Inflammatory M1-like macrophages polarized by NK-4 undergo enhanced phenotypic switching to an anti-inflammatory M2-like phenotype upon co-culture with apoptotic cells

Abstract Background: NK-4 has been used to promote wound healing since the early-1950s; however, the mechanism of action of NK-4 is unknown. In this study, we examined whether NK-4 exerts a regulatory effect on macrophages, which play multiple roles during wound healing from the initial inflammatory phase until the tissue regeneration phase. Results: NK-4 treatment of THP-1 macrophages induced morphological features characteristic of classically-activated M1 macrophages, an inflammatory cytokine profile, and increased expression of the M1 macrophage-associated molecules CD38 and CD86. Interestingly, NK-4 augmented TNF-a production by THP-1 macrophages in combination with LPS, Pam3CSK4, or poly(I:C). Furthermore, NK-4 treatment enhanced THP-1 macrophage phagocytosis. These results indicate that NK-4 drives macrophage polarization toward an inflammatory M1-like phenotype with increased phagocytic activity. Efferocytosis is a crucial event for resolution of the inflammatory phase in wound healing. NK-4-treated THP-1 macrophages co-cultured with apoptotic Jurkat E6.1 (Apo-J) cells switched from an M1-like phenotype to an M2-like phenotype, as seen in the inverted ratio of TNF-a to IL-10 produced in response to LPS. We identified two separate mechanisms that are involved in this phenotypic switch. First, recognition of phosphatidylserine molecules on Apo-J cells by THP-1 macrophages downregulates TNF-a production. Second, phagocytosis of Apo-J cells by THP-1 macrophages and activation of PI3K/Akt signaling pathway upregulates IL-10 production. Conclusion: It is postulated that the phenotypic switch from a proinflammatory M1-like phenotype to an anti-inflammatory M2-like phenotype is dysregulated due to impaired efferocytosis of apoptotic neutrophils at the wound site. Our results demonstrate that NK-4 improves efferocytosis, suggesting its potential as a therapeutic strategy to resolve sustained inflammation in chronic wounds.

Download Full-text

Inflammatory M1-like macrophages polarized by NK-4 undergo enhanced phenotypic switching to an anti-inflammatory M2-like phenotype upon co-culture with apoptotic cells

10.21203/rs.3.rs-66231/v2 ◽

2020 ◽

Author(s):

Keizo Kohno ◽

Satomi Koya-Miyata ◽

Akira Harashima ◽

Takahiko Tsukuda ◽

Masataka Katakami ◽

...

Keyword(s):

Wound Healing ◽

Chronic Wounds ◽

Macrophage Polarization ◽

Phenotypic Switching ◽

Apoptotic Cells ◽

Phenotypic Switch ◽

Inflammatory Phase ◽

Macrophage Phagocytosis ◽

M1 Macrophage ◽

Anti Inflammatory

Abstract Background: NK-4 has been used to promote wound healing since the early-1950s; however, the mechanism of action of NK-4 is unknown. In this study, we examined whether NK-4 exerts a regulatory effect on macrophages, which play multiple roles during wound healing from the initial inflammatory phase until the tissue regeneration phase.Results: NK-4 treatment of THP-1 macrophages induced morphological features characteristic of classically-activated M1 macrophages, an inflammatory cytokine profile, and increased expression of the M1 macrophage-associated molecules CD38 and CD86. Interestingly, NK-4 augmented TNF-a production by THP-1 macrophages in combination with LPS, Pam3CSK4, or poly(I:C). Furthermore, NK-4 treatment enhanced THP-1 macrophage phagocytosis of latex beads. These results indicate that NK-4 drives macrophage polarization toward an inflammatory M1-like phenotype with increased phagocytic activity. Efferocytosis is a crucial event for resolution of the inflammatory phase in wound healing. NK-4-treated THP-1 macrophages co-cultured with apoptotic Jurkat E6.1 (Apo-J) cells switched from an M1-like phenotype to an M2-like phenotype, as seen in the inverted ratio of TNF-a to IL-10 produced in response to LPS. We identified two separate mechanisms that are involved in this phenotypic switch. First, recognition of phosphatidylserine molecules on Apo-J cells by THP-1 macrophages downregulates TNF-a production. Second, phagocytosis of Apo-J cells by THP-1 macrophages and activation of PI3K/Akt signaling pathway upregulates IL-10 production.Conclusion: It is postulated that the phenotypic switch from a proinflammatory M1-like phenotype to an anti-inflammatory M2-like phenotype is dysregulated due to impaired efferocytosis of apoptotic neutrophils at the wound site. Our results demonstrate that NK-4 improves phagocytosis of apoptotic cells, suggesting its potential as a therapeutic strategy to resolve sustained inflammation in chronic wounds.

