Mardiyyah/TAPT_data_V2_split · Datasets at Hugging Face

PMCID stringclasses 24 values	Title stringclasses 24 values	Sentences stringlengths 2 40.7k
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Cells were stained with BioTracker 405 Blue Mitochondrial Dye to visualize mitochondria (a, d, g), while the GFP channel (502 nm excitation) highlights caspase‐3/7 activation (b, e, h).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	At the 24 h time point, caspase‐3/7 activation begins to emerge, indicated by the fluorescence signal in the GFP channel, particularly in cells treated with M1‐derived EPs (e).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	The scale bar represents 20 μm.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Caspase 3/7 was triggered by M1 RAW 264.7 cell‐derived EPs: The induction of cell death at 48 h in MDA‐MB‐231 cells incubated with M0‐derived EPs (a–c), M1‐derived EPs (d–f), and M2‐derived EPs (g–i).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Cells were stained with BioTracker 405 Blue Mitochondrial Dye (a, d, g), and the GFP column at 502 nm excitation (b, e, h) shows the caspase 3/7 activation.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	The arrows in (e) may indicate cytoplasmic blebs/cell death in small dots around cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	The indicated scale bar is 20 μm.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	After observing the effects of M1 macrophage‐derived EPs against TNBC, an experiment was performed to investigate interactions via SEM.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Samples were prepared using a sterile silicon chip and placed in a 6‐well cell culture plate.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	After 24 h, EPs were found attaching themselves to MDA‐MB‐231 cells (Figure S1b).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Figure S1a shows no morphology change in MDA‐MB‐231 cells when EPs were not added; MDA‐MB‐231 cells clumped together in a ball shape when they were attached to a silicon chip without EPs.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	While Figure S1b shows cell division, we can differentiate morphological differences in Figure S1a,b. In addition, EPs were observed to be spreading on MDA‐MB‐231 cells, with some potential areas of attachment or anchoring.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	EPs were observed spreading on MDA‐MB‐231 cells, which seemed to be anchoring to cancer cells (Figure S2b,c, arrows).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Next, XTT analysis was performed to assess the cytotoxicity of M1‐derived EPs on MDA‐MB‐231 cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	XTT assay results showed only 47% MDA‐MB‐231 cell viability after 24 h of treatment with M1 RAW 264.7 cell‐derived EPs (Figure 8).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	An MDA‐MB‐231 cell viability of 206% in the sample containing M2 RAW 264.7 cell‐derived EPs was observed, which supports the published literature that suggests that the M2 macrophage‐like phenotype bolsters cancer cell growth .
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	On the other hand, M1‐derived EPs appeared to inhibit the growth of MDA‐MB‐231 cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Taken together, these results indicate that M1 macrophage‐derived EPs may induce cell death in TNBC cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	MDA‐MB‐231 XTT cell viability assay: MDA‐MB‐231 cells were exposed to EPs derived from M0 (Control), M1, and M2 RAW 264.7 cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	N = 3 independent experiments normalized to the control.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Error bars = SEM, connecting bars denote a p‐value < 0.05 and were considered statistically significant.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	This study indicates the potential for M1 macrophage‐derived EPs to favor tumor interactions and that M1 macrophage‐derived EVs could exhibit anti‐cancer effects.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Several studies have explored the potential role of M1 macrophage‐derived EVs' role in TNBC therapy as a potential cytotoxic agent in vitro.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	For example, macrophage‐derived exosome membranes have been used to load doxorubicin after removing the luminal content and conjugating with PLGA to target c‐Met to show its ability to target TNBC tumors in vivo .
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	However, the effect of EPs derived from macrophages with their native luminal content was never explored.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	This study identifies and characterize EPs with sizes less than 200 nm in each sample by following MISEV guidelines [6, 7].
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	A nitrite concentration of 45 μM in collected cell culture media of M1 polarized RAW 264.7 cells was measured [40, 41].
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	M1 RAW 264.7 cell‐derived EPs with luminal content showed cytotoxicity against MDA‐MB‐231 cells within 48 h by showing time‐dependent increments in caspase‐3/7 expression compared to the M0 uninduced RAW 264.7 cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	This time‐dependent increase in caspase‐3/7 activations in a controlled manner may indicate the EP's ability to release their content in a controlled fashion [42, 43].
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Moreover, this study further confirms the ability of M1 RAW 264.7 cell‐derived EPs to decrease cell viability by 53% in 24 h (Figure 8).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	On the other hand, M2 RAW 264.7 cell‐derived EPs increased the cell viability of MDA‐MB‐231 cells by 206% within 24 h (Figure 8), showing their potential tumor‐supporting effect [22, 23].
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	In addition, we captured the interaction of EPs with cancer cells that, to our knowledge, have yet to be visualized via SEM.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	The reason for this anchoring is unclear, but it could be the nature of M1 RAW 264.7 cell‐derived EPs to stimulate the change in morphology of MDA‐MB‐231 cells (Figures S1 and S2).
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Another reason could be the fusion with the cell membrane of MDA‐MB‐231 cells, which helps EPs internalize or release their content in the MDA‐MB‐231 cell cytosol .
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	This study revealed that M1 RAW 264.7 cell‐derived EPs can interact with MDA‐MB‐231 cells and induce cell death via caspase 3/7 activation.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Anchoring of EPs on TNBC cells has been suggested, which indicates that the other mechanisms of M1 macrophage‐derived EPs could be used to interact with tumor cells .
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	Furthermore, this study paves the way for exploring the content of macrophage‐derived EPs in detail, which can impact the fate of MDA‐MB‐231 cells.
