(2023) 24:31
Kim et al. BMC Molecular and Cell Biology
https://doi.org/10.1186/s12860-023-00493-2

BMC Molecular andCell Biology

Open Access

RESEARCH

Simple, low‑cost, and well‑performing
method, the outgrowth technique,for the isolation of cells from nasal polyps
Jonghui Kim1* , Karla Hegener1 , Claudia Hagedorn2 , Daniel Weidinger3 , Kashin Jamal Jameel3 ,Inga Marte Charlott Seuthe1 , Sabine Eichhorn1 , Florian Kreppel2 , Jonas Jae‑Hyun Park1†   and Jürgen Knobloch3†   

Abstract
Background Epithelial cells are an important part of the pathomechanism in chronic rhinosinusitis with nasal polyps.
It is therefore essential to establish a robust method for the isolation and culture of epithelial cells from nasal polyps
to enable further research. In this study, the feasibility of the outgrowth technique for the isolation of the epithelial
cells from the nasal polyps was evaluated.
Results Using the outgrowth technique, epithelial cells could be isolated from all tissue samples. Isolated epithelial
cells showed a proliferation rate of approximately 7- to 23-fold every 6 days up to the 3rd passage. Over 97% of iso‑lated cells were shown to be cytokeratin- and p63-positive, and over 86% of them were Ki-67–positive in flow cytom‑
etry. Interleukin-33 and periostin were detectable in the supernatant.
Conclusions We introduce a simple, low-cost, and well-performing method for isolating epithelial cells from nasal
polyps with the outgrowth technique.
Keywords Nasal polyps, CRSwNP, Epithelial cells, Isolation, Outgrowth techniqueBackground
Nasal polyps can be developed in the course of chronicrhinosinusitis, which is characterized as the inflammation of the mucous of the nasal and paranasal sinuses.
Chronic rhinosinusitis with nasal polyps (CRSwNP) is†

Jonas Jae-Hyun Park and Jürgen Knobloch contributed equally to this work.

*Correspondence:
Jonghui Kim
jonghui.kim@uni-wh.de1
Department of Oto‑Rhino‑Laryngology – Head and Neck Surgery, St.Josefs-Hospital Hagen, University of Witten/Herdecke, Dreieckstraße 15,58097 Hagen, Germany
2
Chair of Biochemistry and Molecular Medicine, Center for Biomedical
Education and Research (ZBAF), University of Witten/Herdecke,
Stockumer Straße 10, 58453 Witten, Germany
3
Medical Clinic III for Pneumology, Allergology and Sleep Medicine,
Bergmannsheil University Hospital, Ruhr-University Bochum,
Bürkle‑de‑La‑Camp‑Platz 1, 44789 Bochum, Germany

generally characterized as type 2 inflammation in western countries [1]. Much evidence, such as the infiltration of eosinophilic granulocytes in nasal polyps, hassuggested that CRSwNP is a type 2 inflammatory disease [1]. Furthermore, the recently approved therapies
with dupilumab (anti-interleukin-4/anti-interleukin-13),
mepolizumab (anti-interleukin-5), and omalizumab(anti-immunoglobulin E) in patients with nasal polyposis
are highly effective [2–4], so it is strongly indicated thatthe regulation of interleukin-4, interleukin-5, and immunoglobulin E via the type 2–inflammation track may playa crucial role in the pathogenesis.
The epithelial cells seem to be an important part of theentire pathomechanism of CRSwNP [5]. The epithelial
cells of nasal polyps are a distinct pathological cell typeand differ from healthy nasal epithelial cells. As such,basal cell hyperplasia, features of epithelial-mesenchymaltransition, and deregulation of epithelial differentiation© The Author(s) 2023. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which
permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to theoriginal author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images orother third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line
to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory
regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of thislicence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecom‑mons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Kim et al. BMC Molecular and Cell Biology(2023) 24:31

