Review
Antifibrotic and Anti-Inflammatory Actions of α-Melanocytic
Hormone: New Roles for an Old Player
Roshan Dinparastisaleh 1 and Mehdi Mirsaeidi 2,*
Citation: Dinparastisaleh, R.;
Mirsaeidi, M. Antifibrotic and
Anti-Inflammatory Actions of
α-Melanocytic Hormone: New Roles
for an Old Player. Pharmaceuticals
2021, 14, 45.
https://doi.org/
10.3390/ph14010045
Received: 25 December 2020
Accepted: 6 January 2021
Published: 8 January 2021
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
1 Division of Pulmonary and Critical Care Medicine, Johns Hopkins University, Baltimore, MD 21218, USA;
rdin...@jhmi.edu
2 Division of Pulmonary and Critical Care, University of Miami, Miami, FL 33146, USA
* Correspondence:
msm...@med.miami.edu; Tel.:
+1-305-243-1377
Abstract: The melanocortin system encompasses melanocortin peptides, five receptors, and two
endogenous antagonists. Besides pigmentary effects generated by α-Melanocytic Hormone (αMSH), new physiologic roles in sexual activity, exocrine secretion, energy homeostasis, as well
as immunomodulatory actions, exerted by melanocortins, have been described recently. Among
the most common and burdensome consequences of chronic inflammation is the development
of fibrosis. Depending on the regenerative capacity of the affected tissue and the quality of the
inflammatory response, the outcome is not always perfect, with the development of some fibrosis.
Despite the heterogeneous etiology and clinical presentations, fibrosis in many pathological states
follows the same path of activation or migration of fibroblasts, and the differentiation of fibroblasts
to myofibroblasts, which produce collagen and α-SMA in fibrosing tissue. The melanocortin agonists
might have favorable effects on the trajectories leading from tissue injury to inflammation, from
inflammation to fibrosis, and from fibrosis to organ dysfunction. In this review we briefly summarized
the data on structure, receptor signaling, and anti-inflammatory and anti-fibrotic properties of αMSH and proposed that α-MSH analogues might be promising future therapeutic candidates for
inflammatory and fibrotic diseases, regarding their favorable safety profile.
Keywords: α-MSH; melanocortins; α-MSH analogues; anti-inflammatory; anti-fibrotic; MC1R;
lung fibrosis
1. Introduction
The melanocortin system encompasses melanocortins, five transmembrane G proteincoupled receptors, plus two endogenous antagonists: the agouti-signaling protein and
agouti-related peptide. The melanocortins comprise adrenocorticotropic hormone (ACTH),
α-, β-, and γ-melanocyte-stimulating hormones (α-, β-, γ-MSHs), which are generated from
proopiomelanocortin (POMC) processing. Besides steroidogenesis and pigmentary effects
caused by ACTH and α-MSH, respectively, new roles in energy homeostasis, reproductive
system functions, exocrine glands secretion, immunomodulatory, and anti-inflammatory,
have been explained [1,2].
Our knowledge of immunomodulatory effects of melanocortins has progressed, since
the first clinical experiment of ACTH in arthritis and rheumatic fever patients conducted by
Hench et al., in 1949 [3]. Prior to identification and cloning of melanocortin receptor family
(MCR), ACTH-induced improvement in subjects with arthritis was presumed to be due
to activation of the hypothalamus-pituitary-adrenal (HPA) axis and cortisol production.
Today, 53 years later, Getting et al. showed that melanocortin-3 receptor (MC3R) signaling,
triggered by ACTH, regardless of steroid synthesis, was likewise responsible for ACTH
effectiveness in inflammatory arthritis and proposed MC3R agonists as novel therapeutics
for chronic inflammatory diseases [4]. This study opened new perspectives on the role
of melanocortins in inflammation, and considering their receptors as potential targets for
future anti-inflammatory therapies.
Pharmaceuticals 2021, 14, 45.
https://doi.org/10.3390/ph14010045 https://www.mdpi.com/journal/pharmaceuticals
Pharmaceuticals 2021, 14, 45 2 of 20
One of the most common and burdensome consequences of chronic inflammation
is the development of fibrosis. Depending on the regenerative capacity of the involved
tissue and the quality of the inflammatory response, the outcome is not always perfect,
with development of some fibrosis. Disease states in which fibrosis is the leading cause of
mortality and morbidity encompass a broad range of illnesses. These include pulmonary
fibrosis, liver fibrosis and cirrhosis, chronic kidney disease, myocardial infarction, systemic
autoimmune diseases such as systemic sclerosis, etc. Despite the heterogeneous etiology
and clinical presentations, fibrosis in many pathological states follow the same path of
activation or migration of fibroblasts, and the differentiation of fibroblasts to myofibroblasts,
which produce collagen and α-smooth muscle actin (α-SMA) in fibrosing tissue [5–7].
In this review we reported anti-inflammatory, anti-fibrotic and regenerative properties
of melanocortins.
2. Melanocortins
2.1. Ligands
The melanocortins are derived from processing of POMC [8]. Interestingly, cloning
and sequencing has revealed POMC in “lamprey”, the most ancient vertebrate, that shares
similarities to those of higher vertebrates, implying that POMC has an ancient linage,
likely dating back millions of years (Reviewed in [9]). Besides pituitary, melanocytes,
keratinocytes, and central nervous system (CNS) where POMC was discovered, POMC
messenger RNA (mRNA) has been recognized in immune cells such as monocytes and lymphocytes which suggests immunomodulatory roles for POMC-derived peptides [4,10,11].
Proprotein convertase-1 (PC1) catalyzes cleavage of POMC to generate ACTH, and PC2
mediates MSH synthesis in pars intermedia of pituitary, CNS, skin and hair follicles [12].