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	In the future, M1 macrophage‐derived EPs could be combined with commercially available therapeutics to characterize the potential for synergistic therapeutic effects .
PMC12217045	M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.	The authors declare no conflicts of interest.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	The liver is the largest solid organ in the body, yet it remains incompletely characterized.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Here we present a spatial proteogenomic atlas of the healthy and obese human and murine liver combining single-cell CITE-seq, single-nuclei sequencing, spatial transcriptomics, and spatial proteomics.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	By integrating these multi-omic datasets, we provide validated strategies to reliably discriminate and localize all hepatic cells, including a population of lipid-associated macrophages (LAMs) at the bile ducts.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	We then align this atlas across seven species, revealing the conserved program of bona fide Kupffer cells and LAMs.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	We also uncover the respective spatially resolved cellular niches of these macrophages and the microenvironmental circuits driving their unique transcriptomic identities.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	We demonstrate that LAMs are induced by local lipid exposure, leading to their induction in steatotic regions of the murine and human liver, while Kupffer cell development crucially depends on their cross-talk with hepatic stellate cells via the evolutionarily conserved ALK1-BMP9/10 axis.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	The immense advances in single-cell transcriptomics have enabled a better understanding of the cellular composition of different organs across species.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	However, we still lack information regarding how these cells are organized in their distinct microenvironmental niches.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Moreover, the specific cell-cell interactions determining the identity of individual cells within tissues remain to be defined (Guilliams and Scott, 2017; Lindeboom et al., 2021).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	While the spatial organization of hepatocytes within the liver is understood (Halpern et al., 2017), that of non-parenchymal liver cells remains unclear.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	This is the case for the mouse liver, but even more so for the human liver, where the identity and the precise localization of most hepatic cells is unknown.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Moreover, the link between the transcriptome and the proteome has not been studied, resulting in a lack of reliable surface markers to identify these cells by flow cytometry and confocal microscopy.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Here, we used proteogenomic techniques including cellular indexing of transcriptomes and epitomes by sequencing (CITE-seq) and spatial approaches to identify all cells and their specific locations within the healthy and obese livers of mice and humans.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	By doing so, we have developed strategies for the identification and further study of hepatic cells.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Demonstrating the usefulness of this approach, we identify the evolutionary conserved and spatially restricted signals driving the distinct hepatic macrophage phenotypes.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	To generate a proteogenomic atlas of the liver, we first examined the optimal method for retrieving all hepatic cells.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Using the murine liver, we compared single-cell RNA sequencing (scRNA-seq) using cells isolated via ex vivo or in vivo enzymatic digestion with single-nuclei RNA sequencing (snRNA-seq) (Figures S1A–S1C).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	We did not observe any differences in the number of genes/cell between the two digestion methods (Figures S1D and S1E), but snRNA-seq typically yielded a lower number of genes/cell (Figures S1D and S1F).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	This did not prevent distinct cell types from being identified in the snRNA-seq dataset as both scRNA-seq and snRNA-seq identified highly expressed genes in each population.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	However, expression was often higher in the scRNA-seq (Figure S1F).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Additionally, we observed a signature of digestion-associated genes and snRNA-seq-associated genes across cell types (Figure S1F).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	While in terms of genes/cell snRNA-seq is inferior to scRNA-seq, this method best recapitulated the cell frequencies observed in vivo (Figures S1D–S1N).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	As each method has advantages and disadvantages, the optimal method to use depends on the biological question being addressed.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Here, as we sought to generate a proteogenomic atlas of all hepatic cells, we thus opted to use a combination of all protocols.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	To investigate mRNA and protein expression at single-cell resolution, we used CITE-seq (Stoeckius et al., 2017), staining a selection of the scRNA-seq samples with 107–161 oligo-conjugated antibodies (Figure 1A).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Data were pooled together for a single analysis where, with TotalVI (Gayoso et al., 2021), both the protein and mRNA profiles were considered for clustering (Figure 1B).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Analysis of the differentially expressed genes and proteins (DEGs/DEPs; Figures S2A and S2B; Table S1) identified 17 cell types (Figure 1B), which were differentially represented with each isolation method (Figure S2C).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Addition of antibodies in the CITE-seq analysis enabled surface markers for all cells to be identified, including VSIG4 and FOLR2 for Kupffer cells (KCs) (Figure S2B), without affecting the quality of the transcriptomic data (Figure 2D).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	This analysis identified one subset of KCs.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	However, 2 subpopulations termed KC1 and KC2 have recently been described (Blériot et al., 2021; Simone et al., 2021).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	As our CITE-seq analysis identified that the markers used to identify KC2s, namely CD206 and ESAM, are largely expressed by liver sinusoidal endothelial cells (LSECs) (Figure S2E), we next sought to determine if we had previously removed any potential KC2s in our initial QC steps as LSEC-KC doublets.