Comprehensive overview of COVID-19-related respiratory failure: focus on cellular interactions

The pandemic outbreak of coronavirus disease 2019 (COVID-19) has created health challenges in all parts of the world. Understanding the entry mechanism of this virus into host cells is essential for effective treatment of COVID-19 disease. This virus can bind to various cell surface molecules or receptors, such as angiotensin-converting enzyme 2 (ACE2), to gain cell entry. Respiratory failure and pulmonary edema are the most important causes of mortality from COVID-19 infections. Cytokines, especially proinflammatory cytokines, are the main mediators of these complications. For normal respiratory function, a healthy air–blood barrier and sufficient blood flow to the lungs are required. In this review, we first discuss airway epithelial cells, airway stem cells, and the expression of COVID-19 receptors in the airway epithelium. Then, we discuss the suggested molecular mechanisms of endothelial dysfunction and blood vessel damage in COVID-19. Coagulopathy can be caused by platelet activation leading to clots, which restrict blood flow to the lungs and lead to respiratory failure. Finally, we present an overview of the effects of immune and non-immune cells and cytokines in COVID-19-related respiratory failure.

The SARS-CoV-2 pathogenesis process involves a variety of cells, which can interact with each other and with cytokines, leading to the destruction of the air-blood barrier and causing coagulopathy. This can eventually lead to pulmonary edema, ARDS and/or MODS, and finally death. Our goal in this article is to provide a comprehensive overview of the cells and processes involved in COVID-19-related respiratory failure. Firstly, we discuss the pulmonary epithelium and SARS-CoV-2 entry mechanism, followed by the influence of COVID-19 on endothelial cells and platelets, and finally we suggest how all these factors interact together, to understand the pathophysiology of pulmonary complications induced by SARS-CoV-2 infection.

Pulmonary epithelium
The airway epithelium is vital for its varied functions, such as warming and humidifying the inhaled air, clearing and defending the airways from inspired pathogens, and forming the epithelial half of the air-blood barrier. The most important point of SARS-COVID-2 entry is the respiratory tract, and epithelial cells are the most important targets of this virus. Therefore, it is necessary to review the various cell types in the respiratory epithelium ( Fig. 1).
A pseudostratified epithelium covers the upper airways, consisting of basal cells, ciliated cells as the main cell type, goblet cells, Clara or club cells, and a limited number of neuroendocrine cells [33,34]. Columnar ciliated cells have 200-300 individual cilia and are the main cell type in this epithelium. Their hair-like projections move backwards and forwards at a rapid frequency (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20) and help move mucus upwards through the tract. Goblet cells are also columnar, like ciliated cells. They produce and secrete mucin, a glycoprotein, into the airway for trapping inhaled particles and pathogens, also for and moisturizing the epithelium [33,35]. Clara or club cells are cube-shaped cells that secret proteins (e.g., CCSP) and surfactant proteins (SP)-A, SP-B, and SP-D. These proteins are involved in the composition of lung fluid [33,36].
In distal airways and bronchioles, the Clara cells become more abundant and the lung epithelium gradually becomes more columnar in nature. The height of the epithelium decreases and eventually becomes squamous within the alveoli. Alveolar type 1 and 2 (AT1, AT2) cells are two important cell types in the alveolar epithelium. They are involved in gas exchange and surfactant production, respectively [37,38].

Airway stem cells
The most important cause of respiratory failure and alveolar edema in COVID-19 is destruction of the integrity of the pulmonary epithelium. Airway stem cells have a crucial role in maintaining and repairing the epithelial integrity. The self-renewing stem cells are able to generate several different types of daughter cells [39,40]. Owing to the difficulties of direct studies in human lung, there is insufficient knowledge about the human lung epithelial stem cell niche. Knowledge on the repair and maintenance of lung function has been largely based on animal studies. In animals, airway stem cells are found in three niches within the tract: upper airway submucosal glands, small airway branching points, and the bronchoalveolar duct junction (BADJ). The cells in these niches are capable of generating various cell types [40].  (1) and endocytosis (2). In both pathways, spike (S) protein must bind to host cell receptors such as ACE2, NRP1, CLR, MGL, L-SIGN, DC-SIGN, TLRs, and GRP78. In endocytosis-mediated entry, following binding to cell receptor, virus entry into the host cell occurs and the S protein is activated in endosomes by furin cleavage. Fusion occurs by cathepsin-L action, and virus genetic material is released into cytosol, entering the virus through direct fusion mediated by proteases such as TMPRSS2 and/or furin. S protein interacts with a host cell receptor and becomes activated. Eventually, the membranes are merged and the virus releases its genetic material (RNA) via the formed pore into the cytosol. SARS-CoV-2 RNA is replicated and transcribed by host organelles such as ribosomes, Golgi apparatus, rough endoplasmic reticulum (rER), etc. Finally, the virus spreads to other cells and tissues. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; NRP1, neuropilin-1; ACE2, angiotensin-converting enzyme 2; MGL, macrophage galactose-type lectin; CLR, C-lectin type receptors; L-SIGN, homolog dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin related; DC-SIGN, dendritic cell-specific intracellular adhesion molecule-3-grabbing non-integrin; TLRs, Toll-like receptors; GRP78, non-immune receptor glucose-regulated protein 78; TMPRSS2, transmembrane protease, serine 2 host range in the Coronaviridae family [45]. Therefore, coronavirus infection of target cells depends on interactions of S proteins with cellular receptors [46]; for example, MHV uses murine biliary glycoproteins in the immunoglobulin superfamily; HCoV-229E and TGEV use human and porcine aminopeptidase N (APN), respectively; FIPV and FeCoV use feline APN, which can also be utilized by HCoV-229E and TGEV; and BCoV and HCoV-OC43 use N-acetyl-9-O-acetyl neuraminic acid moieties. Expression of the cloned receptor glycoproteins in cells of a foreign species can render them susceptible to infection with coronavirus virions [45]. Thus, coronavirus-receptor interactions are an important determinant of the species specificity of coronavirus infection. SARS-CoV-2 enters the host cells via endocytosis of a receptor-virus complex, or by direct membrane fusion (Fig. 1). In both pathways, the SARS-CoV-2 S protein, a trimeric class I transmembrane glycoprotein, must bind to cell receptors like ACE2 in the host. Respiratory epithelial and endothelial cells, monocytes, and alveolar macrophages show broad expression of ACE2 [47,48].
