This file was generated on 2026-01-12 by Nikola Makdissil A GENERAL INFORMATION 1. Title of the dataset: Image data related to publication "Size-dependent plastic exposure disrupts macrophage function and tissue-specific metabolism" 2. Brief description of the research project and its aims: Plastic pollution is an emerging yet understudied environmental risk to the immune system. Once ingested, nano- and microplastic particles (MNPs) can translocate from the gut to internal organs, with macrophages serving as primary targets. Kupffer cells (KCs), the liver-resident macrophages, play a central role in immune surveillance and metabolism, yet their response to MNPs remains unclear. Here, using a chronic plastic exposure model in mice, we identify KCs as the primary hepatic reservoir for MNPs. Long-term exposure alters their transcriptional profile and impairs phagocytic function, leading to metabolic dysregulation of hepatocytes. Microplastics, but not nanoplastics, reduce KC-mediated clearance of circulating cells and bacteria. Under diet-induced obesity, microplastics exacerbates hepatic lipid accumulation, while nanoplastics impair systemic glucose metabolism. Although the blood-brain barrier limits microplastic infiltration, a small fraction of ingested nanoplastics reaches the brain, where it is taken up by microglia, the brain-resident macrophages. However, we observe no signs of neuroinflammation or behavioral deficits. These findings demonstrate that chronic MNP exposure disrupts macrophage function in a size-dependent manner, with distinct consequences for liver and systemic metabolism, while the brain remains largely protected. Understanding tissue-specific vulnerabilities to MNPs is crucial for assessing their long-term health impact. 3. Author Information A. Investigator Contact Information Name: Dr. Nikola Makdissi Institution: Life & Medical Sciences (LIMES) Institute, University of Bonn Address: Carl-Troll-Str. 31, 53113 Bonn Email: nmakdissi@uni-bonn.de ORCID: 0000-0002-9345-038X Name: Dr. Maria Francesca Viola Institution: Life & Medical Sciences (LIMES) Institute, University of Bonn Address: Carl-Troll-Str. 31, 53113 Bonn Email: mviola@uni-bonn.de ORCID: 0000-0003-0660-9506 B. Project Supervisor (Principal Investigator) Contact Information Name: Prof. Dr. Elvira Mass Institution: Life & Medical Sciences (LIMES) Institute, University of Bonn Address: Carl-Troll-Str. 31, 53113 Bonn Email: elvira.mass@uni-bonn.de ORCID: 0000-0003-2318-2356 C. In case of questions related to this dataset, please contact: Name: Prof. Dr. Elvira Mass Institution: Life & Medical Sciences (LIMES) Institute, University of Bonn Address: Carl-Troll-Str. 31, 53113 Bonn Email: elvira.mass@uni-bonn.de ORCID: 0000-0003-2318-2356 4. Date of data collection range, from 2019-10-01 to 2025-12-20: 5. Information about funding sources that supported the collection of the data: European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program(ERC-StG NanoGlia Grant Agreement No. 851257, EMBO fellowship (ATLF 873-2023),GRK1873/2. Further information regarding the funding grants is available in the publication 6. Language of the dataset: English 7. Geographic location of data collection: Bonn, Germany B DATA & FILE OVERVIEW 1. File List: “ORO_NMPingestion.zip” contains all Oil Red O (ORO) staining images acquired using a 10× objective. All images were used to quantify hepatic lipid content, which is reported as percentage of ORO-positive area (% ORO⁺ area) in the figures of the associated publication. All mice were on a C57BL/6Jrcc background. Within “ORO_NMPingestion.zip”, the following 3 folders are included 1.the folder “ChronicNMP_ORO” contains ORO-stained liver sections from mice that received PBS, nanoplastics (NP; 100 mg/kg), or microplastics (MP; 100 mg/kg) by oral gavage for 12 weeks. 2.The folder “4WeeksNMP_ORO” contains ORO-stained liver sections from mice that received PBS, NP (100 mg/kg), or MP (100 mg/kg) by oral gavage for 4 weeks. 