Mesenchymal Stem Cells (MSCs) as a Novel Therapeutic Option for nCOVID-19—A Review

The novel Coronavirus Disease (nCOVID-19) is a highly contagious viral infection which emerged as “Pneumonia of Unknown Etiology” at Hubei province of Wuhan city in China. The health authorities provided a considerable empirical evidence after this outbreak and it was notified that the causative virus, named Novel Coronavirus (subsequently SARS-CoV-2) is the culprit for progressively exerting grim effects not only on individual patients but also on the International public health, with high mutational tendencies. WHO declared nCOVID-19 as a Pandemic on 11 th March 2020. The spike glycoprotein of pilot study, we emphasize the need for conducting more clinical trials with ethical consideration to prove the efficacy and safety of MSCs in combating nCOVID-19 infection and its complications.


Introduction
Many species of animal and human hosts have been affected by coronaviruses and the family of coronaviruses remains relatively obscure probably because of no severe human diseases attributing to this virus. However, in the year 2003, it became clear that these coronaviruses were responsible for severe acute respiratory syndrome (SARS) epidemic. Since then, two new human respiratory coro-Open Journal of Regenerative Medicine naviruses namely Middle East respiratory syndrome related coronavirus (MERS-CoV) and nCOVID-19 have been described. In the year 1968, scientists named this virus as "coronavirus" because of its crown like morphological appearance and it belongs to the family coronaviridae with two subfamilies namely, the coronaviruses and the toroviruses. Later, it was found that these viruses were also responsible in causing enteric diseases in cattle and possibly in humans.

Etiology of nCOVID-19
There are 2 speculations regarding the origin of SARS-CoV-2 -1) natural selection of animal host before zoonotic transfer and 2) natural selection in humans following zoonotic transfer [4]. The zoonotic source is not confirmed however genomic analysis suggested bats as the main reservoir. The isolated β-CoV shows 88% similarity to the sequence of two bat-derived SARS-like coronaviruses, bat-SL-CoVZC45 and bat-SL-CoVZXC21 [5]. People with history of visit to Wuhan's seafood market contracted this infection but in due course, it was revealed that in some individuals no record of seafood market visit had also contracted infection suggesting its capability of human to human transmission capability [6]. This can occur via direct contact with an infected person, exposed to coughing, sneezing, respiratory droplets or an aerosol which enter human lungs via inhalation through nose or mouth and via indirect contact i.e. as fomite transmission. Recent case series has put forward possibility of vertical transmission of SARS-CoV-2 from infected mother to her new born; however it is not conclusive [7] [8] [9].

Coronavirus Classification/Taxonomy
Based on the serological cross-reactivity characteristics, these coronaviruses are divided into three genera, i.e., group I to III [10] [11]. This was also confirmed by genome sequencing analysis [12]. The Group-I coronavirus includes animal pathogens such as transmissible gastroenteritis virus (TGEV), Porcine epidemic diarrhoea virus (PEDV), feline infectious peritonitis virus (FIPV) as well as human coronaviruses HCoV-229E, HKU1; Group-II coronavirus includes pathogens of veterinary relevance such as Bovine coronavirus (BCoV), equine coronaviruses, porcine hemagglutinating encephalomyelitis virus as well as human coronavirus OC43, NL63 and Group-III includes only avian coronavirus such as IBV, pheasant coronavirus and turkey coronaviruses [13]. But, the Group-II viruses which infects mice and rats were also found to be as coronavirus prototype that were found responsible in causing infections like enteric diseases, hepatitis, encephalitis, respiratory diseases and chronic demyelination [14]. Many controversies exist about the SARS-CoV that whether it belongs to a new group of coronaviruses or a distant member of group [15]. The sequences of coronaviruses detected in Columbia livia (feral pigeon), Anser anser (Greylag goose) and Anas Open Journal of Regenerative Medicine platyrhynchos (mallard) and their phylogenetic analysis also confirms that these viruses belong to group-III [16]. There were two prototypes of human coronaviruses found to cause common cold in humans namely OC43 and 229E previous to the emergence of SARS-CoV [17] and determined to have long speculations about human coronaviruses with serious health issues such as multiple sclerosis [18], hepatitis [19] and enteric diseases in new borns [20].  Figure 1). The membrane protein (M) which spans three times the viral membrane has a short N-terminal ectodomain and a cytoplasmic tail. The small membrane protein (E) is found to be highly hydrophobic in nature [23] and this spans twice the N and C terminals on the interior part of the virion [24]. There are still many other minor proteins present in the viral structure yet undetected and the genomes of all the coronaviruses were found to have similar structural characteristics [25]. For all coronaviruses, the structural proteins are encoded in order of S-E-M-N within the one third of the genome and each group of coronaviruses encodes a group of unique small proteins while these are non-essential proteins and have been found to serve as accessory proteins to interact or interfere with the host innate immune responses which has not been demonstrated for any of these proteins [26] [27]. Untranslated regions of coronaviruses (UTRs) on both 5' and 3' ends of the genome were believed to interact with the host and control the RNA replication process and the viral transcription has been reviewed in recent studies [28].