Each melanocortin peptide is rectified from a different region of POMC. γ-MSH is obtained from the amino-terminus, while α-MSH and ACTH are cleaved from the middle
part. β-MSH, β-LPH, γ-LPH, and β-Endorphin are processed from the carboxy-terminal
region of POMC [13]. Each of the melanocortin ligands shares the conserved sequence of
His-Phe-Arg-Trp (HFRW), which functions as a pharmacophore for MCR signaling [14,15].
ACTH is a 39 amino acid polypeptide, best recognized for its role in physiological
stress response [16]. ACTH stimulates the glucocorticoid production by triggering cholesterol conversion to pregnenolone in the cortex of adrenal gland [17]. This sequence is
mediated by binding to melanocortin receptor 2 (MC2R), stimulation of the membranebound adenylate cyclase and calcium influx [18,19]. With the emerging evidence showing
the expression of MC2R in tissues other than adrenal cortex, recent literature have shed
light on new roles of ACTH, including lipolytic activity in adipocytes [20], reducing lipid
content of cells by knockdown of MC2R and inhibition of peroxisome proliferator-activated
receptor gamma 2 (PPARγ2) [21], suppressing leptin expression [22], playing a role in
the differentiation of mesenchymal cells [23], regulation of bone mass [24], a role in the
maintenance and repair of the vascular extracellular matrix [25], controlling thymocyte
homeostasis [26], and amelioration of tumor necrosis factor (TNF)-induced acute kidney
injury [27].
α-MSH is composed of 13 amino acids and is best known for its pigmentary effects in
skin [28], but also has been indicated to exert anti-inflammatory and microbicidal effects
discussed below [29–34].
β-MSH and γ-MSH are much less understood, compared with α-MSH and ACTH. It
is shown that intraventricular infusion of γ2-MSH suppresses LPS-induced inflammatory
responses [35]. Getting et al. demonstrated that natural and synthetic ligands for MC3R
(γ2-MSH and synthetic agonist MTII, respectively) in a murine model of experimental gout,
inhibit aggregation of chemokine C-X-C motif ligand 1 (CXCL1), polymorphonuclear cells
(PMNs), and suppress production of interleukin-1 beta (IL-1β), evoked by monosodium
urate crystals in the peritoneal cavity [36]. Similar anti-inflammatory actions of γ-MSH
and β-MSH were detected in other investigations [37–39].
Pharmaceuticals 2021, 14, 45 3 of 20
2.2. Receptors
Effects of melanocortin system ligands are mediated by five transmembrane G proteincoupled receptors, which are named based on the sequence of their cloning. Sequence
comparison of MCRs reveals 38 to 60% identity between these receptors [40]. Table 1 demonstrates the diverse tissue distribution of agonists and antagonists of melanocortin receptors.
These G protein-coupled receptors (GPCRs) when activated, lead to activation of adenylyl
cyclase, which catalyzes the conversion of ATP to cyclic adenosine monophosphate (cAMP)
in cytoplasm. cAMP activates protein kinases (mainly protein kinase A, which ends in the
phosphorylation of cAMP response element-binding protein (CREBP) [40].
Table 1. Melanocortin receptors; tissue distribution, known agonists and antagonists, and their biological effects.
Receptor Tissue Distribution Species Agonist Biologic Effects Antagonist Biologic Effects
MC1R
Present in melanocytes/skin Human [41,42]
α-MSH, ACTH,
β-MSH, γ-MSH
Pigmentation,
anti-inflammatory
[43,44]
Agouti Suppresses melanine
production [45,46]
Present in grey matter Rat [47]
NDP-MSH Anti-inflammatory
[48–50]
BMS-470539
Present in monocyte, macrophage
(including alveolar), lymphocyte,
neutrophil
Human, murine
[51,52]
AP-1189
MC2R
Present in adrenal cortex; absent
in liver, lung, thyroid and kidney
Rhesus
macaque [41]
ACTH Induce
steroidogenesis [18]
GPS1574 Inhibits
steroidogenesis [53,54]
Present in chondrocyte and
osteoblast Human [55,56]
MC3R
Present in brain, placenta; absent
in adrenal, kidney, liver Rat [57]
γ-MSH ≥ ACTH =
β-MSH = α-MSH
Energy equilibrium,
cardiovascular
[58–60]
SHU-9119
Inhibits
anti-inflammatory
effects of γMSH [36]
Present in lung Murine [61]
MTII Anti-inflammatory
[61–66]
D-Trp8-γMSH
Present in macrophage and
monocytes
Murine
[51,52,55–57,67]
AP-214 AVM-127
inhibits
α-MSH-induced
penile erection [68]
AP-1189 (biased
agonist)
MC4R
Present in brain; absent in lung,
liver, kidney, adrenal Rat/Canine [69]
α-MSH = ACTH >
β-MSH > γ-MSH
Energy balance,
erectile function,
cardiovascular
effects [43,70]
AgRP
Inhibitory
cardiovascular effects,
increases food intake
[71,72]
Present in thalamus and
hypothalamus Rat [73]
THIQ
Anti-inflammatory,
inhibits food intake
[74–76]
ML00253764
Ro27-3225
PT-141 Induces erection
[77,78]
MC5R
Present in lung, skeletal muscle,
brain; absent in adrenal
Murine/Human
[79]
α-MSH, ACTH,
β-MSH,
Anti-inflammatory
[80]
Present in B-lymphocytes Mouse [81]
Present in T-lymphocytes Mouse [82] PG-901 Inhibits glucose
uptake [83]
3. Anti-Inflammatory Effects of α-MSH
During the last four decades, many investigations have established that α-MSH has
strong anti-inflammatory effects.
3.1. Fever and Multiple Organ Dysfunction Syndrome (MODS)
In 1981, Glyn and Lipton showed that five micrograms of intravenous (IV) or intracerebroventricular α-MSH reduces fever produced by leukocytic pyrogen in rabbits [84].
Pharmaceuticals 2021, 14, 45 4 of 20
Thereafter, other studies further confirmed the antipyretic effects of α-MSH in guinea pigs
and squirrel monkeys [85,86].