Download Full-text

Inflammatory M1-like macrophages polarized by NK-4 undergo enhanced phenotypic switching to an anti-inflammatory M2-like phenotype upon co-culture with apoptotic cells

Journal of Inflammation ◽

10.1186/s12950-020-00267-z ◽

2021 ◽

Vol 18 (1) ◽

Author(s):

Keizo Kohno ◽

Satomi Koya-Miyata ◽

Akira Harashima ◽

Takahiko Tsukuda ◽

Masataka Katakami ◽

...

Keyword(s):

Wound Healing ◽

Chronic Wounds ◽

Macrophage Polarization ◽

Phenotypic Switching ◽

Apoptotic Cells ◽

Phenotypic Switch ◽

Inflammatory Phase ◽

M1 Macrophage ◽

Tnf Α ◽

Anti Inflammatory

Abstract Background NK-4 has been used to promote wound healing since the early-1950s; however, the mechanism of action of NK-4 is unknown. In this study, we examined whether NK-4 exerts a regulatory effect on macrophages, which play multiple roles during wound healing from the initial inflammatory phase until the tissue regeneration phase. Results NK-4 treatment of THP-1 macrophages induced morphological features characteristic of classically-activated M1 macrophages, an inflammatory cytokine profile, and increased expression of the M1 macrophage-associated molecules CD38 and CD86. Interestingly, NK-4 augmented TNF-α production by THP-1 macrophages in combination with LPS, Pam3CSK4, or poly(I:C). Furthermore, NK-4 treatment enhanced THP-1 macrophage phagocytosis of latex beads. These results indicate that NK-4 drives macrophage polarization toward an inflammatory M1-like phenotype with increased phagocytic activity. Efferocytosis is a crucial event for resolution of the inflammatory phase in wound healing. NK-4-treated THP-1 macrophages co-cultured with apoptotic Jurkat E6.1 (Apo-J) cells switched from an M1-like phenotype to an M2-like phenotype, as seen in the inverted ratio of TNF-α to IL-10 produced in response to LPS. We identified two separate mechanisms that are involved in this phenotypic switch. First, recognition of phosphatidylserine molecules on Apo-J cells by THP-1 macrophages downregulates TNF-α production. Second, phagocytosis of Apo-J cells by THP-1 macrophages and activation of PI3K/Akt signaling pathway upregulates IL-10 production. Conclusion It is postulated that the phenotypic switch from a proinflammatory M1-like phenotype to an anti-inflammatory M2-like phenotype is dysregulated due to impaired efferocytosis of apoptotic neutrophils at the wound site. Our results demonstrate that NK-4 improves phagocytosis of apoptotic cells, suggesting its potential as a therapeutic strategy to resolve sustained inflammation in chronic wounds.

Download Full-text

The effect of coal-derived humic substances and their silver-containing bionanocomposites on arginine balance in peritoneal macrophages of intact mice

Bulletin of Siberian Medicine ◽

10.20538/1682-0363-2021-4-71-78 ◽

2022 ◽

Vol 20 (4) ◽

pp. 71-78

Author(s):

E. S. Trofimova ◽

M. V. Zykova ◽

M. G. Danilets ◽

A. A. Ligacheva ◽

E. Yu. Sherstoboev ◽

...