In addition to ACE2, several molecules have been suggested to serve as alternative receptors for SARS-CoV and SARS-CoV-2. These include C-type lectins, DC-SIGN and L-SIGN. Lectins are involved in the recognition of a broad range of pathogens and mediate intercellular adhesion. Although lectins and phosphatidylserine receptors enhance viral entry, they are nonspecific and do not support efficient infection by SARS-CoV or SARS-CoV-2 in the absence of ACE2, and thus "attachment factors" would better describe these molecules. Similarly, CD147, a transmembrane glycoprotein expressed ubiquitously in epithelial and immune cells, was proposed to be an alternative receptor for SARS-CoV and SARS-CoV-2 infection [49]. Although a modest increase in viral entry was observed with higher levels of CD147, and although its upregulation was observed in obesity and diabetes [49], which are potential risk factors for severe COVID-19, the role of CD147 in SARS-CoV-2 infection has been disputed on the basis of the inability of CD147 to bind the S protein [50]. Two groups identified neuropilin 1 (NRP1) as a host factor for SARS-CoV-2 [51,52]. Although NRP1 is expressed in olfactory and respiratory epithelial cells, its expression is low in ciliated cells, the primary target cells for SARS-CoV-2 in the airway, while it is high in goblet cells, which are not susceptible to SARS-CoV-2. Nonetheless, NRP1 was shown to enhance TMPRSS2-mediated entry (see the next section) of wild-type SARS-CoV-2 but not that of mutant virus that lacks the multibasic furin-cleavage site [49]. NRP1 was also shown to bind S1 through the multibasic furin-cleavage site and to promote S1 shedding and expose the S2′ site to TMPRSS2 [53]. Recently, the structure of ACE2 in complex with a neutral amino acid transporter, B0AT1, was analyzed by cryogenic electron microscopy in the presence and absence of the SARS-CoV-2 S protein [54]. ACE2 was previously shown to be essential for B0AT1 expression in the small intestine [55]. While B0AT1 is expressed in the gastrointestinal tract and kidney, it is not present in the lung. However, a B0AT1 homolog in the lung might contribute to SARS-CoV-2 infection. Additional studies are warranted to confirm the role of NRP1 and B0AT1 in SARS-CoV-2 infection [49].
Toll-like receptors (TLRs) recognize pathogen-associated molecular patterns and induce the production of type I interferons. Of these, TLR3, TLR7, TLR8, and TLR9 mount antiviral immune responses: TLR3 recognizes double-stranded RNA viruses, TLR9 recognizes unmethylated CpG in viral DNA, and, relevant to coronaviruses, TLR7   and TLR8 bind G/U-rich single-stranded viral RNA [49]. Many interferon-stimulated gene products were identified as important for SARS-CoV-2 replication, but only a few of them are involved in the entry steps: interferon-induced transmembrane proteins (IFITMs) [56,57] and lymphocyte antigen 6 family member E (LY6E) [58,59]. Recently, IFITM2 was shown to restrict SARS-CoV-2 entry [56]. IFITM proteins prevent viruses from traversing the endosomal membrane to access cellular cytoplasm by an unclear mechanism. Such a restriction can be bypassed if SARS-CoV were directed to enter cells exclusively at the plasma membrane [60]. LY6E is a glycophosphatidylinositol-anchored cell surface protein and was shown to inhibit or promote replication of some viruses [49]. Recently, LY6E was shown to impair infection by SARS-CoV, SAR-CoV-2, and MERS-CoV by inhibiting the S-protein-mediated membrane fusion, and mice lacking LY6E expression in immune cells were highly susceptible to mouse hepatitis virus, also a coronavirus [59]. Unlike IFITM-mediated restriction, LY6E-mediated inhibition was not overcome by TMPRSS2 expression (58). Further study is warranted to clarify the distinct roles of LY6E in regulating infection with SARS-CoV-2 and other viruses.
