Calcium and the Ca-ATPase SPCA1 modulate plasma membrane abundance of ZIP8 and ZIP14 to regulate Mn(II) uptake in brain microvascular endothelial cells

Steimle, Brittany L.; Bailey, Danielle K.; Smith, Frances M.; Rosenblum, Shaina L.; Kosman, Daniel J.

doi:10.1016/j.jbc.2022.102211

Cited by 8 publications

(15 citation statements)

References 58 publications

(134 reference statements)

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“…Blocking LTCC decreases cardiac iron levels Tsushima et al (1999), Oudit et al (2006) Therefore, this increased calcium influx from LTCC and TTCC could be contributing to iron uptake indirectly, through increased iron transporter membrane localization (Steimle et al, 2022). Another possible method for NTBI uptake into cardiomyocytes are the ZIP proteins, ZIP8 and ZIP14.…”

Section: Iron Import Mechanismsmentioning

confidence: 99%

Inflammation, dysregulated iron metabolism, and cardiovascular disease

Rosenblum

2023

Front. Aging

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Iron is an essential trace element associated with both pathologic deficiency and toxic overload. Thus, systemic and cell iron metabolism are highly controlled processes regulated by protein expression and localization, as well as turnover, through the action of cytokines and iron status. Iron metabolism in the heart is challenging because both iron overload and deficiency are associated with cardiac disease. Also associated with cardiovascular disease is inflammation, as many cardiac diseases are caused by or include an inflammatory component. In addition, iron metabolism and inflammation are closely linked. Hepcidin, the master regulator of systemic iron metabolism, is induced by the cytokine IL-6 and as such is among the acute phase proteins secreted by the liver as part of the inflammatory response. In an inflammatory state, systemic iron homeostasis is dysregulated, commonly resulting in hypoferremia, or low serum iron. Less well characterized is cardiac iron metabolism in general, and even less is known about how inflammation impacts heart iron handling. This review highlights what is known with respect to iron metabolism in the heart. Expression of iron metabolism-related proteins and processes of iron uptake and efflux in these cell types are outlined. Evidence for the strong co-morbid relationship between inflammation and cardiac disease is also reviewed. Known connections between inflammatory processes and iron metabolism in the heart are discussed with the goal of linking inflammation and iron metabolism in this tissue, a connection that has been relatively under-appreciated as a component of heart function in an inflammatory state. Therapeutic options connecting inflammation and iron balance are emphasized, with the main goal of this review being to bring attention to alterations in iron balance as a component of inflammatory diseases of the cardiovascular system.

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Section: Iron Import Mechanismsmentioning

confidence: 99%

Inflammation, dysregulated iron metabolism, and cardiovascular disease

Rosenblum

2023

Front. Aging

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“…Peripheral actin was quantified by drawing a line of 0.875 inches (6.405µm) across cell membranes in ImageJ. 2-5 membrane regions of interest were quantified per cell per image and aligned as an XY graph in GraphPad prism as previously described [30][31][32] . Along the line at which the membrane starts is considered the "peak value," and then the neighboring 300nm was assigned as the cortical actin region 33 .…”

Section: Phalloidin-texas Redmentioning

confidence: 99%

“…The line was drawn so that the membrane interface represented the middle of the line to generate a bell-curve, averaging 2-5 membrane regions of interest per cell per image (Fig. 5D) 30 . This analysis revealed that the shFXN (orange) trace had notable differences in F-actin distribution compared to the EVEC controls (black).…”

Section: Shfxn Have Decreased Levels Of Polymerized Total and Periphe...mentioning

confidence: 99%

Frataxin-deficient human brain microvascular endothelial cells lose polymerized actin and are paracellularly permeable –implications for blood-brain barrier integrity in Friedreich’s Ataxia

Smith

Kosman

2023

Preprint

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Friedreich's Ataxia (FRDA) is the most prevalent inherited ataxia; the disease results from loss of Frataxin, an essential mitochondrial iron trafficking protein. FRDA presents as neurodegeneration of the dorsal root ganglion and cerebellar dentate nuclei, followed by brain iron accumulation in the latter. End stage disease includes cardiac fibrosis that contributes to hypertrophic cardiomyopathy. The microvasculature plays an essential barrier role in both the brain and heart, thus an investigation of this tissue system in FRDA is essential to the delineation of the cellular dysfunction in this genetic disorder. Here, we investigate brain microvascular endothelial cell integrity in FRDA in a model of the blood-brain barrier (BBB). We used lentiviral mediated shRNA delivery to generate a novel FRDA model in immortalized human brain microvascular endothelial cells (hBMVEC) that compose the microcapillaries of the BBB. We verified known cellular pathophysiologies of FXN knockdown including increased oxidative stress, loss of energy metabolism, and increased cell size. Furthermore, we investigated cytoskeletal architecture including the abundance and organization of filamentous actin, and barrier physiology via transendothelial electrical resistance and fluorescent tracer flux. shFXN hBMVEC display the known FRDA cell morbidity including increased oxidative stress, decreased energy metabolism, and an increase in cell size. We demonstrate that shFXN hBMVEC have less overall filamentous actin, and that filamentous actin is lost at the cell membrane and cortical actin ring. Consistent with loss of cytoskeletal structure and anchorage, we found decreased barrier strength and increased paracellular tracer flux in the shFXN hBMVEC transwell model. We identified that insufficient FXN levels in the hBMVEC BBB model causes changes in cytoskeletal architecture and increased barrier permeability, cell pathologies that may be related to patient brain iron accumulation, neuroinflammation, neurodegeneration, and stroke. Our findings implicate other barrier cells, e.g., the cardiac microvasculature, likely contributory also to disease pathology in FRDA.

