DOI: 10.5061/dryad.866t1g22t
title: Data from: Adipose tissue explant culture using PDMS flow chambers: an alternative to static explant culture

description: As obesity continues to rise, it is important we can effectively study
 adipose tissue. However, adipose tissue is notoriously difficult to
 culture ex vivo due to fragility, bouyancy, and high nutrient demant. To
 address these challenges, we developed a novel approach using
 polydimethylsiloxane (PDMS) flow chambers attached to a micro peristaltic
 pump for culture of primary adipose tissue explants. Our results
 demonstrate that adipose tissue remains viable and functional in flow
 chambers, therefore providing an alternative to static explant culture in
 a dish. 
description: # Data from: Adipose tissue explant culture using PDMS flow chambers: an
 alternative to static explant culture Dataset DOI:
 [10.5061/dryad.866t1g22t](10.5061/dryad.866t1g22t) ## Description of the
 data and file structure All experiments were performed at UCLA in Los
 Angeles, CA. Flow chambers for culture of primary adipose tissue explants
 were designed in house and fabricated using polydimethylsiloxane (PDMS)
 and 0.4 micron polyethylene terephthalate (PET) membranes. Once produced,
 chambers were placed in 37˚C tissue culture incubators and attached to a
 mini peristaltic pump to allow for media perfusion. Tissue explants in
 flow chambers were assessed for viability using resazurin assays and by
 measurement of lactate dehydrogenase in media effluent. Explants were then
 treated with isoproterenol and insulin to test for induction and
 inhibition of lipolysis, respectively, by colorimetric glycerol assay of
 media effluent. Finally, explants were collected after incubation in
 chambers and assessed for maintenance of adipocyte structure using
 confocal microscopy, and maintenance of adipose gene expression by
 RT-qPCR. Images and gene expression data were also obtained from static
 cultured explants in a dish for comparison.  ### Files and variables ####
 File: Cohen_et_al_compiled_data.xlsx **Description:** Microsoft excel file
 containing compiled data from all quantitative experiments in human and
 mouse tissues: colorimetric resazurin (alamarblue) viability assay,
 luminescent LDH assay, colorimetric glycerol release assay, and RT-qPCR
 cells with value "null" were not applicable to analysis or not
 collected at the listed time point ##### Variables (Tab 1,2) Effluent =
 reduced alamarblue chamber effluent Input = alamarblue media (1:10
 dilution in RPMI complete media) before infusion to chamber % reduction =
 percent reduced resazurin calculated using equation provided by the
 manufacturer protocol ##### Variables (Tab 3) Iso = 10µM isoproterenol
 treatment Ins = 100nM insulin treatment Time 0: media before entering
 chamber (blank; subtracted from all values) ##### Variables (Tab 4) TX-100
 = 0.2% Triton TX-100 detergent in RPMI complete media ##### Variables (Tab
 5,6,7) M199 = Media 199 with earle's salts and
 penicillin/streptomycin, serum free RPMI = RPMI media +10% FBS +1%
 Glutamax +1% sodium pyruvate + 0.1% gentamicin Pre: sample before
 processing for culture Post: 72 hours in chamber (except tab 6: 120 hours)
 Dish: 72 hours in culture dish (except tab 6: 120 hours) RQ: Relative
 quantification CT/CQ: critical threshold/quantification cycle  #### File:
 72hr_Post_Dish.jpg **Description:** Human adipose tissue imaged via
 confocal microscopy after 72 hours in culture plate. Red = Perilipin 1
 (adipocytes), Blue = DAPI (nuclei), Green = CD45 (immune cells) #### File:
 120hr_Post_Chamber.jpg **Description:** Human adipose tissue imaged via
 confocal microscopy after 120 hours in flow chamber. Purple = Perilipin 1
 (adipocytes), Blue = DAPI (nuclei), Green = CD45 (immune cells) #### File:
 216hr_Post_Chamber.jpg **Description:**  Human adipose tissue imaged via
 confocal microscopy after 216 hours in flow chamber. Purple = Perilipin 1
 (adipocytes), Blue = DAPI (nuclei), Green = CD45 (immune cells) #### File:
 72hr_Post_Chamber.jpg **Description:** Human adipose tissue imaged via
 confocal microscopy after 72 hours in flow chamber. Red = Perilipin 1
 (adipocytes), Blue = DAPI (nuclei), Green = CD45 (immune cells) #### File:
 216hr_Pre_Chamber_CD45_only_zoom.jpg **Description:** Magnified confocal
 image of CD45+ immune cells in adipose tissue before flow chamber.
 Green=CD45, Blue=DAPI (nuclei) #### File: 72hr_Pre.jpg
 **Description:** **:** Human adipose tissue imaged via confocal microscopy
 before processing for culture. Red = Perilipin 1 (adipocytes), Blue = DAPI
 (nuclei), Green = CD45 (immune cells) #### File:
 216hr_Post_Chamber_CD45_only_zoom.png **Description:** Magnified confocal
 image of CD45+ immune cells in adipose tissue after 216 hours in flow
 chamber. Green = CD45, Blue = DAPI (nuclei) #### File: 120hr_Post_Dish.jpg
 **Description:** Human adipose tissue imaged via confocal microscopy after
 120 hours in culture dish. Purple = Perilipin 1 (adipocytes), Blue = DAPI
 (nuclei), Green = CD45 (immune cells) #### File: 216hr_Post_Dish.jpg
 **Description:** Human adipose tissue imaged via confocal microscopy after
 216 hours in culture dish. Purple = Perilipin 1 (adipocytes), Blue = DAPI
 (nuclei), Green = CD45 (immune cells) #### File: 120hr_Pre.jpg
 **Description:**  Human adipose tissue imaged via confocal microscopy
 before processing for culture. Purple = Perilipin 1 (adipocytes), Blue =
 DAPI (nuclei), Green = CD45 (immune cells) #### File: 216hr_Pre.jpg
 **Description:** Human adipose tissue imaged via confocal microscopy
 before processing for culture. Purple = Perilipin 1 (adipocytes), Blue =
 DAPI (nuclei), Green = CD45 (immune cells) ## Human subjects data Human
 adipose tissue remnant samples were collected by the UCLA Translational
 Pathology Core Laboratory after obtaining written informed consent to use
 samples for research purposes which may result in publication
 (IRB-11-2504). Distribution of samples to our laboratory was de-identified
 and included no protected health information. Information provided was as
 follows: age, sex, diagnosis, tissue location, height, weight, BMI.