markers and of key regulators of neurogenic inflammation are indicators for pathological epithelial cells innasal polyps [6–8]. Epithelial cells also play an importantrole in type 2 inflammation by secreting a large numberof pro-inflammatory mediators, such as interleukin-33(IL-33) [9], which on the one hand activate type 2 innate
lymphoid cells and mast cells directly and, on the other
hand, support the conventional dendritic cell–mediatedactivation of Th2 cells and the subsequent production of
immunoglobulin E produced by B cells [5]. As a result,remodeling takes place, with a strong change in the composition of the epithelium, resulting, for example, inbasal cell hyperplasia, ciliary dysfunction, and impaired
mucus production of the epithelial cells [5]. The mechanism of the remodeling process in CRSwNP is, however,not fully understood, but it might be an auspicious drug
target. Therefore, it is mandatory to establish a preclinical method, particularly a cell culture model enabling a

Page 2 of 7

wide range of investigations to better understand the roleof epithelial cells in CRSwNP.
In this study, the feasibility of the outgrowth technique
for the isolation of the epithelial cells from the nasal
polyps was evaluated.

Results
Isolation of nasal epithelial cells from polyp tissue, using
the outgrowth technique, and evaluation of proliferation

Epithelial cells from the polyp tissue were observed byculturing using the outgrowth technique. First, outgrowth from the tissue samples of 3 patients was evaluated. Per sample, 24 small pieces of tissue were taken, as
described below, to isolate epithelial cells. The outgrowthof epithelial cells was evaluated every 2 days. Almost all
small pieces of tissue shed epithelial cells within 8 days
(Fig. 1): 19 of 24 tissue pieces from Polyp 1 yielded cells,
and epithelial cells grew out from all 24 tissue pieces of
Polyp 2 and Polyp 3. It was possible to isolate the epithelial cells from all 3 samples (Fig. 2).
On Day 8 of outgrowth, 320,000 (Polyp 1), 90,000(Polyp 2), and 320,000 (Polyp 3) cells were collected,
respectively. Cells were transferred to a 6-well plate at
5,000 cells per well, and the number of cells from 6 wells
together was counted every 6 days up to the 3rd passage.
Epithelial cells showed an excellent proliferation rate of
approximately 7- to 23-fold up to the 3rd passage (Fig. 3).
Flow cytometric analysis to identify epithelial cells
and evaluate differentiation and proliferation

Fig. 1 Epithelial cells grew out of polyp tissue. Outgrowth
was observed in most tissues (19/24) in the first 8 days

To identify epithelial cells in flow cytometric analysis,
cells were stained with anti-cytokeratin antibody. In addition, the cells were simultaneously stained with anti-p63

Fig. 2 Cumulative number of tissue pieces from which cells grew out. Tissues from three patients were taken for isolation of epithelial cells. 24
small-cut pieces of tissue per sample from each patient (two pieces per well in a 12-well plate) were used for outgrowth of epithelial cells. The
cumulative number of tissue samples from which cells grew out was assessed every 2 days. Cells grew out from almost all tissue pieces (67/72)
within 8 days – 19 of 24 tissue pieces from Polyp 1 yielded cells. Epithelial cells grew out from all tissue pieces of Polyp 2 and Polyp 3

Kim et al. BMC Molecular and Cell Biology(2023) 24:31

Page 3 of 7

Fig. 3 Fold increase in cell number up to the 3rd passage. Per well, 5,000 cells were seeded in a 6-well plate with 2 ml BEGM (30,000 cells in total).
The medium was renewed every 2 days. The number of total cells from the 6-well plate was counted after 6 days. Epithelial cells showed a good
proliferation rate, up to the 3rd passage, of approximately 7- to 23-fold. On Day 6 of each passage, approximately 205,000, 687,500, and 315,000 cells
were counted from the Polyp 1 tissue; approximately 550,000, 332,500, and 240,000 cells from Polyp 2 tissue; and approximately 400,000, 535,000,
and 375,000 cells from Polyp 3 tissue, respectively

and anti-Ki-67 to assess differentiation and proliferation.
Positive and negative populations were predefined bycompensation setup. Of the cells, 97.02% ± 0.2194% werecytokeratin and p63 positive (Fig. 4A), indicating that theisolated cells are undifferentiated epithelial cells. In addition, 86.45% ± 2.532% of cells were Ki-67 positive (Fig. 4B),
which reflects good proliferation of the cells (Fig. 3).
Functionality test by determining type 2–relevant proteins,using ELISA