In another study, Bitto et al. showed that intraperitoneal injection of NDP-α-MSH
(340 µk/kg) in LPS-induced MODS model significantly decreased expression of tumor
necrosis factor-α (TNF-α), increased expression of IL-10, and reduced serum levels of TNFα and improved survival [87]. Other investigations have further confirmed the therapeutic
efficacy of α-MSH in septic shock, systemic inflammatory response syndrome, and cardiac
arrest [64,88–92].
3.2. α-MSH and the Respiratory System
Recent analyses have recognized expression of the MC1R and MC3R in alveolar
macrophages in mice [61,93]. α-MSH inhibited leukocyte migration to the lungs in a
lypopolysaccharide-induced acute lung injury in rats [94]. Deng et al. investigated the
protective properties of 25 µg of IV α-MSH at zero, 8, and 16 h after clamping renal arteries
for 40 min and reperfusion in a mice acute lung injury model. This study reported that αMSH administration improved tissue injury, inhibited production of intracellular adhesion
molecule-1 (ICAM-1) and TNF-α in lungs, and turned off inflammatory transcription
factors and stress-induced genes [95]. In agreement with previous literature, Miao and coworkers found that 17 mg/kg IV α-MSH has anti-apoptotic effects on vascular endothelial
cells in rat model of acute respiratory distress syndrome [96].
In another study, Colombo et al. examined the effects of 100 µg intraperitoneal (IP)
α-MSH analogue (NDP-α-MSH) on a bleomycin-induced ALI model. After instillation of
1 mg bleomycin into the trachea, ten of the bleomycin recipients received α-MSH analogue.
NDP-α-MSH offset bleomycin-induced edema in lung tissue (lung weight in control vs.
α-MSH group, 5.8 ± 0.5 vs. 3.9 ± 0.1, respectively, p < 0.05) and mediated transcriptional
modifications in genes involved in fluid handling, including activation of Na+/K+ ATPase
and ENaC (epithelial sodium channels). Moreover, decreased expression of TNF-α, IL-6,
transforming growth factor-β, and iNOS was noticed [97].
Raap et al. sensitized mice by three IP injections of 10 µg ovalbumin (OVA) on days 1,
14, and 21. Injections of α-MSH (1 mg/kg body weight) were performed 30 min before sensitization or allergen aerosolization. Mice received 2 allergen challenges implemented by
an exposure to diluted 1% OVA on days 27 and 28. Significant decreases in the serum levels
of allergen-specific immunoglobulin E (p < 0.05), IgG1 (p < 0.05), and IgG2a (p < 0.001)
were identified in intervention group compared to controls [98]. In a similar study, Webering et al. measured α-MSH levels in bronchoalveolar lavage (BAL) fluid of asthmatic
versus non-asthmatic participants in addition of healthy and asthmatic mice (OVA-induced
eosinophilic airway inflammation model). α-MSH was delivered intratracheally and αMSH antibody was injected immediately before each sensitization, for neutralization of
endogenous α-MSH. Results revealed that α-MSH levels were significantly higher in participants with eosinophilic asthma than in healthy subjects. The OVA-induced asthmatic
mice also showed high α-MSH levels in BAL fluid. Additional administration of α-MSH
to OVA-sensitized mice significantly reduced eosinophils and lymphocytosis in BAL as
well as inflammation in airways. This study also revealed MC5R expression in airway
epithelium [99].
Literature have confirmed that C-terminal tripeptide derivatives of α-MSH (e.g.,
KPV (Lys-Pro-Val, the C-terminal sequence of alpha-MSH) and KdPT (Lys-D-Pro-Thr),
possess anti-inflammatory properties with no pigmentary effects [100]. In an immortalized human bronchial epithelial cell model challenged with rhino-syncytial virus, 1 µg
of IV KPV suppressed intracellular and systemic proinflammatory signaling (TNF-α–
dependent NF-κB–driven reporter activity and IL-8, respectively) and reduced the activity
of matrix metalloprotease-9 (MMP-9) responsible for lung remodeling via MC3R, in a
dose-dependent way [101].
Zhang et al. tested the effects of α-MSH in a novel in experimental sarcoid model.
The granuloma model was developed by mycobacterium-challenged peripheral blood
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mononuclear cells (PBMCs) obtained from sarcoidosis patients. Both challenged and nonchallenged PBMCs were treated with 10 µM α-MSH daily or saline. RNA-Seq analysis
on 3rd day after exposure to α-MSH revealed significant decrease in IL-1b, IL-1R, IL-8,
IL-12, chemokine C-C ligand 3 (CCL3), CCL4, CCL5, GM-CSF, IFN-γ, and TNF-α levels
in intervention group. They reported a significantly increase expression of p-CREB in αMSH-treated experimental sarcoidosis model. Furthermore, addition of a highly selective
CREB inhibitor (666-15), significantly counterbalanced the effects of α-MSH, suggesting
that CREB phosphorylation is essential for anti-inflammatory effects of α-MSH [31].
3.3. α-MSH and the Eye
In the immune-privileged eye, melanocortins are involved in regulation of inflammatory pathways (cytoprotection) and promoting the immune tolerance [102]. An experiment
conducted by Lee et al. showed that α-MSH treatment of mice with experimental autoimmune uveoretinitis inhibited inflammation and may re-define some aspects of immune
privilege [103]. Other studies also supported this finding [34,104,105].
It has been shown that intravitreal injection of 3 µL of α-MSH at week 1 and 3 after
streptozotocin-induced hyperglycemia inhibited breakdown of blood-retina barrier and
vascular permeability through MC4R signaling in diabetic retinas [106]. Additionally,
10 µg/3µL of intravitreal α-MSH normalized levels of H2O2, reactive oxygen species (ROS),
and the total antioxidant capacity and corrected the aberrant changes in endothelial nitric
oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), intracellular adhesion
molecule 1 (ICAM-1), and TNF-α expression levels in diabetic retinas [107]. Zhang and
co-workers indicated that anti-inflammatory actions of melanocortins in the eye are caused
by under-expression of inflammatory cytokines (i.e., TNF-α and IL-6) and suppression of
the NFκB-dependent signaling pathway [108].
