VEGF-A: Vascular Endothelial Growth Factor A; bFGF: basic Fibroblast Growth factor

Mast cells (MCs) have a unique role among the innate immune cells because of their unique and extensive mediator profile and their ability to interact with the vasculature. The physiological role of MCs is unclear. We argue that MCs exist in all species with a circulatory system due to the basic triad tissue response that they are able to produce under physiological and pathological conditions alike: extracellular matrix degradation and tissue remodeling, de novo cell proliferation, and de novo angiogenesis. MC-activation occurs not only in life-saving reactions such as inflammation and wound healing but in implantation, the decidua, the placenta, the pregnant uterus, and the fetus too. In fact, MCactivation is essential for successful pregnancy, and helpful for the survival of the newborn up to reproductive age (owing to beneficial actions of the MCs in inflammation and wound healing), and essential as for reproductive capability in adulthood. The fact that MCs since more than 500 million years are preserved in all species that have a circulatory system, i.e., the vertebrates, suggests that the life-saving and life-promoting MCmediated response of tissue remodeling, changing the functional capabilities of the tissue, cell proliferation, and angiogenesis could be MCs’ purpose. The MCs’ action in the pregnant uterus could, hypothetically, be their physiological role.

•    Mast cells; Matrix-degrading enzymes; Heparin; 
Histamine; VEGF-A; bFGF; Inflammatory cytokines;
Cell proliferation; Angiogenesis; Wound healing; 
Decidua; Placenta; Uterus; Fetus; Hypothesis

Norrby K (2021) Reason of Being of Mast Cells a Hypothesis. Ann Vasc Med Res 8(3): 1136

In efforts to elucidate the physiological role of the connective tissue mast cell (MC), first described by Paul Ehrlich some 140 years ago, the study of various genetically modified MCdeficient strains of mice has emerged as the avenue of choice. It is reported that mice with MC-deficiency without defects in Kitsignaling pathway have a remarkably normal immune system, whereas the MC is essential in mediating allergic disease and anaphylaxis, while other cells inside and outside of the immune system apparently share all of the MCs’ other functions related to immunological responses [1]. New and possibly even more relevant models of MC-deficient mice are being developed but still the physiological role of the MC is ambiguous [2].

The notion that the MC is known foremost for its key role in mediating harmful allergic disease and life-threatening anaphylaxis is disturbing as MCs have been present for 500 million years in all groups of vertebrates, all having a circulatory system, well before the development of adaptive immunity [3]. Considering the evolution and the process of natural selection of cells, it would suggest significant life-promoting functions of MCs. MCs are activated in innate immune responses by multiple Keywords • Mast cells; Matrix-degrading enzymes; Heparin; Histamine; VEGF-A; bFGF; Inflammatory cytokines; Cell proliferation; Angiogenesis; Wound healing; Decidua; Placenta; Uterus; Fetus; Hypothesis mechanisms, and have a unique role among the innate immune cells because of their extensive and unique mediator profile and by their ability to interact with the vasculature [4].

Based on current information about the roles MCs play in lifesaving situations such as inflammation and wound healing, but also in the pregnant uterus, the placenta, as well as the growth and development of the fetus, we speculate on why MCs are preserved in all vertebrates. It is worth noting that all these lifesaving or life-promoting conditions show pronounced migration, recruitment, and activation of MCs in the affected tissues. The Mast Cell MCs are important effectors of type 1 allergy but also essential regulators of innate and adaptive immune responses [2,4]. The role of this sentinel cell in host defense is unmistakably important. The MCs are located preferably close to blood vessels in all connective tissues and play a role in vascular biology such as in the control of cardiovascular homeostasis and diseases [5- 9]. MCs possess a great variety of surface receptors that may be activated by a range of agents such as inflammatory mediators, immunoglobulins, proteases, hormones, neuropeptides, bacterial and helminth products. MCs are prevalent in the female reproductive tract and are reactive to several female sex hormones [10-15]. Activated MCs, typically degranulate by expelling intracellular granules into the local microenvironment, but there are also other modes of secretion.

The granular content of MCs comprises multiple bioactive molecules like heparin, histamine, inflammatory cytokines, tryptase and chymase (exclusively expressed by MCs), highly efficacious extracellular matrix-degrading metallo-proteinases, as well as a variety of bioactive molecules, such as growth factors and inflammatory cytokines, and many other molecules exercising diverse effects [16,17].

MCs are the only cells that produce heparin and they are the main repositories of histamine in the body. Heparin has the highest negative charge density of any known biological molecule. It binds to proteins, including many growth factors, facilitating the interaction between growth factors and their receptors. Vascular endothelial growth factor-A, VEGF-A, is a key regulator of physiological and pathological angiogenesis, while basic fibroblast growth factor, bFGF, is not only a potent angiogenic factor but also a powerful inducer of proliferation of a great variety of tissue cells. Both VEGF-A and bFGF are heparinbinding proteins secreted by MCs.

