accumulation into thyrocytes. Pendrin has been proposed as an apical transporter. In order to identify new iodide transporters, we developed a PCR cloning strategy based on NIS sequence homologies. From a human kidney cDNA library, we characterized a gene, located on chromosome 12q23, that encodes a 610 amino acid protein sharing 46% identity (70% similarity) with the human NIS. Functional analysis of the protein expressed in mammalian cells indicates that it catalyzes a passive iodide transport. The protein product was immunohistochemically localized at the apical pole of the thyroid cells facing the colloid lumen. These results suggest that this new identified protein mediates iodide transport from the thyrocyte into the colloid lumen through the apical membrane. It was designated hAIT for human Apical Iodide Transporter.

scholarly journals The molecular landscape of Asian breast cancers reveals clinically relevant population-specific differences

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Jia-Wern Pan ◽  
Muhammad Mamduh Ahmad Zabidi ◽  
Pei-Sze Ng ◽  
Mei-Yee Meng ◽  
Siti Norhidayu Hasan ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Somatic Mutations ◽  
Checkpoint Inhibitors ◽  
Breast Cancers ◽  
Breast Cancer Patients ◽  
Breast Tumours ◽  
Different Populations ◽  
Caucasian Women ◽  
Molecular Landscape ◽  
Immune Score

AbstractMolecular profiling of breast cancer has enabled the development of more robust molecular prognostic signatures and therapeutic options for breast cancer patients. However, non-Caucasian populations remain understudied. Here, we present the mutational, transcriptional, and copy number profiles of 560 Malaysian breast tumours and a comparative analysis of breast cancers arising in Asian and Caucasian women. Compared to breast tumours in Caucasian women, we show an increased prevalence of HER2-enriched molecular subtypes and higher prevalence of TP53 somatic mutations in ER+ Asian breast tumours. We also observe elevated immune scores in Asian breast tumours, suggesting potential clinical response to immune checkpoint inhibitors. Whilst HER2-subtype and enriched immune score are associated with improved survival, presence of TP53 somatic mutations is associated with poorer survival in ER+ tumours. Taken together, these population differences unveil opportunities to improve the understanding of this disease and lay the foundation for precision medicine in different populations.

Intracellular compartmentation of organic anions within renal cells

1993 ◽  
Vol 264 (5) ◽  
pp. R882-R890 ◽  
Author(s):  
D. S. Miller ◽  
D. E. Stewart ◽  
J. B. Pritchard
Keyword(s):  
Organic Anion ◽  
Basolateral Membrane ◽  
Organic Anions ◽  
Epifluorescence Microscopy ◽  
Renal Cells ◽  
Bladder Epithelium ◽  
Opossum Kidney ◽  
Ok Cells ◽  
Intracellular Compartments ◽  
Energy Dependent

Epifluorescence microscopy and video-image analysis were used to measure the distribution of the monovalent organic anion fluorescein (FL) within the cells of three organic anion-secreting renal epithelia: crab urinary bladder (a proximal tubule analogue), opossum kidney (OK) cells in culture, and intact teleost proximal tubules. In all three preparations the intracellular FL distribution was nonuniform. Two distinct intracellular compartments were detected, one being diffuse and cytoplasmic and the other punctate. With low FL concentrations in the medium (1 microM and below) dye accumulation in the punctate compartment exceeded that of the cytoplasm. In crab bladder epithelium FL uptake into both compartments was inhibited by external probenecid, p-aminohippurate (PAH), and LiCl and stimulated by 10-50 microM external glutarate, suggesting that the punctate compartment loaded by a two-step mechanism: transport into the cytoplasm at the basolateral membrane, followed by accumulation at specific intracellular sites. Experiments in which FL was microinjected into OK cells directly demonstrated movement of FL from the cytoplasmic to the punctate compartment. Accumulation in the latter was specific, i.e., inhibitable by coinjected PAH and probenecid, and energy dependent. Together, these findings indicate that during secretion organic anions are sequestered within renal cells. The role of sequestration in overall transport remains to be determined.

