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Cited by 352Cellular Mechanics and InteractionsBone Tissue Engineering MaterialsWound Healing and Treatments

Micro- and Nanopatterned Topographical Cues for Regulating Macrophage Cell Shape and Phenotype

Open access
Luu, Thuy U., Gott, Shannon C., Woo, Bryan W. K., Rao, Masaru P., Liu, Wendy F.·2015·ACS Applied Materials & Interfaces
DOI 10.1021/acsami.5b10589PMC PMC4797644
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Abstract

Controlling the interactions between macrophages and biomaterials is critical for modulating the response to implants. While it has long been thought that biomaterial surface chemistry regulates the immune response, recent studies have suggested that material geometry may in fact dominate. Our previous work demonstrated that elongation of macrophages regulates their polarization toward a pro-healing phenotype. In this work, we elucidate how surface topology might be leveraged to alter macrophage cell morphology and polarization state. Using a deep etch technique, we fabricated titanium surfaces containing micro- and nanopatterned grooves, which have been previously shown to promote cell elongation. Morphology, phenotypic markers, and cytokine secretion of murine bone marrow derived macrophages on different groove widths were analyzed. The results suggest that micro- and nanopatterned grooves influenced macrophage elongation, which peaked on substrates with 400-500 nm wide grooves. Surface grooves did not affect inflammatory activation but drove macrophages toward an anti-inflammatory, pro-healing phenotype. While secretion of TNF-alpha remained low in macrophages across all conditions, macrophages secreted significantly higher levels of anti-inflammatory cytokine, IL-10, on intermediate groove widths compared to cells on other Ti surfaces. Our findings highlight the potential of using surface topography to regulate macrophage function, and thus control the wound healing and tissue repair response to biomaterials.

Integrity signals

Registry

Crossref · 10.1021/acsami.5b10589

Cites

42 of 42 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 29 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    After 36 h, macrophages on unpatterned titanium attached and spread into a flat, pancake-like shape.

    “Macrophages seeded on Ti materials adhered and spread to a flat, pancake shape on unpatterned surfaces after 36 h of culture”
    Quote in the paperSubstantial claim
  • Findingown data

    Arg1 levels differed significantly between cells on 150 nm grooves and those on 300 nm, 500 nm, 1 μm, 2.5 μm, 5 μm, and 10 μm grooves.

    “Arg1 expression in cells on 300 nm, 500 nm, 1 μm, 2.5 μm, 5 μm, and 10 μm grooves was significantly different from expression in cells on 150 nm grooves”
    Quote in the paperSubstantial claim
  • Findingown data

    Macrophage elongation on grooved titanium surfaces showed a biphasic dependence on groove width.

    “revealed a biphasic dependence of elongation on groove width”
    • Also stated in: § 4. Discussion · plain
      “the degree of cell elongation level was biphasic and decreased when cells were cultured on grooves less than 200 nm or greater than 10 μm”
    Quote in the paperSubstantial claim
  • Findingown data

    Across all titanium conditions, macrophage TNF-α release stayed low.

    “secretion of TNF-alpha remained low in macrophages across all conditions”
    • Also stated in: § 3.2 Surface grooves modulate macrophage phenotype · plain
      “secretion of TNF-α remained low in macrophages across all conditions”
    Quote in the paperSubstantial claim
  • Findingown data

    Macrophages cultured on 5 μm grooved titanium surfaces secreted significantly more IL-10 than cells on other titanium surfaces.

    “macrophages cultured on 5 μm surfaces exhibited significantly higher secretion of the anti-inflammatory cytokine, IL-10, when compared to cells on other Ti surfaces”
    • Also stated in: § Abstract · plain
      “macrophages secreted significantly higher levels of anti-inflammatory cytokine, IL-10, on intermediate groove widths compared to cells on other Ti surfaces”
    Quote in the paperSubstantial claim
  • May combine two claims. 19
  • Classified as probably not a substantial claim. 10

  • May combine two claims.