Keyword(s):

Humic Substances ◽

Chronic Wounds ◽

Macrophage Polarization ◽

Antigen Presenting Cells ◽

Peritoneal Macrophages ◽

Research Area ◽

The Body ◽

M1 Macrophage ◽

Anti Inflammatory ◽

Antigen Presenting

Background. Antigen-presenting cells (APCs), especially macrophages, play an important role in the body defense against various pathogens. Their dysfunction and polarization are associated with most inflammatory and autoimmune diseases. The inflammatory process is regulated by activation and / or inhibition of genes differentially expressed by macrophages. Successful correction of inflammation leads firstly to elimination of inflammatory stimuli and then to remodeling and restoration of tissues and organs. It was experimentally confirmed that silvercontaining bionanocomposites based on natural humic substances (HS) obtained from coal of different origin, as well as initial matrices of these HS, are capable of activating pro- and anti-inflammatory properties of macrophages.Aim. To study cytotoxic, pyrogenic, and immunomodulatory properties (arginine balance) of initial HS samples and samples of silver nanoparticles ultradispersed in these HS matrices (HS-AgNPs) in the cell culture of peritoneal macrophages, as well as their effect on pro- and anti-inflammatory properties of APCs.Materials and methods. Cultural and biochemical methods were used in the study.Results. The study showed that the samples CHE-K, CHE-AgNPs, CHS-K, and CHP-K increased M1 macrophage polarization due to stimulation of the NO-synthase activity and inhibition of arginase. The samples CHI-K, CHIAgNPs, CHP-AgNPs, and CHS-AgNPs modulated an alternative M2 or M2-like state of macrophage activation. At the same time, HS are not cytotoxic at effective concentrations, and three out of four studied samples did not contain pyrogenic impurities.Conclusion. The use of HS and their silver-containing bionanocomposites, which have the ability to greatly affect the polarization of antigen-presenting cells, is a promising research area in correction of the inflammatory response for solving an important social and medical problem of treating chronic wounds.

Download Full-text

Neuropeptides, Inflammation, Biofilms, and diabetic Foot Ulcers

Experimental and Clinical Endocrinology & Diabetes ◽

10.1055/a-1493-0458 ◽

2021 ◽

Author(s):

Shaoling Yang ◽

Liye Hu ◽

Rui Han ◽

Yiwen Yang

Keyword(s):

Wound Healing ◽

Diabetic Foot ◽

Macrophage Polarization ◽

Public Health Problem ◽

Limb Amputation ◽

Major Public Health Problem ◽

Inflammatory Phase ◽

Macrophage Phagocytosis ◽

Increased Risk ◽

Antimicrobial Resistance Genes

AbstractA diabetic foot ulcer (DFU) is a serious complication in patients with diabetes mellitus (DM). A DFU is the most common cause of non-traumatic limb amputation, and patients with DFUs have increased mortality rates within 5 years after amputation. DFUs also increase the risk of cardiovascular and cerebrovascular diseases; therefore, with the increasing incidence and prevalence of diabetic foot wounds, DFUs are gradually becoming a major public health problem. The pathophysiology of DFUs is complicated and remains unclear. In recent years, many studies have demonstrated that the pathophysiology of DFUs is especially associated with neuropeptides, inflammation, and biofilms. Neuropeptides, especially substance P (SP) and calcitonin gene-related peptide (CGRP), play an important role in wound healing. SP and CGRP accelerate the healing of cutaneous wounds by promoting neovascularization, inhibiting the release of certain proinflammatory chemokines, regulating macrophage polarization, and so on. However, the expression of SP and CGRP was downregulated in DM and DFUs. DFUs are characterized by a sustained inflammatory phase. Immune cells such as neutrophils and macrophages are involved in the sustained inflammatory phase in DFUs by extracellular traps (NETs) and dysregulated macrophage polarization, which delays wound healing. Furthermore, DFUs are at increased risk of biofilm formation. Biofilms disturb wound healing by inducing a chronic inflammatory response, inhibiting macrophage phagocytosis and keratinocyte proliferation migration, and transferring antimicrobial resistance genes. To understand the relationships among neuropeptides, inflammation, biofilms, and DFUs, this review highlights the recent scientific advances that provide possible pathophysiological insights into the delayed healing of DFUs.