Importantly, activated MCs participate in implantation, placental, uterine and fetal angiogenesis and development [8,18,19], events that are akin to MC-mediated tissue remodeling, cell proliferation and angiogenesis, which are discussed below.

Tissue remodeling and cell activation caused by mast cell-secretion

Secreting MCs in situ bring about tissue remodeling by their potent extracellular-matrix-degrading enzymes. MC-activation also activates connective tissue cells displaying increased metabolism, phagocytosis, collagen synthesis, proliferation, and mobility; moreover, there is additional production of proteases by activated fibroblasts [8,16-18,20-23]. Tissue remodeling opens up to manifold functional alterations in the tissue affected. Whereas the extracellular matrix-degrading effects by activated MCs are obvious the effect of MC-secretion on cell proliferation and angiogenesis may need some further consideration.

Early evidence of mast cell mediated tissue cellproliferation and angiogenesis

MC-products heparin, histamine, and serotonin stimulate the proliferation of quiescent normal human cells in vitro: First, the unfractionated heparin (UFH) medication was shown to mitogenically stimulate quiescent density-inhibited (multilayered, non-dividing) healthy human normal fibroblasts [24].

We then found that two additional MC products, i.e., histamine and serotonin (in rodents), which are permeabilizing factors, individually at very low concentrations, induce the proliferation of quiescent human normal density-inhibited fibroblasts [25]. In fact, these three molecules are the first documented MC-derived mitogens.

MCs in situ induce tissue-cell proliferation: Our findings in vitro led us to develop a quantitative assay for studying proliferation of defined cell types in situ exploiting the extremely thin intact, connective-tissue membranous small-gut mesentery in normal adult rats [26]. We showed that the in situ selectively activated MC rapidly induces potent proliferation of quiescent fibroblastic and mesothelial cells, which constitute the vast majority of the tissue cells present, as well as of epithelial cells following non-immunologic (such as a single exposure to Compound 48/80 i.p., a highly selective activator of the MC in rats) or immunologic activation of MCs in various rodent species and tissues [20,27-29]. This was before it was known that MCs contain bFGF.

MC-histamine is mitogenic in vivo: Using specific histamine receptor antagonists, histamine was shown to be a MC-derived mitogen that acts via the H2-receptor on target cells in vivo and ex vivo (organ culture) in the rat mesentery [30]. In the biological control, the Guinea pig, the MCs of which are non-responsive to Compound 48/80, no cell proliferation is seen in the mesentery following i.p. injection of Compound 48/80, whereas histamine at 10-10M injected i.p. induces significant proliferation of the fibroblasts and mesothelial cells.

MCs induce angiogenesis: a new angiogenesis model: Angiogenesis, the formation of new blood vessels from parent microvessels, is essential for normal body growth whereas it is negligible in most adult normal tissues because of a stringent balance between numerous anti- and pro-angiogenic factors. Of note, angiogenesis occurs physiologically in hormone-responsive female tissues such as the lining of the uterus, the placenta, and the pregnant uterus. Under pathological conditions angiogenesis in adulthood is a hallmark of hypoxia, inflammation, and wound healing.

Introducing a novel rat mesentery angiogenesis model, we demonstrated, for the first time, the existence of a rapid, potent and long-lasting de novo angiogenic reaction following the selective activation of MCs in situ by i.p. injection of Compound 48/80 in the intact adult animal; Guinea pigs as the biological control are unaffected [22,31]. The content of VEGF-A in MCs was at the time not yet known in the literature. This model in intact normal animals is non-surgical and biologically highly relevant. The extremely thin small-gut mesenteric test tissue is vascularized, albeit sparsely, and lacks significant angiogenesis in adulthood. The vasculature is basically 2-dimensional. Using a monoclonal antibody against rat endothelial cells all blood microvessels are identified immunohistochemically in spreads of the intact tissue. Unintentional inflammation-induced angiogenesis is virtually absent as judged by comparing untreated and endotoxinfree saline vehicle-treated animals, a rare feature of in vivo angiogenesis models [32,33]. This guarantees a high degree of sensitivity and discriminatory power since inflammation activates MCs and induces angiogenesis.

MC-histamine is angiogenic: As first reported by our group, histamine released from secreting MCs in situ is angiogenic, acting via the surface histamine receptors H1 and H2 on target cells in the rat mesentery [34]. This finding of histamine-induced angiogenesis via H1 and H2 receptors in vivo was subsequently verified in s.c. mouse tissues, by Dr. H. F. Dvorak and associates [35]. Moreover, histamine synergistically promotes bFGFinduced angiogenesis by enhancing VEGF-A production via H1 receptor in endothelial cells in vitro [36,37].

MC-derived heparin-binding proteins are angiogenic: We have examined the angiogenic effect of five MC-derived heparinbinding proteins using the rat angiogenesis model. As expected, the growth factors VEGF-A and bFGF, but also the inflammatory cytokines IL-1-alpha, IL-8, and TNF-alpha, which are all released by activated MCs, induce significant de novo angiogenesis after being injected i.p. individually at near-physiological dosages in non-angiogenic endotoxin-free saline vehicle. The inflammatory cytokines apparently stimulate the VEGF-A production in the exposed tissue cells and activate the MCs [38-42]..

