Human neuroblastoma (SH-SY5Y) cell culture and differentiation in 3-D collagen hydrogels for cell-based biosensing

Desai, Anant; Kisaalita, William S.; Keith, Charles H.; Wu, Zhongzhi

doi:10.1016/j.bios.2005.07.005

Cited by 53 publications

(55 citation statements)

References 37 publications

(59 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This finding is particularly important as the viability of mammalian cells after immobilization in biocompatible matrices is the key factor for cell-based assays. 15,27,32,35 In the present study, we have shown that Ped-2E9 cells (which grow in suspension) can be immobilized threedimensionally into a biocompatible collagen gel matrix without losing their biological and physiological properties of sensitivity to pathogenic Listeria and Bacillus cells or toxins. This 3D immobilization also reduces the normal 2D assay steps such as centrifugation, critically important for the integration of these cell types in a biosensor platform.…”

Section: Discussionmentioning

confidence: 63%

“…33 Three-dimensional (3D) encapsulation of different cell types in collagen gel has become a standard method in tissue engineering applications. 31,32 Application of mammalian cells in collagen-entrapped form for use as biosensors is still in its infancy, 34,35 but offers significant promise. [34][35][36] Previous studies from our laboratory have shown that a mouse B-cell hybridoma (Ped-2E9)-based cytotoxicity assay can be used to detect the presence of pathogenic Listeria species and the enterotoxins from Bacillus species.…”

mentioning

confidence: 99%

“…31,32 Application of mammalian cells in collagen-entrapped form for use as biosensors is still in its infancy, 34,35 but offers significant promise. [34][35][36] Previous studies from our laboratory have shown that a mouse B-cell hybridoma (Ped-2E9)-based cytotoxicity assay can be used to detect the presence of pathogenic Listeria species and the enterotoxins from Bacillus species. 16,37,38 This cytotoxicity assay can also differentiate the presence of viable pathogenic Listeria species from nonpathogenic ones.…”

mentioning

confidence: 99%

See 2 more Smart Citations

A novel and simple cell-based detection system with a collagen-encapsulated B-lymphocyte cell line as a biosensor for rapid detection of pathogens and toxins

Banerjee

Lenz

Robinson

et al. 2008

Laboratory Investigation

View full text Add to dashboard Cite

Cell-based biosensors (CBBs) are becoming important tools for biosecurity applications and rapid diagnostics in food microbiology for their unique capability of detecting physiologically hazardous materials. A multi-well plate-based biosensor containing B-cell hybridoma, Ped-2E9, encapsulated in type I collagen matrix, was developed for rapid detection of viable cells of pathogenic Listeria, the toxin listeriolysin O, and the enterotoxin from Bacillus species. This sensor measures the alkaline phosphatase release from infected Ped-2E9 cells colorimetrically. Pathogenic L. monocytogenes cells and toxin preparations from L. monocytogenes or B. cereus showed cytotoxicity ranging from 24 to 98% at 3-6 h postinfection. In contrast, nonpathogenic L. innocua (F4247) and B. subtilis induced minimal cytotoxicity, ranging only 0.4-7.6%. Laser scanning cytometry and cryo-nano scanning electron microscopy confirmed the live or dead status of the infected Ped-2E9 cells in gel matrix. This paper presents the first example of a cell-based sensing system using collagen-encapsulated mammalian cells for rapid detection of pathogenic bacteria or toxin, and demonstrates a potential for onsite use as a portable detection system.

show abstract

Section: Discussionmentioning

confidence: 63%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

A novel and simple cell-based detection system with a collagen-encapsulated B-lymphocyte cell line as a biosensor for rapid detection of pathogens and toxins

Banerjee

Lenz

Robinson

et al. 2008

Laboratory Investigation

View full text Add to dashboard Cite

show abstract

“…However, we were able to detect inhibition by pore blockers such as tetrodotoxin, µ-conotoxin TIIIA and clinically used compounds including amitriptyline and tetracaine (Fig 7), as well as inhibition by the state dependent blocker ProTxII through the measurement of Ca 2+ fluxes. This discrepancy may reflect expression of endogenous sodium channels in SH-SY5Y cells at a more physiological membrane potential compared to commonly used over-expression systems such as HEK293 cells [35,36]. In addition, the human Nav in SH-SY5Y cells are co-expressed with functionally relevant β-subunits, which could affect inhibition of Nav activity by state-dependent blockers such as ProTxII.…”

Section: Discussionmentioning

confidence: 99%

Characterisation of Nav types endogenously expressed in human SH-SY5Y neuroblastoma cells