Download Full-text

Reviewing the Physiology of Cutaneous Wound Healing and Evaluating the Effect of Exercise on it: A Narrative Review Article

Annals of Military and Health Sciences Research ◽

10.5812/amh.115926 ◽

2021 ◽

Vol 19 (3) ◽

Author(s):

Fatima Riyahi ◽

Simin Riahy ◽

Mitra Yousefpour

Keyword(s):

Wound Healing ◽

Chronic Wounds ◽

Herbal Medicines ◽

The Body ◽

Cutaneous Wound Healing ◽

Skin Damage ◽

Cutaneous Wound ◽

Inflammatory Phase ◽

Anti Inflammatory

Context: The skin is the most important organ of the body, and maintaining its integrity is important for health. Severe skin damage is life-threatening, and wound healing restores its integrity. One of the main health problems is impaired cutaneous wound healing. According to the importance of wound healing and the fact that unrepaired skin decreases the quality of life, many studies have investigated the effect of some natural and chemical substances on the length and quality of wound healing to find beneficial interventions for rapid and economical treatment. Objectives: This review was conducted to describe the physiology of cutaneous wound healing and some positive and negative factors affecting it with a focus on exercise. Methods: An electronic search without any time limitation was performed on the PubMed, Google Scholar, and Web of Science databases. The keywords were ‘wound’, ‘healing’, and ‘exercise’. Finally, according to the similarities or differences between the results and the relationship with the subject, 53 papers were selected and reviewed. Results: Wound healing is a complex physiological process with four overlapping processes. It seems that disturbance in the inflammatory phase of wound healing is the main factor in the impairment of healing. Traditionally, many chemical and herbal medicines and compounds have been used to speed up wound recovery due to their anti-inflammatory and antioxidative properties. Many studies have evaluated the effect of exercise, as complementary medicine, on wound healing, and they have examined the effect of different protocols of exercise on the speed of wound healing. According to the results of these studies, aerobic exercise, due to its anti-inflammatory and antioxidative effects, is a beneficial method in shortening the length of healing, especially in aged, obese, and diabetic individuals. Conclusions: Exercise as a low-cost intervention is a good strategy in the treatment of impaired and chronic wounds.

Download Full-text

Prior Pro-inflammatory Polarization Changes the Macrophage Response to IL-4

10.1101/2021.04.06.438673 ◽

2021 ◽

Author(s):

Erin M O'Brien ◽

Kara L Spiller

Keyword(s):

