DOI: 10.5061/dryad.d7wm37qb3
title: Metabolomics data from: Dietary caloric input and tumor growth accelerate senescence and modulate liver and adipose tissue crosstalk

description: Metabolic alterations are related to tumorigenesis and other age-related
 diseases that are accelerated by “Westernized” diets. In fact,
 hypercaloric nutrition is associated with the increased incidence of
 cancers and faster aging. Conversely, lifespan-extending strategies, such
 as caloric restriction, impose beneficial effects on both processes. Here,
 we investigated the metabolic consequences of hypercaloric-induced aging
 on tumor growth in female mice. Our findings indicate that a high-fat
 high-sucrose diet increases tumor growth mainly due to the boosted
 oxidation of glucose and fatty acids. Consequently, through an increased
 expression of lactate, IGFBP3, and PTHLH, tumors modulate liver and white
 adipose tissue metabolism. In the liver, the induced tumor increases
 fibrosis and accelerates the senescence process, despite the lower
 systemic pro-inflammatory state. Importantly, the induced tumor induces
 the wasting and browning of white adipose tissue, thereby reversing
 diet-induced insulin resistance. Finally, we suggest that tumor growth
 alters liver-adipose tissue crosstalk that upregulates Fgf21, induces
 senescence, and negatively modulates lipids and carbohydrates metabolism
 even in caloric restricted-fed mice.
description: <strong>Sample preparation</strong>
 In a 2 mL centrifuge tube (Eppendorf), 50 µL of serum and 100 µL
 of ice-cold methanol with internal standard (p-fluoro-DL-phenylalanine and
 furosemide, final concentration of 2.5 µg/mL) were added. The mixture was
 homogenized by vortexing for 10 s, followed by an ultrasound bath with ice
 for 10 min and homogenized again for 10 s. Subsequently, the samples were
 incubated at -20 °C for 15 min for protein precipitation, followed by
 centrifugation at 10,000 xg for 15 min. In a new tube, the supernatant
 (120 µL) was collected and dried in a Turbovap equipment in a water bath
 at 45 °C and N2 gas pressure regulated at 7 psi for approximately 30 min.
 The dried extract was reconstituted in 60 µL of injection solvent
 (methanol:water, 1:1) with internal standard (Caffeine, final
 concentration of 1 µg/mL) and vortexed for 10 s. For solubilization, the
 reconstituted samples were placed in an ultrasonic bath for 5 min and
 centrifuged again at 10,000 x g for 5 min. Finally, an aliquot (50 µL) was
 collected in a 2 mL vial with a 150 µL insert. Prior to analysis, 5 µL of
 each sample was collected and mixed in a vial to create a quality control
 (QC) solution, which was analyzed at the beginning and at periodic
 intervals throughout the injection list.
 <strong>Description of the high-resolution mass
 spectrometry system</strong> <strong>Liquid
 chromatography</strong> Thermo Dionex Ultimate
 3000 coupled to the high-resolution and mass-accuracy mass spectrometer
 Thermo QExactive Plus with electrospray ionization source operating in
 positive and negative analysis modes. General ionization source
 conditions: Sheath gas and auxiliary gas: 45 and 15 arbitrary units,
 respectively; spray voltage: + or - 3600 V; S-lens voltage: 50 V;
 capillary temperature: 300 °C; source temperature: 400 °C. Chromatographic
 conditions: Separation was achieved using a Thermo Scientific® Hypersil
 Gold C18 column (50 x 2.1 mm x 1.9 μm particle diameter) maintained at 40
 °C at a flow rate of 0.350 mL/min of mobile phase and injection of 5 μL of
 sample. The mobile phases (FM) were: (FM A) ultrapure water with 0.1%
 formic acid and 5 mM ammonium formate, and (FM B) methanol with 0.1%
 formic acid. The chromatographic separation was performed using the
 gradient elution mode: 0-1.0 min 15% B; 1.0-10.0 min 100% B; 10.0-15.0 min
 100% B; 15.0-15.1 min 15% B; 15.1-20.0 min 15% B.
 <strong>Mass spectrometry
 experiments</strong> Data acquisitions were
 performed in full scan mode in the m/z range 100-1000 using a resolution
 of 35,000 FWHM (Full Width at Half Maximum), AGC 1e6 and IT 100ms,
 combined with the data dependent acquisition (DDA, from English Data
 Dependent Analysis; ddMS2 TOP 3) experiment using a resolution of 17,500
 FWHW, AGC 1e5, IT 50 ms; NCE 15-35% and isolation window of 1.2
 Da. <strong>Data
 processing</strong> The processing of the raw
 .RAW files obtained from the analyses by Liquid Chromatography coupled to
 High Resolution Mass Spectrometry (LC-EMAR) was performed in the MS-DIAL
 software (RIKEN, version 4.9) for the extraction of the detected signals,
 spectral deconvolution and peak alignments. The parameters used in the
 processing were: tolerance for MS1 of 0.005 Da and MS2 of 0.05 Da; minimum
 peak height of 1.0E6; mass width of 0.05 Da; sigma value of the
 deconvolution window of 0.5; and tolerance for alignment of 0.3 min and
 0.005 Da. A sample blank (extraction solvent) was used to subtract
 interfering signals. For the putative annotation of the main metabolites,
 the experimental MS/MS spectra were compared with data from a laboratory
 spectral library that combined the Mass Bank of North America public
 library and the commercial NIST MSMS 2020, considering a similarity level
 between the spectra greater than 80%. An error of less than 8 ppm was also
 considered for similarity between the experimental and theoretical m/z
 values. The detected ions were exported to Excel (Microsoft Office 2016)
 to obtain a data matrix table. Inconsistent data were filtered based on i)
 the coefficient of variation (CV) of the signal areas detected in the
 quality controls (pooled QC, a mixture of samples and analyzed at periodic
 intervals throughout the injection list), where variables with CV &gt;
 30% were removed; and ii) by evaluating the chromatographic peaks
 (Gaussian profile) obtained in all samples.