scholarly journals The Sodium-Iodide Symporter NIS and Pendrin in Iodide Homeostasis of the Thyroid

Endocrinology ◽  
2009 ◽  
Vol 150 (3) ◽  
pp. 1084-1090 ◽  
Author(s):  
Aigerim Bizhanova ◽  
Peter Kopp
Keyword(s):  
Autosomal Recessive ◽  
Sodium Iodide ◽  
Basolateral Membrane ◽  
Sensorineural Deafness ◽  
Sodium Iodide Symporter ◽  
Pendred Syndrome ◽  
Iodide Uptake ◽  
Anion Transporter ◽  
Thyroid Hormone Biosynthesis ◽  
Biallelic Mutations

Thyroid hormones are essential for normal development and metabolism. Thyroid hormone biosynthesis requires iodide uptake into the thyrocytes and efflux into the follicular lumen, where it is organified on selected tyrosyls of thyroglobulin. Uptake of iodide into the thyrocytes is mediated by an intrinsic membrane glycoprotein, the sodium-iodide symporter (NIS), which actively cotransports two sodium cations per each iodide anion. NIS-mediated transport of iodide is driven by the electrochemical sodium gradient generated by the Na+/K+-ATPase. NIS is expressed in the thyroid, the salivary glands, gastric mucosa, and the lactating mammary gland. TSH and iodide regulate iodide accumulation by modulating NIS activity via transcriptional and posttranscriptional mechanisms. Biallelic mutations in the NIS gene lead to a congenital iodide transport defect, an autosomal recessive condition characterized by hypothyroidism, goiter, low thyroid iodide uptake, and a low saliva/plasma iodide ratio. Pendrin is an anion transporter that is predominantly expressed in the inner ear, the thyroid, and the kidney. Biallelic mutations in the SLC26A4 gene lead to Pendred syndrome, an autosomal recessive disorder characterized by sensorineural deafness, goiter, and impaired iodide organification. In thyroid follicular cells, pendrin is expressed at the apical membrane. Functional in vitro data and the impaired iodide organification observed in patients with Pendred syndrome support a role of pendrin as an apical iodide transporter. This review shows how the sodium-iodide symporter mediates the active transport of iodide at the basolateral membrane of thyrocytes and discusses biallelic mutations in NIS and the effects of pendrin.

scholarly journals Combining RANK/RANKL and ERBB-2 targeting as a novel strategy in ERBB-2-positive breast carcinomas

2019 ◽  
Vol 21 (1) ◽  
Author(s):  
Ilianna Zoi ◽  
Michalis V. Karamouzis ◽  
Evangelia Xingi ◽  
Panagiotis Sarantis ◽  
Dimitra Thomaidou ◽  
...  
Keyword(s):  
Western Blot ◽  
Cell Lines ◽  
Family Members ◽  
Metastatic Potential ◽  
Nuclear Factor Κb ◽  
Proximity Ligation Assay ◽  
Breast Cancers ◽  
Tissue Samples ◽  
Erbb Family ◽  
Pathway Activation