    Findingown data

    Titanium surfaces produced lower Arg1 expression than IL-4/IL-13 treatment, and the level depended on groove size.

    “in comparison to IL-4/IL- 13 treatment, the expression of Arg1 on titanium surfaces was lower, but varied and dependent on groove dimensions”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    As groove width increased, Arg1 expression rose and reached a broad peak on 400 nm-5 μm substrates.

    “increasing the groove width led to an increase in Arg1 expression peaking broadly on substrates containing 400 nm - 5 μm grooves”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    iNOS levels stayed low across all groove widths and were much lower than in macrophages driven to M1 by LPS and IFN-γ.

    “The expression of iNOS remained similarly low across all groove widths (Figure 4D) and much less than macrophages stimulated towards an M1 phenotype by treatment with 10 ng/mL LPS and 10 ng/mL IFN-γ”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Groove widths between 200 nm and 10 μm produced higher cell alignment and significantly higher elongation factor than blank titanium.

    “widths, that were in a range between 200 nm and 10 μm, led to higher cell alignment and significant higher elongation factor”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    On 200 nm grooved surfaces, cells showed fewer actin-cluster structures and weaker overall vinculin staining.

    “cells cultured on 200 nm grooved surfaces exhibited few structures containing actin clusters and less overall vinculin staining”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    On wider grooves, Arg1 expression was somewhat higher than on blank surfaces, despite average elongation not being higher.

    “the degree of Arg1 expression was somewhat elevated on substrates with wider groove sizes, even though the average cell elongation was not elevated relative to unpatterned blank surfaces”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Arg1 levels generally tracked elongation, but on wider grooves (25-50 μm) and flat titanium, Arg1 stayed relatively high even though elongation was lower.

    “The expression of Arg1 was generally correlated with the degree of elongation, although at wider groove dimensions (25 – 50 μm) and flat titanium surfaces where elongation was generally lower, the Arg1 expression remained relatively high”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Macrophages lacked organized stress fibers and instead had diffuse actin staining, with some clustering at the tips of elongated cells on 450 nm and 5 μm grooves.

    “macrophages do not exhibit organized stress fibers. Instead, the cells exhibited diffuse actin staining with some clustering particularly at the tips of elongated cells on 450 nm and 5 μm grooves”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Vinculin staining showed diffuse structures and few focal adhesions.

    “Vinculin staining revealed similarly diffuse structures with few focal adhesions”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Cells need a minimum groove width to elongate; wider grooves cause them to spread in all directions on flat areas with little elongation.

    “there is a minimum groove width that is necessary for cells to elongate, and that wider grooves lead cells to spread in all directions on the flat regions with minimal elongation”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Cell elongation and groove size both influenced macrophage Arg1 expression and IL-10 secretion.

    “macrophage Arg1 expression and IL-10 secretion was modulated by cell elongation, and depended on groove size”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Titanium topography controls macrophage polarization markers and has a moderate but significant effect on cytokine release.

    “We found that Ti surface topography regulates polarization-associated macrophage markers, and moderately but significantly affect their release of cytokines”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Topographical substrates can alter cellular components and might be important for controlling macrophage polarization.

    “topographical substrates are capable of modulating these cellular components, and may indeed play an important role in regulating macrophage polarization”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Three separate biological experiments were performed for cytokine secretion.

    “Three separate biological experiments were performed”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Macrophages lack extensive focal adhesions, but M2-polarized or micropatterned macrophages show brighter actin phalloidin staining.

    “While macrophages do not form extensive focal adhesions, phalloidin staining of the actin cytoskeleton of M2 polarized or micropatterned macrophages is indeed brighter.17”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [17] source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Collectively, earlier work indicates that micro- and nano-patterned topographies affect macrophage behavior in vitro and host responses in vivo.

