Resistin

Manufactured by Merck Group

Sourced in United States, United Kingdom

Resistin is a laboratory equipment product designed to measure the concentration of resistin, a protein involved in insulin resistance. Resistin is a hormone secreted by adipose tissue and plays a role in glucose and lipid metabolism. The product provides a tool for researchers and clinicians to quantify resistin levels, which may be useful in the study of metabolic disorders such as type 2 diabetes.

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14 protocols using resistin

Multiplex Plasma Immune Profiling

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Immune mediators in plasma were measured using a customized multiplex human factor panel (R & D Systems, Minneapolis MN) measuring cytokines (IFNβ, IFNγ, IL-1β, IL-10, IL-12p70, IL-13, IL-15, IL-17A, IL-18, IL-1RA, IL-2, IL-21, IL-4, IL-5, IL-7, TNFα, IL-23, IL-31, IL-22, IL-27), chemokines (CCL2/MCP-1, CCL3/MIP-1α, CCL4/MIP-1β, CCL5/RANTES, CCL11/Eotaxin, CXCL1/GROα, CXCL8/IL-8, CXCL9/MIG, CXCL10/IP-10, CXCL11/I-TAC, CXCL12/SDF-1α, CXCL13/BCA-1), growth factors (BDNF, GM-CSF, HGF, EGF, VEGF, PDGF-BB) and additional molecules (PD-L1, S100). Metabolic hormones were measured using a 3-plex kit measuring insulin, leptin, and PYY (Millipore, Burlington MA). Adipokines were assayed using a 5-plex kit measuring adiponectin, adipsin, lipocalin-2, total PAI-1, and resistin (Millipore, Burlington MA). CRP and IL-6 were measured in UCB plasma using a high-sensitivity ELISA (Life Technologies, Carlsbad CA) per the manufacturer’s instructions.
Supernatants from fetal rhesus macaque monocyte stimulation experiments were analyzed using an NHP XL Cytokine Premixed 36-plex kit (Bio-Techne, Minneapolis MN). Samples were diluted per the manufacturer’s instructions and analyzed in duplicate on the Magpix Instrument (Luminex, Austin, TX). Data were fit using a 5P-logistic regression on xPONENT software.

Sureshchandra S., Doratt B.M., Mendza N., Varlamov O., Rincon M., Marshall N.E, & Messaoudi I. (2023). Maternal obesity blunts antimicrobial responses in fetal monocytes. eLife, 12, e81320.

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Mouse Serum Metabolic Profiling

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Mouse serum samples were harvested by tail bleeding after overnight fast (∼16 h), or at the indicated time point. Serum samples were separated using microvette® CB 300 (Sarstedt). Glucose concentration was determined at the time of collection with a glucometer (BD bioscience). Serum triglycerides, non-esterified free fatty acids (NEFA), cholesterol, high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol were measured at the Phenotyping Core at the Johns Hopkins University School of Medicine. Serum insulin, leptin, resistin, adiponectin, PAI-1, and TNF-α (Millipore) were measured according to kit manufacturer’s instructions.

Peterson J.M., Seldin M.M., Tan S.Y, & Wong G.W. (2014). CTRP2 Overexpression Improves Insulin and Lipid Tolerance in Diet-Induced Obese Mice. PLoS ONE, 9(2), e88535.

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Biochemical and Hormonal Profiling of Plasma

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Plasma amylase and lipase were determined by a kinetic method using commercial kits for amylase (Labtest, Minas Gerais, Brazil) and lipase (Bioclin, Minas Gerais, Brazil). The assays were performed according to the manufacturer's instructions, and their levels were expressed in units per liter. Plasma glucose, triglycerides, and total cholesterol were analyzed using commercial kits (Labtest, Minas Gerais, Brazil), and the levels were expressed in milligrams per deciliter. Plasma alanine amino transferase (ALT) and aspartate amino transferase (AST) activities expressed in units per liter were analyzed by a kinetic method using commercial kits (Labtest, Minas Gerais, Brazil). The hepatic lipids were extracted using the Folch method [17 (link)], and triglycerides and total cholesterol concentrations were determined using commercial kits (Labtest, Minas Gerais, Brazil) and expressed in milligrams per gram (mg/g). Plasma insulin, leptin, ghrelin, resistin (Millipore, Billerica, MA, USA), TNF-α, IL-6, and MCP-1 (R&D Systems, Minneapolis, MN, USA) were measured by enzyme-linked immunosorbent assay (ELISA) performed in duplicate and expressed in nanograms or picograms per milliliter.