3.The folder “ChronicNMP&HFD_ORO” contains ORO-stained liver sections from mice that received PBS, NP (100 mg/kg), or MP (100 mg/kg) by oral gavage for 12 weeks and were additionally placed on a high-fat diet (HFD) after 8 weeks of NMP exposure for a further 8 weeks. “PAS_NMPingestion.zip” contains all Periodic Acid–Schiff (PAS) staining images acquired using a 10× objective. All images were used to quantify hepatic glycogen content, which is reported as optical density (OD) values in the figures of the associated publication. All mice were on a C57BL/6Jrcc background. Within “PAS_NMPingestion.zip”,the following 4 folders are included: 1. the folders “ChronicNMP_PAS_100mgperkg”: contains PAS-stained liver sections from mice that received PBS, NP, or MP at a concentration of 100mg/kg by oral gavage for 12 weeks. 2. The folder “ChronicNMP_PAS_10mgperkg”:contains PAS-stained liver sections from mice that received PBS, NP, or MP at a concentration of 10mg/kg by oral gavage for 12 weeks. 3.the folder “ChronicNMP_PAS_1mgperkg”: contains PAS-stained liver sections from mice that received PBS, NP, or MP at a concentration of 1mg/kg by oral gavage for 12 weeks. 4.The folder “ChronicNMPwashout_PAS_100mgperkg” contains PAS-stained liver sections from mice that received PBS, NP (100 mg/kg), or MP (100 mg/kg) by oral gavage for 12 weeks and were analyzed 6 months after the final gavage. “InsulinIHC_NMP_HFDingestion.zip” contains all insulin immunohistochemistry images acquired using a 10× objective. All images were used to quantify insulin-secreting β-cell area in pancreatic islets, which is reported as percentage of insulin-positive area (% area) in the figures of the associated publication. Mice received PBS, NP (100 mg/kg), or MP (100 mg/kg) by oral gavage for 12 weeks. After 8 weeks of NMP exposure, mice were additionally placed on a high-fat diet (HFD) for the remaining 8 weeks. The folder “IF_NMPingestion_Clec4f-DTR.zip” contains immunofluorescence-stained liver sections from Clec4f-DTR/+ mice that received PBS or MP by oral gavage for 12 weeks. After completion of the plastic exposure, mice were treated with diphtheria toxin (DT). Animals were sacrificed 12 weeks after DT administration, and the images were used to quantify hepatic macrophage numbers (Iba1⁺ cells), which are reported as cells (×10⁴)/µm². The folders “GFAP_PBS.zip”, “GFAP_NP.zip”, and “GFAP_MP.zip” contain immunofluorescence-stained brain sections from mice that received PBS, NP MP, respectively, by oral gavage for 12 weeks. Images were acquired using a 20× objective and were used to quantify GFAP⁺ astrocytes in the brain. The folder “gWAT_HE_NMPingestion.zip” contains all hematoxylin-eosin (HE) staining images of gonadal white adipose tissue (gWAT). All images were used to assess dipocyte size using AdipoQ software. All mice were on a C57BL/6Jrcc background that received PBS, NP, or MP at 100 mg/kg by oral gavage for 12 weeks. 2. Are there multiple versions of the dataset? No 2.1 If yes, name of file(s) that was updated: i. Why was the file updated? ii. When was the file updated? 3. Relationship between files : 4. Additional related data collected that was not included in the current data package: C SHARING/ACCESS INFORMATION 1. Was data derived from another source? No 2. Licenses/restrictions placed on the data: CC-BY 3. Links to publications that cite or use the data: https://doi.org/10.21203/rs.3.rs-6204281/v1 4. Links to other publicly accessible locations of the data : 5. Links/relationships to ancillary datasets : D METHODOLOGICAL INFORMATION 1. Description of methods used for collection/generation of data: “ORO_NMPingestion.zip”: After overnight fixation in 4 % Formaldehyde (Cat# 11586711, ThermoFisher Scientific), livers were dehydrated overnight with incubation in 30 % D-Sucrose in PBS, then embedded in Tissue-Tek O.C.T. Compound (Cat# 4583, Sakura Finetek). Cryo-preserved liver blocks were cut at 12 μm thickness and dried for 1 h, at RT. Sections were rinsed with 60% isopropanol (Cat# 1027812500, Merck). Thereafter, tissue sections were stained using an Oil-red-O Stain Kit (Cat# O0625, Sigma-Aldrich) according to the manufacturer's instructions. The sections were then stained with hematoxylin for 5-10 min to visualize nuclei, rinsed with distilled water, and mounted with Kaiser's glycerol gelatin (pre-heated at 55°C) (Cat# 6474.1, Carl Roth). Images were acquired with an Axio Lab.A1 microscope. “PAS_NMPingestion.zip”: for the Periodic acid Schiff (PAS)-hematoxylin (H) staining, livers were fixed in 4% Formaldehyde (Cat# 11586711, ThermoFisher Scientific) overnight. The fixed tissues were dehydrated, embedded in paraffin, cut (5 µm) and then processed for staining as described below. Prior to staining, tissue sections were