Primary Cultures of Human Vestibular Schwannoma

Nair, Salil; Leung, Hing Y.; Collins, Anne T.; Ramsden, R. T.; Wilson, Janet A.

doi:10.1097/01.mao.0000247811.93453.6a

Cited by 6 publications

(4 citation statements)

References 21 publications

(27 reference statements)

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“…Our protocol was also effective in establishing a robust schwannoma cell culture (80% pure) for the majority of 12 weeks, in contrast to others who noted significant fibroblast contamination when using a minimally manipulative protocol (Nair, Leung, Collins, Ramsden, & Wilson, 2007). Our purity was highest at 2 weeks (87% pure), suggesting that this time point is optimal for use of the culture to investigate schwannoma pathobiology.…”

Section: Discussionmentioning

confidence: 66%

“…Additionally, cultures derived from adult rat sciatic and other peripheral nerves (Mauritz, Grothe, & Haastert, 2004; Morrissey, Kleitman, & Bunge, 1991; Niapour et al, 2010) may have limited applicability to humans, as cultured animal and human SCs can behave differently (Morrissey, Kleitman, & Bunge, 1991). Primary cultures derived from human VS are established in a similar manner as SCs to prevent fibroblast contamination (Nair, Leung, Collins, Ramsden, & Wilson, 2007), using methods that may alter VS pathobiology. Further, those who have successfully cultured VS cells have not characterized these cultures over time (Bush et al, 2012; Neff et al, 2012).…”

Section: Introductionmentioning

confidence: 99%

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Primary culture of human Schwann and schwannoma cells: Improved and simplified protocol

et al. 2014

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Primary culture of human Schwann cells (SCs) and vestibular schwannoma (VS) cells are invaluable tools to investigate SC physiology and VS pathobiology, and to devise effective pharmacotherapies against VS, which are sorely needed. However, existing culture protocols, in aiming to create robust, pure cultures, employ methods that can lead to loss of biological characteristics of the original cells, potentially resulting in misleading biological findings. We have developed a minimally manipulative method to culture primary human SC and VS cells, without the use of selective mitogens, toxins, or time-consuming and potentially transformative laboratory techniques. Schwann cell purity was quantified longitudinally using S100 staining in SC cultures derived from the great auricular nerve and VS cultures followed for 7 and 12 weeks, respectively. SC cultures retained approximately ≥85% purity for 2 weeks. VS cultures retained approximately ≥80% purity for the majority of the span of 12 weeks, with maximal purity of 87% at 2 weeks. The VS cultures showed high level of biological similarity (68% on average) to their respective parent tumors, as assessed using a protein array featuring 41 growth factors and receptors. Apoptosis rate in vitro negatively correlated with tumor volume. Our results, obtained using a faster, simplified culturing method than previously utilized, indicate that highly pure, primary human SC and VS cultures can be established with minimal manipulation, reaching maximal purity at 2 weeks of culture. The VS cultures recapitulate the parent tumors' biology to a great degree, making them relevant models to investigate VS pathobiology.

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Section: Discussionmentioning

confidence: 66%

Section: Introductionmentioning

confidence: 99%

Primary culture of human Schwann and schwannoma cells: Improved and simplified protocol

et al. 2014

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“…[9] Most subsequent published protocols, as our own, utilize a similar protocol of seeding minced tumor fragments onto a culture surface treated with a charged amino acid (poly-L-ornithine, poly-L-lysine) and laminin preparation. [8] We have used this same method to culture schwannomas arising from the facial, trigeminal, vagus, and spinal nerves with similar results.…”

Section: Discussionmentioning

confidence: 94%

“…Most previous VS culture techniques required relatively long processing times and complicated selective culture techniques. [6–8] Here we present a simple, reproducible protocol for primary VS cell culture with complete processing in under 3 hours, with 95% tumor cell purity as determined by immunostaining.…”

Section: Introductionmentioning

confidence: 99%

Primary Culture of Human Vestibular Schwannomas

Schularick

Clark

Hansen

2014

JoVE

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Vestibular schwannomas (VSs) represent Schwann cell (SC) tumors of the vestibular nerve, compromising 10% of all intracranial neoplasms. VSs occur in either sporadic or familial (neurofibromatosis type 2, NF2) forms, both associated with inactivating defects in the NF2 tumor suppressor gene. Treatment for VSs is generally surgical resection or radiosurgery, however the morbidity of such procedures has driven investigations into less invasive treatments. Historically, lack of access to fresh tissue specimens and the fact that schwannoma cells are not immortalized have significantly hampered the use of primary cultures for investigation of schwannoma tumorigenesis. To overcome the limited supply of primary cultures, the immortalized HEI193 VS cell line was generated by transduction with HPV E6 and E7 oncogenes. This oncogenic transduction introduced significant molecular and phenotypic alterations to the cells, which limit their use as a model for human schwannoma tumors. We therefore illustrate a simplified, reproducible protocol for culture of primary human VS cells. This easily mastered technique allows for molecular and cellular investigations that more accurately recapitulate the complexity of VS disease.

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Generation and Use of Merlin-Deficient Human Schwann Cells for a High-Throughput Chemical Genomics Screening Assay

Petrilli

Fernández‐Valle

2018

Methods in Molecular Biology

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Schwannomas are benign nerve tumors that occur sporadically in the general population and in those with neurofibromatosis type 2 (NF2), a tumor predisposition genetic disorder. NF2-associated schwannomas and most sporadic schwannomas are caused by inactivating mutations in Schwann cells in the neurofibromatosis type 2 gene (NF2) that encodes the merlin tumor suppressor. Despite their benign nature, schwannomas and especially vestibular schwannomas cause considerable morbidity. The primary available therapies are surgery or radiosurgery which usually lead to loss of function of the compromised nerve. Thus, there is a need for effective chemotherapies. We established an untransformed merlin-deficient human Schwann cell line for use in drug discovery studies for NF2-associated schwannomas. We describe the generation of human Schwann cells (HSCs) with depletion of merlin and their application in high-throughput screening of chemical libraries to identify compounds that decrease their viability. This NF2-HSC model is amenable for use in independent labs and high-throughput screening (HTS) facilities.

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Primary Cultures of Human Vestibular Schwannoma

Abstract: We describe a method for obtaining short-term, essentially fibroblast-free, primary VS cultures. Such pure VS cultures, retaining in vivo characteristics, are extremely useful as an in vitro model to study the pathobiology of schwannoma cells.

Cited by 6 publications

References 21 publications

Primary culture of human Schwann and schwannoma cells: Improved and simplified protocol

Primary culture of human Schwann and schwannoma cells: Improved and simplified protocol

Primary Culture of Human Vestibular Schwannomas

Generation and Use of Merlin-Deficient Human Schwann Cells for a High-Throughput Chemical Genomics Screening Assay

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