Tissue Repair ◽

Chronic Wounds ◽

Macrophage Polarization ◽

Phenotypic Switching ◽

M2 Macrophages ◽

M1 Macrophages ◽

Macrophage Response ◽

M1 Macrophage ◽

Gene And Protein Expression ◽

M2 Phenotype

Tissue repair is largely regulated by diverse macrophage populations whose functions are timing- and context-dependent. The early phase of healing is dominated by pro-inflammatory macrophages, also known as M1, followed by the emergence of a distinct and diverse population that is collectively referred to as M2. The extent of the diversity of the M2 population is unknown. M2 macrophages may originate directly from circulating monocytes or from phenotypic switching of pre-existing M1 macrophages within the site of injury. The differences between these groups have not been investigated, but have major implications for understanding and treating pathologies characterized by deficient M2 activation, such as chronic wounds, which also exhibit diminished M1 macrophage behavior. This study investigated the influence of prior M1 activation on human macrophage polarization to an M2 phenotype in response to IL-4 treatment in vitro. Compared to unactivated (M0) macrophages, M1 macrophages upregulated several receptors that promote the M2 phenotype, including the primary receptor for IL-4. M1 activation also changed the macrophage response to treatment with IL-4, generating an M2-like phenotype with a distinct gene and protein expression signature compared to M2 macrophages prepared directly from M0 macrophages. Functionally, compared to M0-derived M2 macrophages, M1-derived M2 macrophages demonstrated increased migratory response to SDF-1α, and conditioned media from these macrophages promoted increased recruitment of endothelial cells in transwell assays. Together, these findings indicate the importance of prior M1 activation in regulating subsequent M2 behavior, and suggest that augmentation of M1 behavior may be a therapeutic target in dysfunctional tissue repair.

Download Full-text

GDF3 Protects Mice against Sepsis-Induced Cardiac Dysfunction and Mortality by Suppression of Macrophage Pro-Inflammatory Phenotype

Cells ◽

10.3390/cells9010120 ◽

2020 ◽

Vol 9 (1) ◽

pp. 120 ◽

Cited By ~ 5

Author(s):

Lu Wang ◽

Yutian Li ◽

Xiaohong Wang ◽

Peng Wang ◽

Kobina Essandoh ◽

...

Keyword(s):

Inflammatory Response ◽

Macrophage Polarization ◽

Endotoxic Shock ◽

Receptor Protein ◽

Regulatory Function ◽

Acute Administration ◽

M1 Macrophage ◽

Smad3 Phosphorylation ◽

Anti Inflammatory ◽

Inflammatory Macrophages

Macrophages are critical for regulation of inflammatory response during endotoxemia and septic shock. However, the mediators underlying their regulatory function remain obscure. Growth differentiation factor 3 (GDF3), a member of transforming growth factor beta (TGF-β) superfamily, has been implicated in inflammatory response. Nonetheless, the role of GDF3 in macrophage-regulated endotoxemia/sepsis is unknown. Here, we show that serum GDF3 levels in septic patients are elevated and strongly correlate with severity of sepsis and 28-day mortality. Interestingly, macrophages treated with recombinant GDF3 protein (rGDF3) exhibit greatly reduced production of pro-inflammatory cytokines, comparing to controls upon endotoxin challenge. Moreover, acute administration of rGDF3 to endotoxin-treated mice suppresses macrophage infiltration to the heart, attenuates systemic and cardiac inflammation with less pro-inflammatory macrophages (M1) and more anti-inflammatory macrophages (M2), as well as prolongs mouse survival. Mechanistically, GDF3 is able to activate Smad2/Smad3 phosphorylation, and consequently inhibits the expression of nod-like receptor protein-3 (NLRP3) in macrophages. Accordingly, blockade of Smad2/Smad3 phosphorylation with SB431542 significantly offsets rGDF3-mediated anti-inflammatory effects. Taken together, this study uncovers that GDF3, as a novel sepsis-associated factor, may have a dual role in the pathophysiology of sepsis. Acute administration of rGDF3 into endotoxic shock mice could increase survival outcome and improve cardiac function through anti-inflammatory response by suppression of M1 macrophage phenotype. However, constitutive high levels of GDF3 in human sepsis patients are associated with lethality, suggesting that GDF3 may promote macrophage polarization toward M2 phenotype which could lead to immunosuppression.

