The role of nerve growth factor (NGF) in the pathogenesis of leprosy

Features of the immune response in lesions of the nervous system by the lepra pathogen Mycobacterium leprae (introduction)

Leprosy is a chronic infectious disease caused by Mycobacterium leprae (M. leprae), characterized by clinically diverse lesions of the skin, mucous membranes of the upper respiratory tract, peripheral nervous system, musculoskeletal system, and internal organs [1][2]. The modern Ridley-Jopling classification of leprosy is based on the division of patients depending on the state of their immunologic reactivity to M. leprae, which is reflected in clinical manifestations, as well as in the data of histopathologic, bacterioscopic, and immunobiologic studies. The leprosy process is considered a continuous spectrum of immunopathologic changes between tuberculoid and lepromatous types of leprosy, called polar types, with the distinction of three borderline and undifferentiated forms [3][4]. The World Health Organization (WHO) concluded that clinical classification depending on the bacteriological load was necessary and proposed to distinguish multibacterial leprosy (ML) and low-bacterial leprosy (LL) [2]. In the LL form of the disease, there is an intensive cell-mediated immune response that prevents the proliferation of mycobacteria in contrast to ML leprosy, in which the immune response, on the contrary, contributes to the dissemination of the pathogen in phagosomes of macrophages. In borderline forms, patients show immunologic and histopathologic characteristics approaching polar forms of the disease [5]. The degree of disability of a leprosy patient is classified as grade 0 (sensory disturbances are not detected, deformity of feet, hands, and eyes is not visible), grade 1 (sensory disturbances appear without deformity of feet, hands, or pronounced visual impairment), and grade 2 (with irreversible neurological damage) [8]. Numerous studies prove that more than a quarter of leprosy patients have some degree of disability, and about half of these patients have disabilities associated with severe limb deformities [3][6][9–11].

Nerve cell damage in leprosy results from demyelination of peripheral nerve cells. The resulting neuropathy caused by the localization of leprosy mycobacteria in nerve endings and Schwann cells induces a response mediated by macrophages and other cells and eventually leads to immune-mediated lesions [12][13]. One strategy to detect early peripheral nerve dysfunction in leupra is to recognize Schwann cell behavior by altering markers of myelin sheath synthesis. When Schwann cells encounter damage, as an autonomous defense mechanism they restore the state through remyelination. This process is influenced in particular by factors such as neurotrophins.

Biological aspects and mechanisms of action of nerve growth factor (NGF)

Neurotrophins are a group of closely related polypeptides that stimulate and control neurogenesis in the central and peripheral nervous system [14–16]. Mammalian neurotrophins include four major neurotrophic factors (nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4)), which are similar in chemical structure but differ in the principle of action [17]. To date, the most studied representative of the neurotrophin family is NGF. According to modern concepts, it is a secreted dimeric protein with a molecular weight of 26 kDa, containing 118 amino acid residues [17–19]. NGF was first described by Levi-Montalcini, and then its key role in differentiation, maturation, and preservation of the integrity of sympathetic and sensory neurons was shown [20].

Proteolytic reactions are known to promote the synthesis of mature NGF molecules from their proneurotrophin (proNGF) precursors, which also have biological activity. While mature neurotrophins promote neuronal survival, proneurotrophins have an opposite, proapoptotic effect [21].

NGF receptors play an important role in modulating pain signaling in various physiological and pathological conditions, and genetic and metabolic features (e.g., in diabetes) promote specific proneurotrophin (proNGF) activity in effector cells or neurons [22].

NGF plays an important role in the development and maintenance of cellular phenotypes in the peripheral nervous system, as well as in maintaining the integrity of cholinergic nerves in the central nervous system [23]. NGF is endogenously produced during the development and maturation by several cell types including neurons, Schwann cells, oligodendrocytes, lymphocytes, mast cells, macrophages, keratinocytes, and fibroblasts. Its main function in the nervous system is to participate in inflammatory processes and immune responses [24]. Schwann cells produce NGF in response to axonal degeneration. NGF prevents Schwann cell damage, meaning that low levels of this factor may contribute to the development of neuropathy [25]. However, while NGF levels are dramatically reduced in affected nerve trunks in patients with neuropathic lesions, patients with chronic cutaneous hyperalgesia have locally elevated NGF levels [25]. NGF concentration increases during inflammatory processes in tissues, causing hyperalgesia due to the direct activation of nociceptors, which leads to the activation of the central nervous system and neurogenic inflammation [26]. In turn, this process leads to the release of histamine and an increase in mast cells and other immune system cells. Inflammatory swelling can cause degeneration of nerve fibers. A positive correlation was shown between levels of NGF, NGF-R (neurotrophin receptor LNGFR or p75), and TGF-β (transforming growth factor). This indicates the synergistic properties of the above factors that prevent tissue vulnerability to nerve injury [27]. There is evidence that NGF restores sensitivity and has a proliferative and antiapoptotic effect on keratinocytes and endothelial cells, contributes to the restoration of pain sensitivity, and, consequently, prevents the development of trophic ulcers associated with the loss of nociception [12][25]. NGF was found to be involved in the regulation of tissue formation and healing. During tissue healing, NGF activates processes related to innervation restoration [27][28], stimulates fibroblast migration, and has proliferative and antiapoptotic effects on keratinocytes and endothelial cells [29][30]. Moreover, NGF plays a dual role in neuronal survival and death [31–33]. Studies showed that neurotrophins and their receptors, including NGF, were widely expressed in skeletal tissues, participated in chondrogenesis, osteoblastogenesis, and osteoclastogenesis, as well as in the regulation of tissue formation and healing processes [34].