Carvalho K.M., Marinho Filho J.D., de Melo T.S., Araújo A.J., Quetz J.D., da Cunha M.D., de Melo K.M., da Silva A.A., Tomé A.R., Havt A., Fonseca S.G., Brito G.A., Chaves M.H., Rao V.S, & Santos F.A. (2015). The Resin from Protium heptaphyllum Prevents High-Fat Diet-Induced Obesity in Mice: Scientific Evidence and Potential Mechanisms. Evidence-based Complementary and Alternative Medicine : eCAM, 2015, 106157.

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Multiplex Quantification of Plasma Proteins

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Cytokines, chemokines, and growth factors in undiluted cell culture supernatants and diluted UCB plasma (1:1) were measured using a human 37-plex luminex panel R & D Systems, Minneapolis MN). Metabolic hormones were measured using a 3-plex kit measuring insulin, leptin, and PYY (Millipore, Burlington MA). Adipokines were assayed using a 5-plex kit measuring adiponectin, adipsin, lipocalin-2, total PAI-1, and resistin (Millipore, Burlington MA). Samples were run in duplicates on the Magpix instrument (Luminex, Austin TX). Data were fit using a 5P-logistic regression on the xPONENT software. Values below the limit of detection were designated as half of the lowest limit. Data in pg/mL were tested for normality using Shapiro-Wilk test. Statistical differences in plasma proteins were tested using unpaired t-test with welch’s correction. Differences in protein in supernatants were tested using ordinary one-way ANOVA followed by Holm-Sidak’s multiple comparison tests. All statistical tests were performed on Prism 8 (GraphPad, San Diego, CA).

Sureshchandra S., Mendoza N., Jankeel A., Wilson R.M., Marshall N.E, & Messaoudi I. (2021). Phenotypic and Epigenetic Adaptations of Cord Blood CD4+ T Cells to Maternal Obesity. Frontiers in Immunology, 12, 617592.

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Adipokine Measurements in SWAN Cohort

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Assays were run on stored serum samples collected from the SWAN follow-up visit 6, corresponding to the MPS follow-up (2002–2003). At collection, a 12-hour fasting blood draw was performed within days 2–5 of the menstrual cycle if a woman was still menstruating. For women not regularly menstruating or if a blood sample was not obtainable in the day 2–5 window, a random fasting blood draw was obtained. For women who were postmenopausal, a blood sample was drawn at their clinic visit, scheduled to be on the anniversary of their previous visit. leptin, sOB-R, adiponectin and HMW adiponectin were determined at the University of Michigan in duplicate, using commercially available colorimetric enzyme immunoassay kits according to the manufacturer’s instructions (adiponectin, HMW adiponectin, resistin, and leptin, Millipore, St. Charles, MO and soluble leptin receptor and MCP-1, R& D systems, Minneapolis, MN). The mean coefficient of variation percent for duplicate samples for each subject and lower limit of detection, respectively, were: adiponectin: 4%, 0.78 ng/mL; HMW adiponectin: 8.1%, 0.5 ng/ml; resistin: 5%, 0.16 ng/mL; leptin: 4%, 0.5 ng/mL; MCP-1: 1.7%, 31.2 pg/mL, and soluble leptin receptor: 3.7%, 0.31 ng/ml.

Ding N., Karvonen-Gutierrez C.A., Herman W.H., Calafat A.M., Mukherjee B, & Park S.K. (2021). Associations of Perfluoroalkyl and Polyfluoroalkyl Substances (PFAS) and PFAS Mixtures with Adipokines in Midlife Women. International journal of hygiene and environmental health, 235, 113777.

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Multiplex Profiling of Immune Mediators

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Immune mediators in plasma were measured using a customized multiplex human factor panel (R & D Systems, Minneapolis MN) measuring cytokines (IFNβ, IFNγ, IL-1β, IL-10, IL-12p70, IL-13, IL-15, IL-17A, IL-18, IL-1RA, IL-2, IL-21, IL-4, IL-5, IL-7, TNFα, IL-23, IL-31, IL-22, IL-27), chemokines (CCL2/MCP-1, CCL3/MIP-1α, CCL4/MIP-1β, CCL5/RANTES, CCL11/Eotaxin, CXCL1/GROα, CXCL8/IL-8, CXCL9/MIG CXCL10/IP-10, CXCL11/I-TAC, CXCL12/SDF-1α, CXCL13/BCA-1, growth factors (BDNF, GM-CSF, HGF, EGF, VEGF, PDGF-BB) and additional molecules (PD-L1, S100). Metabolic hormones were measured using a 3-plex kit measuring insulin, leptin, and PYY (Millipore, Burlington, MA). Adipokines were assayed using a 5-plex kit measuring adiponectin, adipsin, lipocalin-2, total PAI-1, and resistin (Millipore, Burlington, MA). Samples were diluted per the manufacturer’s instructions and run in duplicates on the Magpix Instrument (Luminex, Austin, TX). Data were fit using a 5P-logistic regression on xPONENT software. Group differences were compared using one-way ANOVA for unpaired samples followed by Holm-Sidak multiple test correction.

