Localization of Progenitor/Stem Cells in the Human Tympanic Membrane

Knutsson, Johan; Unge, Magnus von; Rask‐Andersen, Helge

doi:10.1159/000320612

Cited by 45 publications

(40 citation statements)

References 42 publications

(35 reference statements)

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“…Fewer cells were observed to grow out onto the membrane from the annulus explant (Figure 2B). The TM annulus is thought to contain regenerative centers (1,16,17); however, when we attempted to graft annulus tissue into the membrane, the grafts were not as successful. Orienting the annulus tissue in the PET membrane was not as straightforward as it was with the umbo grafts possessing the malleus handle.…”

Section: Resultsmentioning

confidence: 98%

“…Studies have localized areas of high cell turnover in the TM to the center of the membrane (the umbo region) and the outer rim (annulus region), which have been suggested to contain stem cells (1,2) or regenerative centers (3,4). However, there is limited understanding of the biology of the cells responsible for regeneration or their niche within the TM, and in vitro models of the TM are currently unavailable.…”

mentioning

confidence: 99%

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Tympanic Membrane Organ Culture using Cell Culture well Inserts Engrafted with Tympanic Membrane Tissue Explants

2017

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Tissue engineering approaches using growth factors and various materials for repairing chronic perforations of the tympanic membrane are being developed, but there are surprisingly few relevant tissue culture models available to test new treatments. Here, we present a simple three-dimensional model system based on micro-dissecting the rat tympanic membrane umbo and grafting it into the membrane of a cell culture well insert. Cell outgrowth from the graft produced sufficient cells to populate a membrane of similar surface area to the human tympanic membrane within 2 weeks. Tissue grafts from the annulus region also showed cell outgrowth but were not as productive. The umbo organoid supported substantial cell proliferation and migration under the influence of keratinocyte growth medium. Cells from umbo grafts were enzymatically harvested from the polyethylene terephthalate (PET) membrane for expansion in routine culture and cells could be harvested consecutively from the same graft over multiple cycles. We used harvested cells to test cell migration properties and to engraft a porous silk scaffold material as proof-of-principle for tissue engineering applications. This model is simple enough to be widely adopted for tympanic membrane regeneration studies and has promise as a tissue-equivalent model alternative to animal testing.

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Section: Resultsmentioning

confidence: 98%

mentioning

confidence: 99%

Tympanic Membrane Organ Culture using Cell Culture well Inserts Engrafted with Tympanic Membrane Tissue Explants

2017

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“…In response to acute F I G U R E 3 Cell dynamics in the outer epidermal layer of the tympanic membrane. (a) Location and movement of stem/progenitor cells based on lineage tracing experiments and proliferation assays (Frumm et al, 2019;Knutsson, von Unge, & Rask-Andersen, 2011). (b) Migration patterns of keratinocytes based on ink dye labeling experiments in rodents and humans (Jackler, Santa Maria, Varsak, Nguyen, & Blevins, 2015;Michaels & Soucek, 1991).…”

Section: Repair Of the Ear Drummentioning

confidence: 99%

“…Due to this migratory potential of the outer keratin layer, and also because of easier experimental access to the outside of the eardrum, research has focused on progenitor/stem cell localization in the epidermal layer. Immunohistochemical analysis for integrins and cytokeratin 19 in rat and human TM indicated the localized presence of progenitor cells in the epidermal layer covering the manubrium, annulus, and handle of the malleus (Knutsson et al, ; W. Wang, Wang, & Tian, ). This finding was supported by studies investigating cell proliferation using the thymidine analogue BrdU.…”

Section: Introductionmentioning

confidence: 99%

Developmental aspects of the tympanic membrane: Shedding light on function and disease

Mozaffari

Jiang

Tucker

2019

Genesis

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Summary The ear drum, or tympanic membrane (TM), is a key component in the intricate relay that transmits air‐borne sound to our fluid‐filled inner ear. Despite early belief that the mammalian ear drum evolved as a transformation of a reptilian drum, newer fossil data suggests a parallel and independent evolution of this structure in mammals. The term “drum” belies what is in fact a complex three‐dimensional structure formed from multiple embryonic cell lineages. Intriguingly, disease affects the ear drum differently in its different parts, with the superior and posterior parts being much more frequently affected. This suggests a key role for the developmental details of TM formation in its final form and function, both in homeostasis and regeneration. Here we review recent studies in rodent models and humans that are beginning to address large knowledge gaps in TM cell dynamics from a developmental biologist's point of view. We outline the biological and clinical uncertainties that remain, with a view to guiding the indispensable contribution that developmental biology will be able to make to better understanding the TM.

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“…The mucosal clusters express Krt19, while the keratinocyte clusters do not, and restriction of this keratin to the mucosal cells was confirmed by lineage tracing (Figure S5C). Of note, Krt19 has been proposed as a marker of TM epidermal stem cells in human and rat (Knutsson et al 2011;Kim et al 2015).…”

Section: Identifying Cell Populations In the Murine Tmmentioning

confidence: 99%

A hierarchy of migratory keratinocytes maintains the tympanic membrane

Frumm¹,

Yu²,

Chang³

et al. 2019

Preprint

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Although the conductive function of the tympanic membrane (TM) is critical for hearing, it is unknown how the organ maintains cellular homeostasis. Using a combination of single-cell RNA sequencing, lineage tracing, whole-organ explant, and live-cell imaging, we demonstrate that the stem cells of the TM epidermis reside in a distinct location at the superior portion of the TM and, as progeny migrate inferiorly, Pdgfra+ fibroblasts maintain a niche supporting proliferation of committed progenitors, while keratinocytes distal from the niche differentiate. Thus, the TM has a three dimensional differentiation hierarchy of keratinocytes distinct from that at other epidermal sites. The TM represents a physiological context where, in the absence of injury, keratinocytes both transit through a proliferative committed progenitor state and exhibit directional lateral migration.This work forms a foundation for understanding common disorders of the TM and introduces a new model system for the understanding of keratinocyte biology. 1N).The same approaches were utilized to define and locate the cell populations in the fibrous/mucosal fraction, where 12 clusters were discovered. Among these are multiple Vimentin+ (Vim) mesenchymal populations, including: endothelium (Pecam1+), smooth muscle (Acta2+), and Schwann cells (Mbp+) (Figure S3). Four additional clusters of mesenchymal cells were identified, numbered 0, 2, 3, and 5. Inspection of the differentially expressed genes revealed that cluster 0 is Gpx3+, and clusters 2, 3, and 5 are Igfbp3+ (Figure S4A-B). ISH confirmed that Gpx3 and Igfbp3 do indeed mark distinct populations of fibroblasts (Figure S4C). The Gpx3+ cells appear to reside in proximity to bony elements-specifically, in the annular region and near the malleus. The Igfbp3+ cells are more diffusely spread throughout the fibrous layer of the TM.Lastly, Sox2 is a known marker of mucosa, and is expressed in both non-ciliated (clusters 4 and 11)

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Localization of Progenitor/Stem Cells in the Human Tympanic Membrane

Cited by 45 publications

References 42 publications

Tympanic Membrane Organ Culture using Cell Culture well Inserts Engrafted with Tympanic Membrane Tissue Explants

Tympanic Membrane Organ Culture using Cell Culture well Inserts Engrafted with Tympanic Membrane Tissue Explants

Developmental aspects of the tympanic membrane: Shedding light on function and disease

A hierarchy of migratory keratinocytes maintains the tympanic membrane

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