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	To examine this, we generated a UMAP of samples containing CD206 and ESAM in the CITE-seq panel and performed a minimal QC filtering on number of genes and percentage of mitochondrial genes.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	In this UMAP, we found multiple subpopulations including 3 populations of KCs and 2 populations of B cells (Figure S2F).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	To determine if any of these populations could be KC2s, we harnessed the power of CITE-seq to recreate the gating strategy recently proposed to identify KC2s (Blériot et al., 2021).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Converting the CITE-seq data into an FCS file and analyzing this in FlowJo identified 2 of the KC populations to be KC2s expressing CD206 and ESAM, and these cells were present in a similar ratio among KCs as reported by Blériot et al. (Figure S2G).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	However, employing the same gating strategy with the B cells also enabled a B cell2 population to be identified (Figure S2G).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	The KC2 and B cell2 populations also expressed other protein markers associated with LSECs including CD26, CD31, and CD38, suggesting that these may be doublets (Figure S2H).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Consistent with this, we did not uncover any DEGs specifically expressed in the KC2s or B cell2s, rather these cells had an intermediate profile between either KC1s or B cell1s and LSECs (Figure S2I), as would be expected for doublets.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Finally, we perfused the livers with antigen fix to inflate the LSECs to be able to distinguish more readily between KCs and LSECs and performed confocal microscopy.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	3D reconstruction of these images indicates that CD206 expression is observed primarily in the LSECs with which the KCs are intertwined.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	These microscopy images also show that KCs express some CD206, however, consistent with our CITE-seq analysis, this was observed across the KC population rather than in a KC subset further suggesting the existence of only one-KC population (Figure S2J).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	With this in mind, we decided to continue to apply our initial strict QC controls eliminating these potential doublets from further analyses.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Figure S1Cell types identified in transcriptomic studies depend upon cell/nuclei isolation technique used, related to Figure 1Cells were isolated from livers of healthy C57B/l6 mice by either ex vivo or in vivo enzymatic digestion.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Alternatively, livers were snap frozen and nuclei subsequently isolated following tissue homogenization by a sucrose gradient (3 mice per isolation method).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Live cells/intact nuclei were identified and purified using flow cytometry.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	For the cells, either live CD45, live CD45 or live hepatocytes were sorted.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	1 ex vivo digested sample and 1 in vivo digested sample were also stained with a panel of 107 (ex vivo cells) or 161 (in vivo cells) oligo-conjugated antibodies for CITE-seq analysis.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	FACS-purified cells/nuclei were loaded onto the 10× chromium platform and scRNA-seq, CITE-seq, or snRNA-seq performed.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Following clean up and QC, cells from the same mice were pooled together in the same ratios (CD45:CD45:Heps) as found in the tissue as a whole before sorting, different mice were then pooled together and the data were analyzed using scVI.(A–C) UMAPs showing annotations of cell types and proportions of each cell type as a % of total cells in the UMAP isolated using (A) ex vivo digestion; 13,144 cells, (B) in vivo digestion; 24,014 cells and (C) nuclei; 8,583 nuclei.(D) Average number of genes/cell in the annotated mac, B cell, hepatocyte, endothelial, and stromal cell populations following each isolation method.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	p < 0.05 one-way ANOVA with Bonferroni post-test per cell type.(E) Correlation plot showing genes captured within the mac population when the liver is digested with the in vitro versus the in vivo digestion protocol.(F) Correlation plots showing genes captured within the mac, endothelial cell, and hepatocyte populations when cells are isolated using the in vivo digestion protocol or nuclei are isolated.(G–L) Confocal microscopy images to determine true abundance of (G) stromal cells and cholangiocytes (H) endothelial cells, (I) macs, (J) dendritic cells, (K) B cells, and (L) T cells in vivo.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Scale bars, 200 μm.(M) The percentage of each population was calculated based on the percentage of a given population divided by the total number of nuclei.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	A threshold was applied to the DAPI channel (picture 1) in ImageJ (picture 2) and nuclei were automatically counted based on the ImageJ “analyze particles” plugin (size ).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Due to the density of some liver zones, some nuclei were not automatically counted (arrow, picture 3).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Those were then manually counted and added to the total number of nuclei.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	For the populations of interest, cells were counted manually based on specific markers (for example, CD3 for T cells, picture 4).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Counting was performed blinded prior to analysis of the sequencing results.(N) Proportion of indicated cell types as a % of total cells identified in confocal microscopy images.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Data are from 3–7 images per cell type taken from 2–4 mice.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Figure 1A proteogenomic atlas of the healthy murine liver(A) Hepatic cells were isolated from healthy C57B/l6 mice by ex vivo (5 mice, 15 samples) or in vivo (5 mice, 19 samples) enzymatic digestion.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Alternatively, nuclei were isolated by tissue homogenization (4 mice, 12 samples).
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	Live cells/intact nuclei were FACS-purified.
PMC8809252	Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches.	For cells, total live, live CD45, live CD45, live hepatocytes, or myeloid cells (live CD45, CD3, CD19, B220, NK1.1) were sorted.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Cells were stained with BioTracker 405 Blue Mitochondrial Dye to visualize mitochondria (a, d, g), while the GFP channel (502 nm excitation) highlights caspase‐3/7 activation (b, e, h).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