In endocytosis-mediated entry, after binding to a cell receptor, the virus penetrates into the host cells, and the S protein is activated in endosomes by furin cleavage. The endosomes undergo fusion with lysosomes by the action of cathepsin L proteolysis to form the endolysosome stage. Finally, after the fusion of membranes is completed, the viral RNA enters the host cell cytosol. Proteases are responsible for allowing viral entry through direct fusion, including transmembrane protease serine 2 (TMPRSS2)  27:63 or furin. After the S protein interacts with the host cell receptor, S protein activation occurs. Eventually, the membranes are merged and the virus transfer its RNA through the formed pore into the cytosol [61,62]. The RNA of SARS-CoV-2 is translated into structural proteins and nonstructural proteins (NSPs). These proteins are necessary for virus survival, multiplication, and virulence. In addition to the S protein other structural proteins, such as envelope glycoprotein (E), membrane glycoprotein (M), and nucleocapsid (N) proteins, are synthesized as accessory proteins [63].
The replication and transcription of the RNA sequence of SARS-CoV-2 occurs by a large multisubunit viral replicase-transcriptase complex. The structural proteins and genomic material are packaged into a new virus, which is secreted by exocytosis to spread to additional cells. The NSPs have many important functions in virus pathogenesis, such as stimulating the replication enzyme (RNA polymerase), proofreading of the SARS-CoV-2 genome, suppression of interferon (IFN) signaling, blocking the translation of host RNA, and stimulating cytokine expression. SARS-CoV-2 triggers an immune response through proinflammatory cytokine production associated with a weak protective IFN response to viral infection [64][65][66].
When the SARS-CoV-2 enters endothelial cells, it adversely affects these cells. Moreover, endothelial cells can be affected by molecules produced by other cells fighting against the virus. These molecules can bind to their receptors expressed on the endothelial cell surface. Platelet-activating factor (PAF) is one molecule released by a variety of cell types and tissues, but the main source of PAF is leukocytes. PAF mediates the inflammation process. Platelet-activating factor receptor (PAFR) is thought to be formed on the cytoplasmic or nuclear membrane of various cells, in particular endothelial cells, platelets, and leukocytes. The PAF/PAFR complex in endothelial cells may induce increased vascular permeability, hypotension, and expression of cytokines such as CXCL1, interleukin 6 (IL6), tumor necrosis factor-alpha (TNF-α), and IFN-γ [68,69].
TNF-α is another signaling molecule that may affect endothelial cells in COVID-19. Infection by SARS-CoV-2 strongly induces the secretion of TNF-α, which has an essential role in the pathology of acute lung injury. TNF signaling and the production of two TNF receptors (TNFRs, p55 and p75) on the pulmonary endothelium have not been clearly demonstrated in humans. One study reported that the two receptors, predominantly p55, are produced in pulmonary endothelial cells in mice. Reportedly, the administration of intravenous TNF led to surface expression of E-selectin, VCAM-1, and ICAM-1 [70,71]. Moreover, studies have reported that TNF can affect cell-cell junctions. SARS-CoV-2 has many effects on cell junctions, especially in endothelial and epithelial cells. Adherens junctions are specific intercellular adhesive structures mainly formed by cadherin and nectin adhesion receptors. They are essential for organization and flexibility of cell-to-cell attachment. Adherens junctions in endothelial cells are complexes containing vascular endothelial cadherin (VE-cadherin) and nectins. There are many cytosolic partners that interact with them, such as α-and β-catenin for VEcadherin, and afadin for nectins [72,73]. TNF can inhibit the expression of VE-cadherin, and also increase the tyrosine phosphorylation of VE-cadherin that blocks its binding with β-catenin, thus resulting in nonfunctional junctions [74,75]. Furthermore, TNF can affect actin cytoskeleton remodeling, VE-cadherin cytosolic domains, and β-catenin interactions. It has also been reported that TNF can induce caspase activation, which destroys intercellular junctions. In addition to cell junctions, members of the TNF family can activate the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway, resulting in increased proinflammatory gene transcription, cell proliferation, and differentiation [76,77].
Capillary leakage can occur in severe infections or in sepsis, due to disruption of tight junctions between endothelial cells. It has been suggested that hijacking of PALS1, a tight-junction-associated protein, by the SARS-CoV E protein has a central role in lung epithelial disruption in these patients [78]. Studies have reported that PALS1 is also expressed in endothelial cells. Researchers have proposed that the SARS-CoV-2 E protein can interact strongly with PALS1, thereby disrupting the epithelial barrier and leading to severe inflammation [79,80].
Moreover, it has been shown that TNF can disrupt claudin-5 in tight junctions by an NF-κB-dependent process, and activate kinase cascades in human dermal endothelial cells. However, so far, no study has been done on pulmonary endothelium [81]. Another study has reported that TNF can lead to barrier destruction and mislocalization of JAM-A, claudin-4, and claudin-5, which are involved in tight junction structure [82].
In addition to disruption of endothelial cell junctions during inflammation and viral infections, it is known that endothelial cell death may occur following SARS-CoV-2 infection by apoptosis and/or necrosis, resulting in capillary leakage, lung failure, and MODS. Programmed cell death apoptosis can be triggered by two signaling pathways: extrinsic (death receptor-dependent) or intrinsic (mitochondrially mediated). Extrinsic pathway-activated caspase-8 can also activate the intrinsic pathway. Activated caspase-8 can stimulate caspase-3 directly or by promoting mitochondrial cytochrome C release [83,84].
