Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2022 May 18;10(5):23259671221090894.
doi: 10.1177/23259671221090894. eCollection 2022 May.

Changes in Macrophage Polarization During Tendon-to-Bone Healing After ACL Reconstruction With Insertion-Preserved Hamstring Tendon: Results in a Rabbit Model

Shaohua Liu  1 , Jinrong Lin  1 , Zhiwen Luo  1 , Yaying Sun  1 , Chenghui Wang  1 , Shiyi Chen  1 , Xiliang Shang  1 , Jiwu Chen  1   2
Affiliations

Changes in Macrophage Polarization During Tendon-to-Bone Healing After ACL Reconstruction With Insertion-Preserved Hamstring Tendon: Results in a Rabbit Model

Shaohua Liu et al. Orthop J Sports Med. .

Abstract

Background: Decreasing the proinflammatory M1 macrophages or shifting the polarization status from M1 to M2 phenotype is thought to be beneficial for tendon-to-bone healing. In anterior cruciate ligament reconstruction (ACLR), using an insertion-preserved hamstring tendon (IP-HT) graft compared with a free hamstring tendon (FHT) graft has been shown to reduce graft necrosis and improve healing. However, the role of macrophage polarization at the tendon-to-bone interface is unclear.

Hypothesis: ACLR using IP-HT graft would facilitate the phenotype shift from M1 to M2 macrophages at the tendon-to-bone interface.

Study design: Controlled laboratory study.

Methods: Unilateral ACLR was performed on 42 healthy New Zealand White rabbits (study group, 21 rabbits with IP-HT graft; control group, 21 rabbits with FHT graft). At days 1, 3, and 7 and weeks 3, 6, 12, and 24 postoperatively, 3 rabbits in each group were sacrificed to investigate and compare the expression of surrogate markers for M1 macrophages (inducible nitric oxide synthase [iNOS] and tumor necrosis factor α [TNF-α]) and M2 macrophages (CD206 and transforming growth factor β [TGF-β]) via immunohistochemical staining and evaluation.

Results: In the control group, the percentage of iNOS- and TNF-α-positive cells from postoperative day 7 and week 3 increased then decreased by week 6; positive expression of CD206 and TGF-β was weaker and peaked at 3 weeks postoperatively. In the study group, high CD206- and TGF-β-positive expression was observed from weeks 3 to 12 and peaked at week 6, and positive expression of iNOS- and TNF-α was weaker and peaked on day 7. At both 7 days and 3 weeks, the percentages of iNOS- and TNF-α-positive cells in the control group were both significantly higher than in the study group (P ≤ .04 for all). At 6 weeks, the percentages of CD206- and TGF-β-positive cells in the study group were both significantly higher than in the control group (P = .02 and P = .04, respectively).

Conclusion: More expression of surrogate markers for M2 macrophages was observed in the tendon-to-bone healing process after ACLR using IP-HT versus FTP graft.

Clinical relevance: Using IP-HT grafts in ACLR may facilitate postoperative healing by shifting the local status of macrophage polarization at the tendon-to-bone interface.

Keywords: ACL; hamstring tendon; macrophage polarization; tendon-to-bone healing.

PubMed Disclaimer

Conflict of interest statement

This study was funded by National Natural Science Funding (81972062). The authors declared that they have no conflicts of interest in the authorship and publication of this contribution. AOSSM checks author disclosures against the Open Payments Database (OPD). AOSSM has not conducted an independent investigation on the OPD and disclaims any liability or responsibility relating thereto.

