**3. Innate immune receptors involved in phagocytosis**

The interaction of cells of the innate immune response with MTB is based on the contact of PPRs with it and the recognition of PAMPs. The outcome of this encounter will define the response and development of the infection. Among the innate immune PPRs involved in the MTB phagocytosis are: MR, DC-SIGN, Dectin, Mincle TLR, SR, and CR.

#### **3.1 Mannose receptor**

MR (CD206) belongs to the C-type lectin receptors that recognize polysaccharides such as mannose, fucose, and N-acetylglucosamine. MR can be found in monocytederived macrophages (MDMs), and AM and dendritic cells, but it is absent in monocytes [33, 34].

MR is a transmembrane protein constituted by protein domains that recognize carbohydrates, and a cytoplasmatic region enriched with tyrosine and related to phagocytosis [35]. MR binds lipoarabinomannan (LAM), phosphatidylinositol manosides (PIM), mannoproteins, mannans, and arabinomannans from mycobacteria [36, 37]. Cytokine production in response to MTB recognition through MR, in immature monocyte-derived dendritic cells, induces the expression of an anti-inflammatory profile [38]; furthermore, it inhibits the production of ROS and reduces the expression of IL-12 [39, 40]. In addition, the recognition ofmannosylated LAM by MR prevents phagosome-lysosome fusion and prevents phagosome maturation [36].

*Phagocytosis of* Mycobacterium tuberculosis*: A Narrative of the Uptaking and Survival DOI: http://dx.doi.org/10.5772/intechopen.110067*

#### **3.2 DC-SIGN**

Also known as CD209, it is a C-type lectin receptor that can be found in some populations of macrophages and dendritic cells, whereas in AM it is induced after infection with MTB [41, 42]. CD209 is an important link between the innate and acquired immune response, and after the encounter with MTB, it mediates the mycobacterial entry. DC-SIGN identifies glycoproteins, lipomannan (LM), arabinomannan, PIM, and ManLAM from MTB, and discriminates from species with arabinofuranosylterminated LAM (AraLAM) such as *Mycobacterium smegmatis* [43]. MTB ManLAM recognized by DC-SIGN induces the expression of the anti-inflammatory cytokine IL-10, where it also counteracts the TLR-4 response [44]. Moreover, the interaction of DC-SIGN with MTB reduces the expression of IL-12, which has caused a decrease in the activity of T cells [45].

### **3.3 Dectin-1**

Dectin is a group of C-type lectin PPR involved in cellular activation, found in neutrophils, dendritic cells, monocytes, and some clusters of T cells [46]. Dectin-1 recognizes beta-glucans and mannosylated lipids and discriminates between mycobacteria species and its strains, such as MTB Ra, *Mycobacteruim bovis* BCG (BCG)*, Mycobacterium phlei,* and *Mycobacterium abscessus* [47–50]. MTB triggers the production of IL-17A through the response produced by its interaction with Dectin-1 and TLR4 dependent on IL-1 signaling [51]. Also, it has been found that murine macrophages derived from bone marrow, which contain Dectin-1, showed an increased expression of IL-6, TNF alpha, and G-CSF, when infected with virulent mycobacteria such as BCG, *M. smegmatis, M. phlei*, or *Mycobacterium avium* [49]. Dectin-1 contribution seems to be important during MTB infection in splenic dendritic cells; it is involved with the production of IL-12p40 an important subunit to granuloma development [52].

#### **3.4 Mincle**

Macrophage-inducible C-type lectin (Mincle) is a C-type lectin receptor found in leucocytes and macrophages after stimulation [53]. Mincle intervention during MTB infection showed to be fundamental to the recognition of TDM, with an increased production of inflammatory cytokines by macrophages, which contribute to granuloma development [54, 55]. In AM from Mincle deficient mice, the exposure to BCG revealed a reduction in the proinflammatory cytokines, a decreased number of leucocytes in lung tissue, and an increased bacterial count inside and outside the lungs [56]. However, during MTB infection in Mincle-deficient mice, the animals developed a protective immune response TH1, TH17, and a granulomatous response [57].

#### **3.5 TLR**

TLRs are a family of 10 human PPRs involved in recognition and phagocytosis of intra- and extracellular pathogens. TLRs are composed of a transmembrane domain of leucine-rich repeats that identify the PAMPs; in their structure can also be identified the intramembrane domain that allows the assembly of signaling-related components [58].

