Bioengineered 3D Human Kidney Tissue, a Platform for the Determination of Nephrotoxicity

DesRochers, Teresa M.; Suter, Laura; Roth, Adrian; Kaplan, David L.

doi:10.1371/journal.pone.0059219

Cited by 111 publications

(118 citation statements)

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“…In one case, isolation of a specific clone was required to facilitate cyst formation in collagen gels (36). Therefore, to use the LLC-PK1 cell line to consistently induce cyst growth in 3D, we used a hormonal media composition previously shown to produce structural growth of immortalized human renal proximal tubule cells and primary human cortical cells in 3D tissues (37,38). In addition, hormone-supplemented media has historically been used to support LLC-PK1 growth (39).…”

Section: Resultsmentioning

confidence: 99%

“…For 3D tissues, cells were embedded in a previously described mixture of Matrigel and type I collagen (37). Cystic structures were monitored for size by both H&E staining and whole-tissue staining of tissues after 2, 4, 6, and 8 wk of culture (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…LLC-PK1 pig epithelial cells were grown in 2D and 3D matricies as previously described (37). Three-dimensional cultures were stably transfected with shRNA directed against InsP3R1, InsP3R3, PC2, or scrambled control.…”

Section: Methodsmentioning

confidence: 99%

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Cyst formation following disruption of intracellular calcium signaling

Kuo¹,

DesRochers

Kimmerling

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Mutations in polycystin 1 and 2 (PC1 and PC2) cause the common genetic kidney disorder autosomal dominant polycystic kidney disease (ADPKD). It is unknown how these mutations result in renal cysts, but dysregulation of calcium (Ca 2+ ) signaling is a known consequence of PC2 mutations. PC2 functions as a Ca 2+ -activated Ca 2+ channel of the endoplasmic reticulum. We hypothesize that Ca 2+ signaling through PC2, or other intracellular Ca 2+ channels such as the inositol 1,4,5-trisphosphate receptor (InsP3R), is necessary to maintain renal epithelial cell function and that disruption of the Ca 2+ signaling leads to renal cyst development. The cell line LLC-PK1 has traditionally been used for studying PKD-causing mutations and Ca 2+ signaling in 2D culture systems. We demonstrate that this cell line can be used in long-term (8 wk) 3D tissue culture systems. In 2D systems, knockdown of InsP3R results in decreased Ca 2+ transient signals that are rescued by overexpression of PC2. In 3D systems, knockdown of either PC2 or InsP3R leads to cyst formation, but knockdown of InsP3R type 1 (InsP3R1) generated the largest cysts. InsP3R1 and InsP3R3 are differentially localized in both mouse and human kidney, suggesting that regional disruption of Ca 2+ signaling contributes to cystogenesis. All cysts had intact cilia 2 wk after starting 3D culture, but the cells with InsP3R1 knockdown lost cilia as the cysts grew. Studies combining 2D and 3D cell culture systems will assist in understanding how mutations in PC2 that confer altered Ca 2+ signaling lead to ADPKD cysts. primary cilia | polycysin 2 | calcium release T he commonly occurring genetic kidney disorder, autosomal dominant polycystic kidney disease (ADPKD), is the result of mutations in polycystin 1 or 2 (PC1 or PC2). The progressive cyst formation within all segments of the nephron that defines the disorder leads to renal failure requiring treatment by dialysis and/or organ transplantation (1-3). Altered Ca 2+ signaling is one of several pathways that have been implicated in the disease (4, 5). A major limitation toward elucidating the role of Ca 2+ signaling in cyst formation has been the lack of easily manipulated, physiologically relevant experimental methodologies.In the past, ADPKD research has relied largely upon data from mouse models and cells maintained in 2D cell culture. Mouse models have played a significant role in understanding the biology of cyst formation but are unable to fully recapitulate the physiology of disease progression in humans due to the inherent differences between the species including life span, genetics, and environment. Two-dimensional cell culture has the ability to provide information on signaling pathways and response to therapies in a fast, high-throughput manner, but is incapable of replicating the inherent 3D nature of cyst formation. Advances in 3D tissue culture over the past 2 decades have improved the ability to model cyst development in vitro. However, previously published 3D tissue models of ADPKD have relied upon shortter...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Cyst formation following disruption of intracellular calcium signaling

