Summary of Study ST001734

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001109. The data can be accessed directly via it's Project DOI: 10.21228/M8BX1P This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST001734
Study Title	Understanding systemic and local inflammation induced by nasal polyposis: role of the allergic phenotype (part-II)
Study Summary	Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by persistent symptoms associated to the development of nasal polyps. To this day, the molecular mechanisms involved are still not well defined. However, it has been suggested that a sustained inflammation as allergy is involved in its onset. In this pilot study, we aimed to look into the effect of the allergic status of the patient and in their underlying mechanisms. To achieve this, we recruited 22 patients with CRSwNP and classified them in non-allergic and allergic using ImmunoCAP ISAC molecular diagnosis. Plasma samples were analyzed using liquid chromatography coupled to mass spectrometry (LC-MS). Subsequently, the identified changed metabolites from plasma that were commercially available were then analyzed by targeted analysis in some nasal polyps. Additionally, nasal polyp and mucosa tissue samples were examined for eosinophils and neutrophils. We found that 9 out of the 22 patients were sensitized to some aeroallergens (named as allergic). The other 13 patients had no sensitizations (non-allergic). Regarding metabolomics, we found that bilirubin, cortisol, lysophosphatidylcholines (LPCs) 16:0, 18:0 and 20:4 and lysophosphatidylinositol (LPI) 20:4, metabolites that are usually related to a sustained allergic inflammation, were unexpectedly increased in the plasma of non-allergic patients with CRSwNP compared to allergic patients with CRSwNP. LPC 16:0, LPC 18:0 and LPI 20:4 metabolites followed the same trend in the nasal polyp as they did in plasma. Comparison of nasal polyps with nasal mucosa tissue showed a significant increase in eosinophils (p < 0.001) and neutrophils (p < 0.01) in allergic patients with CRSwNP. There were also more eosinophils in the polyps of non-allergic patients with CRSwNP than in their nasal mucosa (p <0.01). The polyps from non-allergic patients with CRSwNP had less eosinophils than the polyps of allergic patients with CRSwNP (p < 0.05). Our data suggests that there is a systemic inflammatory response associated to CRSwNP in the absence of allergy, which could be accountable for the nasal polyp development. Allergic patients with CRSwNP presented a higher number of eosinophils located in nasal polyps suggesting that eosinophilia might be connected to the development of nasal polyps in these patients.
Institute	CEMBIO
Last Name	Delgado Dolset
First Name	María Isabel
Address	Urb. Montepríncipe s/n, Ctra. Boadilla del Monte km 5.3, Madrid, Madrid, 28668, Spain
Email	maria.delgadodolset@beca.ceu.es
Phone	+34 913724700 4665
Submit Date	2021-03-17
Num Groups	2
Total Subjects	22
Raw Data Available	Yes
Raw Data File Type(s)	d
Analysis Type Detail	LC-MS
Release Date	2021-06-17
Release Version	1

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Project:

Project ID:	PR001109
Project DOI:	doi: 10.21228/M8BX1P
Project Title:	LC-MS Nasal Polyp analysis
Project Summary:	Analysis of human samples from patients with nasal polyps with and without allergy.
Institute:	CEMBIO
Last Name:	Delgado Dolset
First Name:	María Isabel
Address:	Urb. Montepríncipe s/n, Ctra. Boadilla del Monte km 5.3, Madrid, Madrid, 28668, Spain
Email:	maria.delgadodolset@beca.ceu.es
Phone:	+34 913724700 4665

Subject:

Subject ID:	SU001811
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Factors:

Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id	local_sample_id	group
SA162664	POL-36	allergic
SA162665	POL-19	allergic
SA162666	POL-15	allergic
SA162667	POL-06	non-allergic
SA162668	POL-14	non-allergic
SA162669	POL-04	non-allergic

Showing results 1 to 6 of 6

Showing results 1 to 6 of 6

Collection:

Collection ID:	CO001804
Collection Summary:	During endoscopic surgical procedures to remove the polyp, 5 mm biopsies of nasal polyp were obtained and kept in RNA later.
Sample Type:	Nasal Polyp tissue
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR001824
Treatment Summary:	Nasal polyp samples were kept in RNA later and stored at -80ºC until preparation

Sample Preparation:

Sampleprep ID:	SP001817
Sampleprep Summary:	RNAlater solvent was removed by washing the tissue 3 times with PBS 1X. Then, the polyp was frozen in liquid nitrogen for 30 seconds. The frozen sample was put in cryoPREPTMCP02 (Covaris, MA, United States) plastic bags and submerged again for 30 seconds in liquid nitrogen. Once the plastic bag was inside the automated cryoPREPTMCP02, two consecutive impact forces of levels 2 and 4 out of 6 were applied. The resultant powder was gathered and weighted. Then, 100 µL of cold methanol:ethanol (1:1, v/v) and 0.5 µL of internal standard (LPC 18:1-d7; 0.01mM) were added per each 10 mg of tissue for metabolite extraction and protein precipitation. Samples were then vortex-mixed and homogenized using Tissue-Lyser LT homogenizer (Qiagen, Germany) for 5 min at 50 Hz, 3 times. Supernatant containing the metabolites was separated from the pellet by centrifugation (2,000 rcf for 10 min at 4 °C). Then, an aliquot of 70 µL was transferred to an LC vial and diluted with 490 µL of mobile phase (5% water: 95% acetonitrile; both with 7.5 mM ammonium acetate and 0.1% acetic acid). All samples were randomized before metabolite extraction and for the corresponding analytical run.

Sampleprep ID:

SP001817

Sampleprep Summary:

RNAlater solvent was removed by washing the tissue 3 times with PBS 1X. Then, the polyp was frozen in liquid nitrogen for 30 seconds. The frozen sample was put in cryoPREPTMCP02 (Covaris, MA, United States) plastic bags and submerged again for 30 seconds in liquid nitrogen. Once the plastic bag was inside the automated cryoPREPTMCP02, two consecutive impact forces of levels 2 and 4 out of 6 were applied. The resultant powder was gathered and weighted. Then, 100 µL of cold methanol:ethanol (1:1, v/v) and 0.5 µL of internal standard (LPC 18:1-d7; 0.01mM) were added per each 10 mg of tissue for metabolite extraction and protein precipitation. Samples were then vortex-mixed and homogenized using Tissue-Lyser LT homogenizer (Qiagen, Germany) for 5 min at 50 Hz, 3 times. Supernatant containing the metabolites was separated from the pellet by centrifugation (2,000 rcf for 10 min at 4 °C). Then, an aliquot of 70 µL was transferred to an LC vial and diluted with 490 µL of mobile phase (5% water: 95% acetonitrile; both with 7.5 mM ammonium acetate and 0.1% acetic acid). All samples were randomized before metabolite extraction and for the corresponding analytical run.

Combined analysis:

Analysis ID	AN002823
Analysis type	MS
Chromatography type	HILIC
Chromatography system	Agilent 1260
Column	Phenomenex Kinetex HILIC 100A (50 x 2.1 mm,2.6um)
MS Type	ESI
MS instrument type	Triple quadrupole
MS instrument name	Agilent 6470 QQQ
Ion Mode	NEGATIVE
Units	ug/mg

Chromatography:

Chromatography ID:	CH002087
Chromatography Summary:	We used HPLC system (1260 Infinity, Agilent Technologies). Metabolites were separated on a Kinetex hydrophilic interaction liquid chromatography (HILIC) silica column (150 mm × 2.1 mm, particle size 100Å, Phenomenex, USA) maintained at 25 ºC. The mobile phases consisted of A) water, and B) acetonitrile, both with 7.5 mM ammonium acetate and 0.1% acetic acid, with a final pH of 4.0 in the aqueous phase. The flow rate was 0.5 mL/min with an injection volume of 5 µl. Gradient started with 5% of A for 2 min, then increased up to 50% until 12 min, and back to initial conditions until 22 min.
Instrument Name:	Agilent 1260
Column Name:	Phenomenex Kinetex HILIC 100A (50 x 2.1 mm,2.6um)
Column Temperature:	25ºC
Flow Rate:	0.5 mL/min
Solvent A:	100% water; 0.1% acetic acid; 7.5 mM ammonium acetate, pH 4
Solvent B:	100% acetonitrile; 0.1% acetic acid; 7.5 mM ammonium acetate
Chromatography Type:	HILIC

MS:

MS ID:	MS002617
Analysis ID:	AN002823
Instrument Name:	Agilent 6470 QQQ
Instrument Type:	Triple quadrupole
MS Type:	ESI
MS Comments:	The MS conditions were: 5500 V of capillary voltage in positive ESI mode, a nebulizer gas flow rate of 11.0 L/min; a source temperature of 250 °C; and a source pressure of 60 psi. The sample tray temperature was maintained at 4 °C. Each transition was optimized adjusting the fragmentor and collision energy voltages. Data were acquired in MassHunter Workstation B.05.00 (Agilent Technologies), and re-processed using MassHunter QQQ Quantitative Analysis B.08.00 (Agilent) where peak areas were integrated. Concentration of metabolites were calculated using calibration curves with the standard addition method.
Ion Mode:	NEGATIVE