3.4. α-MSH and Gastrointestinal System
KPV can weaken the inflammatory responses in colonic epithelium and intestinal
immune cells and lead to reduction in the incidence of inflammatory bowel diseases (IBD)
in vivo [109–112]. Dalmasso et al. investigated effects of 100 µM KPV on mice with experimental colitis induced by dextran sulfate sodium (DSS) and trinitrobenzene sulfonic
acid (TNBS). As an index of neutrophilic infiltration intestinal myeloperoxidase (MPO)
activity was assessed. They proved that treatment with oral KPV decreased MPO activity
by ~50% and these results were confirmed by hematoxylin and eosine (H&E) examination
of colonic slides. They also proposed a non-receptor-dependent immunoregulatory effect
of KPV, mediated by a transporter normally expressed in the small bowel and induced
in colitis [113]. Another similar study showed that KPV treatment leads earlier recovery
and significantly stronger regain of body weight (to 87.8% ± 2.7%, versus 73.9% ± 3.5%
of the original body weight in the intervention and control mice). Interestingly, on day
14 after the administration of DSS, the body weight of KPV-treated mice had standardized to 102.4–0.9%. treatment with KPV, significantly reduced myeloperoxidase activity
(881.7 ± 215.9 U/mg protein) compared to control group (1835.9 ± 283.8 U/mg protein)
(p < 0.05) [114]. Recently, Xiao et al. tested the effects of targeted hyaluronic acid-based
KPV delivery (HA-NP) in experimental ulcerative colitis mouse model. They proposed
that Hyaluronic acid lysine-proline-valine nanoparticles (HA-KPV-NPs) applies combined
mechanisms against ulcerative colitis by both enhancing mucosal healing and regulating
inflammatory responses. Furthermore, oral HA-KPV-NPs encapsulated in a hydrogel
exhibited a higher potency to prevent epithelial injury [115]. Targeted theranostic NDP-αMSH delivery in IBD has further opened up possibilities for therapeutic and selective use
in other inflammatory disease, such as the lung inflammation found in COVID-19 [116].
3.5. α-MSH and Nervous System
Min et al. generated mature monocyte-derived dendritic cells (MoDCs), using TNF-α.
Then, they treated MoDCs with different dosages of α-MSH (10−14–10−6 M) to assess the
Pharmaceuticals 2021, 14, 45 6 of 20
regulatory impact of α-MSH on TNF-α-DCs. The downregulation of CD86, CD83, IL-12
and over-expression of IL-10 was observed in all doses after treatment with α-MSH. They
showed upregulation of Annexin A1 after administration of α-MSH, suggesting an inhibitory effect of α-MSH on TNF-α-induced MoDC maturation via the upregulation of Annexin A1 [117]. In vitro studies favor a possible neuroprotective role for melanocortins, as
they suppressed NF-kB activation in TNF-α-activated Schwann cells or lipopolysaccharideactivated glioma cells [29,118–120]. Mykicki et al. showed that mice treated systemically
with α-MSH in two-day intervals were immune from developing clinical signs of experimental autoimmune encephalitis (EAE), and this effect was associated with reduced inflammatory foci and decreased central nervous system demyelination (p < 0.0001, p = 0.0051).
T-helper 1 and T-helper 17 cells were diminished in the CNS and in the cervical lymphatics from α-MSH-treated animals compared with controls. They found that NDP-MSH
induced functional regulatory T cells through MC1R signaling, leading to alleviation of
EAE in treated mice [121]. Carniglia et al. evaluated the effect of NDP-α-MSH on PPARβ and PPAR-γ expression in rat’s astrocytes and microglial cells. They recognized that
microglial cells of rat express MC4R and treatment with NDP-α-MSH strongly enhances
PPAR-γ expression and decreases PPAR-β expression in both microglia and astrocytes [122].
Wang et al. produced a novel Tat protein(TAT)-human serum albumin (HAS)–α-MSH
fusion protein. They showed that the fusion protein TAT-HSA-α-MSH can successfully
cross the blood-brain barrier after intraperitoneal injection. The NF-κB driven reporter
assay in vitro showed that TAT-HSA-α-MSH strongly suppressed NF-κB in the glioma
cell line. In LPS-induced CNS inflammation in mice, HSA-α-MSH, when given intraperitoneally, markedly attenuated TNF-α production. These results confirmed that the fusion
protein TAT-HSA-α-MSH exerts dominant anti-inflammatory activities in the nervous
system after being delivered into the animal’s brain [29]. Effectiveness of melanocortins in
neuroinflammation is further confirmed by other studies [30,118,119,123,124].
Recently, studies have suggested that melanocortins are safe and effective candidates
for treating subarachnoid hemorrhage- and intracerebral hemorrhage- related complications [125,126].
3.6. α-MSH and Skin
Kleiner et al. showed that the analogues of MC1R and MC5R have regulatory effects
on IgE-mediated allergic inflammation [127]. Other studies showed similar therapeutic
effects of MC1R agonists on atopic dermatitis mouse model, and induction of regulatory
T-cells in vitro and in vivo which led to inhibitory effect on psoriasis progression in a
mouse model [128–130].
3.7. α-MSH and the Musculoskeletal System
Capsoni et al. evaluated the role of melanocortin in regulating production of inflammatory cytokines, metalloproteinases (MMPs), tissue inhibitors of MMPs (TIMPs), iNOS,
and nitric oxide (NO) in response to IL-1β and TNF-α in synovial chondrocytes. They
demonstrated increased TIMP-3 gene expression and downmodulation of TNF-α-induced
stimulation of synovial chondrocytes [131]. Immuno-regulatory and anti-degenerative
actions of melanocortins in joints have been extensively studied [33,132–135].