Kuo¹,

DesRochers

Kimmerling

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

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“…These studies have revealed advantages of 3D tissue models over 2D cell cultures due to differences in cell-cell signaling, ECM interactions, ECM composition, gene expression, and epigenetic gene regulation (23,36). Bioengineered 3D kidney tissues produced in our lab using human cells have been shown to better reflect human renal physiology than 2D cell culture (37). These tissues are composed of human telomerase reverse transcriptase (hTERT) immortalized human renal proximal tubule cells embedded within a hydrogel.…”

mentioning

confidence: 99%

Effects of Shiga Toxin Type 2 on a Bioengineered Three-Dimensional Model of Human Renal Tissue

et al. 2015

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dShiga toxins (Stx) are a family of cytotoxic proteins that can cause hemolytic-uremic syndrome (HUS), a thrombotic microangiopathy, following infections by Shiga toxin-producing Escherichia coli (STEC). Renal failure is a key feature of HUS and a major cause of childhood renal failure worldwide. There are currently no specific therapies for STEC-associated HUS, and the mechanism of Stx-induced renal injury is not well understood primarily due to a lack of fully representative animal models and an inability to monitor disease progression on a molecular or cellular level in humans at early stages. Three-dimensional (3D) tissue models have been shown to be more in vivo-like in their phenotype and physiology than 2D cultures for numerous disease models, including cancer and polycystic kidney disease. It is unknown whether exposure of a 3D renal tissue model to Stx will yield a more in vivo-like response than 2D cell culture. In this study, we characterized Stx2-mediated cytotoxicity in a bioengineered 3D human renal tissue model previously shown to be a predictor of drug-induced nephrotoxicity and compared its response to Stx2 exposure in 2D cell culture. Our results demonstrate that although many mechanistic aspects of cytotoxicity were similar between 3D and 2D, treatment of the 3D tissues with Stx resulted in an elevated secretion of the kidney injury marker 1 (Kim-1) and the cytokine interleukin-8 compared to the 2D cell cultures. This study represents the first application of 3D tissues for the study of Stx-mediated kidney injury.

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“…This transformation allows the cells to maintain their intact chromosomes after every doubling, resulting in stable lines [70][71][72].…”

Section: Conditionally Immortalized Cell Linesmentioning

confidence: 99%

Development of an upscaled bioartificial kidney

Chevtchik¹

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Printed by Gildeprint, Enschede, the Netherlands, Cover design by Natalia Chevtchik and Felix Broens Front page: confocal microscopy image of conditionnaly imrtalized proximal tubule renal epithelial cells (ciPTEC) cultured on polymeric hollow fiber membranes (HFM) with DAPI staining of the nuclei and immunostaining for the tight junction protein ZO-1. The kidneys are composed of hundreds of thousands of filtration units called nephrons (see Figure 1). In the nephron, blood initially passes through the glomerulus where small and middle-size solutes and excess fluids are removed out of the blood by convection. This glomerular filtrate is then transferred to the proximal tubules, which are responsible for reabsorbing essential components of the pre-urine but also for additional removal of a great variety of solutes and wastes from the blood stream, among which, the protein-bound toxins. More details about kidney physiology are explained in chapter 2 of this thesis. DEVELOPMENT OF AN UPSCALED BIOARTIFICIAL KIDNEY Kidney diseaseMore than 10% of the worldwide population is estimated to present a more or less severe form of kidney disease [4][5][6] Chapter 1 4 Renal replacement therapiesThe most common treatment for CKD, ESRD and AKI patients is artificial kidney or dialysis, applied for 2.2 million patients worldwide [10]. This treatment only covers a fraction of the physiological renal function -mostly that of the glomerulus in the normal kidney -and its efficiency in waste removal is incomplete [11, 12]. Indeed, only small water-soluble molecules, inferior to 40 kDa, present in free fraction in the blood, can be eliminated [13] whereas most of the big size and protein bound toxins cannot be removed. Their accumulation is strongly linked to the fatal outcome of the hemodialysed population [14][15][16].These toxins are in large part handled by the proximal tubules in the healthy kidneys [13, 17]. Besides, dialysis is removing a part of the toxin population but is not replacing the kidney endocrine and metabolic physiological functions. The need for a more complete renal replacement therapyThere is a strong need for a device, extracorporeal or implantable, which could fully replace the kidney function. Such a device could be a hybrid combination of polymeric membranes and renal proximal tubule epithelial cells (PTEC), called the bioartificial kidney (BAK). The BAK is conceived to be used in combination with a classical hemofilter [18, 19]. In this way, there is a direct similitude with the natural kidney. First, the glomerular function is replaced by the classical hemodialysis for removal of small size water-soluble molecules.Second, the glomerular filtrate, which comes out of the hemodialysis module, can be processed by the PTEC of the BAK. The principle of application of the BAK is presented in Figure 2.The first BAK prototype was presented by Aebisher et al. in 1987 [21]. Since then, several other prototypes have been proposed by the groups of Humes, Zink and Saito [18,[22][23][24][25][26][27][28][29]. The first...

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Bioengineered 3D Human Kidney Tissue, a Platform for the Determination of Nephrotoxicity

Cited by 111 publications

References 44 publications

Cyst formation following disruption of intracellular calcium signaling

Cyst formation following disruption of intracellular calcium signaling

Effects of Shiga Toxin Type 2 on a Bioengineered Three-Dimensional Model of Human Renal Tissue

Development of an upscaled bioartificial kidney

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