    “Taken together, these studies suggest that micro and nano-patterned topographies modulate macrophage behavior in vitro as well as the host response in vivo”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    This work examined only two-dimensional biomaterial surfaces.

    “the current work has been focused on two-dimensional biomaterial surfaces”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Grooved substrates alter integrin expression and FAK phosphorylation.

    “Grooved substrates also lead to changes in integrin expression and focal adhesion kinase (FAK) phosphorylation.29”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [29] source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    In epithelial cells, focal adhesion size correlates with groove width, rising from 400 to 1500 nm as Si groove widths increased from 330 to 2100 nm at 600 nm depth.

    “The size of focal adhesions in epithelial cells, as detected by immunostaining of vinculin, is often directly correlated with groove width, and increased from 400 nm to 1500 nm as grooves on Si substrates were increased from 330 nm to 2100 nm and were 600 nm deep.25”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [25] source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Moderately roughened TiO2 surfaces promoted a pro-healing phenotype, while rougher surfaces caused inflammatory activation.

    “TiO2 surfaces that were moderately roughened induced polarization towards a pro-healing phenotype, whereas greater roughness led to inflammatory activation.5”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [5] source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    The University of California Irvine Institutional Animal Care and Use Committee approved all animal protocols before the study started.

    “All protocols involving animals were approved by the Institutional Animal Care and Use Committee at the University of California Irvine prior to initiation of the study”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Grooves are known to affect adhesion, adhesion signaling, and cytoskeletal organization in fibroblasts, myocytes, epithelial cells, mesenchymal stem cells, and other cell types.

    “Grooved surfaces have been shown to modulate adhesion, adhesion signaling, and cytoskeletal organization of many cell types including fibroblasts, myocytes, epithelial cells, and mesenchymal stem cells”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Compared with microfibers, nanofibers cause less macrophage adhesion and inflammatory activation and reduce the host response.

    “nanofibers elicit significantly less macrophage adhesion and inflammatory activation, and minimize the host response, when compared to microfibers.14-16”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [14] source

      Not checked in the free run.

    • cites [15] source

      Not checked in the free run.

    • cites [16] source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Arg1 or iNOS positivity was assigned when integrated fluorescence passed a threshold chosen to keep over 95% of non-stimulated glass cells negative.

    “Cells were determined to be positive for Arg1 or iNOS staining if integrated fluorescence for each cell exceeded a threshold value chosen such that more than 95% of non-stimulated cells on glass substrates were negative”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    Fluorescence intensity analysis examined at least 300 cells per condition in three independent biological experiments.

    “At least three hundred cells per condition were examined across three separate biological experiments”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Cells that are elongated typically have focal adhesions oriented along their long axis and a more organized cytoskeleton.

    “Elongated cells generally exhibit focal adhesions that are aligned along the direction of elongation and a more organized cytoskeletal components.24-27”
    Quote in the paperSubstantial claim0 of 4 sources checked
    • cites [24] source

      Not checked in the free run.

    • cites [25] source

      Not checked in the free run.

    • cites [26] source

      Not checked in the free run.

    • cites [27] source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    The study did not separate specific alternative activation subtypes M2a, M2b, and M2c.

    “Our study did not distinguish between specific alternative activation phenotypes (M2a, M2b, and M2c)”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    The experiments used calcium-containing DMEM media.

    “these results were obtained in calcium-containing DMEM media”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    Morphology was assessed in 100 cells per condition across three independent biological experiments.

    “A hundred cells per condition across three separate biological experiments were examined”
    Quote in the paperSubstantial claim
29 of 29

  • The paper relies on this without support.

    The mechanism by which surface topography changes macrophage function is not well understood.

    “However, the mechanism underlying topography-induced changes in macrophage function is still not well understood.”

  • The paper relies on this without support.

    The in vitro response of murine bone marrow derived macrophages to grooved titanium surfaces predicts the in vivo host and wound healing response to biomaterial implants.