At the 24 h time point, caspase‐3/7 activation begins to emerge, indicated by the fluorescence signal in the GFP channel, particularly in cells treated with M1‐derived EPs (e).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

The scale bar represents 20 μm.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Caspase 3/7 was triggered by M1 RAW 264.7 cell‐derived EPs: The induction of cell death at 48 h in MDA‐MB‐231 cells incubated with M0‐derived EPs (a–c), M1‐derived EPs (d–f), and M2‐derived EPs (g–i).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Cells were stained with BioTracker 405 Blue Mitochondrial Dye (a, d, g), and the GFP column at 502 nm excitation (b, e, h) shows the caspase 3/7 activation.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

The arrows in (e) may indicate cytoplasmic blebs/cell death in small dots around cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

The indicated scale bar is 20 μm.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

After observing the effects of M1 macrophage‐derived EPs against TNBC, an experiment was performed to investigate interactions via SEM.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Samples were prepared using a sterile silicon chip and placed in a 6‐well cell culture plate.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

After 24 h, EPs were found attaching themselves to MDA‐MB‐231 cells (Figure S1b).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Figure S1a shows no morphology change in MDA‐MB‐231 cells when EPs were not added; MDA‐MB‐231 cells clumped together in a ball shape when they were attached to a silicon chip without EPs.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