TLRs are found in a variety of human cells, such as dendritic cells and AM. These PPRs can be intracellular (such as TLR-3, TLR-7, TLR-8, and TLR-9) or extracellular (such as TLR-1, TLR-2, TLR-4, TLR-5, and TLR-6). TLR-10 can be found in plasmacytoid dendritic cells and B cells. TLR 10 can be found in plasmacytoid dendritic cells and B cells [59]. During MTB infection, TLR triggers the antibacterial response dependent on vitamin D addition [60]. Multiple mycobacterial Ag can be recognized by TLR receptors. The mycobacterial lipoprotein 19 kDa, phosphor-myo-inositolcapped LAM, lipomannans and PIM, are recognized through TLR-2 [61, 62]. The CpG motives of MTB are recognized by TLR-9 [63]. TLR-4 recognizes the MTB heat shock protein 65 (Hsp-65).

The intracellular signaling of MTB recognition by TLR is dependent on the production of the myeloid differentiation factor 88 (MyD88). However, TLR 2, 4, and 9 deficient mice controlled the inflammation during MTB infection and developed a T cell response [64].

#### **3.6 SR**

Scavenger receptors are a group of transmembrane glycoproteins found on the surface of dendritic cells, some endothelial cells, macrophages, and monocytes. SR are classified in SR sub-group A and SR sub-group B. The A group comprehends MARCO (a macrophage receptor), SR-A1, and SR-A2, whereas the B group includes SR-B1 and CD36 [65]. The absence of SR-A in infected mice with MTB H37Rv prolonged the life of this animal above the average lifespan of a wild type [66]. MARCO recognizes TDM and this receptor, accompanied by CD14 and TLR-2, mediates cytokine production [67]. However, MARCO-deficient mice had no difference in acute and chronic infection with MTB in comparison with the wild type [68]. In contrast, a MARCO polymorphism is associated with an augmented susceptibility to the infection with MTB in Gambian population [69]. *Cd36*−/− macrophages had an increased capacity to destroy *Mycobacterium marinum* and MTB, whereas CD36-deficient mice had a reduced susceptibility to the BCG infection [70].

#### **3.7 CR**

Complement receptors are a group of extracellular receptors that mediate the phagocytosis of non-opsonized, and opsonized bacteria, covered with fragments of proteins of the complement cascade. There are three types of CRs: CR1, CR3, and CR4 located in macrophages, neutrophils, monocytes, NK cells, and lymphocytes. CRs recognize glycopeptolipids from non-opsonized MTB and PIMs [71, 72]. Also, CR3 from monocytes recognize phagocyte microbeads coated with the 85C antigen from BCG and MTB [73]. MTB can be recognized by CR1, CR3, and CR4; however, 80% of the phagocytosis mediated by complement is dependent on the recognition by CR3 [74].

### **4. Evasion of the immune response in macrophages**

Macrophages developed a variety of strategies to destroy bacteria: production of ROS and nitrogen intermediates, iron restriction, use of heavy metals, production of antimicrobial peptides, phagosome acidification, and fusion of the phagosome with the lysosome.

*Phagocytosis of* Mycobacterium tuberculosis*: A Narrative of the Uptaking and Survival DOI: http://dx.doi.org/10.5772/intechopen.110067*

MTB evades and endures the strategies to eliminate bacteria and survive inside the macrophages; this attribute allows them to multiply and increase the population in order to establish the infection or trigger latency. The basic mycobacterial mechanisms to evade the immune response and survive inside the macrophages will be briefly described below.

### **4.1 Phagosome maturation arresting and inhibition of the phagosome acidification**

The phagosome is described as a membrane structure vacuole containing the microorganism; this structure is formed immediately after the phagocytosis. The phagosome maturation is dependent on the actin-mediated movement and is supported by the reactions and delivery of the late and lysosomal constituents [75].

Throughout the establishment of the pathogenic mycobacterial infection, and soon after the bacterial recognition by PPRs, the arresting of the phagosome maturation constitutes a strategy that MTB employs to evade the immune response; specifically, the phagosomal molecule migration pathway is modified in order to avoid the microbicidal activity. During the phagosome maturation, Rab GTPases proteins are recruited to the phagosome membrane; they regulate the membrane fusion and the sorting of lipids and proteins to the organelles. The presence of these molecules is a marker of the phagosome/endosome maturation status. Also, Rab molecules allow identification of the maturity of the structure, specifically Rab5 (which is present on early endosomes) and Rab7 (present on late endosomes) [76–79].