In the extrinsic pathway, the death receptors Fas and TNF receptor are activated, resulting in caspase-8 activation. Caspase-8 stimulates other caspase cascades that ultimately lead to apoptosis [85]. Fas and Fas ligand (FasL) are an important system that mediates extrinsic pathway apoptosis. FasL is released by neutrophils and lymphocytes, especially under inflammatory conditions. Fas is a surface receptor expressed in many types of respiratory cells, including endothelial, alveolar, and bronchial epithelial cells and macrophages. The Fas/FasL system is hyperactivated in patients with ARDS, and it has been reported that activation of this complex in the lungs can lead to the release of TNF-α, transforming growth factor-β1 (TGF-β1), and other proinflammatory cytokines leading to epithelial apoptosis [86]. Another component of the death receptor ligand  family is the transmembrane protein TNF-related apoptosis-inducing ligand (TRAIL) [87]. It has been established that cytotoxic T lymphocytes and natural killer (NK) cells are involved in the immune surveillance of virus-infected or virally transformed cells [88]. These effector cells secrete a membrane-disrupting protein called perforin, and granzymes, structurally related serine proteases, acting as cytotoxic mediators, along with proinflammatory cytokines (IFN-γ and TNFα) [89]. Perforin and granzymes are released by exocytosis to induce apoptosis of the infected or transformed cells [90]. In the granule-exocytosis pathway of lymphocyte-mediated cell death, activated lymphocytes secrete both perforin and granzymes. Monomers of perforin may act together to form the polymeric pore structure in the plasma membrane, causing osmotic cytolysis, a type of necrosis. The pathways responsible for granzyme-induced cell death are unclear. It has been suggested that granzymes penetrate the target cell to begin an internal disintegration pathway, which ultimately causes DNA fragmentation and apoptosis or cell lysis [91,92]. Moreover, the accumulation of cell death receptors such as Fas may occur. In this pathway, the Fas expressed on the target cells binds to the FasL expressed on the killer-cell membrane. This Fas-FasL complex then promotes caspase-dependent apoptosis [93].
The molecular mechanisms of necrosis are not completely clear. It has been suggested that necrosis probably occurs by the release of lysosomal enzymes, generation of ROS, and activation of phospholipases [94]. Elevated levels of ROS can destroy the mitochondrial respiratory chain, inhibiting the transfer of electrons to O 2 , thus further increasing ROS production by a positive feedback loop. A large number of damaged mitochondria will lead to necrosis. In cell death mediated by lysosomal damage, lysosomal enzymes are released into the cytosol, resulting in necrosis [95]. Some studies have reported that FasL and TNF can stimulate both apoptotic and necrotic pathways. Interestingly, although a significant depletion of intracellular ATP has been seen in apoptosis, in necrosis the levels of ATP remained at the normal concentration seen in healthy cells [96,97].

Platelets
Studies have shown that coagulopathy is a common symptom in patients with severe COVID-19. This could be a result of excessive inflammation. Coagulopathy is characterized by dysregulated coagulation parameters, lower platelet count, increased microthrombi, elevated D-dimer, etc. It is reported that patients with COVID-19 who have lower platelet counts and disseminated intravascular coagulation have a poor prognosis and increased mortality [27,28]. In many viral infections, platelets can be activated, and are involved in inflammation and thrombotic processes. The low platelet level in some patients with COVID-19 could be due to increased consumption and thrombus formation by massively activated platelets [98,99].
Collectively, it has been suggested that platelets may have an important role in the pathogenesis of COVID-19 infection. Platelets can internalize the virus, resulting in TLR7 activation and the secretion of complement component 3 (C3) from their alpha granules. C3 stimulates the process of NETosis, which traps any viruses missed by platelets [104,105]. In NETosis, the nuclear material of neutrophils is released into the extracellular space. These structures form a meshwork of chromatin called neutrophil extracellular traps (NETs). In this meshwork, the DNA is coiled around histones and enzymes derived from neutrophil cytoplasmic granules. It has been suggested that NETosis is a type antimicrobial defense mediated by neutrophils. These structures also act as a scaffold for the attachment of RBCs, platelets, and plasma proteins to form a trap for any virus missed by platelets [106,107].
In response to unknown signals from neutrophils, platelets release granulocytemacrophage colony-stimulating factor (GM-CSF) to regulate NETosis. Interestingly, there are reports of increased levels of both GM-CSF and C3 in COVID-19 infections [104,105].
COVID-19-related endothelial cell death can be a result of direct damage by the virus, or indirect damage by cytokines leading to thrombosis, particularly in small vessels with low blood flow. The exposed subendothelial matrix following endothelial cell death causes accumulation and activation of platelets intended to repair the damage [29,108]. When further endothelial cells are destroyed, more platelets are activated that lead to microthrombus formation. It is known that healthy endothelium secretes prostaglandins, nitric oxide (NO), and adenosine to control the extent of thrombus formation by dissolution, and to regulate platelet activation [109,110].
Platelets can also be affected by inflammatory cytokines and high antibody concentrations, which can lead to increased thrombosis. The levels of IL-6 and IL-1β are increased in patients with COVID-19, and it has been reported that IL-6 and IL-1β are involved in platelet hyperactivation [111][112][113][114].