The function of the epithelial cells was assessed by determining the production of a pro-inflammatory cytokine
and protein produced by the epithelium in type 2 disease,
representatively IL-33 and periostin. Cells were able toproduce IL-33 (average ± SEM in pg/ml: 21.39 ± 0.7679 in
24 h, 20.18 ± 1.044 in 48 h, and 21.18 ± 1.491 in 72 h; n = 3)
and periostin (average ± SEM in pg/ml: 147.5 ± 15.92 in 24
h, 186.5 ± 41.11 in 48 h, and 250.3 ± 18.60 in 72 h, n = 3)
(Fig. 5), demonstrating sufficient functioning of the epithelial cells for further study.

Discussion
So far, the isolation of nasal epithelial cells was performed either with brushing of the mucosa or dissociation of nasal tissue with protease [10, 11]. Some
researchers even combine the methods [12]. The brushing technique is easy to perform. Particularly, the donation is possible for volunteers who are not undergoinga surgical procedure; however, the ratio of success withthis method is approximately 63%–90% [10, 13]. The
dissociation technique is performed with tissue materials from the surgery. With the dissociation technique,some difficulties are met. An explicit de-epithelizationof mucosa is not possible, which differs from, for example, the de-epithelialization of skin tissue, which is possible after incubation with Dispase [11, 14]. Therefore,
in the mucosa, not only the epithelium is incubatedwith protease but also the submucosal tissue, which
could deteriorate the purity of the cells. A further disadvantage is the possible alteration of the cell physiology by the protease treatment. A study has shown the
different DNA methylation profile of bronchial epithelial cells isolated by dissociation with pronase compared to those obtained by the brushing technique [15].
Another disadvantage is that the material cost due tothe required proteinase for dissociation, for example,
Dispase [11], is significantly higher than with the outgrowth technique.
The outgrowth technique is known as a method for
isolating adherent cells, such as fibroblasts [16]. Our
outgrowth method showed a good success rate inisolating epithelial cells from nasal polyp tissue (Fig. 2).
The isolated epithelial cells proliferated well up to
the 3rd passage (Fig. 3). The purity of the epithelial
population was high (Fig. 4A) – negative control withfibroblast markers was not performed in our study,
because the epithelial cells of nasal polyps can also
express mesenchymal markers as a result of epithelialmesenchymal transition [7, 17, 18]. Almost all of thecells were undifferentiated and active in proliferation(Fig. 4A and Fig. 4B). The cells were capable ofproducing the typical inflammatory cytokines andproteins of type 2 inflammation, representatively IL-33
and periostin (Fig. 5) in our study, as well as TSLP [19],Kim et al. BMC Molecular and Cell Biology(2023) 24:31

Page 4 of 7

Fig. 4 Flow cytometry analysis of cytokeratin, p63, and Ki-67 expression in isolated cells from 3 tissue samples. A The relative fluorescence intensity
due to CK (PE) and p63 (APC) is plotted on the log scale x- and y-axis, respectively. Of the cells, 97.02% ± 0.2194% are cytokeratin and p63 positive,
indicating that the isolated cells are undifferentiated epithelial cells. B The relative fluorescence intensity due to Ki-67 (FITC) and CK (PE) is plottedon the log scale x- and y-axis, respectively. Of the cells, 86.45% ± 2.532% are Ki-67 positive and are therefore in a proliferative cell cycle, which reflects
good proliferation of the cells. CK: cytokeratin

Fig. 5 Epithelial cell function was assessed by the production of typical cytokines and proteins of type 2 inflammation, IL-33 and periostin,
representatively. Cells were able to produce IL-33 and periostin, demonstrating sufficient functioning of the epithelial cells for further study. IL-33
concentration in pg/ml (n = 3, average ± SEM): 21.39 ± 0.7679 in 24 h, 20.18 ± 1.044 in 48 h, and 21.18 ± 1.491 in 72 h. Periostin concentration in pg/
ml (n = 3, average ± SEM): 147.5 ± 15.92 in 24 h, 186.5 ± 41.11 in 48 h, and 250.3 ± 18.60 in 72 h

Kim et al. BMC Molecular and Cell Biology(2023) 24:31

which trigger a cascade of inflammatory responses as
a pro-inflammatory cytokine [5] and is able to indicate
the intensity of inflammation as a marker of theremodeling [20].
In summary, we established a simple, low-cost, andwell-performing method for isolating epithelial cells
from nasal polyps by using the outgrowth technique.