Synovial fluid α-MSH levels have shown a negative independent correlation with
disease severity in individuals with post-traumatic osteoarthritis and application of local
α-MSH has been suggested as a potential adjuvant therapy [136]. Interestingly, two studies
have revealed a therapeutic property of melanocortins (including ACTH), in osteonecrosis
of bones [137,138].
3.8. Other Anti-Inflammatory Actions
Various studies have proposed that α-MSH protects against ischemia and reperfusion
injuries in kidney, testes, myocardium, intestines, and CNS [139–143].
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The melanocortins show therapeutic characteristics in atherosclerosis by preventing plaque rupture and improving endothelial cell function, which may suggest a novel
therapeutic target for atherosclerosis [135].
Beneficial effects of α-MSH on endothelial production of pro-inflammatory substances
for applications in implantable intravascular devices such as pacemakers, have been
reported [144,145]. One study reported allograft protection in organ recipient mice [146].
Liu et al. attempted interpreting the role of α-MSH in adipose tissue inflammation
and the interactions with forkhead box proteins. By suppressing forkhead box protein
expression, α-MSH could dampen LPS-induced inflammation accompanied with increased
anti-inflammatory mediator release and decreased inflammatory products [147].
4. Mechanisms of Anti-Inflammatory Effects of α-MSH
The mechanisms of anti-inflammatory actions have been extensively studied (briefly
listed below and demonstrated in Figure 1).
4.1. Inhibition of NF-κB
Manna and Aggrawal first described that α-MSH nullified TNF-mediated NF-κB
activation in a concentration-dependent fashion and opposed NF-κB activation induced
by LPS. NF-κB is an evolutionarily conserved transcription factor that regulates immune
responses. Its role as a master regulator of the inflammatory response stems from its critical
function in regulating the expression of hundreds of immune relevant genes, particularly
those encoding proinflammatory mediators, in addition to other genes important for
the development of the immune system. The inhibitory effect of α-MSH appears to
be performed through cAMP production, as inhibitors of adenylyl cyclase and of PKA
antagonized its anti-inflammatory effects [148]. Subsequently, similar effects on various
cell types, including pulmonary epithelial cells, were reported (Reviewed in [4]). Several
mechanisms have been proposed to explain how cAMP interferes with the NF-κB signaling
cascade. These mechanisms include effects of cAMP signaling on IκB kinase activation
and cytoplasmic IκB levels, posttranslational cAMP-induced Rel proteins modification
induced, effects of cAMP on NF-κB dimer composition, etc., (reviewed in [149]). cAMPindependent inhibition of nuclear translocation of NF-κB through MC1R signaling has also
been explained [150].
4.2. Suppression of Proinflammatory Cytokines
α-MSH acts as an anti-inflammatory substance by suppression of proinflammatory
mediators such as TNF-α, interferon-γ (IFN-γ), IL-1, IL-6, IL-8 and induction of cytokine
suppression by production of IL-10 [51,151,152]. The present evidence indicates that α-MSH
has a key role in the regulation of TNF-α and nitric oxide in monocytes and macrophages.
Antibodies against MC1R increased TNF-α in non-challenged macrophages, blunted the
hindering effect of α-MSH, and enhanced TNF-α production in LPS-challenged cells [51].
IL-8 gene transcription demands activation of the combination of both NF-κB and activating
protein-1 (AP-1), or that of NF-κB, and another transcription factor, NFκβ/Interleukin6 [153]. It has been shown that AP-1 in dermal fibroblasts can be modified by α-MSH, and
that this effect appears to be co-stimulus-dependent [154]. Suppression of IL-1 and IL-6
mRNA expression by α-MSH might be due to suppression of the NF-κB signaling pathway,
which is the key factor in production of these proinflammatory cytokines [155]. IL-10 is an
anti-inflammatory cytokine that plays a key role in maintaining the balance of immune
responses and resolves inflammation and blunts unwanted tissue injury [156]. Toll like
receptor (TLR) signaling results in stimulation of NF-κB and mitogen-activated protein
kinase (MAPK) pathways (ERK1/2 and p38), which subsequently lead to production of
IL-10. MAPKs activate mitogen- and stress-activated kinase 1 (MSK1) and MSK2 and
phosphorylate the transcription factors AP-1 and CREB, which subsequently result in IL-10
expression [157]. Additionally, a cAMP signaling cascade can lead to CREB phosphorylation and transcription of a plethora of genes besides IL-10 [149]. Therefore, CREB has
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a central role in the production of IL-10. α-MSH-mediated cAMP cascade signaling and
subsequent CREB phosphorylation is a possible mechanism for IL-10 production.
4.3. Inhibition of Adhesion Molecules
The ability of α-MSH to suppress expression of intercellular adhesion molecule-1
(ICAM-1) has been explained in murine mast cells. The inhibition of vascular cell adhesion
molecule-1 (VCAM-1) and E-selectin expression were also identified in endothelial cells.
Moreover, α-MSH regulates the expression of co-stimulatory molecules essential for antigen
presentation (CD40 and CD86) in monocytes and DCs [107,158–161].
4.4. Suppression of Non-Cytokine Inflammatory Mediators
α-MSH has been reported to suppress proinflammatory non-cytokine regulators
such as nitric oxide (NO), prostaglandin E (PGE), and ROS. The capacity of α-MSH
in damping stimulated nitric oxide synthesis and iNOS expression was first reported
in MC1R expressing mice macrophages [162]. Thereafter, similar results have been described in Raw 264.7 cells, helper T-cells, PBMCs, melanoma cells, mice microglia, and
astrocytes [151,152,163,164]. In Mandrika et al.’s experiment, forskolin (a pharmacological
agent that raises intracellular cAMP) was also able to inhibit nitric oxide production without affecting the translocation of active IκB-free NF-κB, suggesting that cAMP generation
may inhibit NO synthesis, independently of NF-kB signaling [150]. Oktar et al. showed
that non-selective cyclooxynegase (COX) inhibitor indomethacin antagonized the effect
of melanocortins, at a dose which did not influence the high lucigenin (a chemiluminescent probe used to detect superoxide production) chemiluminescence value of stimulated
PMNs. However, the inhibitory effects of α-MSH in lucigenin values were not altered in
cells treated with more selective COX inhibitors, like ketorolac or nimesulide. Although the
mechanisms of interaction between α-MSH and COX are not clear yet, this study proposed
that α-MSH prevents superoxide synthesis PMNs and that both COX1 and COX2 are
involved in this effect [165].