Infected endothelium can also lead to the secretion of tissue factor (TF), a well-known coagulation cascade activator. TF leads to platelet aggregation for clot formation via the stimulation of thrombin and the generation of fibrin fibers. Even if anticoagulant therapy is administered, this may lead to deep vein thrombosis and pulmonary embolism. Following the activation of platelets, they produce functionally active TF [99,115]. The effect of cytokines on platelets and endothelial cells in COVID-19 is not fully understood [116].
It has been found that SARS-CoV-2 infection indirectly induces an increased ability of von Willebrand factor (vWF) to bind to its receptors on platelets [102]. The vWF on the endothelial surface interacts with its receptor (GP Ib-IX-V complex) on platelets. The cytoplasmic GPIb domain underlying the plasma membrane of platelets binds to the actin filaments. It is important to protect the shape and structure of platelet membranes, which is essential for platelet adhesion [117,118]. The binding of vWF to GPIb-IX induces transient platelet adhesion. Platelet signals then activate integrins resulting in stable adhesion. Thrombin is another ligand for GPIb-IX, which is involved in low-dose thrombin activation of platelets, activation of protein kinase C (PKG), and activation of the mitogen-activated protein kinase (MAPK) (ERK1/2 and p38) pathway, and finally secretion of platelet granules [119][120][121][122]. GPVI is known as a collagen receptor, and plays a key role in collagen-mediated platelet activation. In this pathway, phosphatidylinositol 4,5-bisphosphate (PIP2) is hydrolyzed to produce inositol 1,4,5-trisphosphate (IP 3 ) and 1,2-diacylglycerol (DAG) by phospholipase C-2 (PLC2) activity. DAG and IP 3 are known as cellular secondary messengers. DAG activates PKC isoforms, which are involved in the activation of integrins and the secretion of platelet granules [123][124][125]. IP 3 increases the calcium concentration (Ca 2+ ) in the cytosol by triggering the release from the dense tubular system. This depletion of intracellular Ca 2+ storage activates store-operated calcium entry resulting in sustained increases in intracellular calcium [126].
An increase in intracellular calcium is crucial for several intracellular signaling processes. PKC, calpain, and calmodulin are all proteins that are activated by calcium. Moreover, integrin signaling often depends on Ca 2+ levels. Ca 2+ elevation is also required for stable platelet adhesion, granule secretion, procoagulant activity, clot retraction, and almost all platelet functions [125].
Pathogen-associated molecular patterns (PAMPs) are defined as molecules secreted by microorganisms. These can be DNA, proteins, RNA, lipopolysaccharides (LPS), and glycoproteins. Moreover, damage-associated molecular patterns (DAMPs) are also secreted from infected or damaged tissue. DAMPs can be proteins, DNA fragments, or lipid oxidation products. PAMPs and DAMPs can bind to pattern recognition receptors (PRRs), such as nucleotide-binding oligomerization domain (NOD)-like receptors, TLRs, and the advanced glycation end-product (RAGE) receptor. It has been reported that TLRs and NODs can trigger platelet activation. In platelets, the collagen receptor GPVI can activate PLC2, resulting in the formation of DAG and IP3, which can trigger PKC and stimulate Ca 2+ liberation, thus affecting platelet function [127][128][129].
TLR4 is the most important platelet receptor for LPS. It triggers signals that finally result in granule secretion. It has been shown that the activation of the cGMP-PKG pathway mediated by MyD88 has an important role in TLR4-dependent platelet activation. TLR4 can also recognize high-mobility group box 1 (HMGB1), a DAMP protein that activates platelets and triggers granule secretion. HMGB1 is a DNA-binding protein produced by macrophages and monocytes during infection and inflammation [25,130,131].
Thrombin is another ligand binding to platelet receptors. Up to now, at least three thrombin receptors have been identified on the surface of human platelets: GPIb-IX, PAR1, and PAR4 [132,133]. The presence of these three different receptors can increase the sensitivity of platelets to low thrombin levels, and is pivotal to thrombosis formation [134]. It has been reported that GPIb-IX signaling activated by thrombin and vWF mediates platelet activation by a member of the Src kinase family (Lyn or possibly Src) interacting with the Rac1 signaling pathway [135]. Rac1 is a GTPase with an important role in the assembly of subunits of nicotinamide adenine dinucleotide phosphate oxidase (NOX), as well as in ROS production. It also has been suggested that thrombin can activate PLC through a ROS-mediated signaling pathway. Then PLC activation leads to DAG and IP3 production, as described above [135,136].
PAR1 and PAR4 can also activate PLC isozymes to produce PIP2, which is hydrolyzed into IP3 and DAG, which can then promote the production of thromboxane (TXA). It has been reported that high levels of TXA2 can induce granule secretion in platelets. The activation of the Src family kinase-Rac1 signaling pathway is also involved in PAR1/ PAR4-dependent platelet activation and granule secretion, by the mitogen-activated protein kinase (MAPK) pathway [136][137][138][139].
Integrins are also important molecules involved in platelet activation, accumulation, and adhesion. Integrins act as receptors for collagen, fibronectin, laminin, and vitronectin. The integrin α IIb β 3 has been reported to be the most important and abundant member of the family. It is a receptor for fibrinogen, vWF, and several matrix adhesion proteins. Integrin α IIb β 3 is needed for stable adhesion and aggregation of platelets [140,141].