Abstract Background ERBB-2 is overexpressed in about 20% of breast cancers (BCs), indicating poor prognosis. The receptor activator of nuclear factor-κB (RANK) pathway is implicated in ERBB-2 (+) BC. The purpose of this study was to elucidate the underlying molecular mechanism of this interaction and the beneficial impact of dual targeting of RANK and ERBB-2 pathways. Methods We used SKBR3, MCF7, MDA-MB-453, and BT-474 human BC cell lines. We examined RANK and RANKL expression using RT-PCR, Western blot, and immunofluorescence. The evaluation of RANK expression in a cohort of BC patients was performed using immunohistochemistry. The interaction between RANK and ERBB family members was detected using proximity ligation assay (PLA), which enables the visualization of interacting proteins. We used inhibitors of both pathways [trastuzumab (T), pertuzumab (P), denosumab (D)]. NF-κB pathway activation was studied using Western blot. Cell growth and viability was evaluated using XTT, flow cytometry, and clonogenic assay. For cell migration evaluation, scratch assay was performed. Data were analyzed by one-way ANOVA. Results Cell lines express RANK and RANKL. RANK immunostaining was also detected in human BC tissue samples. RANK receptor dimerizes with ERBB family members. RANK/ERBB-2 dimer number seems to be associated with ERBB-2 expression (SKBR3, 5.4; BT-474, 8.2; MCF7, 0.7; MDA-MB-453, 0.3). RANK/ERBB-2 dimers were decreased in the presence of the inhibitors D, T, and P, while they were increased after RANKL (R) treatment in SKBR3 (m, 5.4; D, 1.2; T, 1.9; DT, 0.6; TP, 1; DTP, 0.4; R, 11.8) and BT-474 (m, 8.2; D, 3.1; T, 4.3; DT, 0.7; TP, 3.4; DTP, 3.2; R, 11.6). Combination targeting of SKBR3 further decreased NF-κB pathway activation compared to single targeting. In SKBR3, RANKL and ERBB-2 blockage resulted in reduced cell proliferation, increased apoptosis, and lower metastatic potential compared to mock cells (m) and reversed values in RANKL presence. The combination treatment of SKBR3 with D, T, and P had an advantage in functional traits compared to single targeting. Denosumab suppressed NF-κB signaling and diminished proliferation rate in MDA-MB-453 cells. MCF7 did not correspond to inhibitors. Conclusions The results indicate a novel physical and molecular association between ERBB-2 and RANK pathways that affects ERBB-2 (+) BC growth. We also present data suggesting that the combination of anti-ERBB-2 agents and RANKL inhibitors have a potential direct anti-tumor effect and should be further tested in certain BC patients.

scholarly journals TGN38/41 recycles between the cell surface and the TGN: brefeldin A affects its rate of return to the TGN.

1993 ◽  
Vol 4 (1) ◽  
pp. 93-105 ◽  
Author(s):  
B Reaves ◽  
M Horn ◽  
G Banting
Keyword(s):  
Plasma Membrane ◽  
Specific Binding ◽  
Rat Kidney ◽  
Brefeldin A ◽  
Integral Membrane Protein ◽  
Intracellular Compartments ◽  
Normal Rat ◽  
Golgi Network ◽  
Lumenal Domain ◽  
Kinetics Of

TGN38 and TGN41 are isoforms of an integral membrane protein (TGN38/41) that is predominantly localized to the trans-Golgi network (TGN) of normal rat kidney cells. Polyclonal antisera to TGN38/41 have been used to monitor its appearance at, and removal from, the surface of control and Brefeldin A (BFA)-treated cells. Antibodies that recognize the lumenal domain of TGN38/41 are capable of specific binding to the surface of both control and BFA-treated cells. In both control and BFA-treated cells internalized TGN38/41 is targeted to the TGN; however, there are differences in 1) the morphology of the intracellular structures through which TGN38/41 passes and 2) the kinetics of internalization. These data demonstrate that TGN38/41 cycles between the plasma membrane and the TGN in control and BFA-treated cells and suggest that recycling pathways between the plasma membrane and the TGN exist for predominantly TGN proteins as well as those that normally cycle to other intracellular compartments. They also demonstrate that addition of BFA not only alters the morphology and localization of the TGN but also the kinetics of endocytosis.

scholarly journals Decreased Expression of Thyroglobulin and Sodium Iodide Symporter Genes in Hashimoto’s Thyroiditis

2014 ◽  
Vol 2014 ◽  
pp. 1-5 ◽  
Author(s):  
Anna Popławska-Kita ◽  
Beata Telejko ◽  
Katarzyna Siewko ◽  
Maria Kościuszko-Zdrodowska ◽  
Natalia Wawrusewicz-Kurylonek ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Hashimoto’S Thyroiditis ◽  
Sodium Iodide ◽  
Interleukin 1 ◽  
Thyroid Volume ◽  
Needle Aspiration ◽  
Hashimoto's Thyroiditis ◽  
Sodium Iodide Symporter ◽  
Thyroid Cells ◽  
Iodide Transport