Peripheral nerve injury or any pathologic condition that causes a gap between the target organ and the nerve cell body acts as a signal to induce non-nervous cell populations (e.g., fibroblasts) to produce NGF. Induction of NGF synthesis in these cells is also modulated by cytokines that penetrate the site of nerve injury, where nerve regeneration is initiated [35]. In addition, NGF was shown to play an important role in influencing specific responses to injury through pro-inflammatory effects on neutrophils, eosinophils, mast cells, and T lymphocytes [36]. The interactions of NGF in the tissue microenvironment are complex, and its association with TNF-α, which can induce apoptosis in Schwann cells by binding to specific death receptors, may lead to antagonistic effects. This is because NGF can activate survival signaling in the target cell. The same cytokine can exert antagonistic effects depending on its interaction with specific receptors, and the intracellular cascade is activated upon the activation of these receptors [37]. This supports the assumption that NGF plays a crucial role in the process of myelination of Schwann cells in the peripheral nervous system.

Relationship of NGF with other growth factors

Tumor necrosis factor (TNF-α) interacting with NGF induces the differentiation and maturation of neurons [38]. An association between NGF and transforming growth factor (TGF-β) was found in glial cells of rats and mice with spinal cord injuries [27]. Similar assumptions were made in the study of systemic diseases (e.g., diabetes and osteoarthritis). Facer et al. [39] showed that the action of NGF is associated with the transmembrane receptor TrkA present in subepidermal fibers of the skin. The second receptor p75, so named after its molecular mass of 75 kD, is also transmembrane but with a different mechanism of action. It responds to neuron death, that is, neurodegeneration. The interactions of NGF, as well as other neurotrophins, with these receptors largely determine the fate of the neuron. Apparently, NGF acts as a neurotrophic messenger, and its level is regulated by innervating neurons [20]. It has also been found that nerve endings are not affected and the presence of NGF in keratinocytes correlates with a lack of temperature sensitivity. The use of NGF antibodies is effective in treating hyperalgesia in patients with neuropathy and nerve ending lesions. In addition, physiologic combinations of NGF, NT-3, and glial cell line neurotrophic factor can help restore homeostasis. Thus, they can be used in the treatment of neuropathic pain [25].

All this proves that NGF may play a key role in the pathogenesis of nervous system damage in leprosy. It is known that different levels of NGF are registered in lepromatous and tuberculoid leprosy, higher for the lepromatous form and lower for the tuberculoid form of the disease [40]. In the lepromatous form of leprosy, immunostaining of lesional tissue samples registers significant levels of NGF, indicating a larger and more diffuse focus of nervous system involvement [41][42]. Annand et al. found that it was low NGF values that contributed to the absence of NGF-dependent nociceptive fibers in the damaged skin of leprosy patients [43]. A study by Antunes et al. [9] showed that in patients with a tuberculoid form of leprosy, the level of NGF-R immunoexpression was lower in nerve fibers and Schwann cells compared to controls. The authors found that the phenomenon of hypoesthesia was associated with the decreased expression of NGF-R and glycoprotein-P (PGP). When studying the relationship between clinical forms of leprosy and neuritis episodes, it was found that patients with borderline leprosy were 2.69 times more likely to develop neuritis than patients with the lepromatous form of the disease [44].

Local cellular immunity is able to resist bacterial infections, but inflammation can lead to irreversible tissue damage. For example, nerve damage occurs in approximately 10% of patients with LL, 40% of patients with ML, and is particularly acute in patients with reactive states [45][46].

Higher levels of NGF are associated with lepromatous forms, and increased NGF expression stimulates TGF-β (transforming growth factor) expression, which reduces tissue damage due to nerve injury. TGF-β is an anti-inflammatory agent in tissue repair during nerve and tissue regeneration [27][47]. Higher expression of TGF-β in patients with the lepromatous form of the disease is associated with a higher rate of apoptosis in the lesion foci, especially in Schwann cells [48][49].

The research strategy for detecting peripheral nerve damage at the initial stage in leprosy patients should be to find markers that can be used as a diagnostic tool to detect early nerve damage. Such a marker could be the determination of NGF concentration. Although NGF itself is not involved in myelin formation, other factors such as axonal signaling and TrkA activation are required for this to occur. It was shown that NGF can regulate the myelination process with opposing effects between Schwann cells and oligodendrocytes via axonal signals [50]. When the concentration of NGF in the blood increased, axonal signaling was sent. The axonal signals that control central myelination are probably very similar to those that control peripheral myelination. In addition, using ROC analysis, a threshold value of NGF was determined that can be used to detect early disability in patients with ML leprosy [51].

Conclusion

Thus, studies analyzing patients with different clinical forms of leprosy and different reactive states (e.g., erythema nodosum) may help to better understand the relationship between NGF and the immune response, as well as other factors that contribute to nerve protection and regeneration. Further analysis of NGF levels in tissue and blood between cytokines and immune response is important to better understand the involvement of NGF in the pathophysiology of leprosy and other neurologic lesions. Determination of NGF thresholds will contribute to the early detection of the process of nerve damage when regeneration is still possible.

The work was performed within the framework of the state assignments of the Ministry of Health of the Russian Federation “Development of methods of diagnostics and treatment of leprosy infection based on the principles of personalized medicine” and “Influence of social and medical rehabilitation on improving the quality of life of leprosy patients”.