4.5. Induction of Regulatory T Cells (Tregs)
α-MSH has been reported to induce CD4 and CD25 positive regulatory T cells, which
have a central role in maintenance of immune tolerance [166–168]. α-MSH-treated Tregs
have been shown to inhibit IFN-γ and IL-10 synthesis but increase TGF-β1 synthesis [82].
α-MSH-induced immune regulation arises from converting effector T lymphocytes. These
regulatory cells are CD25+ CD4+, CTLA4+, CD44+, CD62L+, and latency associated peptide (LAP) positive. α-MSH induces TGF-β synthesis but does not exert immunoregulation
in naive T-cells. This indicates that the immunoregulatory actions of α-MSH on T-cells
are confined to antigen-experienced effector T-cells. Thus, it has been suggested that it
is possible to use melanocortins to induce antigen-specific regulatory T-cells which aim
autoimmune diseases [82,168–170]. The immunomodulatory effects of α-MSH in T-cells is
mediated through MC5R, which subsequently activates the Janus kinase 2 (JAK2), signal
transducer and activator of transcription 1 (STAT1), or ERK pathways in immune cells
and lead to cell differentiation and cytokine production [81]. Emerging evidence suggests
a role for CREB in TGF-β/FoxP3–dependent Treg induction and maintenance (reviewed
in [157]).
4.6. Promotion of Efferocytosis
Effective clearance of apoptotic cells by macrophages and other phagocytes is a
fundamental component in homeostasis and resolution of inflammation, termed efferocytosis [171]. For many decades, resolution of inflammation was regarded as a passive process,
simply including removal of inflammatory stimulus, stopping production of inflammatory mediators, and the inhibition of further chemotaxis to injury site. Later, Sehran and
Savill proposed that resolution of inflammation is an active process which also consists
of signaling pathways associated with apoptosis, efferocytosis, and reprogramming of
Pharmaceuticals 2021, 14, 45 9 of 20
macrophages to ensure a regaining of the preinflammatory status [172] Montero–Mendelez
and co-workers evaluated the effect of AP214 (α-MSH analogue with a higher affinity to
MC1R and MC3R) on phagocytosis and apoptotic neutrophils in mouse peritonitis model.
They showed that 400–800 µg/kg body weight of i.p. AP214, can increase phagocytosis of
apoptotic neutrophils by 70 and 30% in In Vitro and In Vivo, respectively. They proposed a
role for MC3R in efferocytosis [65].
Pharmaceuticals 2021, 14, x FOR PEER REVIEW 10 of 21
Figure 1. Simplified role of MC1R signaling in anti-inflammatory actions of α-MSH. MC1R activates adenylyl cyclase and
generates intracellular cAMP, which activates protein kinases (C and A). This leads to activation of MAPK and JAK and
STAT pathways. In the nucleus, the catalytic subunits can phosphorylate different substrates, the best known of which is
the transcription factor CREB. CREB is involved in transcription of anti-inflammatory mediators. Alternatively, protein
kinase activation can lead to increased cytoplasmic inhibitor of κB (IκB) through blocking IκB kinase, inhibition of IκB
ubiquitinylation, etc. This leads to inhibition of NF-κB and decreased expression of downstream proinflammatory genes.
5. α-. MSH and Tissue Repair and Remodeling
Replacement of damaged tissue with new living tissue is referred to as tissue repair
(healing). The basic cellular and molecular mechanisms underlying restoration of tissue
architecture and function after an injury and its failure to heal are still poorly understood,
and treatments are dissatisfying. Defective tissue repair after trauma, surgery, and acute
or chronic disease states affect millions of people worldwide each year and arises from
malregulation of tissue repair responses, including inflammation, angiogenesis, matrix
deposition and degradation, and cell recruitment. Impaired repair can lead to fibrosis and
organ dysfunction. Several possible anti-fibrotic properties of α-MSH are discussed below
(summarized in Figure 2).
Figure 1. Simplified role of MC1R signaling in anti-inflammatory actions of α-MSH. MC1R activates adenylyl cyclase and
generates intracellular cAMP, which activates protein kinases (C and A). This leads to activation of MAPK and JAK and
STAT pathways. In the nucleus, the catalytic subunits can phosphorylate different substrates, the best known of which is
the transcription factor CREB. CREB is involved in transcription of anti-inflammatory mediators. Alternatively, protein
kinase activation can lead to increased cytoplasmic inhibitor of κB (IκB) through blocking IκB kinase, inhibition of IκB
ubiquitinylation, etc. This leads to inhibition of NF-κB and decreased expression of downstream proinflammatory genes.
5. α-MSH and Tissue Repair and Remodeling
Replacement of damaged tissue with new living tissue is referred to as tissue repair
(healing). The basic cellular and molecular mechanisms underlying restoration of tissue
Pharmaceuticals 2021, 14, 45 10 of 20
architecture and function after an injury and its failure to heal are still poorly understood,
and treatments are dissatisfying. Defective tissue repair after trauma, surgery, and acute
or chronic disease states affect millions of people worldwide each year and arises from
malregulation of tissue repair responses, including inflammation, angiogenesis, matrix
deposition and degradation, and cell recruitment. Impaired repair can lead to fibrosis and
organ dysfunction. Several possible anti-fibrotic properties of α-MSH are discussed below
(summarized in Figure 2).