Methods
Ethics statement and patient materials

This study was performed according to the ethicalstandards of the declaration of Helsinki and following
approval of the ethical committees of the universityof Witten/Herdecke (No. 209/2020). Donors gave
informed consent for the research use of their nasal
polyp tissue. Pseudonymisation was carried out, andpatient data were treated confidentially.
Nasal polyps obtained from patients with CRSwNP
and undergoing endoscopic endonasal sinus surgery
are the source of the tissue material. We performeda preoperative clinical endoscopic examination anda computed tomography scan of sinus. The patients
reported typical symptoms of chronic rhinosinusitis,such as rhinorrhea, cephalgia, and obstructed nasal
breathing. Patients with a unilateral finding were
excluded due to possible neoplasia and papilloma.
We include tissues from 6 patients for this study(Table 1). The evaluation of the isolation, proliferation,and flow cytometric analysis was carried out with
Polyp 1, Polyp 2, and Polyp 3. ELISA was performed
with epithelial cells from Polyp 4, Polyp 5, and Polyp 6.
Isolation of epithelial cells from nasal polyp tissue

All isolation and culture procedures were performed ina sterile condition with sterile instruments. Tissues wererinsed with 7.5% povidone-iodine solution (Braunol®,
B.Braun, Germany) and rinsed twice with 0.9% sodium
chloride immediately after harvest from sinus surgery. The

Page 5 of 7

tissue was stored in Dulbecco’s Modified Eagle Medium(Gibco, cat #: 1,437,676, Carlsbad, CA, USA) with 10%
fetal bovine serum (v/v; Bio&SELL, cat #: FBS.S 0615
HI, Feucht, Germany) and 1% penicillin/streptomycin(Bio&SELL, cat #: BS.A 2213, Feucht, Germany) at 4°C.
The following treatment was carried out within 6 h after
surgical removal. The tissue was washed once with Dulbecco’s phosphate-buffered saline without calcium andmagnesium (PBS; Gibco, cat #: 14,190,169, Carlsbad,
CA, USA). Polyps were cut into small pieces on the petri
dish with surgical scissors (approx. 1–1.5 mm diameter)(Fig. 6A). The small pieces were placed in the 12-well culture plate (2 pieces/well) and placed under the safety cabinet without lid for 5–10 min to dry and stick to the bottomof the culture dish. Subsequently, 350 μl of Bronchial Epithelial Growth Medium (BEGM; Lonza, cat #: CC-3170,
Basel, Switzerland) was added carefully without mobilizing
the tissue pieces from the dish (Fig. 6B); the wells should
be completely covered with medium, but the medium volume must be as low as possible to avoid floating of the tissue pieces in the cell culture vessel (approx. 350 µl medium
in our conditions, as mentioned above). The medium wasrenewed every 2 days without mobilizing the tissue pieces.
The incubation lasted for 8 days, and the outgrowth of theepithelial cells from polyp tissues was evaluated microscopically every 2 days when the medium was changed.
The cells were treated with 300 µl trypsin 0.25%/EDTA
0.02% for 2–3 min after rinsing with PBS twice, and thetrypsin was neutralized with the same volume of soybean
trypsin inhibitor (1 mg/ml in PBS; Sigma, cat #: SLCF8902,
Steinheim, Germany). The cells were subsequently washedand centrifuged twice with PBS (300 × g) and then seededon a 6-well plate (5,000 cells per well with 1.5 ml BEGM).
The cells were grown in a humidified incubator with 5%­CO2 at 37°C. The medium was renewed every 2 days. Cells
were trypsinized at 70%–80% confluence and were re-passaged. The cell count was determined with the haemocytometer after mixing cell suspension with 0.1% trypan blue
1:1 (v/v) to identify vital cells.