Aim. The aim of the study was to compare the expression of sodium iodide symporter (NIS), thyroglobulin (Tg), tumor necrosis factor-α(TNFα), and interleukin-1βgenes in patients with Hashimoto’s thyroiditis (HT) and healthy individuals.Subjects and Methods.Thyroid cells were obtained from 39 patients with HT and 15 controls by an ultrasound guided fine needle aspiration biopsy.Results. The patients with HT had lower Tg and NIS mRNA (P=0.002andP=0.001, resp.), as well as higher TNFαmRNA expression (P=0.049) than the controls. In the HT group Tg mRNA expression correlated positively with NIS mRNA expression (R=0.739,P=0.0001) and thyroid volume (R=0.465,P=0.0005), as well as negatively with TNFαmRNA expression (R=-0.490,P=0.001) and anti-peroxidase antibodies (TPOAb) level (R=-0.482,P=0.0002), whereas NIS mRNA expression correlated positively with thyroid volume (R=0.319,P=0.02), as well as negatively with TNFαmRNA expression (R=-0.529,P=0.0006) and TPOAb level (R=-0.422,P=0.001).Conclusions.Our results suggest that decreased Tg and NIS expression in thyroid cells may result in reduced active iodide transport and reduced thyroid volume in patients with HT.

scholarly journals Histone deacetylation of NIS promoter underlies BRAF V600E-promoted NIS silencing in thyroid cancer

2013 ◽  
Vol 21 (2) ◽  
pp. 161-173 ◽  
Author(s):  
Zongjing Zhang ◽  
Dingxie Liu ◽  
Avaniyapuram Kannan Murugan ◽  
Zhimin Liu ◽  
Mingzhao Xing
Keyword(s):  
Thyroid Cancer ◽  
Histone Acetylation ◽  
Underlying Mechanism ◽  
Histone Deacetylation ◽  
Braf V600e ◽  
Epigenetic Mechanism ◽  
Human Thyroid ◽  
Sodium Iodide Symporter ◽  
Thyroid Cells ◽  
Rat Thyroid

The BRAF V600E mutation causes impaired expression of sodium iodide symporter (NIS) and radioiodine refractoriness of thyroid cancer, but the underlying mechanism remains undefined. In this study, we hypothesized that histone deacetylation at the NIS (SLC5A5) promoter was the mechanism. Using the chromatin immunoprecipitation approach, we examined histone acetylation status on the lysine residues H3K9/14, H3K18, total H4, and H4K16 at the NIS promoter under the influence of BRAF V600E. We found that expression of stably or transiently transfected BRAF V600E inhibited NIS expression while the deacetylase inhibitor SAHA stimulated NIS expression in PCCL3 rat thyroid cells. Although BRAF V600E enhanced global histone acetylation, it caused histone deacetylation at the NIS promoter while SAHA caused acetylation in the cells. In human thyroid cancer BCPAP cells harboring homozygous BRAF V600E mutation, BRAF V600E inhibitor, PLX4032, and MEK inhibitor, AZD6244, increased histone acetylation of the NIS promoter, suggesting that BRAF V600E normally maintained histone in a deacetylated state at the NIS promoter. The regions most commonly affected with deacetylation by BRAF V600E were the transcriptionally active areas upstream of the translation start that contained important transcription factor binding sites, including nucleotides −297/−107 in the rat NIS promoter and −692/−370 in the human NIS promoter. Our findings not only reveal an epigenetic mechanism for BRAF V600E-promoted NIS silencing involving histone deacetylation at critical regulatory regions of the NIS promoter but also provide further support for our previously proposed combination therapy targeting major signaling pathways and histone deacetylase to restore thyroid gene expression for radioiodine treatment of thyroid cancer.