Heparin affects angiogenesis depending on its molecular mass: Our pioneering studies show that the systemic effect of heparin molecular mass on angiogenesis is biphasic. Protamine is able to neutralize free heparin. Protamine sulfate injected s.c. suppresses mesenteric MC-mediated angiogenesis significantly in the rat, suggesting an angiogenic role for native heparin proteoglycans [43].Rat MCs release soluble heparin proteoglycans of very high molecular weight (c. 750 000 to c. 900 000-kDa), which in vivo are depolymerized by heparinases. Unfractionated heparin (UFH), with a mean molecular weight (mmw) of c. 15- kDa, is a mixture of disaccharide chains with molecular masses in the 4 to 40-kDa range. UFH is the medicinal counterpart of native heparin and is identical to its native counterpart in composition and structure.

We found that there is a significant fragment-size-dependent effect of s.c. injected heparin fractions (produced from UFH by a naturally occurring heparinase) on angiogenesis mediated by VEGF-A or bFGF in the rat mesentery; linear coefficient of regression r = 0.97 in the examined range of 2.5 to 22-kDa mmw [44-48]; the 22-kDa fraction being selected from UFH. Heparins of large mmw significantly stimulate, UFH tends to stimulate, whereas those of lower mmw inhibit angiogenesis significantly in statistical terms. This implies that post MC-secretion in situ the released heparin stimulates angiogenesis, but as the heparin molecules are successively degraded into smaller fragments by heparinases it becomes anti-angiogenic, suggesting an innate heparin-depolymerization angiogenesis-modulating mechanism [48].

MC- mediated angiogenesis is a complex cell and tissue reaction: Activated MCs promote angiogenesis via cascade-like interacting paracrine pathways that involve: (i) Extracellular matrix degradation and tissue remodeling as tissue cells exhibit increased metabolism, phagocytosis, collagen synthesis, and proliferation. This causes cellular hypoxia and induction of transcription and hypoxia-inducible factors, which activate genes for angiogenic factors (e.g., VEGF-A, bFGF) that act in addition to the angiogenic effect of histamine, VEGF-A, bFGF, and inflammatory cytokines, secreted by the MCs; (ii) Degradation and rearrangement of extracellular matrix releases growth factors bound to the matrix, facilitates cell migration, and affects the function of cells that interact with the matrix; and (iii) Recruitment and activation of other cells, such as macrophages, platelets, fibroblasts, and additional MCs (by the Stem cell factor), which contribute to the strikingly potent and protracted angiogenic response [17,49].

Mast cells and wound healing

It is widely accepted that MCs significantly affect all phases of wound healing —inflammation, proliferation, and remodelingincluding angiogenesis of healing wounds in normal animals [50- 54]. Our findings of (i) MC-mediated and MC-histamine-mediated cell proliferation [20,34], extracellular matrix remodeling, and collagen synthesis [21], and (ii) MC-mediated and MC-histaminemediated angiogenesis in the rat [22,31,34] are consistent with the subsequent discoveries in a mouse model for MC deficiency that both MC-activation and histamine release are required for normal cutaneous wound healing [50].

It is intriguing that the MC is known foremost for its key role in mediating harmful allergic disease and life-threatening anaphylaxis since it has been preserved in all groups of vertebrates for 500 million years, long before the development of adaptive immunity. This would suggest that MCs have significant life-promoting functions. It should be remembered that the MC is unique among all the innate immune cells due to its ability to interact with the vasculature and also its exclusive and extensive mediator profile. The MC is exceptionally reactive to injurious agents as to certain female sex hormones.

The MC is a uniquely endowed cell that when activated releases many powerful preformed and newly synthesized bioactive molecules, such as heparin, the mitogenic and angiogenic factors histamine, bFGF and VEGF-A, several angiogenic inflammatory cytokines, as well as highly efficacious extracellular matrix-degrading enzymes. We argue that MC-secretion, which is able to produce a triad response of extracellular matrix degradation and tissue remodeling (creating altered functional capabilities of the tissue affected), de novo tissue-cell proliferation, and de novo angiogenesis, is of evolutionary significance. This is because the activation of MCs is essential in both physiological and pathological situations e.g. inducing angiogenesis and growth of the decidua, the placenta, the uterus, and the fetus, which is reportedly a prerequisite for successful pregnancy, but also in life-saving inflammatory responses and wound healing. The performance of MCs in the womb during pregnancy thus seems to be a plausible physiological role of MCs. Apparently, this key triad response helps the survival of the newborn up to reproductive age, and to the successful reproduction in adulthood. It is warranted to consider whether this triad MC-mediated response in physiological and pathological conditions alike may account for MCs being preserved in all species that have a circulatory system, i.e., the vertebrates. If so, it could be MCs’ purpose.

The Swedish Medical Council and the Swedish Cancer Foundation financially supported these studies.

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