Collectively, calcium elevation, PKC signaling, SFK signaling, and MAPK signaling, among others, are the platelet signaling pathways involved in granule secretion. However, the complete details of these mechanisms are not clear. It is known that coagulopathy due to SARS-CoV-2 infection has a poor prognosis. Therefore, we propose that further studies focusing on platelets and their receptors in COVID-19 are needed.

Respiratory failure: cellular interactions in the lungs
SARS-CoV-2 infection of lung endothelial cells, which might occur from the luminal or the alveolar interstitial side, triggers endothelial release of cytokines, which cause increased capillary permeability, thereby allowing adhesion and extravasation of neutrophils and monocytes into the alveolar interstitial space. Stimulated by PAMPs and DAMPs, neutrophils and macrophages secrete a multitude of cytokines, procoagulants, and complement, which promote viral attack and clearance but induces further vascular injury enhancing the risk for thrombosis. Several factors might contribute to the prothrombotic environment, thereby promoting intravascular thrombus formation: (a) neutrophils mediate secretion of NETs (neutrophil extracellular traps), and complement enhances platelet aggregation; (b) cytokines trigger secretion of tissue factor (TF) by endothelial cells, and macrophages stimulate coagulation cascade and increases in fibrin clot formation; (c) endothelial damage decreases secretion of anticothrombotic mediators, such as antithrombin (AT) and TF pathway inhibitor (TFPI); (d) lung residential megakaryocytes produce locally available platelets for aggregation; (e) the angiotensin-converting enzyme (ACE)/Ang II (angiotensin II)/AT1 receptor axis is overactivated owing to virus-induced ACE2 downregulation increasing production of PAI1 (plasminogen activator inhibitor 1), reducing plasmin activation and fibrinolysis [142]. Several factors that might contribute to respiratory failure in COVID-19 disease are summarized as follows:

MMP-9-related respiratory failure in COVID-19 disease
A schematic illustration of the cellular interactions in the lungs is shown in Fig. 2.
The endothelial cell layer is lined by a glycocalyx layer, and can release tissue plasminogen activator (t-PA) to inhibit platelet binding and activation of the coagulation cascade. t-PA mediates the conversion of plasminogen into plasmin, with an important role in fibrinolysis. It has been reported that SARS-CoV-2-infected endothelial cells can secrete t-PA. High t-PA concentrations present in COVID-19 samples can carry out spontaneous fibrinolysis [143,144]. It is known that t-PA can enhance neutrophil degranulation, while t-PA administration can increase blood levels of matrix metalloproteinase (MMP)-9 (gelatinase B) [145]. It should be mentioned that MMPs are proteolytic enzymes that degrade extracellular matrix (ECM) components, such as collagen, proteoglycans, and elastin. It has been reported that endothelial cells secretes several MMPs under pathological conditions [146]. MMP-9 is released by neutrophils, monocytes, macrophages, eosinophils, and other inflammatory cells. In acute inflammation, neutrophils undergo chemoattraction to accumulate at the injury site. MMP-9 is released from neutrophils by degranulation and then degrades the basement membrane collagen (especially type IV collagen). Inflammatory cell infiltration is facilitated by MMPs, which then results in leukopenia [147][148][149].

Role of DAMPs in SARS CoV-2-induced respiratory failure
Widespread endothelial dysfunction caused by direct viral infection or immune response results in the secretion of numerous cytokines and DAMPs. DAMPs can directly induce NETosis following influenza virus infection [150,151]. NETs are made up of DNA, histones, and enzymes such as serine proteases [106,107]. The released histones can activate pro-factor VII activating protease (pro-FSAP) to produce factor VII activating protease (FSAP). FSAP contributes to the coagulation and fibrinolysis cascades, by indirectly mediating plasminogen activation. Interestingly FSAP can also degrade histones [152].
NETs can also stimulate FXII activation, and FXIIa is able to directly convert prothrombin to thrombin, which is a key factor in coagulation. Thrombin causes fibrinogen conversion to fibrin, which is amplified by factor XIII activation. Then, the soluble fibrin monomers interact with each other to produce a cross-linked network of long fibrin threads that trap the platelets. Then a spongy mass builds up, hardens, and contracts to form the final blood clot. The coagulation cascade is important both in normal circulatory conditions and in pathological reactions [152,153]. Moreover, NET serine proteases and histones may also play a role in platelet activation [106,154,155].

The role of von Willebrand factor and granulocyte subsets in SARS CoV-2-induced respiratory failure
Von Willebrand factor (vWF) is another factor involved in SARS-CoV-2 coagulopathy. It is secreted by endothelial cells and megakaryocytes. Increased vWF antigen concentrations in plasma have been measured in COVID-19 infections [155,156]. Reportedly, the increased concentration of vWF enhanced the adhesion of platelets onto the collagen surface, and increased the aggregation of platelets. vWF could be a therapeutic target in patients with thrombocytopenia [157]. On the basis of some studies, basophil and eosinophil counts, as well as granulocyte subsets, are altered in severe COVID-19 infection. These cells play a pivotal role in the antiviral response and immunopathology in COVID-19. Basophils are decreased during the acute phase, while the numbers recover during the COVID-19 recovery period. Basophils can enhance B-cell responses during the humoral immune response triggered by IL-4 or IL-6 production. Moreover, it has been shown that basophil count is significantly correlated with the concentration of IgG antibodies against SARS CoV-2 [157]. However, the level of IL-6 is inversely correlated with antibody production [158]. It was also reported that basophils can secrete IL-13 following activation, and IL-13 receptors are expressed on mature human B cells, so basophils may contribute to humoral immune responses and inflammatory allergic reactions [159].