Vetter

Mozar

Durek

et al. 2012

Biochemical Pharmacology

View full text Add to dashboard Cite

Abbreviations: Nav, voltage-gated sodium channel; TTX, tetrodotoxin; ProTxII, β/ω-theraphotoxin-Tp2a; Ca 2+ , calcium ion; DMEM, Dulbecco's Modified Eagle's Medium; FBS, foetal bovine serum; PBS, phosphate buffered saline; PSS, physiological salt solution; HBS, HEPES-buffered saline; AFU, arbitrary fluorescence unit; SEM, standard error of the mean; Fluo-4 AM, Fluo-4 acetoxymethylester; RPMI, Roswell Park Memorial Institute; BSA, bovine serum albumin; CCD, charge-coupled device; DAPI, 4',6-diamidino-2-phenylindole 2 AbstractThe human neuroblastoma cell line SH-SY5Y is a potentially useful model for the identification and characterisation of Nav modulators, but little is known about the pharmacology of their endogenously expressed Navs. The aim of this study was to determine the expression of endogenous Nav α and β subunits in SH-SY5Y cells using PCR and immunohistochemical approaches, and pharmacologically characterise the Nav isoforms endogenously expressed in this cell line using electrophysiological and fluorescence approaches. SH-SY5Y human neuroblastoma cells were found to endogenously express several Nav isoforms including Nav1.2 and Nav1.7. Activation of endogenously expressed Navs with veratridine or the scorpion toxin OD1 caused membrane depolarization and subsequent Ca 2+ influx through voltage-gated L-and N-type calcium channels, allowing Nav activation to be detected with membrane potential and fluorescent Ca 2 dyes. -Conotoxin TIIIA and ProTxII identified Nav1.2 and Nav1.7 as the major contributors of this response. The Nav1.7-selective scorpion toxin OD1 in combination with veratridine produced a Nav1.7-selective response, confirming that endogenously expressed human Nav1.7 in SH-SY5Y cells is functional and can be synergistically activated, providing a new assay format for ligand screening.Key Words: SH-SY5Y; Ca 2+ ; Nav1.7; ProTxII; OD1 3 IntroductionVoltage-gated sodium channels (Nav) are complex transmembrane proteins comprised of a poreforming α subunit and accessory β subunits that play an essential role in the initiation and propagation of action potentials in excitable cells. To date, apart from the related Nax which appears to function as a sodium sensor [1,2], nine isoforms termed Nav1.1 -Nav1.9 have been functionally defined as sodium-selective ion channels [3]. Their distinct tissue distribution and amenability to modulation by toxins and drugs has led to significant interest in Nav as therapeutic targets in a number of poorly treated conditions ranging from epilepsy to cardiac arrhythmias and pain [4]. In recent years Nav1.7 has emerged as an attractive drug target, with expression restricted to a subset of nociceptive neurons that is expected to limit on-target side effects of pharmacological modulators of Nav1.7 [5]. In addition, loss-of-function mutations in humans have highlighted the possibility that Nav1.7 inhibition could produce complete loss of pain sensations without dose-limiting side effects [6]. However, it remains unclear if such an effect can be translated to the clinic...

show abstract

“…A number of studies have been done to reveal the effects of 2D versus 3D cell culture on differentiation, [21][22][23][24][25] drug metabolism, [26][27][28][29][30] gene expression and protein synthesis, [31][32][33][34][35] general cell function, [36][37][38][39][40] increase of in vivo relevance, 31,[41][42][43] morphology, [44][45][46][47] proliferation, 26,[48][49][50][51] response to stimuli, 41,[52][53][54] viability, 36,[55][56][57] and migration. [58][59][60][61] Adding the third dimension to cellular environment provides them with more in vivo-like morphology, behaviors, and intercellular interactions for understanding cytology in more physiological conditions.…”

Section: Introductionmentioning

confidence: 99%

Electrotaxis of lung cancer cells in ordered three-dimensional scaffolds

et al. 2012

View full text Add to dashboard Cite

In this paper, we report a new method to incorporate 3D scaffold with electrotaxis measurement in the microfluidic device. The electrotactic response of lung cancer cells in the 3D foam scaffolds which resemble the in vivo pulmonary alveoli may give more insight on cellular behaviors in vivo. The 3D scaffold consists of ordered arrays of uniform spherical pores in gelatin. We found that cell morphology in the 3D scaffold was different from that in 2D substrate. Next, we applied a direct current electric field (EF) of 338 mV/mm through the scaffold for the study of cells' migration within. We measured the migration directedness and speed of different lung cancer cell lines, CL1-0, CL1-5, and A549, and compared with those examined in 2D gelatin-coated and bare substrates. The migration direction is the same for all conditions but there are clear differences in cell morphology, directedness, and migration speed under EF. Our results demonstrate cell migration under EF is different in 2D and 3D environments and possibly due to different cell morphology and/or substrate

show abstract

Human neuroblastoma (SH-SY5Y) cell culture and differentiation in 3-D collagen hydrogels for cell-based biosensing

Cited by 53 publications

References 37 publications

A novel and simple cell-based detection system with a collagen-encapsulated B-lymphocyte cell line as a biosensor for rapid detection of pathogens and toxins

A novel and simple cell-based detection system with a collagen-encapsulated B-lymphocyte cell line as a biosensor for rapid detection of pathogens and toxins

Characterisation of Nav types endogenously expressed in human SH-SY5Y neuroblastoma cells

Electrotaxis of lung cancer cells in ordered three-dimensional scaffolds

Contact Info

Product

Resources

About