Figures

Figure 1.
Figure 1.
Immunohistochemical staining of inducible nitric oxide synthase (iNOS) in the study groups at postoperative (A) days 1, 3, and 7 and (B) weeks 3, 6, 12, and 24. Scale bars indicate 100 µm. (C) Percentage of iNOS-positive cells at the tendon-to-bone interface; error bars represent SDs. Part A: A-1 day study group; B-3 day study group; C-7 day study group; D-1day control group; E-3 day control group; F-7 day control group. Part B: A-3 week study group; B-6 week study group; C-12 week study group; D-24 week study group; E-3 week control group; F-6 week control group; G-12 week control group; H-24 week control group. *Significant difference between groups (P < .05). ACLR, anterior cruciate ligament reconstruction; FHT, free hamstring tendon; IP-HT, insertion-preserved hamstring tendon.
Figure 2.
Figure 2.
Immunohistochemical staining of tumor necrosis factor α (TNF-α) in the study groups at postoperative (A) days 1, 3, and 7 and (B) weeks 3, 6, 12, and 24. Scale bars indicate 100 µm. (C) Percentage of TNF-α–positive cells at the tendon-to-bone interface; error bars represent SDs. Part A: A-1 day study group; B-3 day study group; C-7 day study group; D-1day control group; E-3 day control group; F-7 day control group. Part B: A-3 week study group; B-6 week study group; C-12 week study group; D-24 week study group; E-3 week control group; F-6 week control group; G-12 week control group; H-24 week control group. *Significant difference between groups (P < .05). ACLR, anterior cruciate ligament reconstruction; FHT, free hamstring tendon; IP-HT, insertion-preserved hamstring tendon.
Figure 3.
Figure 3.
Immunohistochemical staining of CD206 in the study groups at postoperative (A) days 1, 3, and 7 and (B) weeks 3, 6, 12, and 24. Scale bars indicate 100 µm. (C) Percentage of CD206-positive cells at the tendon-to-bone interface; error bars represent SDs. Part A: A-1 day study group; B-3 day study group; C-7 day study group; D-1day control group; E-3 day control group; F-7 day control group. Part B: A-3 week study group; B-6 week study group; C-12 week study group; D-24 week study group; E-3 week control group; F-6 week control group; G-12 week control group; H-24 week control group. *Significant difference between groups (P < .05). ACLR, anterior cruciate ligament reconstruction; FHT, free hamstring tendon; IP-HT, insertion-preserved hamstring tendon.
Figure 4.
Figure 4.
Immunohistochemical staining of transforming growth factor β (TGF-β) in the study groups at postoperative (A) days 1, 3, and 7 and (B) weeks 3, 6, 12, and 24. Scale bars indicate 100 µm. (C) Percentage of TGF-β–positive cells at the tendon-to-bone interface; error bars represent SDs. Part A: A-1 day study group; B-3 day study group; C-7 day study group; D-1day control group; E-3 day control group; F-7 day control group. Part B: A-3 week study group; B-6 week study group; C-12 week study group; D-24 week study group; E-3 week control group; F-6 week control group; G-12 week control group; H-24 week control group. *Significant difference between groups (P < .05). ACLR, anterior cruciate ligament reconstruction; FHT, free hamstring tendon; IP-HT, insertion-preserved hamstring tendon.
Figure 5.
Figure 5.
Percentage of positive cells of inducible nitric oxide synthase (iNOS), CD206, tumor necrosis factor α (TNF-α), and transforming growth factor β (TGF-β) after anterior cruciate ligament reconstruction (ACLR) using (A) insertion-preserved hamstring tendon (IP-HT) graft and (B) free hamstring tendon (FHT) graft at postoperative days 1, 3, and 7 and weeks 3, 6, 12, and 24.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Amitava D, Mithun S, Soma D, et al. Monocyte and macrophage plasticity in tissue repair and regeneration. Am J Pathol. 2015;185(10):2596–2606. - PMC - PubMed
    1. Anca S, Thorsten P, Stefan W, et al. An unrestrained proinflammatory M1 macrophage population induced by iron impairs wound healing in humans and mice. J Clin Invest. 2011;121(3):985–997. - PMC - PubMed
    1. Barrientos S, Stojadinovic OM, Brem H, Tomic-Canic M. Growth factors and cytokines in wound healing. Wound Repair Regen. 2010;16(5):585–601. - PubMed
    1. Budny J, Fox J, Rauh M, Fineberg M. Emerging trends in anterior cruciate ligament reconstruction. J Knee Surg. 2017;30(1):63–69. - PubMed
    1. Chen W, Sun Y, Gu X, et al. Conditioned medium of human bone marrow-derived stem cells promotes tendon-bone healing of the rotator cuff in a rat model. Biomaterials. 2021;271:120714. - PubMed

LinkOut - more resources