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“…The middle of the line was placed at the start of the membrane. 2-5 membrane regions of interest were quanti ed per cell per image and aligned as a histogram as previously described using the "Plot Pro le" function (Ctrl + K) (31)(32)(33). Individual traces were combined in an XY graph in Graphpad prism to obtain the average trace per condition.…”

Section: Phalloidin-texas Red -Membrane-bound and Cortical Actin Stai...mentioning

confidence: 99%

“…The line was drawn so that the middle of the line represents where the cell membrane starts, generating a bell-curve of F-actin staining. This is compiled by averaging 2-5 arbitrary membrane regions of interest per cell per image (31). This analysis revealed that the shFXN (orange) trace had notable differences in F-actin distribution compared to the EVEC controls (black) (Fig.…”

Section: Shfxn Hbmvec Have Decreased Levels Of Polymerized Total and ...mentioning

confidence: 99%

Decreased filamentous actin and tight junction protein expression, and paracellular permeability in Frataxin-deficient human brain microvascular endothelial cells – implications for blood-brain barrier integrity in Friedreich's Ataxia

Smith

Kosman

2023

Preprint

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Background Friedreich’s Ataxia (FRDA) is the most prevalent inherited ataxia. FRDA results from loss of Frataxin (FXN), an essential mitochondrial iron trafficking protein. FRDA starts with an early burst of neurodegeneration of the dorsal root ganglion and cerebellar dentate nuclei, followed by progressive brain iron accumulation in the latter. End stage disease includes cardiac fibrosis that contributes to hypertrophic cardiomyopathy. The microvasculature plays an essential barrier role in both brain and heart homeostasis, thus an investigation of this tissue system in FRDA is essential to the delineation of the cellular dysfunction in this genetic disorder. Here, we investigate brain microvascular endothelial cell integrity in FRDA in a model of the blood-brain barrier (BBB).Methods We used lentiviral delivery of shRNA to knockdown FXN in a novel FRDA model in immortalized human brain microvascular endothelial cells (hBMVEC), which compose the microcapillaries of the BBB. We investigated known cellular pathophysiologies of FXN knockdown including energy metabolism, actin glutathionylation, and cell size. Furthermore, we investigated cytoskeletal architecture including the abundance and organization of filamentous actin (F-actin) and tight junction proteins, and quantified barrier physiology via transendothelial electrical resistance and fluorescent tracer flux.Results shFXN hBMVEC display the known FRDA cell morbidities including decreased energy metabolism, increased actin glutathionylation, and increased cell size. We also quantify a total loss of F-actin, and decreased abundance of F-actin at the cell membrane and in the cortical actin ring of shFXN hBMVEC. Furthermore, our model demonstrates a decreased production of the transmembrane tight junction proteins Claudin-5 and Occludin. Consistent with these phenotypes, we have identified increased paracellular permeability of a shFXN barrier system. These cells start with only 67% barrier integrity of the controls, and flux a paracellular tracer at 800% of physiological levels.Conclusion We identified that insufficient FXN levels in the hBMVEC BBB model causes changes in cytoskeletal architecture and tight junction protein abundance, co-incident with increased barrier permeability. Changes in the integrity of the BBB may be related to patient brain iron accumulation, neuroinflammation, neurodegeneration, and stroke. Furthermore, our findings implicate other barrier cells, e.g., the cardiac microvasculature, likely contributory also to disease pathology in FRDA.

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Calcium and the Ca-ATPase SPCA1 modulate plasma membrane abundance of ZIP8 and ZIP14 to regulate Mn(II) uptake in brain microvascular endothelial cells

Cited by 8 publications

References 58 publications

Inflammation, dysregulated iron metabolism, and cardiovascular disease

Inflammation, dysregulated iron metabolism, and cardiovascular disease

Frataxin-deficient human brain microvascular endothelial cells lose polymerized actin and are paracellularly permeable –implications for blood-brain barrier integrity in Friedreich’s Ataxia

Decreased filamentous actin and tight junction protein expression, and paracellular permeability in Frataxin-deficient human brain microvascular endothelial cells – implications for blood-brain barrier integrity in Friedreich's Ataxia

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