Notch signaling is a juxtacrine signaling pathway, mediated by receptor-ligand interactions between neighboring cells. The notch signaling pathway is dysregulated in many pathological conditions, especially inflammation [160][161][162]. In experimental animal models, the pharmacological blocking of notch signaling has been reported to improve inflammatory conditions [163]. In vitro, it was shown that cytokines can affect notch signaling in basophils, and inhibition of this signaling pathway reduced basophil cytokine production. It was also reported that the notch signaling pathway is involved in the expression of cytokine genes in activated macrophages during inflammation [164]. Notch signaling is also important for peripheral T-cell differentiation and activation [165].
The overexpression of Jagged (a notch ligand) in professional antigen-presenting cells (APCs) has been reported [166]. Although there is not enough evidence for basophils to function as antigen-presenting cells, it is known that B cells are APCs, and can activate T cells [167]. Moreover, basophils can enhance B-cell responses [158].
Because COVID-19 is an inflammatory disease, it may also affect the notch signaling pathway. As a result, basophil cytokine production and basophil-induced B-cell activity would be impaired, affecting both cellular and humeral immunity. This is still a hypothesis, and further studies are needed to confirm the role of the notch signaling pathway in COVID-19.
Eosinophils are another granulocyte subset that contribute to viral infections by producing NO, increasing TLR7, and can also produce eosinophil extracellular traps (EETs, similar to NETs) to limit viral replication [168]. It was also reported that eosinophils are involved in the immune response against rhinovirus infections by stimulating T cells [169]. NO is a major endogenous vasodilator that maintains blood pressure within the normal range and also blocks platelet activation [168,170]. It was reported that the increase in blood eosinophils found in patients with eosinophilic asthma exerted no protective influence on COVID-19 [171]. A variety of eosinophil functions in patients has been reported. Eosinophil EETosis and release of major basic protein (MBP) could contribute to coagulopathy, while in blood vessels, eosinophils could mediate platelet activation and thrombus formation [172]. It has been reported that activated eosinophils can secrete cytokines, such as IL-2 [172], IL-8 [173], IL-12 [174], INF λ [175], etc. Eosinophil degranulation occurs in inflammatory conditions and virus infections. The important components of eosinophil granules include eosinophil-derived neurotoxin (EDN), major basic protein (MBP), eosinophil peroxidase, and eosinophil cationic protein (ECP) [176].
EDN can induce the activation of the dendritic cell TLR2-MyD88 signaling pathway in response to microbial and viral infections [176]. Both macrophages and DCs can secrete IL-12, IL-27, and IL-18. IL-18 has a significant role in ARDS [176][177][178]. IL-12 acts on activated NK and T cells expressing the IL-12 receptor. IL-12, IL-27, IL-15, IL-18, and type I IFN cal all increase the activity of NK cells and induce the expression of IFN-γ. IFN-γ can activate macrophages to destroy pathogens. Moreover, it was shown that IL-12 can trigger the differentiation of T-helper 1 cells. Pulmonary DCs also produce IL-6 at a significantly higher level than other DCs. Some studies have reported that IL-6 is the most important single cytokine in the cytokine storm seen in patients with COVID-19 [179][180][181].
ECP and EDN have both been reported to activate apoptotic pathways in infected cells. Moreover, ECP is strongly associated with tissue necrosis in different organs. In addition, higher concentrations of MBP and ECP occur in patients with eosinophilic asthma. They could contribute to many biological processes, such as the stimulation of perivascular mast cells (MCs) resulting in microvascular permeability [176,182].
Various proinflammatory cytokines are released in patients with COVID-19, especially IL-6. It has been reported that IL-6 is a pivotal mediator of lung damage. Mast cells (MCs) have been found to be important sources of IL-6, IL-1β, VEGF, and TNF, and they play a crucial role in several pulmonary diseases [183].

The role of activated cells of the immune system in SARS CoV-2-induced respiratory failure
NK cells have also been identified as important effector cells for the innate immune response. NK cells are involved in the early defense against viral infections and to deal with malignant transformation. Studies have shown that ROS can act as an inhibitor of NK cells [184,185]. ROS can produced by eosinophils, and their ROS production may be stimulated by immunoglobulins (Igs), adhesion molecules, and cytokine signaling. ROS are essential for optimal defense against viruses, but they may also contribute to viral pathogenesis and host cell destruction. NK cell cytotoxic activity can by decreased by IL-6. Studies have reported that the inhibition of IL-6 and IL-1 can increase the expression of perforin, granzyme, and IFN [186][187][188][189][190].
However, contrary to expectations, some studies found that the percentage of NK cells was dramatically higher in patients suffering from COVID-19 infection admitted to the intensive care unit (ICU) [191]. NK cells can release FasL and TRAIL death ligands, which bind to death receptors on target cells and trigger the extrinsic apoptosis pathway. NK cells can eliminate target cells via both apoptosis and necrosis. NK cells also secrete several cytokines, and are the major source of IFN-γ in the early stage of viral infection, where they coordinate the adaptive and innate immune responses [192,193]. Adaptive immunity depends on lymphocytes, CD8 + and CD4 + T cells, and B cells. Plasma cells are antibody-producing cells that are derived from B cells. They govern the humoral response to foreign antigens by producing Ag-specific B cells and CD4 + T-helper cell interactions. In COVID-19 infections, plasma cells secrete anti-SARS antibodies [194].