    “Our findings highlight the potential of using surface topography to regulate macrophage function, and thus control the wound healing and tissue repair response to biomaterials.”
2 of 2

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • In vitro studies observed an elongation peak on 400-500 nm grooves, which showed the highest expression of IL-10 and Arg1 positive cells, suggesting polarization toward M2 macrophages.

    “In vitro studies observed elongation peak on the 400–500 nm grooves, which showed the highest expression of IL‐10 and Arg1 positive cells, suggesting polarization toward M2 macrophages.”

    From Unraveling and Harnessing the Immune Response at the Cell–Biomaterial Interface for Tissue Engineering Purposes (2024) · cites this paper as [146]

    Results §3.1 reports peak macrophage elongation on 400–500 nm grooves, and Results §3.2 reports broadly elevated Arg1 expression across 400 nm–5 μm grooves. But §3.2 places the highest IL-10 secretion on 5 μm grooves, not 400–500 nm grooves. The paper interprets the grooved surfaces as promoting an anti-inflammatory phenotype. (LLM, conf=0.92)

    Experimental — accuracy not yet measured

  • Aligned patterns with micron-level feature sizes promote M2 macrophage differentiation through forced alignment.

    “Aligned patterns with feature sizes in the micron level were shown to promote M2 macrophage differentiation through forced alignment”

    From Modeling mechanical activation of macrophages during pulmonary fibrogenesis for targeted anti-fibrosis therapy (2024) · cites this paper as [38]

  • Irregular topographies with dimensions similar to aligned patterns produce circularly spreading macrophages with M1 characteristics.

    “irregular topographies with similar dimensions produce circularly spreading macrophages with M1 characteristics”

    From Modeling mechanical activation of macrophages during pulmonary fibrogenesis for targeted anti-fibrosis therapy (2024) · cites this paper as [38]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Conflicting descriptions of elongation on 200 nm grooves

    Minor

    The results say that grooves from 200 nm through 10 μm produced a significantly higher elongation factor than blank titanium. The discussion instead describes macrophages on 200 nm grooves as not elongated.

1 of 1

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Related

Authors
Luu, T. U., Gott, S. C., Woo, B. W. K., Rao, M. P., & Liu, W. F.
Affiliations
  • University of California, Irvine (US)
  • University of California, Riverside (US)
Corresponding author
Liu, Wendy F.
ORCID
Venue
ACS Applied Materials & Interfaces
Publisher
American Chemical Society (ACS)
Published
2015-12-16
Volume
7
Issue
51
Pages
28665-28672
ISSN
  • 1944-8244 (ISSN-L)
  • 1944-8252
DOI
10.1021/acsami.5b10589
PMC
PMC4797644
OpenAlex
W2338968478
Cited by
344
References
42
Citations by year
2026: 29 · 2025: 34 · 2024: 31 · 2023: 36 · 2022: 38 · 2021: 53 · 2020: 31 · 2019: 34 · 2018: 28 · 2017: 18 · 2016: 12
FWCI
10.31
Type
journal article
Language
en
Open access
Locations
  • ACS Applied Materials & Interfaces (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
  • eScholarship (California Digital Library) (submittedVersion)
Funders
  • National Institute of Dental and Craniofacial Research (DP2DE023319)
  • California Institute for Regenerative Medicine (TG2-01152)
  • Edwards Lifesciences
  • National Science Foundation
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular Mechanics and Interactions
  • Physical Sciences › Engineering › Biomedical Engineering › Bone Tissue Engineering Materials
  • Health Sciences › Medicine › Rehabilitation › Wound Healing and Treatments
Keywords
macrophage polarization, surface topography, titanium implants, cell elongation, IL-10, TNF-α, anti-inflammatory phenotype, wound healing, biomaterial-immune interaction, deep etching
MeSH terms
Animals, Cells, Cultured, Macrophages, Mice, Inbred C57BL, Phenotype, Surface Properties, Biomarkers, Cytokines, Cell Shape, Mice

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