Replication of nCOVID-19
Coronaviruses (nCOVID-19/SARS-CoV-2) attach to specific cellular receptors like ACE-2 with its spike protein (S) and this triggers a conformational change in the viral spike protein which mediates a fusion reaction between the virus and the cell membrane resulting in injecting the viral genetic material into the host cell. Upon entry, the 5' end of the RNA genome, ORFs 1a and 1b are translated into polyproteins (pp 1a and pp 1b) and here, the polyproteins pp 1ab are translated through a frameshift mechanism occurring at a high frequency of 25% to 30% [20] [28]. ORF 1a encodes one or two papain like protease enzymes and a picornavirus 3C-like protease which functions in processing the polyproteins into mature replicase proteins [29] and encoded in X domain of ORF 1a and processed by an RNA-dependent RNA polymerase (RdRP) along with helicase enzyme [30] as well as with other enzymatic activities [31]. An additional putative enzymatic activity of cyclic phosphodiesterase encodes the downstream process in ORF 2a and these multiple enzymatic activities play a vital role in metabolism of coronavirus RNA interaction with the host cell process [32]. The interaction between the viral spike protein (S) and the ACE-2 receptor on the host cell surface significantly initiates the infection process and the cryo-EM structure analysis has revealed the binding affinity efficacy of nCOVID-19 (SARS-CoV-2) S protein to ACE-2 receptor of about 10 to 20 times higher than that of SARS-CoV S protein [33] [34] and has higher transmissibility and contagiosity of nCOVID-19 as compared to SARS-CoV [35].
On the basis of genome sequence alignment and homology, SARS-CoV and SARS-CoV-2 (nCOVID-19) are found to have shared a highly conserved receptor-binding domain (RBD) and there is 76% similarity of the domain of S protein sequence [22] [36] and 83% of similarity in their active sites [4]. Due to their much higher binding affinity to the cellular receptors the human to human transmission of the virus is also visibly higher when compared to that of SARS-CoV coronavirus outbreak and additional studies are required in this pandemic situation for better investigation of this possibility of higher transmission among individuals [37]. Based on these evidences, the natural evolution of nCOVID-19 or the SARS-CoV-2 was supported based on its overall molecular mechanisms and also scientists have found that SARS-CoV-2 backbone differed substantially from those known coronaviruses. Additionally, their viral resemblances were found in bats and pangolins. Hence, the scientists concluded that the nCOVID-19 coronavirus is the product of natural evolution ending any deliberate genetic engineering [38].