B cells can be stimulated by either anti-inflammatory or proinflammatory cytokines. After contact with T cells, they produce proinflammatory cytokines such as IL-12, IL-15, and IL-6. IL-6 and IL-12 can form a positive feedback loop between B and T cells [195,196]. It was reported that IL-15 secretion could enhance the migration and cytotoxicity of CD8 + T cells in the brains of MS animal models. It has been demonstrated that regulatory B cells can produce anti-inflammatory cytokines, such as IL-10, TGF-β, and IL-35. Pregnant women show high concentrations of immunosuppressive cytokines (IL-10, TGF-β, and IL-35) and chemokines (CCL22 and CCR4) [197][198][199].
B cells are also important in the initiation of an inflammatory response and GM-CSF secretion. In addition, GM-CSF can be expressed by various cells, including macrophages, T cells, endothelial cells, and NK cells. It was reported that the number of GM-CSF-expressing cells was higher in patients with COVID-19 in the ICU [191,200,201].
GM-CSF can stimulate the differentiation of monocytes into macrophages or into DCs [202]. Both alveolar and interstitial macrophages can be categorized as either M1-like macrophages (M1, classically activated) or M2-like macrophages (M2, alternatively activated) [203]. The M1 and M2 phenotypes have different cytokine profiles and cellular functions. M1 has antimicrobial activity and produces proinflammatory cytokines, such as IL-1β, TNF-α, IL-12, IFNs, and IL-6. On the other hand, M2 contributes to tissue maintenance, repair, and angiogenesis, and secretes anti-inflammatory cytokines. In viral infections such as COVID-19, both types of macrophage are involved. M1 can recognize and destroy viruses in various families, such as Coronaviridae, and promote an inflammatory response. On the other hand M2 plays a role in tissue repair and resolution of lung damage. Therefore, these two phenotypes must be balanced for optimal antiviral immune response, resulting in pathogen elimination without tissue injury [204].
It has also been observed that, after the induction of inflammation, M1 can be stimulated to switch to the M2 phenotype. This conversion is marked by a lower production of inflammatory cytokines and an increased secretion of anti-inflammatory cytokines (IL-10 or TGF-β). The conversion is involved in the amelioration of inflammation and protection of tissue integrity [205].
M2 produces various ECM components (type I and III collagens, and MMPs). It was reported that M2-like macrophages can be converted into collagen-producing fibroblasts by the effects of TGF-β [206]. Pulmonary fibrosis after COVID-19 pneumonia is a common complication in COVID-19 survivors [207].
M1 can stimulate activity of T-helper type 1 cells and promote cell-mediated immune responses. Th1 cells secrete IFN-γ, TNF-β, and IL-2, and can activate macrophages to increase phagocyte-dependent responses [208]. Macrophages and NK cells interact with each other, as an important antimicrobial and antitumor defense mechanism. M1 can increase NK cell cytotoxicity by affecting the IL-1β, IFN-β, and IL-15 pathways [209]. Alveolar macrophages are resident cells within the lung alveoli that act as a major early defense against respiratory infections. They release various anti-inflammatory and proinflammatory cytokines. Alveolar macrophages were found to release IL-1 (low amount), IL-6, TNF (moderate amount), and IL-8 (high amount) [210]. Moreover, in the alveolar epithelium, type 2 pneumocytes play a role in the cytokine storm and respiratory failure. The pneumonocytes produce, release, and reuptake the pulmonary surfactant and surfactant proteins A and D, control the amount of alveolar fluid, and produce some immunomodulatory proteins [211,212].

Conclusion
In COVID-19 infection, virus entry can be mediated by a variety of receptors and proteins. These receptors may be useful in therapeutic approaches, and more studies should be performed. On the other hand, after the destruction of the air-blood barrier caused by apoptosis or necrosis of epithelial and endothelial cells, and disruption of the basement membrane, a variety of cells enter the alveolar space in an abnormal manner, depending on the severity of the damage. Moreover, the damaged endothelial cells are involved in the coagulopathy process. It is suggested that clot formation may restrict blood flow and could lead to respiratory failure. Platelets are another important cell type in this process. Platelets have many surface receptors involved in coagulopathy and inflammation, especially by activating kinase cascades leading to granule secretion. Cytokines are the main mediator in these events. It has been shown that infiltrated inflammatory cells and lung epithelial cells are able to secrete cytokines (IL-1β, IL-2, IL-6, IL-7, IL-8, IL-10, IL-17, and TNFα). Positive feedback loops (especially involving IL-6) finally result in the uncontrolled release of cytokines and a cytokine storm resulting in further injury. In addition, the destruction of the gas exchange barrier leads to flooding of the alveolar space, thus increasing susceptibility to infection and finally causing lung edema and respiratory failure.

ACE2
Angiotensin-converting enzyme 2 APCs Antigen-presenting cells aPKC Atypical protein kinase C ARDS Acute respiratory distress syndrome