Download Full-text

Bacteriophage Therapy of Chronic Nonhealing Wound: Clinical Study

The International Journal of Lower Extremity Wounds ◽

10.1177/1534734619835115 ◽

2019 ◽

Vol 18 (2) ◽

pp. 171-175 ◽

Cited By ~ 14

Author(s):

Pooja Gupta ◽

Hari Shankar Singh ◽

Vijay K. Shukla ◽

Gopal Nath ◽

Satyanam Kumar Bhartiya

Keyword(s):

Wound Healing ◽

Biofilm Formation ◽

Bacterial Infections ◽

Chronic Wounds ◽

Bacterial Count ◽

Complete Healing ◽

Resistant Bacteria ◽

Bacteriophage Therapy ◽

Inflammatory Phase ◽

Drug Resistant Bacteria

Background: A chronic wound usually results due to halt in the inflammatory phase of wound healing. Bacterial infections and biofilm formation are considered to be the basic cause of it. Chronic wounds significantly impair the quality of life. Antibiotics are now failing due to biofilm formation emergence of drug-resistant bacteria. Objective: This study aims to see the effect of bacteriophage therapy in chronic nonhealing wound infected with the following bacteria: Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa. Subject: Patients with chronic nonhealing wound not responding to conventional local debridement and antibiotic therapy were included in the study. The age of patients ranged between 12 and 60 years. Method: A total of 20 patients selected and tissue biopsies and wound swabs were taken for isolation of the bacteria. After confirmation of organism, a cocktail of customized bacteriophages was topically applied over the wound on alternate days till the wound surface became microbiologically sterile. Mean bacterial count and clinical assessment were done and compared at the time of presentation and after 3 and 5 doses of application. Results: A significant improvement was observed in the wound healing, and there were no signs of infection clinically and microbiologically after 3 to 5 doses of topical bacteriophage therapy. Seven patients achieved complete healing on day21 during follow up while in others healthy margins and healthy granulation tissue were observed. Conclusion: Topical bacteriophage application may be quite effective therapy for the treatment of chronic nonhealing wounds.

Download Full-text

The Vitamin D Receptor Regulates Tissue Resident Macrophage Response to Injury

Endocrinology ◽

10.1210/en.2016-1474 ◽

2016 ◽

Vol 157 (10) ◽

pp. 4066-4075 ◽

Cited By ~ 18

Author(s):

Lige Song ◽

Garyfallia Papaioannou ◽

Hengguang Zhao ◽

Hilary F. Luderer ◽

Christine Miller ◽

...

Keyword(s):

Vitamin D ◽

Wound Healing ◽

Vitamin D Receptor ◽

Macrophage Polarization ◽

Critical Role ◽

Dermal Fibroblasts ◽

Wild Type ◽

Macrophage Response ◽

Protein Levels ◽

Inflammatory Phase

Ligand-dependent actions of the vitamin D receptor (VDR) play a pleiotropic role in the regulation of innate and adaptive immunity. The liganded VDR is required for recruitment of macrophages during the inflammatory phase of cutaneous wound healing. Although the number of macrophages in the granulation tissue 2 days after wounding is markedly reduced in VDR knockout (KO) compared with wild-type mice, VDR ablation does not alter macrophage polarization. Parabiosis studies demonstrate that circulatory chimerism with wild-type mice is unable to rescue the macrophage defect in the wounds of VDR KO mice and reveal that wound macrophages are of local origin, regardless of VDR status. Wound cytokine analyses demonstrated a decrease in macrophage colony-stimulating factor (M-CSF) protein levels in VDR KO mice. Consistent with this, induction of M-CSF gene expression by TGFβ and 1,25-dihydroxyvitamin D was impaired in dermal fibroblasts isolated from VDR KO mice. Because M-CSF is important for macrophage self-renewal, studies were performed to evaluate the response of tissue resident macrophages to this cytokine. A decrease in M-CSF induced proliferation and cyclin D1 expression was observed in peritoneal resident macrophages isolated from VDR KO mice, suggesting an intrinsic macrophage abnormality. Consistent with this, wound-healing assays in mice with macrophage-specific VDR ablation demonstrate that a normal wound microenvironment cannot compensate for the absence of the VDR in macrophages and thus confirm a critical role for the macrophage VDR in the inflammatory response to injury.

Download Full-text