deparaffinized at 65 °C for 30 minutes and rehydrated by passing through a series of steps, each for 3 minutes: 2 changes of xylene (Cat# 8749, Carl Rot), followed by 100 %, 95 %, 90 %, 80 %, and 70 % ethanol, and finally distilled water. Afterwards, liver sections were incubated in periodic acid solution (Cat# HP00, Carl Roth) for 10 minutes to oxidize glycogen, rinsed in tap water for 3 minutes, and then briefly rinsed in distilled water. The sections were placed in Schiff’s reagent (Cat# X900.2, Carl Roth) for 15 minutes. Following staining, sections were washed in running tap water for 10 minutes then briefly rinsed in distilled water. Counterstaining was performed using hematoxylin (Cat# T865, Carl Roth) for 45 seconds to visualize nuclei. Excess hematoxylin was removed by washing in tap water for 3 minutes, followed by a brief rinse in distilled water. Sections were passed through distilled water, 70%, 80%, 90%, 95%, and 100% ethanol, each for 3 minutes, and then cleared in 2 changes of xylene. Sections were mounted with Entellan (Cat# 107961, Merck) and covered with glass coverslips. Images were acquired with Axio Lab.A1 microscope (Zeiss). “InsulinIHC_NMP_HFDingestion.zip”: Whole pancreata were fixed in 4% Formaldehyde (Cat# 11586711, ThermoFisher Scientific) overnight. The fixed tissues were dehydrated, embedded in paraffin and 5 µm cuts were performed at intervals of 200 µm. Sections were deparaffinized by incubating at 60°C for 30, then rehydrated by passing through a series of steps, each for 3 minutes: 2 changes of xylene (Cat# 8749, Carl Roth), followed by 100 %, 95 %, 90 %, 80 %, and 70 % ethanol, and finally distilled water. Endogenous peroxidase were blocked using 1.2% hydrogen peroxide in PBS for 10 minutes, then antigen retrieval was performed using sodium citrate buffer containing 10mM C₆H₅Na₃O₇·2H₂O (Cat# 3580.1, Carl Roth) and 0.05% Tween 20 (Cat# A4974, AppliChem) in distilled water, adjusted to pH6.0, for 30 minutes at 95°C, followed by a brief ice-water bath. Slides were washed with 3 times using 0.4% Triton X-100 in PBS, before blocking using 1% BSA and 2% normal goat serum. Samples were then incubated overnight with anti-Insulin (IR00261-2, Agilent), followed by washing and incubation with anti-guinea pig HRP (A18769, Invitrogen) for 2 hours. The ImmPACT NovaRED HRP Substrate Marker was prepared according to manufacturer’s instructions (Cat# SK-4805-NB, Novus Biologicals) and used to stain the slides for 7 minutes, followed by a washing step under running tap water (5 mins). Counterstaining was carried out by dipping the slides in Hematoxylin for 1-2 mins, and rinsing the slides under running tap water. Finally, the slides were dehydrated in ascending alcohol series, as stated above but in the opposite order. DPX Mountant for histology (Cat# 06522, Sigma) was utilized to mount the slides. Slides were using a Zeiss Axioscan.Z1 Slidescanner to image whole sections. “IF_NMPingestion_Clec4f-DTR.zip”, “GFAP_PBS.zip”, “GFAP_NP.zip” and “GFAP_MP.zip”: Organs harvested for immunofluorescence analysis were fixed in 4% Formaldehyde (Cat# 11586711, ThermoFisher Scientific) overnight (brain, liver). Organs were dehydrated overnight in 30 % D-Sucrose (Cat# 10638403, Fisher Scientific) in PBS, then embedded in Tissue-Tek O.C.T. Compound (Cat# 4583, Sakura Finetek). Cryo-preserved blocks were cut and dried for 1 hour at RT. Tissue slices were permeabilized with 0.4% Triton X-100 (Cat# X100, Sigma-Aldrich) in PBS for 1 hour. Thereafter, non-specific binding was blocked with 0.4% Triton X-100 in PBS containing 2% bovine serum albumin (BSA) (Cat# 37525, ThermoFisher Scientific) and 2% donkey serum (Cat# D9663, Sigma Aldrich) or 2 % normal goat serum (ICNA 08642921, VWR) for 2 hours. Samples were incubated at 4°C overnight with primary antibodies, washed with PBS, and then incubated with secondary antibodies for 2 hours at RT. After washing, nuclei were stained with DAPI (Cat# 42281, Biolegend), and samples were mounted using Fluoromount G Mounting Medium (Cat# 00-4958-02, ThermoFisher Scientific). Immunofluorescence images were acquired with a Zeiss LSM Airyscan 880 confocal microscope (Zeiss). “gWAT_HE_NMPingestion.zip”:For Hematoxylin and Eosin (H&E) staining, adipose tissue (gWAT) were fixed in 4% Formaldehyde (Cat# 11586711, ThermoFisher Scientific) overnight. The fixed tissues were dehydrated, embedded