Table 1 The demographic and clinical characteristics of tissue doners in this study
Polyp 1

Polyp 2Polyp 3Polyp 4

Polyp 5

Polyp 6

Age

45

65

45

47

62

58

Sex

Female

Male

Male

Male

FemaleMale

Nasal polyp score(right/left)

2/3

3/1

3/4

2/2

2/2

3/3

Prior sinus surgeries

No

No

No

No

No

Yes

Tissue eosinophilia

Yes

Yes

Yes

Yes

Yes

Yes

Asthma

No

No

No

No

Yes

No

NSAID-exacerbated respiratory
disease

No

No

Yes

No

Yes

No

Kim et al. BMC Molecular and Cell Biology(2023) 24:31

Page 6 of 7

Fig. 6 Tissue is on the petri dish placed and 1 ml PBS is added to keep the tissue from becoming dry A. The tissue is cut into small pieces of approx.
1–1.5 mm with surgical scissors. Small pieces were placed on the 12-well culture plate (2 pieces/well) and stand under the safety cabinet without lid
for 5–10 min to dry and stick on the surface of the culture dish. Approx. 350 μl of BEGM was then given carefully without mobilizing tissue pieces
from the dish (B)Flow cytometry

Enzyme‑linked immunosorbent assay (ELISA)

Flow cytometry with anti-cytokeratin-PE (Miltenyi Biotec,
clone: REA831, Bergisch Gladbach, Germany) and anti-p63(Ventana Medical, clone: 4A4, AZ, USA) was performed to
identify epithelial cells and evaluate differentiation. Simultaneously, the expression of Ki-67 (anti-Ki-67-FITC, Miltenyi
Biotec, clone: REA183, Bergisch Gladbach, Germany) was
determined to evaluate proliferation. Cells from the 2nd or
3rd passage were used for the analysis. Cells were cultured in
a 75 ­cm2 cell culture flask with 10 ml BEGM, renewing the
medium every 2 days. In 300 µl incubation buffer (0.5% BSA
in PBS), 5 × ­105—1 × ­106 cells were collected. Supernatant
was removed after centrifuging at 300 × g for 10 min at 4°C,
and the cells were washed 3 times in total in the same way,by adding 300 µl incubation buffer followed by centrifugation.
(The same washing process was carried out after each step
and will not be mentioned from here on.) Cells were fixed in
500 µl of 4% formaldehyde for 15 min at room temperature.
After removing the formaldehyde, permeabilization was carried out with 300 µl precooled permeabilization buffer (0.5%
Octoxynol-9, 0.68 µM ethylenediaminetetraacetic acid, and
1% bovine serum albumin in distilled aqua) for 10 min on ice.
The permeabilization buffer was removed, and cells wereincubated first with 300 µl unconjugated anti-p63 (dilution
1:200 in incubation buffer) overnight at 4°C and then withsecondary anti-mouse AlexaFluor633-conjugated antibody(dilution 1:200 in incubation buffer; Molecular Probes, cat#: A-21052, Eugene, OR, USA) for 1 h in a darkened chamber at room temperature. Cells were washed and incubated with fluorescent-dye conjugated anti-cytokeratin-PE
and anti-Ki-67-FITC (each dilution 1:200 in incubationbuffer) overnight at 4°C in a darkened chamber. Cells were
washed and resuspended in 100 µl incubation buffer toperform a flow cytometric analysis. Compensation was set
up before analysis, and positive and negative populations
were predefined accordingly.