Endocytic pathway for surfactant protein A in human macrophages: binding, clathrin-mediated uptake, and trafficking through the endolysosomal pathway

2006 ◽  
Vol 290 (2) ◽  
pp. L334-L342 ◽  
Author(s):  
Joy E. Crowther ◽  
Larry S. Schlesinger
Keyword(s):  
Specific Binding ◽  
Lung Surfactant ◽  
Protein A ◽  
Surfactant Protein ◽  
Endocytic Pathway ◽  
Surfactant Protein A ◽  
Coated Pits ◽  
Protein Protein Interaction ◽  
Intracellular Compartments ◽  
Over Time

In the noninflamed lung, surfactant protein A (SP-A) acts as an anti-inflammatory molecule through its effects on macrophage (MΦ) function, modulating cytokine and reactive oxygen and nitrogen intermediate production. The receptors responsible for these effects of SP-A on human MΦ are not clear, although SP-A binding to several proteins has been described. In this study, we demonstrate high-affinity specific binding of SP-A to primary human MΦ. SP-A binding was inhibited by EGTA, indicating calcium dependence. However, mannan did not inhibit SP-A binding, suggesting that binding is mediated by a direct protein-protein interaction that does not involve carbohydrate recognition. Our laboratory has previously shown that SP-A is rapidly endocytosed by human MΦ into discrete vesicles. Although previous work indicates that SP-A is ultimately degraded by murine MΦ over time, the trafficking pathway of SP-A through MΦ after uptake has not been reported and is of potential biological importance. We examined trafficking of SP-A in human MΦ by electron and confocal microscopy and show for the first time that SP-A is endocytosed by primary human MΦ through clathrin-coated pits and colocalizes sequentially over time with the early endosome marker EEA1, late endosome marker lamp-1, and lysosome marker cathepsin D. We conclude that SP-A binds to receptor(s) on human MΦ, is endocytosed by a receptor-mediated, clathrin-dependent process, and trafficks through the endolysosomal pathway. These studies provide further insight into the interactions of SP-A with the MΦ cell surface and intracellular compartments that play important roles in SP-A modulation of lung MΦ biology.

scholarly journals Proteomic analysis of differentially expressed proteins in normal human thyroid cells transfected with PPFP

2012 ◽  
Vol 19 (5) ◽  
pp. 681-694 ◽  
Author(s):  
Xinying Li ◽  
Zhiming Wang ◽  
Jianming Liu ◽  
Cane Tang ◽  
Chaojun Duan ◽  
...  
Keyword(s):  
Western Blot ◽  
Proteomic Analysis ◽  
Colony Formation ◽  
Fusion Gene ◽  
Differentially Expressed ◽  
Human Thyroid ◽  
Peroxisome Proliferator ◽  
Differentially Expressed Proteins ◽  
Thyroid Cells ◽  
Peroxisome Proliferator Activated Receptor

The fusion gene encoding the thyroid-specific transcription factor PAX8 and peroxisome proliferator-activated receptor γ (PPARγ (PPARG)) (designated as the PPFP gene) is oncogenic and implicated in the development of follicular thyroid carcinoma (FTC). The effects of PPFP transfection on the biological characteristics of Nthy-ori 3-1 cells were studied by MTT assay, colony formation, soft-agar colony formation, and scratch wound-healing assays as well as by flow cytometry. Furthermore, the differentially expressed proteins were analyzed on 2-DE maps and identified by MALDI-TOF-MS. Validation of five identified proteins (prohibitin, galectin-1, cytokeratin 8 (CK8), CK19, and HSP27) was determined by western blot analysis. PPFP not only significantly increased the viability, proliferation, and mobility of the Nthy-ori 3-1 cells but also markedly inhibited cellular apoptosis. Twenty-eight differentially expressed proteins were identified, among which 19 proteins were upregulated and nine proteins were downregulated in Nthy-ori 3-1PPFP(Nthy-ori 3-1 cells transfected with PPFP). The western blot results, which were consistent with the proteome analysis results, showed that prohibitin was downregulated, whereas galectin-1, CK8, CK19, and HSP27 were upregulated in Nthy-ori 3-1PPFP. Our results suggest that PPFP plays an important role in malignant thyroid transformation. Proteomic analysis of the differentially expressed proteins in PPFP-transfected cells provides important information for further study of the carcinogenic mechanism of PPFP in FTCs.

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