Sureshchandra S., Marshall N.E., Mendoza N., Jankeel A., Zulu M.Z, & Messaoudi I. (2021). Functional and genomic adaptations of blood monocytes to pregravid obesity during pregnancy. iScience, 24(6), 102690.

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Fractionation and Western Blot Analysis

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Cardiomyocytes and isolated heart tissues were homogenized in lysis buffer (Cell Signaling Tech.) containing protease and phosphatase inhibitors. Fractionated cytoplasmic and nuclear protein lysates (20–40 μg) were separated and applied to SDS-PAGE and transferred onto PVDF membrane (BioRad). Antibodies used were phospho- or total against AMPKα, phospho-AMPKα-T172, (Cell Signaling Technology), histone 3 (H3) (Genetex), NFATc and β-actin (Santa Cruz Biotechnology) and resistin (Millipore). Serca2a antibody is custom made in our lab). β-actin expression verified cytosolic protein loading while H3 served as nuclear specific internal control.

Singh R., Moreno P., Hajjar R.J, & Lebeche D. (2018). A role for calcium in resistin transcriptional activation in diabetic hearts. Scientific Reports, 8, 15633.

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Resistin and Adiponectin Effects on Insulin Secretion

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All chemicals were purchased from Sigma-Aldrich Ireland unless otherwise stated. Culture media and its related components were purchased from Gibco (Glasgow, UK). The BRIN-BD11 cell line was used in this study [24 (link)] and was maintained as previously described [25 (link)].
For experimental treatments, cells were seeded at a density of 1.5 x 10⁵ cells per well in a 24 well plate for insulin secretion assays. Cells were allowed to attach for 24 h before being treated with recombinant rat resistin (Cambridge Biosciences, Cambridge, UK) or rat GACRP30/Adiponectin (Sigma-Aldrich) or ratios of both, for 24 h. Concentrations of 10–20ng ml^-1 of resistin and 5–20nmol l^-1 of globular (g) adiponectin were used. Concentrations were chosen in accordance with previous studies [26 (link), 27 (link)]. Cells between passage 23–33 were used and all experiments were n = 4 unless otherwise stated.

Curran A.M., Ryan M.F., Drummond E., Gibney E.R., Gibney M.J., Roche H.M, & Brennan L. (2016). Uncovering Factors Related to Pancreatic Beta-Cell Function. PLoS ONE, 11(8), e0161350.

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Adipocyte Inflammation Response Assay

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After 15 days induction of differentiation, >85% of the adipocytes exhibited lipid droplet accumulation. Following serum starvation for 12 h, the human adipocytes were treated with a 1 mM FFA cocktail and adipokines, including 10 ng/ml TNF-α (Sigma-Aldrich), 30 ng/ml IL-6 (Sigma-Aldrich), 100 ng/ml leptin (Sigma-Aldrich) and 60 ng/ml resistin (Sigma-Aldrich) for different durations (4, 8 and 24 h) (24 (link)–26 (link)), then cells were treated with TRIzol (Invitrogen; Thermo Fisher Scientific, Inc., Waltham, MA, USA) for the following experiment.

Gu N., You L., Shi C., Yang L., Pang L., Cui X., Ji C., Zheng W, & Guo X. (2016). Expression of miR-199a-3p in human adipocytes is regulated by free fatty acids and adipokines. Molecular Medicine Reports, 14(2), 1180-1186.

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Resistin-induced vascular effects

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For wire myography experiments carried out to assess the effect of resistin, vessels were mounted on the small vessel wire myograph followed by incubation with 40 ng/ml recombinant rat resistin (Sigma-Aldrich, Dorset, UK) for 4 hours at 37 °C. Following incubation, vessels were subject to dose-response curves as described in “Wire myography”.

Small H.Y., McNeilly S., Mary S., Sheikh A.M, & Delles C. (2019). Resistin Mediates Sex-Dependent Effects of Perivascular Adipose Tissue on Vascular Function in the Shrsp. Scientific Reports, 9, 6897.

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