Epithelial cell function was assessed by the production of
typical cytokines of type 2 inflammation, representatively
IL-33 and periostin. Concentrations of IL33 and periostinin culture supernatants were measured by ELISA
according to the manufacturer’s instructions (Invitrogen,cat #: BMS2048TEN, Carlsbad, CA, USA; R&D Systems,
cat #: DY3548B, Minneapolis, MN, USA) and as
described previously [21]. To obtain the supernatants,
5,000 cells were seeded in a 24-well plate with 1 ml
BEGM, and medium was renewed every 2 days until
70%–80% confluence. Then 1 ml of BEGM was added,
and supernatants were collected after 24, 48, and 72 h.
Cells from 2nd or 3rd passage were used for the analysis.
Abbreviations
BEGM	Bronchial Epithelial Growth Medium
CRSwNP	Chronic rhinosinusitis with nasal polyps
ELISA	Enzyme-linked immunosorbent assay
IL-33	Interleukin-33
PBS	Dulbecco’s phosphate-buffered saline
Acknowledgements
The authors are grateful to the operating room nurses (Adriane Di Ninni,
Bernadette Gautzsch, Carsten Goldammer, Chendresa Todorova, Claudia
Jedraszczak, Katrin-Jasmin Schulz, Melanie Janus, Nihal Calti, Nina Morgenroth,
Patrick Neumann, Silvia Leschik and Tarik Ceyhan) of the Department of OtoRhino-Laryngology – Head and Neck Surgery at St.-Josefs-Hospital Hagen,Germany, for their help in tissue collection for this study.
Authors’ contributions
JKim, JJHP, and JKno designed the study. JJHP, IMCS, and SE helped to collectsamples. JKim, KH, CH, FK, KJJ, DW and JKno performed the experimentand analyzed data. JKim drafted the manuscript. JJHP and JKno revised themanuscript and approval of the submitted manuscript. All authors read andapproved the final manuscript.
Funding
Open Access funding enabled and organized by Projekt DEAL. This study
was financed by the general funding for research and teaching of the RuhrUniversity Bochum. The Ruhr-University Bochum played no role in the design

Kim et al. BMC Molecular and Cell Biology(2023) 24:31

of the study and collection, analysis, interpretation of data, and in writing themanuscript.
Availability of data and materials
All data that support the fndings of this study are available from thecorresponding author upon reasonable request.

Declarations
Ethics approval and consent to participate
This study was performed according to the ethical standards of thedeclaration of Helsinki and following approval of the ethical committees of
the university of Witten/Herdecke (No. 209/2020). Donors gave informed
consent for the research use of their nasal polyp tissue. Pseudonymisation was
carried out, and patient data were treated confidentially.
Consent for publication
Not applicable.
Competing interests
The authors declare no competing interests.
Received: 22 December 2022 Accepted: 2 October 2023