Pathogenesis of nCOVID-19 with Human Host
nCOVID-19 patients show clinical manifestations such as fever, non-productive cough, dyspnoea, fatigue, myalgia, normal or decreased leukocyte counts with radiographic evidence of pneumonia. Recent evidences shows anosmia, hyposmia and dysgeusia to be considered in the list of symptoms associated with nCOVID-19 infection [39]. As per several reports, the envelope spike glycoprotein play pivotal role in the entry. It recognizes and binds to its cellular receptor angiotensin I converting enzyme 2 receptor (ACE2R) for SARS-CoV and SARS-CoV-2, CD209L (a C-type lectin, also called L-SIGN) for SARS-CoV, DPP4 for MERS-CoV [40]. Moreover the entry mechanism is also dependent on

Emerging Stem Cell Based Therapy for nCOVID-19
Recent advances in molecular biology and regenerative medicine has promised induction of the activity of micro-molecules such as stem cells, bioactive materials and growth factors promoting improved quality, structure, function and bio-mechanical strength of tissue regeneration and healing process. The cellular therapy induces cellular proliferation, chondrogenesis, angiogenesis and rejuvenation of degenerated tissue to attain micromolecular homeostasis. Open Journal of Regenerative Medicine The art of cellular therapy has revolutionized the field of regenerative medicine towards natural healing cascade. Among cellular therapy, mesenchymal stem cells (MSCs) has homing ability, immunoregulatory nature, anti-inflammatory effects, and multilineage differentiation potential as reported by Ullah et al. [43] and Ramesh R et al. [44] (as shown in Figure 3).
MSCs could act upon two ways in the nCOVID19 treatment, namely immunomodulatory effects and differentiation ability. In nCOVID19 cases, acute respiratory distress is the result of SARS-CoV-2 infecting alveolar type II cells (AT2) and the capillary endothelium as mentioned by Hamming et al. [45]. The   weeks after treatment in terms of haematological and radiological improvement along with 28-days mortality of the patient [50]. The end results of this trial are yet to be published.

Discussion
Mesenchymal stem cells (MSCs) have self-renewal and multi-differential abilities. They are readily accessible and expandable in-vitro with exceptional genomic stability and few ethical issues, marking its importance in cellular therapy, regenerative medicine and tissue rejuvenation and repairment [51]. Tsai MS et al. reported immunomodulatory effects and differentiation abilities of mesenchymal stem cells [52]. The immunomodulatory effects of MSCs are triggered by the activation of toll like receptor in MSCs [53] [54]. Few pilot studies by Leng Z et al. [38] and Liang B et al. [55], using MSCs have produced promising outcome for patients reported positive for SARS-CoV-2 nucleic acid. By utilizing its reparative and immunomodulatory characteristics rightly, it can be a reliable therapeutic strategy, notably in the nCOVID-19 pandemic.
MSCs exert antimicrobial effect indirectly through coordination of the proand anti-inflammatory elements of the immune system or by increasing the phagocytotic activity in the microenvironment; and directly by the secretion of antimicrobial peptides & proteins (AMPs), and by the expression of molecules such as indoleamine 2,3-dioxygenase (IDO) and interleukin (IL)-17 [56].
AMPs-mediated cell killing occur by disrupting membrane integrity, by inhibiting protein, DNA or RNA synthesis, and by interacting with certain intracellular targets. Thus, MSC helps in curbing the microbial agent [57] [58].
Mesenchymal Stem Cell (MSC) therapy inhibit the exaggerated response of the immune system and promotes, regenerates and rejuvenates the microenvironment. After entering the systemic circulation, MSCs invade the lung parenchyma and exerts its action by improving the pulmonary micro alveolar structure and pulmonary compliance which further prevents pulmonary fibrosis [59].
After administration of IV MSCs, due to its immunosuppressive potential, there

Conclusion
To summarise, this review focuses on the MSCs as a novel therapeutic option in managing nCOVID-19 infected patients with severe pneumonia considering its safety and efficacy. The novel cellular therapy against severe novel coronavirus infection (nCOVID-19) renders autologous or allogenic MSCs as a therapeutic option to regulate inflammatory response, maintain functional alveoli microenvironment, promote endogenous regeneration and repair, and natural resistance against it with no or minimal side-effects (as shown in Figure 2). To emphasize, MSCs through its anti-inflammatory and immunomodulatory potential inhibit