Dataset Information

Qualitative and label-free quantitative analysis of protein changes in the lymphatic muscle cells isolated in a hind-limb lymphatic injury model in the sheep

ABSTRACT: We developed a large animal model, with clinically relevant gravitational loading on the lymphatic vasculature, that enables longitudinal tracking of lymphatic function in vivo as well as end-point analysis of vessel function and mechanics. One of two parallel lymphatic vessels in the sheep hind limb was ligated with the contralateral limb acting as an internal control. To determine the changes in the protein expression profiles in the lymphatic muscle cells from the remodeled vessel compared to the control, we expanded isolated LMC in vitro and performed bottom-up proteomic analysis using a Q Exactive Plus mass spectrometer. Label-free quantitative (LFQ) methods directly used the raw spectral data from parallel MS runs to determine relative protein abundances. We used both “MS/MS (MS2) spectral counting” and “precursor MS1 area” methods for label free quantitation and further contrasting the differentially expressed proteomic profiles across the control and remodeled lymphatic muscle samples to determine the alterations in mitochondrial dysfunction and elevated oxidative stress within the lymphatic muscle.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Ovis Aries

TISSUE(S): Lymphatic Endothelium, Smooth Muscle Tissue

DISEASE(S): Disease Free

SUBMITTER: Cristina Clement

LAB HEAD: Brandon Dixon PhD

PROVIDER: PXD014343 | Pride | 2021-01-12

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	2016_08_22E_P068_SA R2.pride.mgf.gz	Mgf
	2016_08_22E_P068_SA.mgf	Mgf
	2016_08_22E_P068_SA.mzxml	Mzxml
	2016_08_22E_P068_SA.pride.mgf.gz	Mgf
	2016_08_22E_P068_SA.raw	Raw

Items per page:

1 - 5 of 240

Publications

Lymphatic remodelling in response to lymphatic injury in the hind limbs of sheep.

Nelson Tyler S TS Nepiyushchikh Zhanna Z Hooks Joshua S T JST Razavi Mohammad S MS Lewis Tristan T Clement Cristina C CC Thoresen Merrilee M Cribb Matthew T MT Ross Mindy K MK Gleason Rudolph L RL Santambrogio Laura L Peroni John F JF Dixon J Brandon JB

Nature biomedical engineering 20191223 6

Contractile activity in the lymphatic vasculature is essential for maintaining fluid balance within organs and tissues. However, the mechanisms by which collecting lymphatics adapt to changes in fluid load and how these adaptations influence lymphatic contractile activity are unknown. Here we report a model of lymphatic injury based on the ligation of one of two parallel lymphatic vessels in the hind limb of sheep and the evaluation of structural and functional changes in the intact, remodelling ...[more]

PMID: 31873209

Similar Datasets

Project description:We have previously reported that the deficiency of p53 alone or in combination with Rb (Rb-/- p53-/-) in adipose-derived MSCs (ASCs) promotes leiomyosarcoma-like tumors in vivo. Here, we hypothesized that the source of MSCs and/or the cell differentiation stage could determine the phenotype of sarcoma development. To investigate whether there is a link between the source of MSCs and sarcoma phenotype, we generated p53-/- and Rb-/-p53-/- MSCs from bone marrow (BM-MSCs). Both genotypes of BM-MSCs initiated leiomyosarcoma formation similar to p53-/- and Rb-/-p53-/- ASCs. In addition, gene expression profiling revealed a link between p53- or Rb-p53-deficient BM-MSCs and ASCs and muscle-associated sarcomagenesis. These data suggest that the tissue source of MSC does not seem a crucial factor in the development of a particular sarcoma phenotype. To analyze whether the differentiation stage defines the sarcoma phenotype, BM-MSCs and ASCs were induced to differentiate towards the osteogenic lineage, and both p53 and Rb were excised using Cre-expressing adenovectors at different stages along osteogenic differentiation. Regardless of the level of osteogenic commitment, the inactivation of Rb and p53 in BM-MSC-derived, but not in ASC-derived, osteogenic progenitors gave rise to osteosarcoma-like tumors which could be serially transplanted. This indicates that the osteogenic differentiation stage of BM-MSCs imposes the phenotype of in vivo sarcoma development, and that BM-MSC-derived osteogenic progenitors rather than undifferentiated BM-MSCs, undifferentiated ASCs or ASC-derived osteogenic progenitors, represent the cell of origin for osteosarcoma development. BM-MSC and ASC cultures were established from 3 mouse strains: (a) WT, (b) p53loxP/loxP and (c) p53loxP/loxP RbloxP/loxP. The corresponding mutant cells were generated by excision of the LoxP-flanked sequences by infection of all MSC cultures with adenoviral vectors expressing the Cre-recombinase gene (Ad-CMV-Cre). NSG mice were inoculated subcutaneously with 5×10^6 mutant cells. Animals were killed when tumors reached 1 cm3 or 150 days after infusion. Some of the obtained tumors were mechanically disaggregated to establish ex vivo MSC-transformed cell lines. Gene expression analysis was performed using BM-MSCs and ASCs from all genotypes, as well as tumor cell lines derived from p53-/- and p53-/-Rb-/- BM-MSC and ASC cultures.