Pharmaceuticals 2021, 14, x FOR PEER REVIEW 13 of 21
Figure 2. Possible anti-fibrotic properties of α-MSH (see text).
6. Future Perspectives
Tissue injury and inflammation are crucial triggers for either regeneration or fibrosis.
Melanocortin agonists might have favorable effects on the processes leading from injury
to tissue inflammation, from inflammation to tissue fibrosis, and from fibrosis to organ
dysfunction. α-MSH may have significant potentials in inflammation control and repairment process in numerous inflammatory lung diseases including sarcoidosis, interstitial
lung disease, and COVID-19 related pulmonary fibrosis, with fewer safety concerns than
other immunomodulatory medications. Validation via further investigation is recommended to prove the therapeutic properties of MSH agonists in lung diseases.
Author Contributions: Conceptualization, M.M.; methodology and data collection, R.D.; writing—original draft preparation, R.D.; writing—review and editing, M.M. All authors have read
and agreed to the published version of the manuscript.
Funding: This research received no external funding.
Institutional Review Board Statement: Not Applicable.
Informed Consent Statement: Not Applicable.
Data Availability Statement: Not Applicable.
Acknowledgments: Authors would like to thank April Mann for critical review and edits.
Conflicts of Interest: Dinparastisaleh declares no conflicts of interest. Mirsaeidi is Advisor Board
of Mallinckrodt and received grant to study sarcoidosis.
References
1. Tao, Y.X. Melanocortin receptors. Biochim. Biophys. Acta Mol. Basis Dis. 2017, 1863, 2411–2413, doi:10.1016/j.bbadis.2017.08.001.
2. Gallo-Payet, N. 60 YEARS OF POMC: Adrenal and extra-adrenal functions of ACTH. J. Mol. Endocrinol. 2016, 56, T135–T156,
doi:10.1530/jme-15-0257.
3. Hench, P.S.; Kendall, E.C.; Slocumb, C.H.; Polley, H.F. The effect of a hormone of the adrenal cortex (17-hydroxy-11-dehydrocorticosterone: Compound E) and of pituitary adrenocortical hormone in arthritis: Preliminary report. Ann. Rheum. Dis. 1949,
8, 97–104, doi:10.1136/ard.8.2.97.
4. Wang, W.; Guo, D.Y.; Lin, Y.J.; Tao, Y.X. Melanocortin Regulation of Inflammation. Front. Endocrinol. 2019, 10, 683,
doi:10.3389/fendo.2019.00683.
5. Smith, P.C.; Martínez, C.; Martínez, J.; McCulloch, C.A. Role of Fibroblast Populations in Periodontal Wound Healing and Tissue Remodeling. Front. Physiol. 2019, 10, 270, doi:10.3389/fphys.2019.00270.
Figure 2. Possible anti-fibrotic properties of α-MSH (see text).
Over the past few decades, transforming growth factor-β (TGF-β) may have been
the best studied cytokine in fibrosis and has been a prototypical “profibrotic” mediator [173]. TGF-β1 regulates fibroblast recruitment to sites of tissue injury and mediates
fibroblast-to-myofibroblast differentiation [174]. Bohm et al. showed that α-MSH reduces
the extracellular levels of procollagens I, III, and V by 70% in human dermal fibroblasts
and strongly reverses the encouraging actions of TGF-β1 on the extracellular matrix (ECM)
collagen levels, with the most predominant effects on collagens I and III and more blunted
effects on collagen V [175]. α-MSH inhibits IL-1β-mediated IL-8 secretion [154], exerts
cytoprotective effects [176], and suppresses experimentally induced cutaneous fibrosis in
dermal fibroblasts [177]. Kokot et al. developed an animal model of scleroderma induced
by subcutaneous injection of bleomycin and treated the mice with 5 µg/day of subcutaneous α-MSH for 21 days. The administration of α-MSH inhibited expression of type I
and type III collagens induced by bleomycin [177]. Bleomycin-treated mice with defective
α-MSH and MC1R signaling showed increased cutaneous collagen type I mRNA levels
accompanying cutaneous fibrosis [178]. In addition, MC1R mRNA expression levels in
keloid fibroblast cell lines were reduced to less than 50%, in comparison with the normal
fibroblasts [157], and α-MSH administration to keloid fibroblasts did not inhibit TGF-β1-
mediated collagen synthesis and myofibroblast differentiation as much as in the control
group, seemingly because of defective expression of MC1R in keloid fibroblasts [179]. de
Souza et al. demonstrated that intraperitoneal administration of 1 mg/kg α-MSH immediately before skin wounding significantly reduces the quantity of leukocytes, mast cells, and
fibroblasts at the site of injury. α-MSH reduced scar area and enhanced the orientation of
the collagen fibers, suggesting it may command the healing process to a more regeneration
and less scar formation pathways [180].
Hepatic fibrosis arises from escalating deposition of extracellular matrix components
in the hepatic parenchyma due to recurring tissue injury. Lee et al. developed a mouse
model of hepatic fibrosis with administration of carbon tetrachloride (CCl4) for 10 weeks.
α-MSH expression vector was delivered via electro-permeabilization after full-blown liver
fibrosis. Histologic examination and assessment of extracellular matrix contents of the
Pharmaceuticals 2021, 14, 45 11 of 20
livers revealed that transfected animals markedly reversed CCL4-induced fibrosis, compared to untreated animals (collagen content in intervention group was 23.7 ± 4.7 vs.