description: # Metabolomics data from: Dietary caloric input and tumor growth
 accelerate senescence and modulate liver and adipose tissue crosstalk
 [https://doi.org/10.5061/dryad.d7wm37qb3](https://doi.org/10.5061/dryad.d7wm37qb3) #### Description of the data and file structure In the attempt to evaluate whether the different treatments were altering circulating metabolites. For this, we prepared a pool of the sera from the different groups and proceeded with a metabolomic analysis of these samples. Sample preparation:  In a 2 mL centrifuge tube (Eppendorf), 50 µL of serum and 100 µL of ice-cold methanol with internal standard (p-fluoro-DL-phenylalanine and furosemide, final concentration of 2.5 µg/mL) were added. The mixture was homogenized by vortexing for 10 s, followed by an ultrasound bath with ice for 10 min and homogenized again for 10 s. Subsequently, the samples were incubated at -20 °C for 15 min for protein precipitation, followed by centrifugation at 10,000 xg for 15 min. In a new tube, the supernatant (120 µL) was collected and dried in a Turbovap equipment in a water bath at 45 °C and N2 gas pressure regulated at 7 psi for approximately 30 min. The dried extract was reconstituted in 60 µL of injection solvent (methanol:water, 1:1) with internal standard (Caffeine, final concentration of 1 µg/mL) and vortexed for 10 s. For solubilization, the reconstituted samples were placed in an ultrasonic bath for 5 min and centrifuged again at 10,000 x g for 5 min. Finally, an aliquot (50 µL) was collected in a 2 mL vial with a 150 µL insert. Prior to analysis, 5 µL of each sample was collected and mixed in a vial to create a quality control (QC) solution, which was analyzed at the beginning and at periodic intervals throughout the injection list. The processing of the raw .RAW files obtained from the analyses by Liquid Chromatography coupled to High Resolution Mass Spectrometry (LC-EMAR) was performed in the MS-DIAL software (RIKEN, version 4.9) for the extraction of the detected signals, spectral deconvolution and peak alignments. The parameters used in the processing were: tolerance for MS1 of 0.005 Da and MS2 of 0.05 Da; minimum peak height of 1.0E6; mass width of 0.05 Da; sigma value of the deconvolution window of 0.5; and tolerance for alignment of 0.3 min and 0.005 Da. A sample blank (extraction solvent) was used to subtract interfering signals. For the putative annotation of the main metabolites, the experimental MS/MS spectra were compared with data from a laboratory spectral library that combined the Mass Bank of North America public library and the commercial NIST MSMS 2020, considering a similarity level between the spectra greater than 80%. An error of less than 8 ppm was also considered for similarity between the experimental and theoretical m/z values. The detected ions were exported to Excel (Microsoft Office 2016) to obtain a data matrix table. Inconsistent data were filtered based on i) the coefficient of variation (CV) of the signal areas detected in the quality controls (pooled QC, a mixture of samples and analyzed at periodic intervals throughout the injection list), where variables with CV &gt; 30% were removed; and ii) by evaluating the chromatographic peaks (Gaussian profile) obtained in all samples. ### Files and variables #### File: CHOW\_TUMOR.raw **Description:** Metabolomics raw data from a pool of sera from eight 12-weeks old C57BL/6J female mice fed a standard diet* ad libitum* for 24 weeks, where before the beginning of the 23rd week received an subcutaneous injection of 10,000 B16F10 cells to develop a cancer.  #### File: CHOW\.raw **Description:** Metabolomics raw data from a pool of sera from eight 12-weeks old C57BL/6J female mice fed a standard diet *ad libitum* for 24 weeks. #### File: CR\_TUMOR.raw **Description:** Metabolomics raw data from a pool of sera from four 12-weeks old C57BL/6J female mice submitted to 30% caloric restriction of a standard diet for 24 weeks, where before the beginning of the 23rd week received an subcutaneous injection of 10,000 B16F10 cells to develop a cancer.  #### File: CR.raw **Description:** Metabolomics raw data from a pool of sera from five 12-weeks old C57BL/6J female mice submitted to 30% caloric restriction of a standard diet for 24 weeks. #### File: HFHS.raw **Description:** Metabolomics raw data from a pool of sera from four 12-weeks old C57BL/6J female mice fed a High-fat, High-sucrose diet* ad libitum* for 24 weeks, where before the beginning of the 23rd week received an subcutaneous injection of 10,000 B16F10 cells to develop a cancer.  #### File: HFHS\_TUMOR.raw **Description:** Metabolomics raw data from a pool of sera from six 12-weeks old C57BL/6J female mice fed a High-fat, High-sucrose diet* ad libitum* for 24 weeks #### File: QC\_TOTAL\_5.raw **Description:** QUALITY CONTROL of the HFHS_TUMOR.raw #### File: QC\_TOTAL1.raw **Description:** QUALITY CONTROL of the CHOW.raw #### File: QC\_TOTAL2.raw **Description:** QUALITY CONTROL of the CR.raw #### File: QC\_TOTAL3.raw **Description:** QUALITY CONTROL of the HFHS.raw #### File: QC\_TOTAL4.raw **Description:** QUALITY CONTROL of the CR_TUMOR.raw ## Code/software Any spreadsheet reader can access data.