The recruiting of Rab effectors, the endosomal tethering molecule (EEA1), and the phosphatidylinositol 3 kinase hVPS34 to mycobacteria-infected phagosomes are inhibited by mycobacterial PIM and LAM, leading to an arresting of the phagolysosome development [80–82]. Also, MTB ManLAM inhibits the augmentation of Ca2+ in the cytosol, avoiding the phosphatidylinositol 3-phosphate fusion with calmodulin at the phagosomal membrane, driving the inhibition of the recruitment of GTPases to the phagosome [81].

The mycobacterial antigens—early secretory antigen target 6 (ESAT-6), culture filtrate protein 10 (CFP10), the eukaryotic-like serine/threonine protein kinase G (PknG), and the SecA1 and SecA2—arrest the phagosome maturation and contribute to the mycobacterial survival inside the macrophages [83–85]. *M. avium* keeps the phagosomal pH between 6.2 and 6.5, due to the exclusion of the proton ATPase in phagosomal acidification [86]. MTB protein tyrosine phosphatase (PtpA) contributes to the survival of the bacteria inside the phagosome, as a consequence of the inhibition of the complex V-ATPase + H with the phagosomal membrane [8].

MTB permits the V-ATPase catalytic subunit A proteasome degradation because of ubiquitination signaling, while also regulating the reduction of the phagosome pH [87]. Glycolipid TDM recognition by the receptor Mincle induces the blockage of signaling involved in the phagosomal formation [88].

#### **4.2 Resistance to reactive nitrogen species and reactive oxygen species**

Reactive nitrogen species (RNS) and ROS are short-lived chemical compounds that mediate and contribute to the innate immune response through microbicidal mechanisms [89]. The ROS generation is dependent on the phagosomal acidification. Among the effects of the oxidative stress due to the ROS activity, can be described the oxidation of lipids, proteins and DNA damage. During the MTB infection, the sigma

factor and the stress response factor SigH, produced during ROS and RNS action, contribute to the infection [90–92]. The mycobacterial mycothiol has an antioxidant activity and keeps the cell reduced. The MTB mutation of the gene that encodes for the mycothiol synthase, *mshD,* had an increased susceptibility to H202 [93, 94]. MTB Cu, Zn superoxide dismutase SodC contributes to the resistance to the oxidative burst produced by the ROS of macrophages; also, the MTB *sodC* mutant was sensitive to the superoxide and was susceptible to IFN-gamma too [95]. Similarly, the alkyl hydroperoxide reductase (AphC) contributes to the resistance to ROS of the innate immune response [96].

MTB exposed to NO had a bacteriostatic effect and induced the expression of genes related to dormancy [9]. The expression of inducible nitric oxide synthase (iNOS) confers alveolar macrophages with the ability to kill MTB, and the latency of MTB in macrophages from healthy subjects was dependent on the production of the NO [97]. MTB controls the production of ROS by the increased expression of host histamine receptor H1 (HRH1), by regulating the GRK2-p38MAPK signaling pathway [98].

#### **4.3 Apoptosis and autophagy evasion**

Cell apoptosis is a hosting strategy to destroy the intracellular niche of the bacteria. The evasion of apoptosis is related to the mycobacterial virulence. The avirulent strains like *Mycobacterium kansasii*, *M. tuberculosis* H37Ra, and BCG induced more human alveolar macrophages apoptosis, whereas *Mycobacterium bovis*, *M. tuberculosis* H37Rv, and the MTB clinical isolated, named as BMC 96.1, did not [99]. Virulent MTB stimulates the cell necrosis of macrophages by the mitochondrial inner membrane rupture, favoring the release of the microorganism [100].

The autophagy leads to the destruction of damaged cell parts resulting in the cell survival. In MTB infection, the autophagy development conducts a defense mechanism against it. In macrophages infected with MTB or BCG, the autophagy induces the phagolysosomal formation and mycobacterial death [101]. Finally, the foamy phenotype in macrophages protects the cell and reduces autophagy of MTB-infected macrophages [102].