59.7 ± 5.0 µg/mg in untreated animals, p < 0.01). The over-expression of TGF-β1, collagen
1, fibronectin, TNF-α, ICAM-1, and VCAM-1 mRNA were reported in the experimental models of hepatic fibrosis. Gene therapy with α-MSH significantly attenuated this
over-expression. They further showed that the intervention reversed established hepatic cirrhosis by increasing MMP activity and decrease in their tissue inhibitors (TIMP),
suggesting that extracellular matrix metabolism modification might play a role in the
tissue repair properties of α-MSH [181]. Wang et al. introduced a liver fibrosis model
induced by chronic thioacetamide (selective hepatotoxin) administration and investigated
the effects of α-MSH gene therapy on tissue remodeling. Hepatic ECM collagen content
in the treated animals was 32.2 ± 6.2 µg/mg while it was 71.6 ± 10.0 µg/mg in control
group (p < 0.01). Treatment significantly inhibited TGF-β1, procollagen I, TNF-α, ICAM1, VCAM-1 and TIMP-1 mRNA over-expression in intervention group. They proposed
that the collagenolytic actions of α-MSH can be due to MMP and TIMP balance modulation [182]. Lonati et al. aimed to experiment if treatment with melanocortin adjusts
tissue remodeling after performing partial hepatectomy (PH) or sham procedure in rats.
Immediately prior to surgery the intervention group received a single dose of NDP-MSH,
while controls received only saline. RT-PCR analyses demonstrated that NDP-MSH altered
the expression of a substantial proportion of transcripts, including multiple cytokines and
their receptors. The critical signaling pathway IL-6/STAT/SOC was significantly enhanced
by the α-MSH agonist [183]. Another older study had shown the regenerative effects of
α-MSH on hepatectomized rats [184].
Xu and co-workers investigated the anti-fibrotic properties of an α-MSH analogue
(STY39) on a bleomycin-induced lung fibrosis murine model. Mice received STY39 (0.625,
1.25, or 2.5 mg/kg, IP) once daily for 2 weeks. multiple items associated with inflammatory
pathways, extracellular matrix (ECM) components, myofibroblast proliferation, and tissue
remodeling were assessed. They found that α-MSH analogue predominantly improved
the survival rates of animals with severe bleomycin-induced lung fibrosis, opposed weight
loss, reduced the expression of types I and III procollagen mRNA, blunted myofibroblast
differentiation and proliferation, and reduced pulmonary fibrosis. Further evaluation
showed that STY39 administration inhibited neutrophil migration into the lungs, inhibited
the production of local TNF-α, IL-6, macrophage inflammatory protein 2, and TGF-β, and
modified MMP-1/TIMP-1 ratio [185].
Lee et al. evaluated the anti-fibrotic properties of an α-MSH agonist (STY39) on a
cyclosporine-induced tubulointerstitial fibrosis rat model. STY39 counterbalanced the
Bax and TGF-α increase and induced synthesis of anti-apoptotic Bcl2 protein, as well as
inhibition of inflammation and tubulointerstitial renal fibrosis [186].
Verhaagen et al. have explained the effects of α-MSH in nerve regeneration [187]. This
is also confirmed by Dekker et al., who injected 10 µg of α-MSH into rats every 48 h after a
sciatic nerve crush and tested the number of myelinated axons in cross sections of sciatic
nerve at several time points and observed that α-MSH increased the number and diameter
of axons after nerve injury [188]. Later, the effectiveness of α-MSH on peripheral nerve
regeneration was further established [189–191].
Bonfiglio et al. investigated the effects of KPV on corneal wound re-epithelization
in rabbits and the potential role of nitric oxide. Denuded corneas of rabbits were treated
four times a day with KPV 1, 5, or 10 mg/mL (30 mL) or sodium nitroprusside (NO donor)
instantaneously after corneal abrasion while control group only received normal saline.
Then, 60 hours later, 100% of the corneas treated with KPV and SP were fully re-epithelized
while none from untreated rabbits were re-epithelized. They concluded that the availability
of nitric oxide might be of specific importance in therapeutic efficacy of topical KPV in
experimental corneal abrasion model [161]. Pavan et al. evaluated the influence of topical
α-MSH on the healing of corneal wound healing in rats. Topical α-MSH eye-drop in a
concentration of 1 × 10−4 mg/mL improved corneal wound healing significantly, while
Pharmaceuticals 2021, 14, 45 12 of 20
non from control group were healed [192]. Zhang and co-workers tested the anti-fibrotic
effect of α-MSH on TGF-β1-stimulated human Tenon’s capsule fibroblasts (HTFs) since
these fibroblasts play a central role in the initiation and handling of wound healing and
tissue remodeling after trabeculectomy. α-MSH inhibited the proliferation of TGF-β1-
induced HTFs in a concentration-dependent fashion and demonstrated inhibitory effect
on the mRNA expression of type I collagen, TNF-α, ICAM-1, and VCAM-1, which were
upregulated by TGF-β1. They proposed an opposite effect of α-MSH on the disparity
between MMPs and TIMPs compared with TGF-β1 [193].
6. Future Perspectives
Tissue injury and inflammation are crucial triggers for either regeneration or fibrosis.
Melanocortin agonists might have favorable effects on the processes leading from injury to
tissue inflammation, from inflammation to tissue fibrosis, and from fibrosis to organ dysfunction. α-MSH may have significant potentials in inflammation control and repairment
process in numerous inflammatory lung diseases including sarcoidosis, interstitial lung
disease, and COVID-19 related pulmonary fibrosis, with fewer safety concerns than other
immunomodulatory medications. Validation via further investigation is recommended to
prove the therapeutic properties of MSH agonists in lung diseases.
Author Contributions: Conceptualization, M.M.; methodology and data collection, R.D.; writing
—original draft preparation, R.D.; writing—review and editing, M.M. All authors have read and
agreed to the published version of the manuscript.
Funding: This research received no external funding.
Acknowledgments: Authors would like to thank April Mann for critical review and edits.
Conflicts of Interest: Dinparastisaleh declares no conflict of interest. Mirsaeidi is Advisor Board of
Mallinckrodt and received grant to study sarcoidosis.
References
1. Tao, Y.X. Melanocortin receptors. Biochim. Biophys. Acta Mol. Basis Dis. 2017, 1863, 2411–2413. [CrossRef] [PubM