References
1. Bachert C, Marple B, Schlosser RJ, Hopkins C, Schleimer RP, Lambrecht BN,
et al. Adult chronic rhinosinusitis. Nat Rev Dis Primer. 2020;6(1):86.
2. Bachert C, Han JK, Desrosiers M, Hellings PW, Amin N, Lee SE, et al.
Efficacy and safety of dupilumab in patients with severe chronicrhinosinusitis with nasal Polyps (LIBERTY NP SINUS-24 and LIBERTY NP
SINUS-52): results from two multicentre, randomised, double-blind,placebo-controlled, parallel-group phase 3 trials. Lancet Lond Engl.
2019;394(10209):1638–50.
3. Gevaert P, Omachi TA, Corren J, Mullol J, Han J, Lee SE, et al. Efficacy andsafety of omalizumab in nasal polyposis: 2 randomized phase 3 trials. J
Allergy Clin Immunol. 2020;146(3):595–605.
4. Han JK, Bachert C, Fokkens W, Desrosiers M, Wagenmann M, Lee SE, et al.
Mepolizumab for chronic rhinosinusitis with nasal Polyps (SYNAPSE):a randomised, double-blind, placebo-controlled, phase 3 trial. Lancet
Respir Med. 2021;9(10):1141–53.
5. Bankova LG, Barrett NA. Epithelial cell function and remodeling in nasal
polyposis. Ann Allergy Asthma Immunol Off Publ Am Coll Allergy Asthma
Immunol. 2020;124(4):333–41.
6. Ordovas-Montanes J, Dwyer DF, Nyquist SK, Buchheit KM, Vukovic M, Deb
C, et al. Allergic inflammatory memory in human respiratory epithelial
progenitor cells. Nature. 2018;560(7720):649–54.
7. Könnecke M, Burmeister M, Pries R, Böscke R, Bruchhage KL, Ungefroren
H, et al. Epithelial-Mesenchymal transition in chronic rhinosinusitis: dif‑ferences revealed between epithelial cells from nasal Polyps and inferior
turbinates. Arch Immunol Ther Exp (Warsz). 2017;65(2):157–73.
8. Cornet M, Kristina K, Bas R, Koos Z, van Danielle E, de Esther G, et al.
Expression profiling of nasal Polyp epithelial cells identifies two distinct
phenotypes and suggests a role for neurogenic inflammation. Clin Transl
Allergy. 2015;5(4):O5.
9. Shaw JL, Fakhri S, Citardi MJ, Porter PC, Corry DB, Kheradmand F, et al.
IL-33–Responsive innate lymphoid cells are an important source of IL-13
in chronic rhinosinusitis with nasal Polyps. Am J Respir Crit Care Med.
2013;188(4):432–9.
10. Stokes AB, Kieninger E, Schögler A, Kopf BS, Casaulta C, Geiser T, et al.
Comparison of three different brushing techniques to isolate andculture primary nasal epithelial cells from human subjects. Exp Lung Res.
2014;40(7):327–32.
11. Fulcher ML, Gabriel S, Burns KA, Yankaskas JR, Randell SH. Well-dif‑ferentiated human airway epithelial cell cultures. Methods Mol Med.
2005;107:183–206.
12. Manna V, Caradonna S. Isolation, expansion, differentiation, and histologicalprocessing of human nasal epithelial cells. STAR Protoc. 2021;2(4): 100782.

Page 7 of 7

13. Hussain R, Hugosson S, Roomans GM. Isolation and culture of primary
human nasal epithelial cells from anesthetized nasal epithelia. Acta
Otolaryngol (Stockh). 2014;134(3):296–9.
14. Kim J, Guhl S, Babina M, Zuberbier T, Artuc M. Integration of the humandermal mast cell into the organotypic co-culture skin model. Methods
Mol Biol Clifton NJ. 2020;2163:91–107.
15. Clifford RL, Patel J, MacIsaac JL, McEwen LM, Johnson SR, Shaw D, et al.
Airway epithelial cell isolation techniques affect DNA methylation profiles
with consequences for analysis of asthma related perturbations to DNA
methylation. Sci Rep. 2019;9(1):14409.
16. Normand J, Karasek MA. A method for the isolation and serial
propagation of keratinocytes, endothelial cells, and fibroblasts froma single punch biopsy of human skin. In Vitro Cell Dev Biol Anim.
1995;31(6):447–55.
17. Chiarella E, Lombardo N, Lobello N, Aloisio A, Aragona T, Pelaia C,
et al. Nasal polyposis: insights in epithelial-mesenchymal transitionand differentiation of Polyp mesenchymal stem cells. Int J Mol Sci.
2020;21(18):E6878.
18. Li X, Li C, Zhu G, Yuan W, Xiao ZA. TGF-β1 induces epithelial-mesenchymaltransition of chronic sinusitis with nasal Polyps through MicroRNA-21. Int
Arch Allergy Immunol. 2019;179(4):304–19.
19. Kim J, Hegener K, Hagedorn C, Jamal Jameel K, Weidinger D, Seuthe
IMC, et al. Long-term cryopreservation of nasal Polyp tissue in a biobank
for the isolation and culture of primary epithelial cells. Int J Mol Sci.
2023;24(7):6383.
20. Xu M, Chen D, Zhou H, Zhang W, Xu J, Chen L. The role of periostin in theoccurrence and progression of eosinophilic chronic sinusitis with nasal
Polyps. Sci Rep. 2017;7(1):9479.
21. Jamal Jameel K, Gallert WJ, Yanik SD, Panek S, Kronsbein J, Jungck D, et al.
Biomarkers for comorbidities modulate the activity of T-cells in COPD. Int
J Mol Sci. 2021;22(13):7187.

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