While Figure S1b shows cell division, we can differentiate morphological differences in Figure S1a,b. In addition, EPs were observed to be spreading on MDA‐MB‐231 cells, with some potential areas of attachment or anchoring.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

EPs were observed spreading on MDA‐MB‐231 cells, which seemed to be anchoring to cancer cells (Figure S2b,c, arrows).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Next, XTT analysis was performed to assess the cytotoxicity of M1‐derived EPs on MDA‐MB‐231 cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

XTT assay results showed only 47% MDA‐MB‐231 cell viability after 24 h of treatment with M1 RAW 264.7 cell‐derived EPs (Figure 8).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

An MDA‐MB‐231 cell viability of 206% in the sample containing M2 RAW 264.7 cell‐derived EPs was observed, which supports the published literature that suggests that the M2 macrophage‐like phenotype bolsters cancer cell growth .

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

On the other hand, M1‐derived EPs appeared to inhibit the growth of MDA‐MB‐231 cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Taken together, these results indicate that M1 macrophage‐derived EPs may induce cell death in TNBC cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

MDA‐MB‐231 XTT cell viability assay: MDA‐MB‐231 cells were exposed to EPs derived from M0 (Control), M1, and M2 RAW 264.7 cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

N = 3 independent experiments normalized to the control.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Error bars = SEM, connecting bars denote a p‐value < 0.05 and were considered statistically significant.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

This study indicates the potential for M1 macrophage‐derived EPs to favor tumor interactions and that M1 macrophage‐derived EVs could exhibit anti‐cancer effects.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Several studies have explored the potential role of M1 macrophage‐derived EVs' role in TNBC therapy as a potential cytotoxic agent in vitro.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

For example, macrophage‐derived exosome membranes have been used to load doxorubicin after removing the luminal content and conjugating with PLGA to target c‐Met to show its ability to target TNBC tumors in vivo .

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

However, the effect of EPs derived from macrophages with their native luminal content was never explored.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

This study identifies and characterize EPs with sizes less than 200 nm in each sample by following MISEV guidelines [6, 7].

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

A nitrite concentration of 45 μM in collected cell culture media of M1 polarized RAW 264.7 cells was measured [40, 41].

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

M1 RAW 264.7 cell‐derived EPs with luminal content showed cytotoxicity against MDA‐MB‐231 cells within 48 h by showing time‐dependent increments in caspase‐3/7 expression compared to the M0 uninduced RAW 264.7 cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

This time‐dependent increase in caspase‐3/7 activations in a controlled manner may indicate the EP's ability to release their content in a controlled fashion [42, 43].

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Moreover, this study further confirms the ability of M1 RAW 264.7 cell‐derived EPs to decrease cell viability by 53% in 24 h (Figure 8).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

On the other hand, M2 RAW 264.7 cell‐derived EPs increased the cell viability of MDA‐MB‐231 cells by 206% within 24 h (Figure 8), showing their potential tumor‐supporting effect [22, 23].

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

In addition, we captured the interaction of EPs with cancer cells that, to our knowledge, have yet to be visualized via SEM.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

The reason for this anchoring is unclear, but it could be the nature of M1 RAW 264.7 cell‐derived EPs to stimulate the change in morphology of MDA‐MB‐231 cells (Figures S1 and S2).

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Another reason could be the fusion with the cell membrane of MDA‐MB‐231 cells, which helps EPs internalize or release their content in the MDA‐MB‐231 cell cytosol .

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

This study revealed that M1 RAW 264.7 cell‐derived EPs can interact with MDA‐MB‐231 cells and induce cell death via caspase 3/7 activation.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Anchoring of EPs on TNBC cells has been suggested, which indicates that the other mechanisms of M1 macrophage‐derived EPs could be used to interact with tumor cells .

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

Furthermore, this study paves the way for exploring the content of macrophage‐derived EPs in detail, which can impact the fate of MDA‐MB‐231 cells.

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

In the future, M1 macrophage‐derived EPs could be combined with commercially available therapeutics to characterize the potential for synergistic therapeutic effects .

PMC12217045

M1 Macrophage-Derived Extracellular Particles Induce Cell Death in MDA-MB-231 Cells.

The authors declare no conflicts of interest.

PMC8809252