in paraffin, cut (5 µm) and then processed for staining as described below. Prior to staining, tissue sections were deparaffinized at 65 °C for 30 minutes and rehydrated by passing through a series of steps, each for 3 minutes: 2 changes of xylene (Cat# 8749, Carl Rot), followed by 100 %, 95 %, 90 %, 80 %, and 70 % ethanol, and finally distilled water. afterwards, gWAT sections were incubated in hematoxylin (Cat# T865.2, Carl Roth) for 90 seconds, then rinsed in tap water to stabilize the colour. The sections were placed in eosin (Cat# X883.2, Carl Roth) for 3 minutes, followed by a brief rinse in distilled water. The sections were dehydrated, mounted with Entellan (Cat# 107961, Merck), and covered with glass coverslips. Images were acquired with an Axio Lab.A1 (Zeiss) microscope with a 10X objective. 2. Methods for processing the data: “ORO_NMPingestion.zip”: For ORO quantification, 5 images of each sample with a 10X objective were taken and quantification was performed using QuPath software (version 0.5.1). “PAS_NMPingestion.zip”: For analysis of acquired images Fiji analysis software (v2.1.0/1.53c) was used. First, the colour threshold was set for the images and they were converted to 8-bit images. After setting an auto threshold, they were reverted, the colour was deconvoluted and signal intensity for the PAS image plane was measured. “InsulinIHC_NMP_HFDingestion.zip”: Images were analyzed with QuPath 0.6.0. Images were first thresholded to pancreas area, then 20% of the images were used to manually train a pixel classifier to identify insulin-expressing β-islets. The pixel classifier was applied to each section, the relative stained area of the pancreas was thus calculated and averaged throughout sections of the same sample. “IF_NMPingestion_Clec4f-DTR.zip”, “GFAP_PBS.zip”, “NP_PBS.zip”, and “MP_PBS.zip”: IF images were processed with (Fiji is just) ImageJ 2 (v2.14.0). “gWAT_HE_NMPingestion.zip” : H&E-stained gWAT sections were assessed for adipocyte size using AdipoQ software as previously described by Sieckmann et al.(https://doi.org/10.1091/MBC.E21-11-0592). Briefly, the image was preprocessed by segmenting into the fore- and background, creating a mask of each adipocyte. Subsequently, the mask was quantified and parameters such as area of the object and number of objects are determined. This allowed to calculate the frequency distribution for different adipocyte sizes between conditions. 3. Instrument- and/or software-specific information needed to interpret the data: Immunofluorescence images were acquired with a Zeiss LSM Airyscan 880 confocal microscope (Zeiss) and processed with (Fiji is just) ImageJ 2 (v2.14.0). PAS images were acquired with Axio Lab.A1 microscope (Zeiss) and processed with (Fiji is just) ImageJ 2 (v2.14.0). ORO images were acquired with Axio Lab.A1 microscope (Zeiss) and processed with QuPath1050 software (version 0.5.1) The insulin IHC images were acquired with Zeiss Axio Scan.Z1 Slide scanner and processed with QuPath (version0.6.0) The gWAT HE images were acquired with Zeiss Axio Scan.Z1 Slide Scanner and processed with AdipoQ software. 4. People involved in sample collection, processing, analysis and/or submission: Nikola Makdissi, Maria Francesca Viola and Elvira Mass contributed to the PAS staining and analysis Maria Francesca Viola, Virginia Aliprandi and Elvira Mass contributed to the insulin immunohistochemistry staining and analysis Nikola Makdissi, Maria Francesca Viola, Nele Kronau and Elvira Mass contributed to the ORO staining and analysis Nikola Makdissi, Elvira Mass, Katharina Sieckmann, and Dagmar Wachten contributed to the AdipoQ analysis of gWAT Nikola Makdissi, Maria Francesca Viola, Marina Mayer and Elvira Mass contributed to the GFAP staining in the Brian Nikola Makdissi, Maria Francesca Viola and Elvira Mass contributed to the analysis of NMP ingestion in Clec4f-DTR mice 5. Describe any quality-assurance procedures performed on the data: 6. Standards and calibration information : 7. Environmental/experimental conditions : E DATA-SPECIFIC INFORMATION FOR: “IF_NMPingestion_Clec4f-DTR.zip” DTR: diphtheria toxin receptor 1. Variable list including full names and definitions (please spell out abbreviated words) of column headings for tabular data: 2. Units of measurement used: 3. Missing data codes/symbols: 4. Specialized formats or other abbreviations used: